p2x7  (Alomone Labs)


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    Structured Review

    Alomone Labs p2x7
    Motor neurons in primary culture express <t>P2X7</t>
    P2x7, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/p2x7/product/Alomone Labs
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    p2x7 - by Bioz Stars, 2022-05
    95/100 stars

    Images

    1) Product Images from "P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway"

    Article Title: P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway

    Journal: Journal of neurochemistry

    doi: 10.1111/jnc.12286

    Motor neurons in primary culture express P2X7
    Figure Legend Snippet: Motor neurons in primary culture express P2X7

    Techniques Used:

    2) Product Images from "Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling"

    Article Title: Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

    Journal: Cardiovascular Research

    doi: 10.1093/cvr/cvx213

    P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Activity Assay

    Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Expressing, Western Blot, Immunostaining, Mouse Assay, Staining, Fluorescence, Marker

    P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.

    Techniques Used: Expressing, In Vivo, Immunostaining, Mouse Assay, Staining

    Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =
    Figure Legend Snippet: Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =

    Techniques Used: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    3) Product Images from "MicroRNA-22 Controls Aberrant Neurogenesis and Changes in Neuronal Morphology After Status Epilepticus"

    Article Title: MicroRNA-22 Controls Aberrant Neurogenesis and Changes in Neuronal Morphology After Status Epilepticus

    Journal: Frontiers in Molecular Neuroscience

    doi: 10.3389/fnmol.2018.00442

    Increased miR-22 levels in the contralateral hippocampus during epilepsy in mice. (A) Schematic showing intraamygdala injection of kainic acid (KA) into the ipsilateral amygdala to induce status epilepticus (SE). (B) Graph showing increased levels of miR-22 in the contralateral dentate gyrus (DG) 2 weeks following SE ( n = 4 per group, Student’s t -test: * t = 3.483, p = 0.0131). (C) In situ hybridization showing miR-22 in the DG of the contralateral hippocampus in non-epileptic control vehicle-injected mice and in mice subjected to SE. Of note, miR-22 levels seem to be mainly increased in the subgranular zone in the DG of the contralateral hippocampus during epilepsy (6 weeks following SE). Scale bar = 50 μm. (D) MiR-22 expression in the hippocampus in a patient suffering from temporal lobe epilepsy (TLE). MiR-22-positive cells were mainly localized to the DG (arrows and magnification). snRNA U6 was used as positive control. Scale bar = 1 mm. (E) Graph showing decreased miR-22 levels in the hippocampus ( n = 4 per group Student’s t -test: ** t = 4.955, p = 0.0026). (F) In situ hybridization showing decreased miR-22 levels in the subgranular zone of the DG 6 weeks following intra-cerebro-ventricular (i.c.v.) injection of Ant22. Scale bar = 50 μm. (G) Decreased P2X7 receptor expression in the contralateral hippocampus of epileptic mice 14 days following SE when compared to control mice ( n = 4 per group, Students t -test: * t = 2.734, p = 0.034). (H) Lower BzATP-provoked currents detected by patch clamp from green fluorescent protein (GFP)-positive cells present in the contralateral DG of the hippocampus of epileptic P2rx7 -GFP expressing mice when compared to GFP-positive cells from the ipsilateral DG 14 days following SE ( n = 19 cells for ipsi- and 17 cells from contralateral hippocampus from nine mice per group, Student’s t -test, * t = 2.673, p = 0.0115). Scale bar = 50 μm. (I) Increased P2X7 receptor expression mainly localized to the subgranular zone of the DG of the contralateral hippocampus (arrows) in mice treated with Ant22 6 weeks following SE when compared to scramble-treated epileptic mice. n = 5, Scale bar = 50 μm.
    Figure Legend Snippet: Increased miR-22 levels in the contralateral hippocampus during epilepsy in mice. (A) Schematic showing intraamygdala injection of kainic acid (KA) into the ipsilateral amygdala to induce status epilepticus (SE). (B) Graph showing increased levels of miR-22 in the contralateral dentate gyrus (DG) 2 weeks following SE ( n = 4 per group, Student’s t -test: * t = 3.483, p = 0.0131). (C) In situ hybridization showing miR-22 in the DG of the contralateral hippocampus in non-epileptic control vehicle-injected mice and in mice subjected to SE. Of note, miR-22 levels seem to be mainly increased in the subgranular zone in the DG of the contralateral hippocampus during epilepsy (6 weeks following SE). Scale bar = 50 μm. (D) MiR-22 expression in the hippocampus in a patient suffering from temporal lobe epilepsy (TLE). MiR-22-positive cells were mainly localized to the DG (arrows and magnification). snRNA U6 was used as positive control. Scale bar = 1 mm. (E) Graph showing decreased miR-22 levels in the hippocampus ( n = 4 per group Student’s t -test: ** t = 4.955, p = 0.0026). (F) In situ hybridization showing decreased miR-22 levels in the subgranular zone of the DG 6 weeks following intra-cerebro-ventricular (i.c.v.) injection of Ant22. Scale bar = 50 μm. (G) Decreased P2X7 receptor expression in the contralateral hippocampus of epileptic mice 14 days following SE when compared to control mice ( n = 4 per group, Students t -test: * t = 2.734, p = 0.034). (H) Lower BzATP-provoked currents detected by patch clamp from green fluorescent protein (GFP)-positive cells present in the contralateral DG of the hippocampus of epileptic P2rx7 -GFP expressing mice when compared to GFP-positive cells from the ipsilateral DG 14 days following SE ( n = 19 cells for ipsi- and 17 cells from contralateral hippocampus from nine mice per group, Student’s t -test, * t = 2.673, p = 0.0115). Scale bar = 50 μm. (I) Increased P2X7 receptor expression mainly localized to the subgranular zone of the DG of the contralateral hippocampus (arrows) in mice treated with Ant22 6 weeks following SE when compared to scramble-treated epileptic mice. n = 5, Scale bar = 50 μm.

    Techniques Used: Mouse Assay, Injection, In Situ Hybridization, Expressing, Positive Control, Patch Clamp

    4) Product Images from "Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling"

    Article Title: Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

    Journal: Cardiovascular Research

    doi: 10.1093/cvr/cvx213

    P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Activity Assay

    Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Expressing, Western Blot, Immunostaining, Mouse Assay, Staining, Fluorescence, Marker

    P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.

    Techniques Used: Expressing, In Vivo, Immunostaining, Mouse Assay, Staining

    Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =
    Figure Legend Snippet: Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =

    Techniques Used: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    5) Product Images from "Staphylococcus aureus Activates the NLRP3 Inflammasome in Human and Rat Conjunctival Goblet Cells"

    Article Title: Staphylococcus aureus Activates the NLRP3 Inflammasome in Human and Rat Conjunctival Goblet Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0074010

    Purinergic receptors P2X4, P2X7, and TLR2 are expressed in the rat conjunctiva and rat goblet cell cultures. All three receptors were identified by red immunofluorescent staining. UEA stains goblet cell secretory products green, allowing the identification of goblet cells within the conjunctiva. Rabbit isotype controls were negative. Arrows indicate location of goblet cells.
    Figure Legend Snippet: Purinergic receptors P2X4, P2X7, and TLR2 are expressed in the rat conjunctiva and rat goblet cell cultures. All three receptors were identified by red immunofluorescent staining. UEA stains goblet cell secretory products green, allowing the identification of goblet cells within the conjunctiva. Rabbit isotype controls were negative. Arrows indicate location of goblet cells.

    Techniques Used: Staining

    The purinergic receptors P2X4, P2X7, and TLR2 are functional in cultured rat goblet cells. Rat goblet cells were loaded with fura-2 and then stimulated with ATP and intracellular calcium response measured. A typical trace from one experiment with ATP (5 mM, A ) or lipoteichoic acid (LTA, 10 µg/ml, B ) is representative of 3 animals. Representative photographs of a single field of cells after treatment with ATP or LTA, with warmer colors indicating intracellular Ca 2+ increase, are shown as insets. Peak [Ca 2+ ] i calculated for ATP (0.1 µM–5 mM) from 3 experiments, is shown in C . Peak [Ca 2+ ] i for goblet cells, also preincubated with LTA for 5 h and then loaded with fura-2 for 1 additional h before addition of ATP (5 mM), calculated from 3 experiments, is shown in D . Results are expressed as mean ± SEM. * indicates significance of p
    Figure Legend Snippet: The purinergic receptors P2X4, P2X7, and TLR2 are functional in cultured rat goblet cells. Rat goblet cells were loaded with fura-2 and then stimulated with ATP and intracellular calcium response measured. A typical trace from one experiment with ATP (5 mM, A ) or lipoteichoic acid (LTA, 10 µg/ml, B ) is representative of 3 animals. Representative photographs of a single field of cells after treatment with ATP or LTA, with warmer colors indicating intracellular Ca 2+ increase, are shown as insets. Peak [Ca 2+ ] i calculated for ATP (0.1 µM–5 mM) from 3 experiments, is shown in C . Peak [Ca 2+ ] i for goblet cells, also preincubated with LTA for 5 h and then loaded with fura-2 for 1 additional h before addition of ATP (5 mM), calculated from 3 experiments, is shown in D . Results are expressed as mean ± SEM. * indicates significance of p

    Techniques Used: Functional Assay, Cell Culture

    Purinergic receptors P2X4, P2X7 and TLR2 are expressed in rat goblet cell cultures. All three receptors were identified by western blot. Lanes 1–3 represent separate animals. B is the positive control of rat brain. The β-actin blot is for both P2X4 and P2X7 receptors blots.
    Figure Legend Snippet: Purinergic receptors P2X4, P2X7 and TLR2 are expressed in rat goblet cell cultures. All three receptors were identified by western blot. Lanes 1–3 represent separate animals. B is the positive control of rat brain. The β-actin blot is for both P2X4 and P2X7 receptors blots.

    Techniques Used: Western Blot, Positive Control

    6) Product Images from "P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway"

    Article Title: P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway

    Journal: Journal of neurochemistry

    doi: 10.1111/jnc.12286

    Motor neurons in primary culture express P2X7
    Figure Legend Snippet: Motor neurons in primary culture express P2X7

    Techniques Used:

    7) Product Images from "P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway"

    Article Title: P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway

    Journal: Journal of neurochemistry

    doi: 10.1111/jnc.12286

    Motor neurons in primary culture express P2X7
    Figure Legend Snippet: Motor neurons in primary culture express P2X7

    Techniques Used:

    8) Product Images from "Staphylococcus aureus Activates the NLRP3 Inflammasome in Human and Rat Conjunctival Goblet Cells"

    Article Title: Staphylococcus aureus Activates the NLRP3 Inflammasome in Human and Rat Conjunctival Goblet Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0074010

    Purinergic receptors P2X4, P2X7, and TLR2 are expressed in the rat conjunctiva and rat goblet cell cultures. All three receptors were identified by red immunofluorescent staining. UEA stains goblet cell secretory products green, allowing the identification of goblet cells within the conjunctiva. Rabbit isotype controls were negative. Arrows indicate location of goblet cells.
    Figure Legend Snippet: Purinergic receptors P2X4, P2X7, and TLR2 are expressed in the rat conjunctiva and rat goblet cell cultures. All three receptors were identified by red immunofluorescent staining. UEA stains goblet cell secretory products green, allowing the identification of goblet cells within the conjunctiva. Rabbit isotype controls were negative. Arrows indicate location of goblet cells.

    Techniques Used: Staining

    The purinergic receptors P2X4, P2X7, and TLR2 are functional in cultured rat goblet cells. Rat goblet cells were loaded with fura-2 and then stimulated with ATP and intracellular calcium response measured. A typical trace from one experiment with ATP (5 mM, A ) or lipoteichoic acid (LTA, 10 µg/ml, B ) is representative of 3 animals. Representative photographs of a single field of cells after treatment with ATP or LTA, with warmer colors indicating intracellular Ca 2+ increase, are shown as insets. Peak [Ca 2+ ] i calculated for ATP (0.1 µM–5 mM) from 3 experiments, is shown in C . Peak [Ca 2+ ] i for goblet cells, also preincubated with LTA for 5 h and then loaded with fura-2 for 1 additional h before addition of ATP (5 mM), calculated from 3 experiments, is shown in D . Results are expressed as mean ± SEM. * indicates significance of p
    Figure Legend Snippet: The purinergic receptors P2X4, P2X7, and TLR2 are functional in cultured rat goblet cells. Rat goblet cells were loaded with fura-2 and then stimulated with ATP and intracellular calcium response measured. A typical trace from one experiment with ATP (5 mM, A ) or lipoteichoic acid (LTA, 10 µg/ml, B ) is representative of 3 animals. Representative photographs of a single field of cells after treatment with ATP or LTA, with warmer colors indicating intracellular Ca 2+ increase, are shown as insets. Peak [Ca 2+ ] i calculated for ATP (0.1 µM–5 mM) from 3 experiments, is shown in C . Peak [Ca 2+ ] i for goblet cells, also preincubated with LTA for 5 h and then loaded with fura-2 for 1 additional h before addition of ATP (5 mM), calculated from 3 experiments, is shown in D . Results are expressed as mean ± SEM. * indicates significance of p

    Techniques Used: Functional Assay, Cell Culture

    Purinergic receptors P2X4, P2X7 and TLR2 are expressed in rat goblet cell cultures. All three receptors were identified by western blot. Lanes 1–3 represent separate animals. B is the positive control of rat brain. The β-actin blot is for both P2X4 and P2X7 receptors blots.
    Figure Legend Snippet: Purinergic receptors P2X4, P2X7 and TLR2 are expressed in rat goblet cell cultures. All three receptors were identified by western blot. Lanes 1–3 represent separate animals. B is the positive control of rat brain. The β-actin blot is for both P2X4 and P2X7 receptors blots.

    Techniques Used: Western Blot, Positive Control

    9) Product Images from "Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling"

    Article Title: Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

    Journal: Cardiovascular Research

    doi: 10.1093/cvr/cvx213

    P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Activity Assay

    Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Expressing, Western Blot, Immunostaining, Mouse Assay, Staining, Fluorescence, Marker

    P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.

    Techniques Used: Expressing, In Vivo, Immunostaining, Mouse Assay, Staining

    Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =
    Figure Legend Snippet: Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =

    Techniques Used: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    10) Product Images from "Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling"

    Article Title: Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

    Journal: Cardiovascular Research

    doi: 10.1093/cvr/cvx213

    P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Activity Assay

    Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Expressing, Western Blot, Immunostaining, Mouse Assay, Staining, Fluorescence, Marker

    P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.

    Techniques Used: Expressing, In Vivo, Immunostaining, Mouse Assay, Staining

    Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =
    Figure Legend Snippet: Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =

    Techniques Used: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    11) Product Images from "MicroRNA-22 Controls Aberrant Neurogenesis and Changes in Neuronal Morphology After Status Epilepticus"

    Article Title: MicroRNA-22 Controls Aberrant Neurogenesis and Changes in Neuronal Morphology After Status Epilepticus

    Journal: Frontiers in Molecular Neuroscience

    doi: 10.3389/fnmol.2018.00442

    Increased miR-22 levels in the contralateral hippocampus during epilepsy in mice. (A) Schematic showing intraamygdala injection of kainic acid (KA) into the ipsilateral amygdala to induce status epilepticus (SE). (B) Graph showing increased levels of miR-22 in the contralateral dentate gyrus (DG) 2 weeks following SE ( n = 4 per group, Student’s t -test: * t = 3.483, p = 0.0131). (C) In situ hybridization showing miR-22 in the DG of the contralateral hippocampus in non-epileptic control vehicle-injected mice and in mice subjected to SE. Of note, miR-22 levels seem to be mainly increased in the subgranular zone in the DG of the contralateral hippocampus during epilepsy (6 weeks following SE). Scale bar = 50 μm. (D) MiR-22 expression in the hippocampus in a patient suffering from temporal lobe epilepsy (TLE). MiR-22-positive cells were mainly localized to the DG (arrows and magnification). snRNA U6 was used as positive control. Scale bar = 1 mm. (E) Graph showing decreased miR-22 levels in the hippocampus ( n = 4 per group Student’s t -test: ** t = 4.955, p = 0.0026). (F) In situ hybridization showing decreased miR-22 levels in the subgranular zone of the DG 6 weeks following intra-cerebro-ventricular (i.c.v.) injection of Ant22. Scale bar = 50 μm. (G) Decreased P2X7 receptor expression in the contralateral hippocampus of epileptic mice 14 days following SE when compared to control mice ( n = 4 per group, Students t -test: * t = 2.734, p = 0.034). (H) Lower BzATP-provoked currents detected by patch clamp from green fluorescent protein (GFP)-positive cells present in the contralateral DG of the hippocampus of epileptic P2rx7 -GFP expressing mice when compared to GFP-positive cells from the ipsilateral DG 14 days following SE ( n = 19 cells for ipsi- and 17 cells from contralateral hippocampus from nine mice per group, Student’s t -test, * t = 2.673, p = 0.0115). Scale bar = 50 μm. (I) Increased P2X7 receptor expression mainly localized to the subgranular zone of the DG of the contralateral hippocampus (arrows) in mice treated with Ant22 6 weeks following SE when compared to scramble-treated epileptic mice. n = 5, Scale bar = 50 μm.
    Figure Legend Snippet: Increased miR-22 levels in the contralateral hippocampus during epilepsy in mice. (A) Schematic showing intraamygdala injection of kainic acid (KA) into the ipsilateral amygdala to induce status epilepticus (SE). (B) Graph showing increased levels of miR-22 in the contralateral dentate gyrus (DG) 2 weeks following SE ( n = 4 per group, Student’s t -test: * t = 3.483, p = 0.0131). (C) In situ hybridization showing miR-22 in the DG of the contralateral hippocampus in non-epileptic control vehicle-injected mice and in mice subjected to SE. Of note, miR-22 levels seem to be mainly increased in the subgranular zone in the DG of the contralateral hippocampus during epilepsy (6 weeks following SE). Scale bar = 50 μm. (D) MiR-22 expression in the hippocampus in a patient suffering from temporal lobe epilepsy (TLE). MiR-22-positive cells were mainly localized to the DG (arrows and magnification). snRNA U6 was used as positive control. Scale bar = 1 mm. (E) Graph showing decreased miR-22 levels in the hippocampus ( n = 4 per group Student’s t -test: ** t = 4.955, p = 0.0026). (F) In situ hybridization showing decreased miR-22 levels in the subgranular zone of the DG 6 weeks following intra-cerebro-ventricular (i.c.v.) injection of Ant22. Scale bar = 50 μm. (G) Decreased P2X7 receptor expression in the contralateral hippocampus of epileptic mice 14 days following SE when compared to control mice ( n = 4 per group, Students t -test: * t = 2.734, p = 0.034). (H) Lower BzATP-provoked currents detected by patch clamp from green fluorescent protein (GFP)-positive cells present in the contralateral DG of the hippocampus of epileptic P2rx7 -GFP expressing mice when compared to GFP-positive cells from the ipsilateral DG 14 days following SE ( n = 19 cells for ipsi- and 17 cells from contralateral hippocampus from nine mice per group, Student’s t -test, * t = 2.673, p = 0.0115). Scale bar = 50 μm. (I) Increased P2X7 receptor expression mainly localized to the subgranular zone of the DG of the contralateral hippocampus (arrows) in mice treated with Ant22 6 weeks following SE when compared to scramble-treated epileptic mice. n = 5, Scale bar = 50 μm.

    Techniques Used: Mouse Assay, Injection, In Situ Hybridization, Expressing, Positive Control, Patch Clamp

    12) Product Images from "Synthetic cathinone MDPV enhances reward function through purinergic P2X7 receptor-dependent pathway and increases P2X7 gene expression in nucleus accumbens"

    Article Title: Synthetic cathinone MDPV enhances reward function through purinergic P2X7 receptor-dependent pathway and increases P2X7 gene expression in nucleus accumbens

    Journal: Drug and alcohol dependence

    doi: 10.1016/j.drugalcdep.2018.12.022

    P2X7 antagonist A438079 inhibits reduction in brain reward threshold produced by MDPV. Percentage change in baseline reward threshold + S.E.M. shown in (A, C, E) and maximum response rate shown in (B, D, F). (A-B) MDPV (0.5 mg/kg) reduces brain reward threshold without affecting maximal response rate. N=4-7. ** p
    Figure Legend Snippet: P2X7 antagonist A438079 inhibits reduction in brain reward threshold produced by MDPV. Percentage change in baseline reward threshold + S.E.M. shown in (A, C, E) and maximum response rate shown in (B, D, F). (A-B) MDPV (0.5 mg/kg) reduces brain reward threshold without affecting maximal response rate. N=4-7. ** p

    Techniques Used: Produced

    13) Product Images from "P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway"

    Article Title: P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway

    Journal: Journal of neurochemistry

    doi: 10.1111/jnc.12286

    Motor neurons in primary culture express P2X7
    Figure Legend Snippet: Motor neurons in primary culture express P2X7

    Techniques Used:

    14) Product Images from "Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling"

    Article Title: Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

    Journal: Cardiovascular Research

    doi: 10.1093/cvr/cvx213

    P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Activity Assay

    Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Expressing, Western Blot, Immunostaining, Mouse Assay, Staining, Fluorescence, Marker

    P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.

    Techniques Used: Expressing, In Vivo, Immunostaining, Mouse Assay, Staining

    Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =
    Figure Legend Snippet: Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =

    Techniques Used: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    15) Product Images from "MicroRNA-22 Controls Aberrant Neurogenesis and Changes in Neuronal Morphology After Status Epilepticus"

    Article Title: MicroRNA-22 Controls Aberrant Neurogenesis and Changes in Neuronal Morphology After Status Epilepticus

    Journal: Frontiers in Molecular Neuroscience

    doi: 10.3389/fnmol.2018.00442

    Increased miR-22 levels in the contralateral hippocampus during epilepsy in mice. (A) Schematic showing intraamygdala injection of kainic acid (KA) into the ipsilateral amygdala to induce status epilepticus (SE). (B) Graph showing increased levels of miR-22 in the contralateral dentate gyrus (DG) 2 weeks following SE ( n = 4 per group, Student’s t -test: * t = 3.483, p = 0.0131). (C) In situ hybridization showing miR-22 in the DG of the contralateral hippocampus in non-epileptic control vehicle-injected mice and in mice subjected to SE. Of note, miR-22 levels seem to be mainly increased in the subgranular zone in the DG of the contralateral hippocampus during epilepsy (6 weeks following SE). Scale bar = 50 μm. (D) MiR-22 expression in the hippocampus in a patient suffering from temporal lobe epilepsy (TLE). MiR-22-positive cells were mainly localized to the DG (arrows and magnification). snRNA U6 was used as positive control. Scale bar = 1 mm. (E) Graph showing decreased miR-22 levels in the hippocampus ( n = 4 per group Student’s t -test: ** t = 4.955, p = 0.0026). (F) In situ hybridization showing decreased miR-22 levels in the subgranular zone of the DG 6 weeks following intra-cerebro-ventricular (i.c.v.) injection of Ant22. Scale bar = 50 μm. (G) Decreased P2X7 receptor expression in the contralateral hippocampus of epileptic mice 14 days following SE when compared to control mice ( n = 4 per group, Students t -test: * t = 2.734, p = 0.034). (H) Lower BzATP-provoked currents detected by patch clamp from green fluorescent protein (GFP)-positive cells present in the contralateral DG of the hippocampus of epileptic P2rx7 -GFP expressing mice when compared to GFP-positive cells from the ipsilateral DG 14 days following SE ( n = 19 cells for ipsi- and 17 cells from contralateral hippocampus from nine mice per group, Student’s t -test, * t = 2.673, p = 0.0115). Scale bar = 50 μm. (I) Increased P2X7 receptor expression mainly localized to the subgranular zone of the DG of the contralateral hippocampus (arrows) in mice treated with Ant22 6 weeks following SE when compared to scramble-treated epileptic mice. n = 5, Scale bar = 50 μm.
    Figure Legend Snippet: Increased miR-22 levels in the contralateral hippocampus during epilepsy in mice. (A) Schematic showing intraamygdala injection of kainic acid (KA) into the ipsilateral amygdala to induce status epilepticus (SE). (B) Graph showing increased levels of miR-22 in the contralateral dentate gyrus (DG) 2 weeks following SE ( n = 4 per group, Student’s t -test: * t = 3.483, p = 0.0131). (C) In situ hybridization showing miR-22 in the DG of the contralateral hippocampus in non-epileptic control vehicle-injected mice and in mice subjected to SE. Of note, miR-22 levels seem to be mainly increased in the subgranular zone in the DG of the contralateral hippocampus during epilepsy (6 weeks following SE). Scale bar = 50 μm. (D) MiR-22 expression in the hippocampus in a patient suffering from temporal lobe epilepsy (TLE). MiR-22-positive cells were mainly localized to the DG (arrows and magnification). snRNA U6 was used as positive control. Scale bar = 1 mm. (E) Graph showing decreased miR-22 levels in the hippocampus ( n = 4 per group Student’s t -test: ** t = 4.955, p = 0.0026). (F) In situ hybridization showing decreased miR-22 levels in the subgranular zone of the DG 6 weeks following intra-cerebro-ventricular (i.c.v.) injection of Ant22. Scale bar = 50 μm. (G) Decreased P2X7 receptor expression in the contralateral hippocampus of epileptic mice 14 days following SE when compared to control mice ( n = 4 per group, Students t -test: * t = 2.734, p = 0.034). (H) Lower BzATP-provoked currents detected by patch clamp from green fluorescent protein (GFP)-positive cells present in the contralateral DG of the hippocampus of epileptic P2rx7 -GFP expressing mice when compared to GFP-positive cells from the ipsilateral DG 14 days following SE ( n = 19 cells for ipsi- and 17 cells from contralateral hippocampus from nine mice per group, Student’s t -test, * t = 2.673, p = 0.0115). Scale bar = 50 μm. (I) Increased P2X7 receptor expression mainly localized to the subgranular zone of the DG of the contralateral hippocampus (arrows) in mice treated with Ant22 6 weeks following SE when compared to scramble-treated epileptic mice. n = 5, Scale bar = 50 μm.

    Techniques Used: Mouse Assay, Injection, In Situ Hybridization, Expressing, Positive Control, Patch Clamp

    16) Product Images from "Staphylococcus aureus Activates the NLRP3 Inflammasome in Human and Rat Conjunctival Goblet Cells"

    Article Title: Staphylococcus aureus Activates the NLRP3 Inflammasome in Human and Rat Conjunctival Goblet Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0074010

    Purinergic receptors P2X4, P2X7, and TLR2 are expressed in the rat conjunctiva and rat goblet cell cultures. All three receptors were identified by red immunofluorescent staining. UEA stains goblet cell secretory products green, allowing the identification of goblet cells within the conjunctiva. Rabbit isotype controls were negative. Arrows indicate location of goblet cells.
    Figure Legend Snippet: Purinergic receptors P2X4, P2X7, and TLR2 are expressed in the rat conjunctiva and rat goblet cell cultures. All three receptors were identified by red immunofluorescent staining. UEA stains goblet cell secretory products green, allowing the identification of goblet cells within the conjunctiva. Rabbit isotype controls were negative. Arrows indicate location of goblet cells.

    Techniques Used: Staining

    The purinergic receptors P2X4, P2X7, and TLR2 are functional in cultured rat goblet cells. Rat goblet cells were loaded with fura-2 and then stimulated with ATP and intracellular calcium response measured. A typical trace from one experiment with ATP (5 mM, A ) or lipoteichoic acid (LTA, 10 µg/ml, B ) is representative of 3 animals. Representative photographs of a single field of cells after treatment with ATP or LTA, with warmer colors indicating intracellular Ca 2+ increase, are shown as insets. Peak [Ca 2+ ] i calculated for ATP (0.1 µM–5 mM) from 3 experiments, is shown in C . Peak [Ca 2+ ] i for goblet cells, also preincubated with LTA for 5 h and then loaded with fura-2 for 1 additional h before addition of ATP (5 mM), calculated from 3 experiments, is shown in D . Results are expressed as mean ± SEM. * indicates significance of p
    Figure Legend Snippet: The purinergic receptors P2X4, P2X7, and TLR2 are functional in cultured rat goblet cells. Rat goblet cells were loaded with fura-2 and then stimulated with ATP and intracellular calcium response measured. A typical trace from one experiment with ATP (5 mM, A ) or lipoteichoic acid (LTA, 10 µg/ml, B ) is representative of 3 animals. Representative photographs of a single field of cells after treatment with ATP or LTA, with warmer colors indicating intracellular Ca 2+ increase, are shown as insets. Peak [Ca 2+ ] i calculated for ATP (0.1 µM–5 mM) from 3 experiments, is shown in C . Peak [Ca 2+ ] i for goblet cells, also preincubated with LTA for 5 h and then loaded with fura-2 for 1 additional h before addition of ATP (5 mM), calculated from 3 experiments, is shown in D . Results are expressed as mean ± SEM. * indicates significance of p

    Techniques Used: Functional Assay, Cell Culture

    Purinergic receptors P2X4, P2X7 and TLR2 are expressed in rat goblet cell cultures. All three receptors were identified by western blot. Lanes 1–3 represent separate animals. B is the positive control of rat brain. The β-actin blot is for both P2X4 and P2X7 receptors blots.
    Figure Legend Snippet: Purinergic receptors P2X4, P2X7 and TLR2 are expressed in rat goblet cell cultures. All three receptors were identified by western blot. Lanes 1–3 represent separate animals. B is the positive control of rat brain. The β-actin blot is for both P2X4 and P2X7 receptors blots.

    Techniques Used: Western Blot, Positive Control

    17) Product Images from "P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway"

    Article Title: P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway

    Journal: Journal of neurochemistry

    doi: 10.1111/jnc.12286

    Motor neurons in primary culture express P2X7
    Figure Legend Snippet: Motor neurons in primary culture express P2X7

    Techniques Used:

    18) Product Images from "P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway"

    Article Title: P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway

    Journal: Journal of neurochemistry

    doi: 10.1111/jnc.12286

    Motor neurons in primary culture express P2X7
    Figure Legend Snippet: Motor neurons in primary culture express P2X7

    Techniques Used:

    19) Product Images from "P2X receptor overexpression induced by soluble oligomers of amyloid beta peptide potentiates synaptic failure and neuronal dyshomeostasis in cellular models of Alzheimer’s disease"

    Article Title: P2X receptor overexpression induced by soluble oligomers of amyloid beta peptide potentiates synaptic failure and neuronal dyshomeostasis in cellular models of Alzheimer’s disease

    Journal: Neuropharmacology

    doi: 10.1016/j.neuropharm.2017.10.027

    SO-Aβ induces changes in the mRNA expression levels of P2X subunits. A) Quantification of relative mRNA for P2X subunits at 12 h of treatment with Aβ (0.5 μM) revealed that the subunits P2X1, 2, 5 and 7 had a significant increase in their expression (P2X1: 1.25 ± 0.007, P = 0.0009; P2X2: 1.23 ± 0.06, P = 0.029; P2X5: 1.21 ± 0.06, P = 0.039; P2X7: 1.42 ± 0.06, P = 0.019; P2X3: 0.95 ± 0.09, P = 0.636; P2X4: 1.21 ± 0.07, P = 0.056; P2X6: 1.03 ± 0.22, P = 0.918). B) Quantification of the relative expression of P2X at 24 h of treatment (Aβ, 0.5 μM) showing that at this time point only P2X2 remained significantly increased (P2X2: 1.2 ± 0.04, P = 0.022; P2X1: 1.1 ± 0.06, P = 0.152; P2X3: 0.9 ± 0.08, P = 0.286; P2X4: 1.16 ± 0.07, P = 0.110; P2X5: 1.13 ± 0.05, P = 0.096; P2X6: 0.95 ± 0.15, P = 0.783; P2X7: 1.21 ± 0.13, P = 0.193), n = 5. *P
    Figure Legend Snippet: SO-Aβ induces changes in the mRNA expression levels of P2X subunits. A) Quantification of relative mRNA for P2X subunits at 12 h of treatment with Aβ (0.5 μM) revealed that the subunits P2X1, 2, 5 and 7 had a significant increase in their expression (P2X1: 1.25 ± 0.007, P = 0.0009; P2X2: 1.23 ± 0.06, P = 0.029; P2X5: 1.21 ± 0.06, P = 0.039; P2X7: 1.42 ± 0.06, P = 0.019; P2X3: 0.95 ± 0.09, P = 0.636; P2X4: 1.21 ± 0.07, P = 0.056; P2X6: 1.03 ± 0.22, P = 0.918). B) Quantification of the relative expression of P2X at 24 h of treatment (Aβ, 0.5 μM) showing that at this time point only P2X2 remained significantly increased (P2X2: 1.2 ± 0.04, P = 0.022; P2X1: 1.1 ± 0.06, P = 0.152; P2X3: 0.9 ± 0.08, P = 0.286; P2X4: 1.16 ± 0.07, P = 0.110; P2X5: 1.13 ± 0.05, P = 0.096; P2X6: 0.95 ± 0.15, P = 0.783; P2X7: 1.21 ± 0.13, P = 0.193), n = 5. *P

    Techniques Used: Expressing

    P2X2R immunocytochemistry of hippocampal neurons treated with SO-Aβ. A-I) Representative image of hippocampal neuron (10 DIV) in control conditions stained with MAP-2 (red) and P2X2 receptor subunit (green, scale bar 20 μm). A-II) Magnification of control neuron with immunostaining for P2X2R (green). A-III) SO-Aβ treated neuron and staining in the same conditions as Aa. A-IV) Magnification of SO-Aβ treated neuron with immunostaining for P2X2R (green). B) Quantification of P2X2R immunoreactivity in SO-Aβ treated neurons with respect to control conditions (n = 4, * vs control condition). C-I) Representative image of a hippocampal neuron (10 DIV) in control conditions stained with MAP-2 (red) and P2X7 receptor subunit (green, scale bar 20 μm). C-II) Magnification of control neuron with immunostaining for P2X7R (green). C-III) SO-Aβ treated neuron and staining in the same conditions as Aa. C-IV) Magnification of SO-Aβ treated neuron with immunostaining for P2X7R (green). D) Quantification of P2X7R immunoreactivity in SO-Aβ treated neurons with respect to control conditions (n = 4).
    Figure Legend Snippet: P2X2R immunocytochemistry of hippocampal neurons treated with SO-Aβ. A-I) Representative image of hippocampal neuron (10 DIV) in control conditions stained with MAP-2 (red) and P2X2 receptor subunit (green, scale bar 20 μm). A-II) Magnification of control neuron with immunostaining for P2X2R (green). A-III) SO-Aβ treated neuron and staining in the same conditions as Aa. A-IV) Magnification of SO-Aβ treated neuron with immunostaining for P2X2R (green). B) Quantification of P2X2R immunoreactivity in SO-Aβ treated neurons with respect to control conditions (n = 4, * vs control condition). C-I) Representative image of a hippocampal neuron (10 DIV) in control conditions stained with MAP-2 (red) and P2X7 receptor subunit (green, scale bar 20 μm). C-II) Magnification of control neuron with immunostaining for P2X7R (green). C-III) SO-Aβ treated neuron and staining in the same conditions as Aa. C-IV) Magnification of SO-Aβ treated neuron with immunostaining for P2X7R (green). D) Quantification of P2X7R immunoreactivity in SO-Aβ treated neurons with respect to control conditions (n = 4).

    Techniques Used: Immunocytochemistry, Staining, Immunostaining

    20) Product Images from "P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway"

    Article Title: P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway

    Journal: Journal of neurochemistry

    doi: 10.1111/jnc.12286

    Motor neurons in primary culture express P2X7
    Figure Legend Snippet: Motor neurons in primary culture express P2X7

    Techniques Used:

    21) Product Images from "P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway"

    Article Title: P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway

    Journal: Journal of neurochemistry

    doi: 10.1111/jnc.12286

    Motor neurons in primary culture express P2X7
    Figure Legend Snippet: Motor neurons in primary culture express P2X7

    Techniques Used:

    22) Product Images from "Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling"

    Article Title: Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

    Journal: Cardiovascular Research

    doi: 10.1093/cvr/cvx213

    P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Activity Assay

    Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Expressing, Western Blot, Immunostaining, Mouse Assay, Staining, Fluorescence, Marker

    P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.

    Techniques Used: Expressing, In Vivo, Immunostaining, Mouse Assay, Staining

    Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =
    Figure Legend Snippet: Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =

    Techniques Used: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    23) Product Images from "Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling"

    Article Title: Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

    Journal: Cardiovascular Research

    doi: 10.1093/cvr/cvx213

    P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Activity Assay

    Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Expressing, Western Blot, Immunostaining, Mouse Assay, Staining, Fluorescence, Marker

    P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.

    Techniques Used: Expressing, In Vivo, Immunostaining, Mouse Assay, Staining

    Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =
    Figure Legend Snippet: Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =

    Techniques Used: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    24) Product Images from "P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway"

    Article Title: P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway

    Journal: Journal of neurochemistry

    doi: 10.1111/jnc.12286

    Motor neurons in primary culture express P2X7
    Figure Legend Snippet: Motor neurons in primary culture express P2X7

    Techniques Used:

    25) Product Images from "P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway"

    Article Title: P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway

    Journal: Journal of neurochemistry

    doi: 10.1111/jnc.12286

    Motor neurons in primary culture express P2X7
    Figure Legend Snippet: Motor neurons in primary culture express P2X7

    Techniques Used:

    26) Product Images from "P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway"

    Article Title: P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway

    Journal: Journal of neurochemistry

    doi: 10.1111/jnc.12286

    Motor neurons in primary culture express P2X7
    Figure Legend Snippet: Motor neurons in primary culture express P2X7

    Techniques Used:

    27) Product Images from "Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling"

    Article Title: Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

    Journal: Cardiovascular Research

    doi: 10.1093/cvr/cvx213

    P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Activity Assay

    Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Expressing, Western Blot, Immunostaining, Mouse Assay, Staining, Fluorescence, Marker

    P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.

    Techniques Used: Expressing, In Vivo, Immunostaining, Mouse Assay, Staining

    Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =
    Figure Legend Snippet: Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =

    Techniques Used: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    28) Product Images from "Staphylococcus aureus Activates the NLRP3 Inflammasome in Human and Rat Conjunctival Goblet Cells"

    Article Title: Staphylococcus aureus Activates the NLRP3 Inflammasome in Human and Rat Conjunctival Goblet Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0074010

    Purinergic receptors P2X4, P2X7, and TLR2 are expressed in the rat conjunctiva and rat goblet cell cultures. All three receptors were identified by red immunofluorescent staining. UEA stains goblet cell secretory products green, allowing the identification of goblet cells within the conjunctiva. Rabbit isotype controls were negative. Arrows indicate location of goblet cells.
    Figure Legend Snippet: Purinergic receptors P2X4, P2X7, and TLR2 are expressed in the rat conjunctiva and rat goblet cell cultures. All three receptors were identified by red immunofluorescent staining. UEA stains goblet cell secretory products green, allowing the identification of goblet cells within the conjunctiva. Rabbit isotype controls were negative. Arrows indicate location of goblet cells.

    Techniques Used: Staining

    The purinergic receptors P2X4, P2X7, and TLR2 are functional in cultured rat goblet cells. Rat goblet cells were loaded with fura-2 and then stimulated with ATP and intracellular calcium response measured. A typical trace from one experiment with ATP (5 mM, A ) or lipoteichoic acid (LTA, 10 µg/ml, B ) is representative of 3 animals. Representative photographs of a single field of cells after treatment with ATP or LTA, with warmer colors indicating intracellular Ca 2+ increase, are shown as insets. Peak [Ca 2+ ] i calculated for ATP (0.1 µM–5 mM) from 3 experiments, is shown in C . Peak [Ca 2+ ] i for goblet cells, also preincubated with LTA for 5 h and then loaded with fura-2 for 1 additional h before addition of ATP (5 mM), calculated from 3 experiments, is shown in D . Results are expressed as mean ± SEM. * indicates significance of p
    Figure Legend Snippet: The purinergic receptors P2X4, P2X7, and TLR2 are functional in cultured rat goblet cells. Rat goblet cells were loaded with fura-2 and then stimulated with ATP and intracellular calcium response measured. A typical trace from one experiment with ATP (5 mM, A ) or lipoteichoic acid (LTA, 10 µg/ml, B ) is representative of 3 animals. Representative photographs of a single field of cells after treatment with ATP or LTA, with warmer colors indicating intracellular Ca 2+ increase, are shown as insets. Peak [Ca 2+ ] i calculated for ATP (0.1 µM–5 mM) from 3 experiments, is shown in C . Peak [Ca 2+ ] i for goblet cells, also preincubated with LTA for 5 h and then loaded with fura-2 for 1 additional h before addition of ATP (5 mM), calculated from 3 experiments, is shown in D . Results are expressed as mean ± SEM. * indicates significance of p

    Techniques Used: Functional Assay, Cell Culture

    Purinergic receptors P2X4, P2X7 and TLR2 are expressed in rat goblet cell cultures. All three receptors were identified by western blot. Lanes 1–3 represent separate animals. B is the positive control of rat brain. The β-actin blot is for both P2X4 and P2X7 receptors blots.
    Figure Legend Snippet: Purinergic receptors P2X4, P2X7 and TLR2 are expressed in rat goblet cell cultures. All three receptors were identified by western blot. Lanes 1–3 represent separate animals. B is the positive control of rat brain. The β-actin blot is for both P2X4 and P2X7 receptors blots.

    Techniques Used: Western Blot, Positive Control

    29) Product Images from "Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling"

    Article Title: Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

    Journal: Cardiovascular Research

    doi: 10.1093/cvr/cvx213

    P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Activity Assay

    Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Expressing, Western Blot, Immunostaining, Mouse Assay, Staining, Fluorescence, Marker

    P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.

    Techniques Used: Expressing, In Vivo, Immunostaining, Mouse Assay, Staining

    Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =
    Figure Legend Snippet: Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =

    Techniques Used: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    30) Product Images from "Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling"

    Article Title: Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

    Journal: Cardiovascular Research

    doi: 10.1093/cvr/cvx213

    P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Activity Assay

    Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Expressing, Western Blot, Immunostaining, Mouse Assay, Staining, Fluorescence, Marker

    P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.

    Techniques Used: Expressing, In Vivo, Immunostaining, Mouse Assay, Staining

    Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =
    Figure Legend Snippet: Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =

    Techniques Used: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    31) Product Images from "Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling"

    Article Title: Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

    Journal: Cardiovascular Research

    doi: 10.1093/cvr/cvx213

    P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Activity Assay

    Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Expressing, Western Blot, Immunostaining, Mouse Assay, Staining, Fluorescence, Marker

    P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.

    Techniques Used: Expressing, In Vivo, Immunostaining, Mouse Assay, Staining

    Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =
    Figure Legend Snippet: Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =

    Techniques Used: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    32) Product Images from "P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway"

    Article Title: P2X7 receptor-induced death of motor neurons by a peroxynitrite/FAS-dependent pathway

    Journal: Journal of neurochemistry

    doi: 10.1111/jnc.12286

    Motor neurons in primary culture express P2X7
    Figure Legend Snippet: Motor neurons in primary culture express P2X7

    Techniques Used:

    33) Product Images from "Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling"

    Article Title: Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

    Journal: Cardiovascular Research

    doi: 10.1093/cvr/cvx213

    P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Activity Assay

    Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Expressing, Western Blot, Immunostaining, Mouse Assay, Staining, Fluorescence, Marker

    P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.

    Techniques Used: Expressing, In Vivo, Immunostaining, Mouse Assay, Staining

    Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =
    Figure Legend Snippet: Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =

    Techniques Used: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    34) Product Images from "Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling"

    Article Title: Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

    Journal: Cardiovascular Research

    doi: 10.1093/cvr/cvx213

    P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 receptor activity occurs exclusively in HUVEC under atheroprone flow. Representative single cell ATP-induced calcium responses in atheroprotected ( A ) or atheroprone ( B, C ) flow conditioned HUVEC after thapsigargin pre-treatment ± the P2X7 inhibitor 10 µM A438079 (P2X7i) or siRNA control and P2X7 knockdown ( C ) and analysed by measuring the average area under the curve ( n = 4, 175 cells per donor, * indicates P ≤ 0.05 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Activity Assay

    Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.
    Figure Legend Snippet: Expression of ATP-gated P2X7 receptors is enhanced under atheroprone flow. ( A ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the ibidi flow system ( n = 6, ** indicates P ≤ 0.01 using a paired t -test). P2X7 is shown by an arrow, validated by P2X7 knockdown, shown in Supplementary material online , Figure 4 B. ( B ) Western blot and densitometry of P2X7 in flow conditioned HUVEC using the orbital shaker system ( n = 4, ** indicates P ≤ 0.01 using a paired t -test). ( C ) Representative en face immunostaining for P2X7 (red) on wildtype C57BL/6 mice at atheroprotective (outer curvature) and atheroprone (inner curvature) regions of the aorta (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). P2X7 expression was analysed by measuring the relative fluorescent intensity at sites protected or prone to atherosclerosis. Surface expression was measured by measuring P2X7 fluorescence at sites co-localized with the endothelial cell surface marker CD31. Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG performed on the descending aorta ( n = 5, * indicates P ≤ 0.05 and *** indicates P ≤ 0.001 using a paired t -test). Values are mean ± S.E.M.

    Techniques Used: Expressing, Western Blot, Immunostaining, Mouse Assay, Staining, Fluorescence, Marker

    P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.
    Figure Legend Snippet: P2X7 regulates E-selectin expression at sites susceptible to atherosclerosis in vivo . Representative en face immunostaining for E-selectin (red) on wildtype or P2X7 −/− BALB/c mice at atheroprotective (outer curvature) and atheroprone (inner curvature) sites of the aorta ( n = 4–5) (scale = 25 µm). Endothelial cells were identified by staining with CD31 (green) and cell nuclei were stained using TO-PRO (blue). E-selectin expression was analysed by measuring the relative fluorescent intensity at sites of protected or prone to atherosclerosis ( n = 4–5, * indicates P ≤ 0.05, ** indicates P ≤ 0.01 and *** indicates P ≤ 0.001 using a two-way ANOVA). Relative fluorescent intensities for P2X7 were corrected against the relative fluorescent intensity of IgG. Values are mean ± S.E.M.

    Techniques Used: Expressing, In Vivo, Immunostaining, Mouse Assay, Staining

    Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =
    Figure Legend Snippet: Atheroprone flow mediated inflammatory signalling is regulated by P2X7 responses. qPCR analysis of E-selectin ( A ) and IL-8 ( B ) in HUVEC conditioned with atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 5, * indicates P ≤ 0.05 using a two-way ANOVA). ( C ) IL-8 release measured by ELISA in the supernatant of HUVEC conditioned under atheroprotective or atheroprone flow using the ibidi system ± the P2X7 inhibitor 10 µM A438079 (P2X7i) ( n = 9, * indicates P ≤ 0.05, *** indicates p =

    Techniques Used: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

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    Alomone Labs rabbit polyclonal anti human p2x7 receptor atto 488
    TLR4 signaling influences <t>P2X7</t> receptor distribution. Representative confocal microphotographs showing (A) HuC/D (magenta) and P2X7Rs (yellow) distribution and (B) analysis of P2X7Rs fluorescence intensities in WT and TLR4 -/- preparations ( N = 5 mice/group). Cell nuclei were stained with TOTO-3 (blue). Bars = 22 μm. ∗ P
    Rabbit Polyclonal Anti Human P2x7 Receptor Atto 488, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Alomone Labs rabbit polyclonal p2x7 c terminus antibody
    The N terminus is important for the sensitivity of P2X7 receptor activity to cholesterol depletion. A , amino acid sequence alignment of the N-terminal region of human P2X1, P2X2, and P2X7 receptors, with conserved amino acids highlighted in  gray , P2X1 aa20–23 and equivalent residues in P2X2  underlined , and the P2X7 residues Lys 17  and Val 18  in  red. B , time course of dye uptake mediated by the K17R/V18M mutant and wild type P2X7, alone, pretreated with MCD and loaded with cholesterol ( chol ), normalized to the untreated WT P2X7.  C , bar graph of the relative rate of dye uptake normalized to the untreated WT P2X7. Dye uptake mediated by the K17R/V18M mutant is not potentiated by MCD, but is inhibited by cholesterol loading (WT:  n  = 14 alone,  n  = 11 MCD,  n  = 3 cholesterol; mutants:  n  = 12 alone,  n  = 11 MCD,  n  = 3 cholesterol; *,  p
    Rabbit Polyclonal P2x7 C Terminus Antibody, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal p2x7 c terminus antibody/product/Alomone Labs
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    96
    Alomone Labs anti p2x7 antibody
    <t>P2X7</t> inhibition reduced the ATP release with simultaneous decrease of some pro-inflammatory cytokine levels in mouse serum and changed the redox metabolism in C6 tumors. a An ATP release is significantly lower in tumors treated with BBG compared to the control tumors ( n = 4). The significance of the differences was determined using Student’s t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. b Decline NOS-2 expression in BBG-treated tumors ( n = 4). c Decreased expression of FOXP3 (T reg lymphocytes) and CD68 (macrophages) markers in BBG-treated tumors evaluated using flow cytometry ( n = 5). The significance of the differences was determined using Student’s t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. d The level of the pro-inflammatory and anti-inflammatory cytokines, in mouse serum and glioma tumors using cytokine array for multiplex protein detection ( n = 3). e P2X7 inhibition caused a decrease of GSSG/GSH ratio with a slight decline of SOD activity in BBG-treated tumors ( n = 4). The significance of the differences was determined using Student’s t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control
    Anti P2x7 Antibody, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TLR4 signaling influences P2X7 receptor distribution. Representative confocal microphotographs showing (A) HuC/D (magenta) and P2X7Rs (yellow) distribution and (B) analysis of P2X7Rs fluorescence intensities in WT and TLR4 -/- preparations ( N = 5 mice/group). Cell nuclei were stained with TOTO-3 (blue). Bars = 22 μm. ∗ P

    Journal: Frontiers in Pharmacology

    Article Title: Toll-Like Receptor 4 Modulates Small Intestine Neuromuscular Function through Nitrergic and Purinergic Pathways

    doi: 10.3389/fphar.2017.00350

    Figure Lengend Snippet: TLR4 signaling influences P2X7 receptor distribution. Representative confocal microphotographs showing (A) HuC/D (magenta) and P2X7Rs (yellow) distribution and (B) analysis of P2X7Rs fluorescence intensities in WT and TLR4 -/- preparations ( N = 5 mice/group). Cell nuclei were stained with TOTO-3 (blue). Bars = 22 μm. ∗ P

    Article Snippet: LMMP whole mount preparations or ileal cryosections were then incubated overnight at room temperature with the following antibodies: chicken polyclonal anti-mouse glial fibrillary acidic protein (GFAP; 1:100; Abcam, Milan, Italy), rabbit polyclonal anti-human GFAP (1:200; Merck Millipore Corporation, Milan, Italy), mouse biotin-conjugated anti-human HuC/D (1:50; Thermo Fisher Scientific, Milan, Italy), rabbit polyclonal anti-mouse neuronal nitric oxide synthase (nNOS, 1:100; Thermo Fisher Scientific, Milan, Italy), rabbit polyclonal anti-human inducible NOS (iNOS; 1:50, Santa Cruz Biotechnology, Milan, Italy), rabbit polyclonal anti-human vasoactive intestinal peptide (VIP, 1:100; GenWay Biotech, Milan, Italy), rabbit polyclonal anti-human P2X7 receptor-ATTO-488 (1:100; Alomone labs, Jerusalem, Israel), rabbit polyclonal anti-human P2Y1 receptor (1:100; Alomone labs, Jerusalem, Israel), rabbit monoclonal anti-human S100β (1:100; Merck Millipore Corporation, Milan, Italy) and guinea pig polyclonal anti-mouse substance P (1:100; Abcam, Milan, Italy).

    Techniques: Fluorescence, Mouse Assay, Staining

    The N terminus is important for the sensitivity of P2X7 receptor activity to cholesterol depletion. A , amino acid sequence alignment of the N-terminal region of human P2X1, P2X2, and P2X7 receptors, with conserved amino acids highlighted in  gray , P2X1 aa20–23 and equivalent residues in P2X2  underlined , and the P2X7 residues Lys 17  and Val 18  in  red. B , time course of dye uptake mediated by the K17R/V18M mutant and wild type P2X7, alone, pretreated with MCD and loaded with cholesterol ( chol ), normalized to the untreated WT P2X7.  C , bar graph of the relative rate of dye uptake normalized to the untreated WT P2X7. Dye uptake mediated by the K17R/V18M mutant is not potentiated by MCD, but is inhibited by cholesterol loading (WT:  n  = 14 alone,  n  = 11 MCD,  n  = 3 cholesterol; mutants:  n  = 12 alone,  n  = 11 MCD,  n  = 3 cholesterol; *,  p

    Journal: The Journal of Biological Chemistry

    Article Title: Plasma Membrane Cholesterol as a Regulator of Human and Rodent P2X7 Receptor Activation and Sensitization *

    doi: 10.1074/jbc.M114.574699

    Figure Lengend Snippet: The N terminus is important for the sensitivity of P2X7 receptor activity to cholesterol depletion. A , amino acid sequence alignment of the N-terminal region of human P2X1, P2X2, and P2X7 receptors, with conserved amino acids highlighted in gray , P2X1 aa20–23 and equivalent residues in P2X2 underlined , and the P2X7 residues Lys 17 and Val 18 in red. B , time course of dye uptake mediated by the K17R/V18M mutant and wild type P2X7, alone, pretreated with MCD and loaded with cholesterol ( chol ), normalized to the untreated WT P2X7. C , bar graph of the relative rate of dye uptake normalized to the untreated WT P2X7. Dye uptake mediated by the K17R/V18M mutant is not potentiated by MCD, but is inhibited by cholesterol loading (WT: n = 14 alone, n = 11 MCD, n = 3 cholesterol; mutants: n = 12 alone, n = 11 MCD, n = 3 cholesterol; *, p

    Article Snippet: Rabbit polyclonal P2X7 C terminus antibody was obtained from Alomone Labs (Jerusalem, Israel), and anti-mouse and anti-rabbit HRP-conjugated secondary antibodies were from Thermo Fisher Scientific and Bio-Rad, respectively.

    Techniques: Activity Assay, Sequencing, Mutagenesis, Relative Rate

    P2X7 inhibition reduced the ATP release with simultaneous decrease of some pro-inflammatory cytokine levels in mouse serum and changed the redox metabolism in C6 tumors. a An ATP release is significantly lower in tumors treated with BBG compared to the control tumors ( n = 4). The significance of the differences was determined using Student’s t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. b Decline NOS-2 expression in BBG-treated tumors ( n = 4). c Decreased expression of FOXP3 (T reg lymphocytes) and CD68 (macrophages) markers in BBG-treated tumors evaluated using flow cytometry ( n = 5). The significance of the differences was determined using Student’s t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. d The level of the pro-inflammatory and anti-inflammatory cytokines, in mouse serum and glioma tumors using cytokine array for multiplex protein detection ( n = 3). e P2X7 inhibition caused a decrease of GSSG/GSH ratio with a slight decline of SOD activity in BBG-treated tumors ( n = 4). The significance of the differences was determined using Student’s t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control

    Journal: Purinergic Signalling

    Article Title: P2X7 receptor: the regulator of glioma tumor development and survival

    doi: 10.1007/s11302-021-09834-2

    Figure Lengend Snippet: P2X7 inhibition reduced the ATP release with simultaneous decrease of some pro-inflammatory cytokine levels in mouse serum and changed the redox metabolism in C6 tumors. a An ATP release is significantly lower in tumors treated with BBG compared to the control tumors ( n = 4). The significance of the differences was determined using Student’s t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. b Decline NOS-2 expression in BBG-treated tumors ( n = 4). c Decreased expression of FOXP3 (T reg lymphocytes) and CD68 (macrophages) markers in BBG-treated tumors evaluated using flow cytometry ( n = 5). The significance of the differences was determined using Student’s t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. d The level of the pro-inflammatory and anti-inflammatory cytokines, in mouse serum and glioma tumors using cytokine array for multiplex protein detection ( n = 3). e P2X7 inhibition caused a decrease of GSSG/GSH ratio with a slight decline of SOD activity in BBG-treated tumors ( n = 4). The significance of the differences was determined using Student’s t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control

    Article Snippet: Anti-P2X7 antibody (Alomone Labs, Israel) was diluted as recommended in the manufacturer’s instruction and incubated with tissue overnight at 4 °C.

    Techniques: Inhibition, Expressing, Flow Cytometry, Multiplex Assay, Activity Assay

    P2X7 receptor effects on human glioma development. a Western blot analysis of p-p38 MAPK, CD-133 and HSPA1 expression in U-138 human glioma cells ( n = 3). b In silico analysis of P2X7 expression level and survival of glioma patients using information from the TCGA database. c In silico analysis of P2X7 expression level in different WHO astrocytoma grades compared to normal tissue using NCBI dataset. d A transwell migration assay of LN-229 and U-251 glioma cells treated with 2 μM KN-62 for 24 h ( n = 3). e A crystal violet assay demonstrating a synergistic effect of 2 μM KN-62 with BCNU/TMZ co-treatment on LN-229, U-251, and U-138 glioma cell growth for 24 h ( n = 4). The significance of the differences was determined with repeated-measures one-way ANOVA with Duncan multiple range post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. all groups

    Journal: Purinergic Signalling

    Article Title: P2X7 receptor: the regulator of glioma tumor development and survival

    doi: 10.1007/s11302-021-09834-2

    Figure Lengend Snippet: P2X7 receptor effects on human glioma development. a Western blot analysis of p-p38 MAPK, CD-133 and HSPA1 expression in U-138 human glioma cells ( n = 3). b In silico analysis of P2X7 expression level and survival of glioma patients using information from the TCGA database. c In silico analysis of P2X7 expression level in different WHO astrocytoma grades compared to normal tissue using NCBI dataset. d A transwell migration assay of LN-229 and U-251 glioma cells treated with 2 μM KN-62 for 24 h ( n = 3). e A crystal violet assay demonstrating a synergistic effect of 2 μM KN-62 with BCNU/TMZ co-treatment on LN-229, U-251, and U-138 glioma cell growth for 24 h ( n = 4). The significance of the differences was determined with repeated-measures one-way ANOVA with Duncan multiple range post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. all groups

    Article Snippet: Anti-P2X7 antibody (Alomone Labs, Israel) was diluted as recommended in the manufacturer’s instruction and incubated with tissue overnight at 4 °C.

    Techniques: Western Blot, Expressing, In Silico, Transwell Migration Assay, Crystal Violet Assay

    Characterization of P2X7 receptor in glioma C6 cells in vitro. a Western blot analysis of P2X7 protein expression in C6 cells after P2X7 silencing for 72 h ( n = 3). b Calcium signal level in glioma C6 cells after P2X7 downregulation and BzATP stimulation (300 µM) ( n = 4). Gray lines represent the signals from the biological replicates; black lines represent the signal averaged among the individual replicates. Insert shows effect of P2X7 downregulation on the signal strength, one-way t -test: * P ≤ 0.05. c Measurement of bioluminescence originated from released ATP after P2X7 activation/inhibition in C6 cells ( n = 3). The statistical significance of the differences was determined with one-way ANOVA with Bonferroni post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. control group

    Journal: Purinergic Signalling

    Article Title: P2X7 receptor: the regulator of glioma tumor development and survival

    doi: 10.1007/s11302-021-09834-2

    Figure Lengend Snippet: Characterization of P2X7 receptor in glioma C6 cells in vitro. a Western blot analysis of P2X7 protein expression in C6 cells after P2X7 silencing for 72 h ( n = 3). b Calcium signal level in glioma C6 cells after P2X7 downregulation and BzATP stimulation (300 µM) ( n = 4). Gray lines represent the signals from the biological replicates; black lines represent the signal averaged among the individual replicates. Insert shows effect of P2X7 downregulation on the signal strength, one-way t -test: * P ≤ 0.05. c Measurement of bioluminescence originated from released ATP after P2X7 activation/inhibition in C6 cells ( n = 3). The statistical significance of the differences was determined with one-way ANOVA with Bonferroni post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. control group

    Article Snippet: Anti-P2X7 antibody (Alomone Labs, Israel) was diluted as recommended in the manufacturer’s instruction and incubated with tissue overnight at 4 °C.

    Techniques: In Vitro, Western Blot, Expressing, Activation Assay, Inhibition

    P2X7 activation increased C6 cell proliferation, viability, and cell adhesion in vitro. Quantitative data are presented as signal relative to control. a MTS test of long-term C6 cell viability after 100 µM BzATP stimulation of control cells and cells with P2X7 downregulation using RNAi ( n = 3). The significance of the differences was determined with paired t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. b Representative Western blot analysis of pro-survival and pro-proliferative proteins: p-p38 (Thr180/Tyr182), pAkt (Ser473), tAkt, CD133, HSPA1, and HSPA5 in C6 cell lysates after 24-h BzATP stimulation ( n = 3). c Upper panel: MTS test of C6 cell viability after 200 µM BCNU (carmustine) treatment for 24 h. Co-treatment with 100 µM BzATP and 200 µM BCNU increased viability of C6 cells ( n = 3). The significance of the differences was determined with one-way ANOVA with Bonferroni post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. all groups. Bottom panel: a crystal violet assay demonstrating a synergistic effect of BBG with BCNU/TMZ co-treatment on C6 cell growth for 24 h ( n = 6). The significance of the differences was determined with repeated measures one-way ANOVA with Duncan’s multiple range post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. all groups. d P2X7 activation affects C6 cell adhesion to extracellular matrix components: collagen I, collagen IV, fibronectin. C6 cells showed higher adhesion potential after 24-h BzATP stimulation. 24-h co-treatment/treatment with 100 nM BBG significantly decreased C6 cell adhesion to ECM components ( n = 6). The significance of the differences was determined with one-way ANOVA with Bonferroni post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. control group. e P2X7 RNAi downregulation declined C6 cell adhesion to ECM components upon BzATP stimulation ( n = 4). The significance of the differences was determined with paired t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control

    Journal: Purinergic Signalling

    Article Title: P2X7 receptor: the regulator of glioma tumor development and survival

    doi: 10.1007/s11302-021-09834-2

    Figure Lengend Snippet: P2X7 activation increased C6 cell proliferation, viability, and cell adhesion in vitro. Quantitative data are presented as signal relative to control. a MTS test of long-term C6 cell viability after 100 µM BzATP stimulation of control cells and cells with P2X7 downregulation using RNAi ( n = 3). The significance of the differences was determined with paired t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. b Representative Western blot analysis of pro-survival and pro-proliferative proteins: p-p38 (Thr180/Tyr182), pAkt (Ser473), tAkt, CD133, HSPA1, and HSPA5 in C6 cell lysates after 24-h BzATP stimulation ( n = 3). c Upper panel: MTS test of C6 cell viability after 200 µM BCNU (carmustine) treatment for 24 h. Co-treatment with 100 µM BzATP and 200 µM BCNU increased viability of C6 cells ( n = 3). The significance of the differences was determined with one-way ANOVA with Bonferroni post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. all groups. Bottom panel: a crystal violet assay demonstrating a synergistic effect of BBG with BCNU/TMZ co-treatment on C6 cell growth for 24 h ( n = 6). The significance of the differences was determined with repeated measures one-way ANOVA with Duncan’s multiple range post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. all groups. d P2X7 activation affects C6 cell adhesion to extracellular matrix components: collagen I, collagen IV, fibronectin. C6 cells showed higher adhesion potential after 24-h BzATP stimulation. 24-h co-treatment/treatment with 100 nM BBG significantly decreased C6 cell adhesion to ECM components ( n = 6). The significance of the differences was determined with one-way ANOVA with Bonferroni post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. control group. e P2X7 RNAi downregulation declined C6 cell adhesion to ECM components upon BzATP stimulation ( n = 4). The significance of the differences was determined with paired t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control

    Article Snippet: Anti-P2X7 antibody (Alomone Labs, Israel) was diluted as recommended in the manufacturer’s instruction and incubated with tissue overnight at 4 °C.

    Techniques: Activation Assay, In Vitro, Western Blot, Crystal Violet Assay

    Effect of P2X7 inhibition using BBG on glioma C6 tumor development in vivo. a Brilliant Blue G administration led to reduction of glioma tumor mass. Representative images of control and BBG-treated C6 glioma tumors ( n = 9 for control group and n = 10 for BBG-treated group). The significance of the differences was determined using Student’s t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. b Representative Western blot showing the significant decrease of P2X7 expression in tumor homogenates after BBG treatment. Representative image of P2X7 immunodetection in C6 glioma tumors. c P2X7 inhibition resulted in diminished activation of matrix metalloproteinase-2 in glioma tumors. The significance of the differences was determined with Student’s t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. d Western blot analysis of known proteins involved in cell adhesion and EMT signaling in C6 tumor homogenates ( n = 4). e Western blot analysis of known proteins involved in tumor aggressiveness in C6 tumor homogenates ( n = 4)

    Journal: Purinergic Signalling

    Article Title: P2X7 receptor: the regulator of glioma tumor development and survival

    doi: 10.1007/s11302-021-09834-2

    Figure Lengend Snippet: Effect of P2X7 inhibition using BBG on glioma C6 tumor development in vivo. a Brilliant Blue G administration led to reduction of glioma tumor mass. Representative images of control and BBG-treated C6 glioma tumors ( n = 9 for control group and n = 10 for BBG-treated group). The significance of the differences was determined using Student’s t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. b Representative Western blot showing the significant decrease of P2X7 expression in tumor homogenates after BBG treatment. Representative image of P2X7 immunodetection in C6 glioma tumors. c P2X7 inhibition resulted in diminished activation of matrix metalloproteinase-2 in glioma tumors. The significance of the differences was determined with Student’s t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. d Western blot analysis of known proteins involved in cell adhesion and EMT signaling in C6 tumor homogenates ( n = 4). e Western blot analysis of known proteins involved in tumor aggressiveness in C6 tumor homogenates ( n = 4)

    Article Snippet: Anti-P2X7 antibody (Alomone Labs, Israel) was diluted as recommended in the manufacturer’s instruction and incubated with tissue overnight at 4 °C.

    Techniques: Inhibition, In Vivo, Western Blot, Expressing, Immunodetection, Activation Assay

    Activation of P2X7 receptor led to elevated ROS production and mitochondrial membrane depolarization in C6 cells. Quantitative data are presented as signal relative to control. a DCF-DA fluorescence measurement in C6 glioma cells. Cells stimulated with 100 µM BzATP for 1 h were characterized by increased ROS production compared to control in C6 cells ( n = 3). The significance of the differences was determined with one-way ANOVA with Bonferroni post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. control group. b Potential-dependent staining of mitochondria in C6 cells using JC-1. 1-h BzATP (100 µM) stimulation led to considerable mitochondrial membrane depolarization ( n = 3). The significance of the differences was determined with one-way ANOVA with Bonferroni post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. control group. c Representative images of MitoTracker Red CMXRos accumulation in C6 cells stimulated with 100 µM BzATP for 1 h. d DCF-DA fluorescence measurement in C6 glioma cells with RNA interference of P2X7 expression ( n = 4). Downregulation of P2X7 led to decreased ROS production in C6 cells after 1-h BzATP (100 µM) stimulation. The significance of the differences was determined using paired t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. e Potential-dependent staining of mitochondria using JC-1 dye in C6 cells with RNA interference of P2X7 expression after 1 h of 100 μM BzATP pre-treatment ( n = 3). The amount of depolarized mitochondria was higher in control cells when compared to that in cells with P2X7 downregulation. The significance of the differences was determined using paired t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control

    Journal: Purinergic Signalling

    Article Title: P2X7 receptor: the regulator of glioma tumor development and survival

    doi: 10.1007/s11302-021-09834-2

    Figure Lengend Snippet: Activation of P2X7 receptor led to elevated ROS production and mitochondrial membrane depolarization in C6 cells. Quantitative data are presented as signal relative to control. a DCF-DA fluorescence measurement in C6 glioma cells. Cells stimulated with 100 µM BzATP for 1 h were characterized by increased ROS production compared to control in C6 cells ( n = 3). The significance of the differences was determined with one-way ANOVA with Bonferroni post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. control group. b Potential-dependent staining of mitochondria in C6 cells using JC-1. 1-h BzATP (100 µM) stimulation led to considerable mitochondrial membrane depolarization ( n = 3). The significance of the differences was determined with one-way ANOVA with Bonferroni post hoc test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. control group. c Representative images of MitoTracker Red CMXRos accumulation in C6 cells stimulated with 100 µM BzATP for 1 h. d DCF-DA fluorescence measurement in C6 glioma cells with RNA interference of P2X7 expression ( n = 4). Downregulation of P2X7 led to decreased ROS production in C6 cells after 1-h BzATP (100 µM) stimulation. The significance of the differences was determined using paired t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control. e Potential-dependent staining of mitochondria using JC-1 dye in C6 cells with RNA interference of P2X7 expression after 1 h of 100 μM BzATP pre-treatment ( n = 3). The amount of depolarized mitochondria was higher in control cells when compared to that in cells with P2X7 downregulation. The significance of the differences was determined using paired t -test: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 vs. the respective control

    Article Snippet: Anti-P2X7 antibody (Alomone Labs, Israel) was diluted as recommended in the manufacturer’s instruction and incubated with tissue overnight at 4 °C.

    Techniques: Activation Assay, Fluorescence, Staining, Expressing

    P2X7 receptor mediates CION-induced microglial activation and anxiodepressive-like behaviors. (A and B) Western blot analysis reveals significant upregulation of P2X7 receptor (P2X7R) level on day 14 after CION in the ipsilateral hippocampal CA1 area of rats. ** P

    Journal: bioRxiv

    Article Title: Asymmetric activation of microglia in the hippocampus drives anxiodepressive consequences of trigeminal neuralgia

    doi: 10.1101/2022.04.16.488241

    Figure Lengend Snippet: P2X7 receptor mediates CION-induced microglial activation and anxiodepressive-like behaviors. (A and B) Western blot analysis reveals significant upregulation of P2X7 receptor (P2X7R) level on day 14 after CION in the ipsilateral hippocampal CA1 area of rats. ** P

    Article Snippet: We used the following primary antibodies: rabbit anti-P2X7R (1:2000, #APR-004, Alomone labs, Jerusalem, Israel), goat-anti IBA-1 (1:1000, #ab5076, Abcam, Cambridge, MA), and rabbit-anti IL-1β (1:500, rabbit, PeproTech, Rocky Hill, NJ).

    Techniques: Activation Assay, Western Blot