ionomycin  (Alomone Labs)


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  • 91
    Name:
    Ionomycin calcium salt
    Description:
    Ionomycin is a narrow spectrum antibiotic for gram positive bacteria In mammalian cells Ionomycin acts as a selective Ca2 ionophore3 In addition Ionomycin induces hydrolysis of phosphoinositides activation of PKC in human T cells as well as apoptosis in various cell lines
    Catalog Number:
    I-700
    Price:
    23.0
    Category:
    Small Molecule
    Source:
    Streptomyces conglobatus.
    Applications:
    0
    Purity:
    >99% (HPLC)
    Size:
    1 mg
    Format:
    Lyophilized/solid.
    Formula:
    C41H70O9Ca
    Molecular Weight:
    747.08
    Molecule Name:
    (4R,6S,8S,10Z,12R,14R,16E,18R,19R,20S,21S)-11,19,21-Tri hydroxy-4,6,8,12,14,18,20-heptamethyl-22-[(2S,2'R,5S,5'S)-octahydro-5'-[(1R)-1-hydroxyethyl]-2,5'-dimethyl[2,2'-bifuran]-5-yl]-9-oxo-10,16-docosadienoic acid calcium salt.
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    Structured Review

    Alomone Labs ionomycin
    Ionomycin calcium salt
    Ionomycin is a narrow spectrum antibiotic for gram positive bacteria In mammalian cells Ionomycin acts as a selective Ca2 ionophore3 In addition Ionomycin induces hydrolysis of phosphoinositides activation of PKC in human T cells as well as apoptosis in various cell lines
    https://www.bioz.com/result/ionomycin/product/Alomone Labs
    Average 91 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    ionomycin - by Bioz Stars, 2021-09
    91/100 stars

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    Related Articles

    Mass Spectrometry:

    Article Title: TRPV4-mediates oscillatory fluid shear mechanotransduction in mesenchymal stem cells in part via the primary cilium
    Article Snippet: .. Exposure time was kept below 600 ms and was kept constant between control and treatment groups, allowing image acquisition every 1.29 s. Following each experiment, 10 µM ionomycin (Alomone Labs, Jerusalem, Israel, http://www.alomone.com ) was applied to the cells as a positive control for sensitivity of the indicator to calcium. ..

    Positive Control:

    Article Title: TRPV4-mediates oscillatory fluid shear mechanotransduction in mesenchymal stem cells in part via the primary cilium
    Article Snippet: .. Exposure time was kept below 600 ms and was kept constant between control and treatment groups, allowing image acquisition every 1.29 s. Following each experiment, 10 µM ionomycin (Alomone Labs, Jerusalem, Israel, http://www.alomone.com ) was applied to the cells as a positive control for sensitivity of the indicator to calcium. ..

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  • 94
    Alomone Labs anti kcnq1
    The chronic effect of PMA on I Ks current in HEK 293B cell. (a) The representative I Ks current under the PMA (100 nM) treatment. (b) The current–voltage relationship for the tail currents under the PMA (100 nM) treatment. (c) The summary data for the tail currents under the PMA (100 nM) treatment at +40 mV prepulse. (d) The normalized I–V relationship for I Ks current. The solid lines represent fits to a Boltzmann function. (e) The representative immunoblot and summary data for the <t>KCNQ1</t> channel membrane protein under the PMA (100 nM) treatment (** P
    Anti Kcnq1, 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/anti kcnq1/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti kcnq1 - by Bioz Stars, 2021-09
    94/100 stars
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    91
    Alomone Labs phrixotoxin 1
    Influences of voltage-gated potassium (K v ) channel inhibition on progenitor c ell proliferation . Proliferation of hNPCs was analyzed via BrdU incorporation assay. (A): Progenitor cell proliferation was measured colorimetrically after 72 h of K v channel inhibition and normalized to control values without addition of inhibitor. Electrophysiologically determined inhibitory doses (IC 50 /IC 80 ) of 4-aminopyridine (4-AP), <t>phrixotoxin-1</t> (PTX), ammonium chloride (NH 4 Cl), tetraethylammonium chloride (TEA), quinidine (QND) and α-dendrotoxin (DTX) were applied. Progenitor cell proliferation was significantly reduced by inhibition of I A with 4-AP, PTX, NH 4 Cl as well as by unspecific blockers like TEA and higher doses of QND. In contrast, the I K antagonist DTX increased proliferation of hNPCs (n≥4, 3 tissue preparations; one-way ANOVA, followed by Tukey's post-hoc test, *p
    Phrixotoxin 1, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/phrixotoxin 1/product/Alomone Labs
    Average 91 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    phrixotoxin 1 - by Bioz Stars, 2021-09
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    94
    Alomone Labs rabbit anti p75
    Effect of 8-OH-DPAT on the protein expression of molecules from the BDNF signaling pathway in the HC, determined by Western Blot. (A) mBDNF. (B) TrkB. (C) proBDNF. (D) <t>p75</t> in hyposerotonergic (PCPA-treated) or control (vehicle-treated) mice receiving the 5-HT 1 A agonist 8-OH-DPAT (striped bars) or NaCl (white bars). (E) Representative membrane showing signal for p75 (75 kDa) and tubuline (52 kDa). 1: Control-NaCl. 2: Control-DPAT. 3: PCPA-NaCl. 4: PCPA-DPAT. Data are expressed as mean ± S.E.M., n = 6 per experimental group. ∗ p
    Rabbit Anti P75, 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 anti p75/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit anti p75 - by Bioz Stars, 2021-09
    94/100 stars
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    95
    Alomone Labs rabbit anti trpv4
    <t>TRPV4</t> is required for matrix stiffness-induced dermal myofibroblast differentiation. A : HDFs were plated (10,000 cells per well) on collagen-coated (10 μg/ml) hydrogels with varying degrees of stiffness (0.5 and 8 kPa) for 48 h under vehicle-treated or GSK219-treated (5 μM) conditions. TRPV4 antagonist, GSK219, blocks matrix stiffness-induced human dermal myofibroblast differentiation. Representative photomicrographic images are shown for α-SMA (green), F-actin (red), and nucleus (blue). Original magnification, ×40. B and C : quantified results are expressed as means ± SE *** P
    Rabbit Anti Trpv4, 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/rabbit anti trpv4/product/Alomone Labs
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit anti trpv4 - by Bioz Stars, 2021-09
    95/100 stars
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    Image Search Results


    The chronic effect of PMA on I Ks current in HEK 293B cell. (a) The representative I Ks current under the PMA (100 nM) treatment. (b) The current–voltage relationship for the tail currents under the PMA (100 nM) treatment. (c) The summary data for the tail currents under the PMA (100 nM) treatment at +40 mV prepulse. (d) The normalized I–V relationship for I Ks current. The solid lines represent fits to a Boltzmann function. (e) The representative immunoblot and summary data for the KCNQ1 channel membrane protein under the PMA (100 nM) treatment (** P

    Journal: Channels

    Article Title: Specific protein kinase C isoform exerts chronic inhibition on the slowly activating delayed-rectifier potassium current by affecting channel trafficking

    doi: 10.1080/19336950.2021.1882112

    Figure Lengend Snippet: The chronic effect of PMA on I Ks current in HEK 293B cell. (a) The representative I Ks current under the PMA (100 nM) treatment. (b) The current–voltage relationship for the tail currents under the PMA (100 nM) treatment. (c) The summary data for the tail currents under the PMA (100 nM) treatment at +40 mV prepulse. (d) The normalized I–V relationship for I Ks current. The solid lines represent fits to a Boltzmann function. (e) The representative immunoblot and summary data for the KCNQ1 channel membrane protein under the PMA (100 nM) treatment (** P

    Article Snippet: The following primary antibodies were used as follows: anti-Na/K-ATPase (Proteintech, 1:500) and anti-KCNQ1 (Alomone, 1:700).

    Techniques:

    The chronic effect of Ang II on I Ks current in HEK 293B cell. (a) The representative I Ks current evoked by voltage protocol shown in inset under the Ang II (100 nM) treatment. (b) The current–voltage relationship for the tail currents under the Ang II (100 nM) treatment. (c) The summary data for the tail currents under the Ang II (100 nM) treatment at +40 mV prepulse. (d) The normalized I–V relationship for I Ks current. The solid lines represent fits to a Boltzmann function. (e) The representative immunoblot and summary data for the KCNQ1 channel membrane protein under the Ang II (100 nM) treatment (** P

    Journal: Channels

    Article Title: Specific protein kinase C isoform exerts chronic inhibition on the slowly activating delayed-rectifier potassium current by affecting channel trafficking

    doi: 10.1080/19336950.2021.1882112

    Figure Lengend Snippet: The chronic effect of Ang II on I Ks current in HEK 293B cell. (a) The representative I Ks current evoked by voltage protocol shown in inset under the Ang II (100 nM) treatment. (b) The current–voltage relationship for the tail currents under the Ang II (100 nM) treatment. (c) The summary data for the tail currents under the Ang II (100 nM) treatment at +40 mV prepulse. (d) The normalized I–V relationship for I Ks current. The solid lines represent fits to a Boltzmann function. (e) The representative immunoblot and summary data for the KCNQ1 channel membrane protein under the Ang II (100 nM) treatment (** P

    Article Snippet: The following primary antibodies were used as follows: anti-Na/K-ATPase (Proteintech, 1:500) and anti-KCNQ1 (Alomone, 1:700).

    Techniques:

    Influences of voltage-gated potassium (K v ) channel inhibition on progenitor c ell proliferation . Proliferation of hNPCs was analyzed via BrdU incorporation assay. (A): Progenitor cell proliferation was measured colorimetrically after 72 h of K v channel inhibition and normalized to control values without addition of inhibitor. Electrophysiologically determined inhibitory doses (IC 50 /IC 80 ) of 4-aminopyridine (4-AP), phrixotoxin-1 (PTX), ammonium chloride (NH 4 Cl), tetraethylammonium chloride (TEA), quinidine (QND) and α-dendrotoxin (DTX) were applied. Progenitor cell proliferation was significantly reduced by inhibition of I A with 4-AP, PTX, NH 4 Cl as well as by unspecific blockers like TEA and higher doses of QND. In contrast, the I K antagonist DTX increased proliferation of hNPCs (n≥4, 3 tissue preparations; one-way ANOVA, followed by Tukey's post-hoc test, *p

    Journal: PLoS ONE

    Article Title: Characterization of Voltage-Gated Potassium Channels in Human Neural Progenitor Cells

    doi: 10.1371/journal.pone.0006168

    Figure Lengend Snippet: Influences of voltage-gated potassium (K v ) channel inhibition on progenitor c ell proliferation . Proliferation of hNPCs was analyzed via BrdU incorporation assay. (A): Progenitor cell proliferation was measured colorimetrically after 72 h of K v channel inhibition and normalized to control values without addition of inhibitor. Electrophysiologically determined inhibitory doses (IC 50 /IC 80 ) of 4-aminopyridine (4-AP), phrixotoxin-1 (PTX), ammonium chloride (NH 4 Cl), tetraethylammonium chloride (TEA), quinidine (QND) and α-dendrotoxin (DTX) were applied. Progenitor cell proliferation was significantly reduced by inhibition of I A with 4-AP, PTX, NH 4 Cl as well as by unspecific blockers like TEA and higher doses of QND. In contrast, the I K antagonist DTX increased proliferation of hNPCs (n≥4, 3 tissue preparations; one-way ANOVA, followed by Tukey's post-hoc test, *p

    Article Snippet: Different antagonists (all from Sigma-Aldrich GmbH if not stated otherwise) were dissolved in this bathing solution: 4-aminopyridine (4-AP, 0.1–10 mM), phrixotoxin-1 (PTX, 1–1000 nM, Alomone Labs, Jerusalem, Israel), ammonium chloride (NH4 Cl, 1–100 mM), quinidine (QND, 0.1–100 µM), α-dendrotoxin (DTX, 1–1000 nM), margatoxin (MTX, 0.1–50 nM) and tetraethylammonium chloride (TEA, 1–100 mM).

    Techniques: Inhibition, BrdU Incorporation Assay

    Cell viability after inhibition of voltage-gated potassium (K v ) channels. Determination of cell viability in proliferating hNPCs via 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium salt (MTT) assay. (A): Cell viability was measured colorimetrically after 72 h of K v channel inhibition with different concentrations of 4-aminopyridine (4-AP), phrixotoxin-1 (PTX), ammonium chloride (NH 4 Cl), tetraethylammonium chloride (TEA), quinidine (QND) and α-dendrotoxin (DTX) and normalized to control values without addition of inhibitor. (B): Viability of hNPCs was significantly reduced by electrophysiologically determined inhibitory doses (IC 50 /IC 80 ) of 4-AP, PTX and NH 4 Cl, which specifically blocked I A , as well as by TEA and higher doses of QND, which inhibited both current components (n≥4, 3 tissue preparations; one-way ANOVA, followed by Tukey's post-hoc test, ***p

    Journal: PLoS ONE

    Article Title: Characterization of Voltage-Gated Potassium Channels in Human Neural Progenitor Cells

    doi: 10.1371/journal.pone.0006168

    Figure Lengend Snippet: Cell viability after inhibition of voltage-gated potassium (K v ) channels. Determination of cell viability in proliferating hNPCs via 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium salt (MTT) assay. (A): Cell viability was measured colorimetrically after 72 h of K v channel inhibition with different concentrations of 4-aminopyridine (4-AP), phrixotoxin-1 (PTX), ammonium chloride (NH 4 Cl), tetraethylammonium chloride (TEA), quinidine (QND) and α-dendrotoxin (DTX) and normalized to control values without addition of inhibitor. (B): Viability of hNPCs was significantly reduced by electrophysiologically determined inhibitory doses (IC 50 /IC 80 ) of 4-AP, PTX and NH 4 Cl, which specifically blocked I A , as well as by TEA and higher doses of QND, which inhibited both current components (n≥4, 3 tissue preparations; one-way ANOVA, followed by Tukey's post-hoc test, ***p

    Article Snippet: Different antagonists (all from Sigma-Aldrich GmbH if not stated otherwise) were dissolved in this bathing solution: 4-aminopyridine (4-AP, 0.1–10 mM), phrixotoxin-1 (PTX, 1–1000 nM, Alomone Labs, Jerusalem, Israel), ammonium chloride (NH4 Cl, 1–100 mM), quinidine (QND, 0.1–100 µM), α-dendrotoxin (DTX, 1–1000 nM), margatoxin (MTX, 0.1–50 nM) and tetraethylammonium chloride (TEA, 1–100 mM).

    Techniques: Inhibition, MTT Assay

    Pharmacological inhibition of K v currents in hNPCs. Biophysically separated A-type (I A ) and delayed-rectifying (I K ) K v currents in proliferating hNPCs were differentially inhibited by the 4-aminopyridine (4-AP, i), phrixotoxin-1 (PTX, ii), ammonium chloride (NH 4 Cl, iii), tetraethylammonium chloride (TEA, iv), quinidine (QND, v) and α-dendrotoxin (DTX, vi). (A): Peak amplitudes of I A were measured during a depolarizing voltage step from −130 mV to 0 mV between 0 and 20 ms (inset). (B): I K was determined between 280 and 300 ms of a 100 mV depolarization step following a −40 mV prepulse during the application of different antagonist concentrations (insets). (C): Both current values were normalized to the non-inhibited peak amplitudes. Dose-response relationships were fitted with the Hill equation and IC 50 values were determined (see Tab. 2 ). Note that PTX selectively and 4-AP preferentially inhibited I A , while DTX selectively blocked I K .

    Journal: PLoS ONE

    Article Title: Characterization of Voltage-Gated Potassium Channels in Human Neural Progenitor Cells

    doi: 10.1371/journal.pone.0006168

    Figure Lengend Snippet: Pharmacological inhibition of K v currents in hNPCs. Biophysically separated A-type (I A ) and delayed-rectifying (I K ) K v currents in proliferating hNPCs were differentially inhibited by the 4-aminopyridine (4-AP, i), phrixotoxin-1 (PTX, ii), ammonium chloride (NH 4 Cl, iii), tetraethylammonium chloride (TEA, iv), quinidine (QND, v) and α-dendrotoxin (DTX, vi). (A): Peak amplitudes of I A were measured during a depolarizing voltage step from −130 mV to 0 mV between 0 and 20 ms (inset). (B): I K was determined between 280 and 300 ms of a 100 mV depolarization step following a −40 mV prepulse during the application of different antagonist concentrations (insets). (C): Both current values were normalized to the non-inhibited peak amplitudes. Dose-response relationships were fitted with the Hill equation and IC 50 values were determined (see Tab. 2 ). Note that PTX selectively and 4-AP preferentially inhibited I A , while DTX selectively blocked I K .

    Article Snippet: Different antagonists (all from Sigma-Aldrich GmbH if not stated otherwise) were dissolved in this bathing solution: 4-aminopyridine (4-AP, 0.1–10 mM), phrixotoxin-1 (PTX, 1–1000 nM, Alomone Labs, Jerusalem, Israel), ammonium chloride (NH4 Cl, 1–100 mM), quinidine (QND, 0.1–100 µM), α-dendrotoxin (DTX, 1–1000 nM), margatoxin (MTX, 0.1–50 nM) and tetraethylammonium chloride (TEA, 1–100 mM).

    Techniques: Inhibition, Mass Spectrometry

    Effect of 8-OH-DPAT on the protein expression of molecules from the BDNF signaling pathway in the HC, determined by Western Blot. (A) mBDNF. (B) TrkB. (C) proBDNF. (D) p75 in hyposerotonergic (PCPA-treated) or control (vehicle-treated) mice receiving the 5-HT 1 A agonist 8-OH-DPAT (striped bars) or NaCl (white bars). (E) Representative membrane showing signal for p75 (75 kDa) and tubuline (52 kDa). 1: Control-NaCl. 2: Control-DPAT. 3: PCPA-NaCl. 4: PCPA-DPAT. Data are expressed as mean ± S.E.M., n = 6 per experimental group. ∗ p

    Journal: Frontiers in Cellular Neuroscience

    Article Title: Differential Hippocampal Expression of BDNF Isoforms and Their Receptors Under Diverse Configurations of the Serotonergic System in a Mice Model of Increased Neuronal Survival

    doi: 10.3389/fncel.2019.00384

    Figure Lengend Snippet: Effect of 8-OH-DPAT on the protein expression of molecules from the BDNF signaling pathway in the HC, determined by Western Blot. (A) mBDNF. (B) TrkB. (C) proBDNF. (D) p75 in hyposerotonergic (PCPA-treated) or control (vehicle-treated) mice receiving the 5-HT 1 A agonist 8-OH-DPAT (striped bars) or NaCl (white bars). (E) Representative membrane showing signal for p75 (75 kDa) and tubuline (52 kDa). 1: Control-NaCl. 2: Control-DPAT. 3: PCPA-NaCl. 4: PCPA-DPAT. Data are expressed as mean ± S.E.M., n = 6 per experimental group. ∗ p

    Article Snippet: Western Blotting Membranes were incubated for 1 h with blocking solution (5% milk in TBST) and then probed overnight at 4°C with mouse anti-BDNF (1:2000; Icosagen; 327-100 clone 3C11), rabbit anti-p75 (1:700; Alomone Labs; ANT-007), rabbit anti-TrkB (1:700; Alomone Labs; ANT-019), and rabbit anti-proBDNF (1:250; Abcam; ab72440) in TBST. β-III Tubulin was used as a loading control (1:2500; R & D Systems).

    Techniques: Expressing, Western Blot, Mouse Assay

    Protein expression of molecules from the BDNF signaling pathway determined by Western Blot and representative membranes. (A) mBDNF. (B) TrkB. (C) proBDNF. (D) p75 in hyposerotonergic (PCPA-treated) or control (vehicle-treated) mice for 4 weeks. (E–H) Representative membrane showing signal for TrkB, p75, pro-BDNF, and mBDNF in Control and PCPA-treatd mice. Data are expressed as mean ± S.E.M., n = 11 (control) and 12 (PCPA). ∗ p

    Journal: Frontiers in Cellular Neuroscience

    Article Title: Differential Hippocampal Expression of BDNF Isoforms and Their Receptors Under Diverse Configurations of the Serotonergic System in a Mice Model of Increased Neuronal Survival

    doi: 10.3389/fncel.2019.00384

    Figure Lengend Snippet: Protein expression of molecules from the BDNF signaling pathway determined by Western Blot and representative membranes. (A) mBDNF. (B) TrkB. (C) proBDNF. (D) p75 in hyposerotonergic (PCPA-treated) or control (vehicle-treated) mice for 4 weeks. (E–H) Representative membrane showing signal for TrkB, p75, pro-BDNF, and mBDNF in Control and PCPA-treatd mice. Data are expressed as mean ± S.E.M., n = 11 (control) and 12 (PCPA). ∗ p

    Article Snippet: Western Blotting Membranes were incubated for 1 h with blocking solution (5% milk in TBST) and then probed overnight at 4°C with mouse anti-BDNF (1:2000; Icosagen; 327-100 clone 3C11), rabbit anti-p75 (1:700; Alomone Labs; ANT-007), rabbit anti-TrkB (1:700; Alomone Labs; ANT-019), and rabbit anti-proBDNF (1:250; Abcam; ab72440) in TBST. β-III Tubulin was used as a loading control (1:2500; R & D Systems).

    Techniques: Expressing, Western Blot, Mouse Assay

    TRPV4 is required for matrix stiffness-induced dermal myofibroblast differentiation. A : HDFs were plated (10,000 cells per well) on collagen-coated (10 μg/ml) hydrogels with varying degrees of stiffness (0.5 and 8 kPa) for 48 h under vehicle-treated or GSK219-treated (5 μM) conditions. TRPV4 antagonist, GSK219, blocks matrix stiffness-induced human dermal myofibroblast differentiation. Representative photomicrographic images are shown for α-SMA (green), F-actin (red), and nucleus (blue). Original magnification, ×40. B and C : quantified results are expressed as means ± SE *** P

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: TRPV4 ion channel is a novel regulator of dermal myofibroblast differentiation

    doi: 10.1152/ajpcell.00187.2016

    Figure Lengend Snippet: TRPV4 is required for matrix stiffness-induced dermal myofibroblast differentiation. A : HDFs were plated (10,000 cells per well) on collagen-coated (10 μg/ml) hydrogels with varying degrees of stiffness (0.5 and 8 kPa) for 48 h under vehicle-treated or GSK219-treated (5 μM) conditions. TRPV4 antagonist, GSK219, blocks matrix stiffness-induced human dermal myofibroblast differentiation. Representative photomicrographic images are shown for α-SMA (green), F-actin (red), and nucleus (blue). Original magnification, ×40. B and C : quantified results are expressed as means ± SE *** P

    Article Snippet: Immunoblotting was performed using the following primary antibodies: rabbit anti-phospho-Smad2/3, rabbit anti-Smad2/3, rabbit anti-phospho-Akt, rabbit anti-Akt (1:2,000; Cell Signaling, Beverly, MA); mouse anti-α-SMA (1:20,000; Sigma, St. Louis, MO); goat anti-collagen type I (1:700; EMD Millipore, Billerica, MA); rabbit anti-TRPV4 (1:700; Alomone, Jerusalem, Israel), goat anti-β-actin (EMD Millipore); rabbit anti-GAPDH (1:3,000; Santa Cruz, Dallas, TX), followed by incubation with secondary antibodies goat anti-mouse, goat anti-rabbit, and donkey anti-goat (1:5,000; Jackson ImmunoResearch, West Grove, PA).

    Techniques:

    TRPV4 calcium channel is functional in MDFs and in HDF cells. A and B : FlexStation 3 recording of Calcium 5 dye-loaded WT MDF monolayers shows that TRPV4 agonist GSK101 induces Ca 2+ influx in a concentration-dependent manner, which is completely absent in TRPV4 KO MDFs. C : quantitation of results (means ± SE) from A and B . All experiments were repeated 3 times in quadruplicate. D : HDFs (10,000 cells per well) were seeded on collagen-coated (10 μg/ml) 96-well plastic plate. Ca 2+ influx is shown by relative fluorescence units (RFUs) measuring ΔF/F (Max-Min). A23 (2 μM), a calcium ionophore, was used as a positive control. FlexStation 3 recording of Calcium 5 dye-loaded HDF monolayers shows that TRPV4 agonist GSK101 induces Ca 2+ influx in a concentration-dependent manner. E : quantitation of results from D (means ± SE). F : HDFs (10,000 cells per well) were seeded on collagen-coated (10 μg/ml) 96-well plastic plate. Concentration-dependent inhibition of TRPV4-elicited Ca 2+ influx in HDFs by its selective antagonist, GSK219. TRPV4-elicited Ca 2+ influx was generated by GSK101 (10 nM). All experiments were repeated 3 times in quadruplicate. G : quantitation of results from F (means ± SE). H : TRPV4-elicited calcium rise is not dependent on intracellular pools and/or regulators of calcium. HDF monolayers were compared for their intracellular calcium rise after TRPV4 activation by its specific agonist, GSK101 (10 nM), with or without selective inhibitors to TGFβRI (SD208, 10 μM), the inositol 1,4,5-triphosphate receptor [xestospongin C (Xesto), 10 μM], ryanodine receptors [ryanodine (RyD), 10 μM], or sarco/endoplasmic reticulum calcium transport ATPase [cyclopiazonic acid (CPA), 10 μM]. Results are expressed as means ± SE.

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: TRPV4 ion channel is a novel regulator of dermal myofibroblast differentiation

    doi: 10.1152/ajpcell.00187.2016

    Figure Lengend Snippet: TRPV4 calcium channel is functional in MDFs and in HDF cells. A and B : FlexStation 3 recording of Calcium 5 dye-loaded WT MDF monolayers shows that TRPV4 agonist GSK101 induces Ca 2+ influx in a concentration-dependent manner, which is completely absent in TRPV4 KO MDFs. C : quantitation of results (means ± SE) from A and B . All experiments were repeated 3 times in quadruplicate. D : HDFs (10,000 cells per well) were seeded on collagen-coated (10 μg/ml) 96-well plastic plate. Ca 2+ influx is shown by relative fluorescence units (RFUs) measuring ΔF/F (Max-Min). A23 (2 μM), a calcium ionophore, was used as a positive control. FlexStation 3 recording of Calcium 5 dye-loaded HDF monolayers shows that TRPV4 agonist GSK101 induces Ca 2+ influx in a concentration-dependent manner. E : quantitation of results from D (means ± SE). F : HDFs (10,000 cells per well) were seeded on collagen-coated (10 μg/ml) 96-well plastic plate. Concentration-dependent inhibition of TRPV4-elicited Ca 2+ influx in HDFs by its selective antagonist, GSK219. TRPV4-elicited Ca 2+ influx was generated by GSK101 (10 nM). All experiments were repeated 3 times in quadruplicate. G : quantitation of results from F (means ± SE). H : TRPV4-elicited calcium rise is not dependent on intracellular pools and/or regulators of calcium. HDF monolayers were compared for their intracellular calcium rise after TRPV4 activation by its specific agonist, GSK101 (10 nM), with or without selective inhibitors to TGFβRI (SD208, 10 μM), the inositol 1,4,5-triphosphate receptor [xestospongin C (Xesto), 10 μM], ryanodine receptors [ryanodine (RyD), 10 μM], or sarco/endoplasmic reticulum calcium transport ATPase [cyclopiazonic acid (CPA), 10 μM]. Results are expressed as means ± SE.

    Article Snippet: Immunoblotting was performed using the following primary antibodies: rabbit anti-phospho-Smad2/3, rabbit anti-Smad2/3, rabbit anti-phospho-Akt, rabbit anti-Akt (1:2,000; Cell Signaling, Beverly, MA); mouse anti-α-SMA (1:20,000; Sigma, St. Louis, MO); goat anti-collagen type I (1:700; EMD Millipore, Billerica, MA); rabbit anti-TRPV4 (1:700; Alomone, Jerusalem, Israel), goat anti-β-actin (EMD Millipore); rabbit anti-GAPDH (1:3,000; Santa Cruz, Dallas, TX), followed by incubation with secondary antibodies goat anti-mouse, goat anti-rabbit, and donkey anti-goat (1:5,000; Jackson ImmunoResearch, West Grove, PA).

    Techniques: Functional Assay, Concentration Assay, Quantitation Assay, Fluorescence, Positive Control, Inhibition, Generated, Activation Assay

    TRPV4 is expressed in primary normal human (HDFs) and mouse (MDFs) dermal fibroblasts, and stimulation of HDFs or MDFs with TGFβ1 produced a dose-dependent increase in total cellular TRPV4 protein expression. A : RT-PCR analysis demonstrating that TRPV4 mRNAs are present in HDFs. H 2 O (no RNA) and No RT (no reverse transcriptase) samples were used as negative control, whereas RNA extracted from human lung fibroblast cells (19Lu) was used as positive control. B : representative immunoblots show TRPV4 protein expression in HDFs treated with indicated doses of TGFβ1 for 48 h. Total actin was used as loading control. C : quantitative analysis of total cellular TRPV4 protein expression in TGFβ1-treated HDFs. Results shown are means ± SE from 3 independent experiments (** P

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: TRPV4 ion channel is a novel regulator of dermal myofibroblast differentiation

    doi: 10.1152/ajpcell.00187.2016

    Figure Lengend Snippet: TRPV4 is expressed in primary normal human (HDFs) and mouse (MDFs) dermal fibroblasts, and stimulation of HDFs or MDFs with TGFβ1 produced a dose-dependent increase in total cellular TRPV4 protein expression. A : RT-PCR analysis demonstrating that TRPV4 mRNAs are present in HDFs. H 2 O (no RNA) and No RT (no reverse transcriptase) samples were used as negative control, whereas RNA extracted from human lung fibroblast cells (19Lu) was used as positive control. B : representative immunoblots show TRPV4 protein expression in HDFs treated with indicated doses of TGFβ1 for 48 h. Total actin was used as loading control. C : quantitative analysis of total cellular TRPV4 protein expression in TGFβ1-treated HDFs. Results shown are means ± SE from 3 independent experiments (** P

    Article Snippet: Immunoblotting was performed using the following primary antibodies: rabbit anti-phospho-Smad2/3, rabbit anti-Smad2/3, rabbit anti-phospho-Akt, rabbit anti-Akt (1:2,000; Cell Signaling, Beverly, MA); mouse anti-α-SMA (1:20,000; Sigma, St. Louis, MO); goat anti-collagen type I (1:700; EMD Millipore, Billerica, MA); rabbit anti-TRPV4 (1:700; Alomone, Jerusalem, Israel), goat anti-β-actin (EMD Millipore); rabbit anti-GAPDH (1:3,000; Santa Cruz, Dallas, TX), followed by incubation with secondary antibodies goat anti-mouse, goat anti-rabbit, and donkey anti-goat (1:5,000; Jackson ImmunoResearch, West Grove, PA).

    Techniques: Produced, Expressing, Reverse Transcription Polymerase Chain Reaction, Negative Control, Positive Control, Western Blot

    TRPV4 is required for TGFβ1-induced dermal myofibroblast differentiation. A : WT and TRPV4 KO MDFs were plated on collagen-coated (10 μg/ml) plastic wells and incubated with or without TGFβ1 (2 ng/ml) for 48 h. Representative fluorescence micrographs (original magnification, ×40) showing that myofibroblast differentiation is reduced in fibroblasts from TRPV4 KO mice (colocalization of α-SMA and F-actin, orange). B : quantification of photomicrographs from A using ImageJ software. Results are expressed as means ± SE *** P

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: TRPV4 ion channel is a novel regulator of dermal myofibroblast differentiation

    doi: 10.1152/ajpcell.00187.2016

    Figure Lengend Snippet: TRPV4 is required for TGFβ1-induced dermal myofibroblast differentiation. A : WT and TRPV4 KO MDFs were plated on collagen-coated (10 μg/ml) plastic wells and incubated with or without TGFβ1 (2 ng/ml) for 48 h. Representative fluorescence micrographs (original magnification, ×40) showing that myofibroblast differentiation is reduced in fibroblasts from TRPV4 KO mice (colocalization of α-SMA and F-actin, orange). B : quantification of photomicrographs from A using ImageJ software. Results are expressed as means ± SE *** P

    Article Snippet: Immunoblotting was performed using the following primary antibodies: rabbit anti-phospho-Smad2/3, rabbit anti-Smad2/3, rabbit anti-phospho-Akt, rabbit anti-Akt (1:2,000; Cell Signaling, Beverly, MA); mouse anti-α-SMA (1:20,000; Sigma, St. Louis, MO); goat anti-collagen type I (1:700; EMD Millipore, Billerica, MA); rabbit anti-TRPV4 (1:700; Alomone, Jerusalem, Israel), goat anti-β-actin (EMD Millipore); rabbit anti-GAPDH (1:3,000; Santa Cruz, Dallas, TX), followed by incubation with secondary antibodies goat anti-mouse, goat anti-rabbit, and donkey anti-goat (1:5,000; Jackson ImmunoResearch, West Grove, PA).

    Techniques: Incubation, Fluorescence, Mouse Assay, Software