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R&D Systems elisa kits
Concentrations of TNF-α, IL-10, and PGE 2 in IPEC-J2 cell culture supernatants. Supernatants were collected from the indicated cultures at 3 h after <t>F4</t> + ETEC challenge. The concentrations of (A) TNF-α, (B) IL-10, and (C) PGE 2 were determined by <t>ELISA.</t> Data are presented as means ± SEM of three independent experiments. * P
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1) Product Images from "A Selected Lactobacillus rhamnosus Strain Promotes EGFR-Independent Akt Activation in an Enterotoxigenic Escherichia coli K88-Infected IPEC-J2 Cell Model"

Article Title: A Selected Lactobacillus rhamnosus Strain Promotes EGFR-Independent Akt Activation in an Enterotoxigenic Escherichia coli K88-Infected IPEC-J2 Cell Model

Journal: PLoS ONE

doi: 10.1371/journal.pone.0125717

Concentrations of TNF-α, IL-10, and PGE 2 in IPEC-J2 cell culture supernatants. Supernatants were collected from the indicated cultures at 3 h after F4 + ETEC challenge. The concentrations of (A) TNF-α, (B) IL-10, and (C) PGE 2 were determined by ELISA. Data are presented as means ± SEM of three independent experiments. * P
Figure Legend Snippet: Concentrations of TNF-α, IL-10, and PGE 2 in IPEC-J2 cell culture supernatants. Supernatants were collected from the indicated cultures at 3 h after F4 + ETEC challenge. The concentrations of (A) TNF-α, (B) IL-10, and (C) PGE 2 were determined by ELISA. Data are presented as means ± SEM of three independent experiments. * P

Techniques Used: Cell Culture, Enzyme-linked Immunosorbent Assay

2) Product Images from "Acidosis Drives Damage-associated Molecular Pattern (DAMP)-induced Interleukin-1 Secretion via a Caspase-1-independent Pathway *"

Article Title: Acidosis Drives Damage-associated Molecular Pattern (DAMP)-induced Interleukin-1 Secretion via a Caspase-1-independent Pathway *

Journal: The Journal of Biological Chemistry

doi: 10.1074/jbc.M113.478941

Effect of extracellular pH on IL-1β processing in THP-1 cells. LPS-primed THP-1 cells were treated with vehicle ( Veh ), CPPD (250 μg/ml), or MSU (250 μg/ml) for 1 h at pH 6.2 or 7.4. Processing of IL-1β was observed by Western blotting of the supernatants ( A ), with quantification of levels released by ELISA ( B ). The effect of pH and DAMPs on cell death was measured by the release of LDH ( C ). ELISA and LDH data are means ± S.E. of three separate experiments. Western blots are representative of three separate experiments.
Figure Legend Snippet: Effect of extracellular pH on IL-1β processing in THP-1 cells. LPS-primed THP-1 cells were treated with vehicle ( Veh ), CPPD (250 μg/ml), or MSU (250 μg/ml) for 1 h at pH 6.2 or 7.4. Processing of IL-1β was observed by Western blotting of the supernatants ( A ), with quantification of levels released by ELISA ( B ). The effect of pH and DAMPs on cell death was measured by the release of LDH ( C ). ELISA and LDH data are means ± S.E. of three separate experiments. Western blots are representative of three separate experiments.

Techniques Used: Western Blot, Enzyme-linked Immunosorbent Assay

DAMP-induced release of 20-kDa IL-1β is caspase-1-independent in primary mixed glia. LPS-primed mixed glia were pretreated with the caspase-1 inhibitor Ac-YVAD-CHO ( YVAD ; 100 μ m ) or the cathepsin D inhibitor pepstatin A ( PepA ; 50 μ m ) for 15 min prior to CPPD treatment (250 μg/ml, 1 h) at pH 7.4 ( A and C ) or pH 6.2 ( B and D ). Processing of IL-1β was observed by Western blotting of the supernatants ( blots ), with quantification of levels released by ELISA ( graphs ). ELISA data are means ± S.E. of between five and six separate experiments. Western blots are representative of three separate experiments. Statistical analysis was performed using a one-way ANOVA with a Bonferroni post hoc test. ns , not significant; **, p
Figure Legend Snippet: DAMP-induced release of 20-kDa IL-1β is caspase-1-independent in primary mixed glia. LPS-primed mixed glia were pretreated with the caspase-1 inhibitor Ac-YVAD-CHO ( YVAD ; 100 μ m ) or the cathepsin D inhibitor pepstatin A ( PepA ; 50 μ m ) for 15 min prior to CPPD treatment (250 μg/ml, 1 h) at pH 7.4 ( A and C ) or pH 6.2 ( B and D ). Processing of IL-1β was observed by Western blotting of the supernatants ( blots ), with quantification of levels released by ELISA ( graphs ). ELISA data are means ± S.E. of between five and six separate experiments. Western blots are representative of three separate experiments. Statistical analysis was performed using a one-way ANOVA with a Bonferroni post hoc test. ns , not significant; **, p

Techniques Used: Western Blot, Enzyme-linked Immunosorbent Assay

CPPD-induced 20-kDa IL-1β is NLRP3-independent in primary mixed glia. Mixed glial cultures prepared from WT and NLRP3 KO mice were treated with CPPD (250 μg/ml, 1 h) at pH 6.2. Processing of IL-1β was observed by Western blotting of the supernatants ( blot ), with quantification of levels released by ELISA ( graph ). ELISA data are means ± S.E. of three separate experiments. Western blots are representative of three separate experiments. Statistical analysis was performed using a one-way ANOVA with a Bonferroni post hoc test. ns , not significant; **, p
Figure Legend Snippet: CPPD-induced 20-kDa IL-1β is NLRP3-independent in primary mixed glia. Mixed glial cultures prepared from WT and NLRP3 KO mice were treated with CPPD (250 μg/ml, 1 h) at pH 6.2. Processing of IL-1β was observed by Western blotting of the supernatants ( blot ), with quantification of levels released by ELISA ( graph ). ELISA data are means ± S.E. of three separate experiments. Western blots are representative of three separate experiments. Statistical analysis was performed using a one-way ANOVA with a Bonferroni post hoc test. ns , not significant; **, p

Techniques Used: Mouse Assay, Western Blot, Enzyme-linked Immunosorbent Assay

DAMP-induced release of 20-kDa IL-1β is caspase-1-independent. LPS-primed THP-1 cells were incubated at pH 6.2 and treated with the caspase-1 inhibitor Ac-YVAD-CHO ( YVAD ; 100 μ m ; A ), the cathepsin B inhibitor CA-074 methyl ester ( Ca074Me ; 50 μ m ; B ), or the cathepsin D inhibitor pepstatin A ( PepA ; 50 μ m ; C ) for 15 min prior to CPPD treatment (250 μg/ml, 1 h). Processing of IL-1β was observed by Western blotting of the supernatants ( blots ), with quantification of levels released by ELISA ( graphs ). ELISA data are means ± S.E. of three separate experiments. Western blots are representative of three separate experiments. Statistical analysis was performed using a one-way ANOVA with a Bonferroni post hoc test. ns , not significant; *, p
Figure Legend Snippet: DAMP-induced release of 20-kDa IL-1β is caspase-1-independent. LPS-primed THP-1 cells were incubated at pH 6.2 and treated with the caspase-1 inhibitor Ac-YVAD-CHO ( YVAD ; 100 μ m ; A ), the cathepsin B inhibitor CA-074 methyl ester ( Ca074Me ; 50 μ m ; B ), or the cathepsin D inhibitor pepstatin A ( PepA ; 50 μ m ; C ) for 15 min prior to CPPD treatment (250 μg/ml, 1 h). Processing of IL-1β was observed by Western blotting of the supernatants ( blots ), with quantification of levels released by ELISA ( graphs ). ELISA data are means ± S.E. of three separate experiments. Western blots are representative of three separate experiments. Statistical analysis was performed using a one-way ANOVA with a Bonferroni post hoc test. ns , not significant; *, p

Techniques Used: Incubation, Western Blot, Enzyme-linked Immunosorbent Assay

Lactic acid-induced release of 20-kDa IL-1β occurs independently of caspase-1 in primary mixed glia. LPS-primed mixed glia were pretreated with 100 μ m Ac-YVAD-CHO ( YVAD ; A ) or 50 μ m pepstatin A ( PepA ; C ) for 15 min prior to lactic acid treatment (25 m m , 8 h). Mixed glial cultures prepared from WT and NLRP3 KO mice were treated with 25 m m lactic acid (8 h; B ). Processing of IL-1β was observed by Western blotting of the supernatants ( blots ), with quantification of levels released by ELISA ( graphs ). ELISA data are means ± S.E. of between three and six separate experiments. Western blots are representative of three separate experiments. Statistical analysis was performed using a one-way ANOVA with a Bonferroni post hoc test. ns , not significant; **, p
Figure Legend Snippet: Lactic acid-induced release of 20-kDa IL-1β occurs independently of caspase-1 in primary mixed glia. LPS-primed mixed glia were pretreated with 100 μ m Ac-YVAD-CHO ( YVAD ; A ) or 50 μ m pepstatin A ( PepA ; C ) for 15 min prior to lactic acid treatment (25 m m , 8 h). Mixed glial cultures prepared from WT and NLRP3 KO mice were treated with 25 m m lactic acid (8 h; B ). Processing of IL-1β was observed by Western blotting of the supernatants ( blots ), with quantification of levels released by ELISA ( graphs ). ELISA data are means ± S.E. of between three and six separate experiments. Western blots are representative of three separate experiments. Statistical analysis was performed using a one-way ANOVA with a Bonferroni post hoc test. ns , not significant; **, p

Techniques Used: Mouse Assay, Western Blot, Enzyme-linked Immunosorbent Assay

Lactic acid induces release of IL-1α from primary mixed glia. LPS-primed WT mixed glia were treated with 25 m m lactic acid for 8 h, and IL-1α processing and release was measured ( A ). LPS-primed mixed glia from WT and NLRP3 KO mice were treated with lactic acid (25 m m ; 8 h; B ). LPS-primed WT mixed glia were pretreated with calpain inhibitor III ( Cal ; 50 μ m ) for 15 min prior to lactic acid treatment (25 m m , 8 h; C ). Processing of IL-1α was observed by Western blotting of the supernatants ( blots ), with quantification of levels released by ELISA ( graphs ). ELISA data are means ± S.E. of between three and six separate experiments. Western blots are representative of three separate experiments. Statistical analysis was performed using Student's t test ( A ) or one-way ANOVA with a Bonferroni post hoc test ( B and C ). ns , not significant; *, p
Figure Legend Snippet: Lactic acid induces release of IL-1α from primary mixed glia. LPS-primed WT mixed glia were treated with 25 m m lactic acid for 8 h, and IL-1α processing and release was measured ( A ). LPS-primed mixed glia from WT and NLRP3 KO mice were treated with lactic acid (25 m m ; 8 h; B ). LPS-primed WT mixed glia were pretreated with calpain inhibitor III ( Cal ; 50 μ m ) for 15 min prior to lactic acid treatment (25 m m , 8 h; C ). Processing of IL-1α was observed by Western blotting of the supernatants ( blots ), with quantification of levels released by ELISA ( graphs ). ELISA data are means ± S.E. of between three and six separate experiments. Western blots are representative of three separate experiments. Statistical analysis was performed using Student's t test ( A ) or one-way ANOVA with a Bonferroni post hoc test ( B and C ). ns , not significant; *, p

Techniques Used: Mouse Assay, Western Blot, Enzyme-linked Immunosorbent Assay

Lactic acid induces the release of 20-kDa IL-1β from primary mixed glia. LPS-primed mixed glia were treated with lactic acid (25 m m , 8 h) or ATP (5.5 m m , 1 h). Processing of IL-1β was observed by Western blotting of the supernatants ( A ), with quantification of levels of IL-1β in cell lysates ( B ) and released into the supernatants ( C ) measured by ELISA. The effect of lactic acid and ATP on cell death was measured by the release of LDH ( D ). ELISA data are means ± S.E. of six separate experiments. Western blots are representative of three separate experiments. LDH data are means ± S.E. of three separate experiments. Statistical analysis was performed using a one-way ANOVA with a Bonferroni post hoc test. ns , not significant; ***, p
Figure Legend Snippet: Lactic acid induces the release of 20-kDa IL-1β from primary mixed glia. LPS-primed mixed glia were treated with lactic acid (25 m m , 8 h) or ATP (5.5 m m , 1 h). Processing of IL-1β was observed by Western blotting of the supernatants ( A ), with quantification of levels of IL-1β in cell lysates ( B ) and released into the supernatants ( C ) measured by ELISA. The effect of lactic acid and ATP on cell death was measured by the release of LDH ( D ). ELISA data are means ± S.E. of six separate experiments. Western blots are representative of three separate experiments. LDH data are means ± S.E. of three separate experiments. Statistical analysis was performed using a one-way ANOVA with a Bonferroni post hoc test. ns , not significant; ***, p

Techniques Used: Western Blot, Enzyme-linked Immunosorbent Assay

Effect of extracellular pH on IL-1β processing in primary mixed glia. LPS-primed mixed glia were treated for 1 h with ATP (5.5 m m ), MSU (250 μg/ml), or CPPD (250 μg/ml) at pH 7.4 ( A ) or pH 6.2 ( B ). Processing of IL-1β was observed by Western blotting of the supernatants ( blots ), with quantification of levels released by ELISA ( graphs ). The effect of pH and DAMPs on cell death was measured by the release of LDH ( C ). ELISA and LDH data are means ± S.E. of between five and six separate experiments. Western blots are representative of three separate experiments. Statistical analysis was performed using Student's t test. ns , not significant; *, p
Figure Legend Snippet: Effect of extracellular pH on IL-1β processing in primary mixed glia. LPS-primed mixed glia were treated for 1 h with ATP (5.5 m m ), MSU (250 μg/ml), or CPPD (250 μg/ml) at pH 7.4 ( A ) or pH 6.2 ( B ). Processing of IL-1β was observed by Western blotting of the supernatants ( blots ), with quantification of levels released by ELISA ( graphs ). The effect of pH and DAMPs on cell death was measured by the release of LDH ( C ). ELISA and LDH data are means ± S.E. of between five and six separate experiments. Western blots are representative of three separate experiments. Statistical analysis was performed using Student's t test. ns , not significant; *, p

Techniques Used: Western Blot, Enzyme-linked Immunosorbent Assay

3) Product Images from "Green Tea Polyphenols Reduced Fat Deposits in High Fat-Fed Rats via erk1/2-PPAR?-Adiponectin Pathway"

Article Title: Green Tea Polyphenols Reduced Fat Deposits in High Fat-Fed Rats via erk1/2-PPAR?-Adiponectin Pathway

Journal: PLoS ONE

doi: 10.1371/journal.pone.0053796

GTPs and selective inhibitor of erk1/2 alleviated high glucose-induced adiponectin decrease. One hundred fifty mg VAT were cultured in DMEM with high glucose (33 mmol/L) and cotreated with GTPs (4 µg/ml) for 48 hrs or pretreated with PD98059 for 1 hr. The supernatant of cell culture medium was collected for ELISA of secreted adiponectin. (A) The mRNA level of adiponectin, comparative Ct method with GADPH as reference was adopted. (B) The secreted adiponectin in the supernatant of culture medium is in ng/mL. High glucose incubation (H) down-regulated the mRNA expression and secretion of adiponectin, the effects could be attenuated by GTPs treatment (GH) or PD98059(PD). (* P
Figure Legend Snippet: GTPs and selective inhibitor of erk1/2 alleviated high glucose-induced adiponectin decrease. One hundred fifty mg VAT were cultured in DMEM with high glucose (33 mmol/L) and cotreated with GTPs (4 µg/ml) for 48 hrs or pretreated with PD98059 for 1 hr. The supernatant of cell culture medium was collected for ELISA of secreted adiponectin. (A) The mRNA level of adiponectin, comparative Ct method with GADPH as reference was adopted. (B) The secreted adiponectin in the supernatant of culture medium is in ng/mL. High glucose incubation (H) down-regulated the mRNA expression and secretion of adiponectin, the effects could be attenuated by GTPs treatment (GH) or PD98059(PD). (* P

Techniques Used: Cell Culture, Enzyme-linked Immunosorbent Assay, Incubation, Expressing

4) Product Images from "TGF-? Signaling Regulates Neuronal C1q Expression and Developmental Synaptic Refinement"

Article Title: TGF-? Signaling Regulates Neuronal C1q Expression and Developmental Synaptic Refinement

Journal: Nature neuroscience

doi: 10.1038/nn.3560

TGF-β is necessary and sufficient for neuronal C1q upregulation in vitro (A) ACM cytokine profiling by ELISA. N=3 independent ACM batches. (B) Immunodepletion of TGF-β, but not other cytokines, significantly reduced ACM-induced C1q upregulation (one-way ANOVA, n= 3 experiments, ***p
Figure Legend Snippet: TGF-β is necessary and sufficient for neuronal C1q upregulation in vitro (A) ACM cytokine profiling by ELISA. N=3 independent ACM batches. (B) Immunodepletion of TGF-β, but not other cytokines, significantly reduced ACM-induced C1q upregulation (one-way ANOVA, n= 3 experiments, ***p

Techniques Used: In Vitro, Enzyme-linked Immunosorbent Assay

5) Product Images from "CD4+ T cells are trigger and target of the glucocorticoid response that prevents lethal immunopathology in toxoplasma infection"

Article Title: CD4+ T cells are trigger and target of the glucocorticoid response that prevents lethal immunopathology in toxoplasma infection

Journal: The Journal of Experimental Medicine

doi: 10.1084/jem.20122300

Mortality in T. gondii –infected GR lck-Cre mice is dependent on CD4 + T cells that also drive the induction of the GC response. (A–C) T. gondii –infected mice were treated i.p. with anti-CD4, anti-CD8, or control mAb on days −1, 3, 5, 8, and 12 ( n = 3–5 mice). Survival was monitored daily (A), and serum levels of AST and CK (B) and IL-12p40, IFN-γ, and IL-10 (C) were measured on day 8. Bars represent mean ± SEM of values for individual animals ( n = 3–12). (D) Survival of T. gondii –infected RAG KO mice ( n = 4) and those adoptively transferred with naive GR lck-Cre ( n = 6) or WT ( n = 9) CD4 + T cells. (E) Serum corticosterone levels in animals described in A. The results shown in A–E are representative of three experiments performed. (F–H) Serum corticosterone levels in naive and toxoplasma-infected mice deficient in the genes indicated (F), WT animals treated with anti-TNF and anti–IFN-γ Ab alone or in combination with anti–IL-6 Ab (G), and RAG −/− and OT-II RAG −/− mice (H). Bars represent mean ± SEM of the ELISA values for individual mice ( n = 4–10) pooled from two independent experiments. (I) Corticosterone levels in uninfected and 8-d T. gondii –infected RAG −/− animals, one group of which received naive WT CD4 + T cells. Bars represent mean ± SEM of ELISA values for individual mice ( n = 5–8) from one representative out of three performed. *, P
Figure Legend Snippet: Mortality in T. gondii –infected GR lck-Cre mice is dependent on CD4 + T cells that also drive the induction of the GC response. (A–C) T. gondii –infected mice were treated i.p. with anti-CD4, anti-CD8, or control mAb on days −1, 3, 5, 8, and 12 ( n = 3–5 mice). Survival was monitored daily (A), and serum levels of AST and CK (B) and IL-12p40, IFN-γ, and IL-10 (C) were measured on day 8. Bars represent mean ± SEM of values for individual animals ( n = 3–12). (D) Survival of T. gondii –infected RAG KO mice ( n = 4) and those adoptively transferred with naive GR lck-Cre ( n = 6) or WT ( n = 9) CD4 + T cells. (E) Serum corticosterone levels in animals described in A. The results shown in A–E are representative of three experiments performed. (F–H) Serum corticosterone levels in naive and toxoplasma-infected mice deficient in the genes indicated (F), WT animals treated with anti-TNF and anti–IFN-γ Ab alone or in combination with anti–IL-6 Ab (G), and RAG −/− and OT-II RAG −/− mice (H). Bars represent mean ± SEM of the ELISA values for individual mice ( n = 4–10) pooled from two independent experiments. (I) Corticosterone levels in uninfected and 8-d T. gondii –infected RAG −/− animals, one group of which received naive WT CD4 + T cells. Bars represent mean ± SEM of ELISA values for individual mice ( n = 5–8) from one representative out of three performed. *, P

Techniques Used: Infection, Mouse Assay, AST Assay, Enzyme-linked Immunosorbent Assay

T. gondii infection elicits a GC response, and the lack of GR signaling in T cells results in acute mortality of toxoplasma-infected mice. (A) C57BL/6 mice were infected i.p. with an average of 15 ME49 cysts, and serum corticosterone, IFN-γ, IL-12p70 and p40, IL-27p28, and IL-10 levels were measured on the days indicated. Symbols represent mean ± SEM of the ELISA values for the individual animals ( n = 3–12) at each time point pooled from three independent experiments. (B) Survival of homozygote GR lck-Cre , heterozygote GR fl/+lck-Cre , and littermate control animals after infection. The survival curves shown are from one representative of 10 experiments performed, two of which included GR fl/+lck-Cre mice. (C) Parasite burdens in PECs and spleen on day 8 after infection as determined by plaque assay. Bars represent mean ± SEM number of PFU per organ ( n = 3–5 mice). (D) Weight loss and serum levels of AST and CK in infected animals. Results shown are means ± SEM for values for the individual mice ( n = 4–5). No distinguishing histopathological changes were detected in lung, heart, liver, and kidney at this day 8 time point. Data presented in C–E are representative of two experiments performed. *, P
Figure Legend Snippet: T. gondii infection elicits a GC response, and the lack of GR signaling in T cells results in acute mortality of toxoplasma-infected mice. (A) C57BL/6 mice were infected i.p. with an average of 15 ME49 cysts, and serum corticosterone, IFN-γ, IL-12p70 and p40, IL-27p28, and IL-10 levels were measured on the days indicated. Symbols represent mean ± SEM of the ELISA values for the individual animals ( n = 3–12) at each time point pooled from three independent experiments. (B) Survival of homozygote GR lck-Cre , heterozygote GR fl/+lck-Cre , and littermate control animals after infection. The survival curves shown are from one representative of 10 experiments performed, two of which included GR fl/+lck-Cre mice. (C) Parasite burdens in PECs and spleen on day 8 after infection as determined by plaque assay. Bars represent mean ± SEM number of PFU per organ ( n = 3–5 mice). (D) Weight loss and serum levels of AST and CK in infected animals. Results shown are means ± SEM for values for the individual mice ( n = 4–5). No distinguishing histopathological changes were detected in lung, heart, liver, and kidney at this day 8 time point. Data presented in C–E are representative of two experiments performed. *, P

Techniques Used: Infection, Mouse Assay, Enzyme-linked Immunosorbent Assay, Plaque Assay, AST Assay

6) Product Images from "Homeostatic expansion of autoreactive immunoglobulin-secreting cells in the Rag2 mouse model of Omenn syndrome"

Article Title: Homeostatic expansion of autoreactive immunoglobulin-secreting cells in the Rag2 mouse model of Omenn syndrome

Journal: The Journal of Experimental Medicine

doi: 10.1084/jem.20091928

Residual autoreactive B cells in Rag2 R229Q mice contribute to tissue damage in autoimmune-like manifestations. (A) Characterization of kidney infiltrates and correlation with clinical symptoms. Kidney sections from a mouse with proteinuria (score 4: ≥ 2000 mg/dl; top) and a mouse without renal disease (bottom). Kidneys from mice with proteinuria show infiltrates positive for staining with CD3, B220, IgM and IgG. Bars: (left) 100 µM; (middle) 50 µM; (right) 60 µM. (B) The presence of IgG anti–double-strand DNA autoantibodies was analyzed in the sera of WT ( n = 70) and Rag2 R229Q ( n = 97) mice by ELISA and IFA in five independent experiments. Bars indicate the frequency of positive sera in mice aged 8–24 wk. (C) Ig targeting tissues are detected in the sera of Rag2 R229Q mice. Frozen sections obtained from, gut, thyroid, and adrenal gland of Rag2/Il2rc double-KO mice were incubated with sera obtained from WT and Rag2 R229Q mice. Staining with anti–mouse IgG Ab followed by incubation with secondary, anti–mouse IgG peroxidase-conjugated antibody was performed. Representative images from 1 out of 10 mice tested/group. Bars, 100 µM. (D) FACS analysis of λ light chain expression in gated B220 + IgM + cells. Results are representative of 11 mice/group analyzed in three independent experiments. (E) Serum BAFF in Rag2 R229Q ( n = 26) and WT littermates ( n = 15), measured by ELISA in two independent experiments. **, P ≤ 0.01; ***, P ≤ 0.001.
Figure Legend Snippet: Residual autoreactive B cells in Rag2 R229Q mice contribute to tissue damage in autoimmune-like manifestations. (A) Characterization of kidney infiltrates and correlation with clinical symptoms. Kidney sections from a mouse with proteinuria (score 4: ≥ 2000 mg/dl; top) and a mouse without renal disease (bottom). Kidneys from mice with proteinuria show infiltrates positive for staining with CD3, B220, IgM and IgG. Bars: (left) 100 µM; (middle) 50 µM; (right) 60 µM. (B) The presence of IgG anti–double-strand DNA autoantibodies was analyzed in the sera of WT ( n = 70) and Rag2 R229Q ( n = 97) mice by ELISA and IFA in five independent experiments. Bars indicate the frequency of positive sera in mice aged 8–24 wk. (C) Ig targeting tissues are detected in the sera of Rag2 R229Q mice. Frozen sections obtained from, gut, thyroid, and adrenal gland of Rag2/Il2rc double-KO mice were incubated with sera obtained from WT and Rag2 R229Q mice. Staining with anti–mouse IgG Ab followed by incubation with secondary, anti–mouse IgG peroxidase-conjugated antibody was performed. Representative images from 1 out of 10 mice tested/group. Bars, 100 µM. (D) FACS analysis of λ light chain expression in gated B220 + IgM + cells. Results are representative of 11 mice/group analyzed in three independent experiments. (E) Serum BAFF in Rag2 R229Q ( n = 26) and WT littermates ( n = 15), measured by ELISA in two independent experiments. **, P ≤ 0.01; ***, P ≤ 0.001.

Techniques Used: Mouse Assay, Staining, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Incubation, FACS, Expressing

Characterization of effector T cell subsets in Rag2 R229Q mice. (A) Cytokine production profile of splenic CD4 + T cells. Total splenocytes were stimulated for a total of 5 h in the presence of PMA and ionomycin. Cytokine production was assessed by flow cytometry with intracellular staining. Plots are gated on CD4 + T cells. Numbers indicate the percentage of cells in the respective quadrants. Data represent one of the three independent experiments with consistent results. (B) Serum levels of IL-21 (WT, n = 30; Rag2 R229Q , n = 50) and IFN-γ (WT, n = 30; Rag2 R229Q , n = 50) cytokines in WT and Rag2 R229Q mice (8–24 wk old) as analyzed by ELISA in three independent experiments. (C) Expression of CXCR5 and ICOS by CD4 + splenic T cells from WT and Rag2 R229Q mice. Numbers in quadrants indicate percentage of cells in each. (D) Expression of indicated molecules in sorted splenic CD4 + T cell population assessed by qRT-PCR. Samples were run in duplicate, and the target mRNA was normalized to Ppia mRNA. The RNA contents are shown as arbitrary units (A.U.). Means ± SD of six mice/group. Data represent one of the three independent experiments with consistent results. *, P
Figure Legend Snippet: Characterization of effector T cell subsets in Rag2 R229Q mice. (A) Cytokine production profile of splenic CD4 + T cells. Total splenocytes were stimulated for a total of 5 h in the presence of PMA and ionomycin. Cytokine production was assessed by flow cytometry with intracellular staining. Plots are gated on CD4 + T cells. Numbers indicate the percentage of cells in the respective quadrants. Data represent one of the three independent experiments with consistent results. (B) Serum levels of IL-21 (WT, n = 30; Rag2 R229Q , n = 50) and IFN-γ (WT, n = 30; Rag2 R229Q , n = 50) cytokines in WT and Rag2 R229Q mice (8–24 wk old) as analyzed by ELISA in three independent experiments. (C) Expression of CXCR5 and ICOS by CD4 + splenic T cells from WT and Rag2 R229Q mice. Numbers in quadrants indicate percentage of cells in each. (D) Expression of indicated molecules in sorted splenic CD4 + T cell population assessed by qRT-PCR. Samples were run in duplicate, and the target mRNA was normalized to Ppia mRNA. The RNA contents are shown as arbitrary units (A.U.). Means ± SD of six mice/group. Data represent one of the three independent experiments with consistent results. *, P

Techniques Used: Mouse Assay, Flow Cytometry, Cytometry, Staining, Enzyme-linked Immunosorbent Assay, Expressing, Quantitative RT-PCR

7) Product Images from "Chitinase 3-like 1 promotes macrophage recruitment and angiogenesis in colorectal cancer"

Article Title: Chitinase 3-like 1 promotes macrophage recruitment and angiogenesis in colorectal cancer

Journal: Oncogene

doi: 10.1038/onc.2011.498

Enhanced chemotaxis of macrophages and angiogenesis through ERK1/2 and JNK signaling but not p38 in colon cancer cells (A) Western blot analysis of phospho-ERK, -JNK, -p38, and total-ERK, -JNK, -p38 expression in SW480 cells. SW480 cells were transfected with empty- (EV), CHI3L1 expression- (CH), negative control miRNA- (NC), or two kinds of CHI3L1 specific miRNAs (miRNA 1, miRNA 2)- vector for 48 hrs. CHI3L1 overexpression in SW480 enhanced ERK and JNK phosphorylation but not p38. CHI3L1 knockdown diminished ERK and JNK phosphorylation but not p38. The fold expression of phospho-ERK/JNK/p38 normalized by total ERK/JNK/p38 was shown relative to the control (EV or NC). (B) The protein secretions of IL-8 and MCP-1 from SW480 cells assessed by ELISA. Incubation with ERK inhibitor (PD98059) or JNK inhibitor (SP600125) significantly diminished the protein secretions of IL-8 and MCP-1 from SW480 cells, but not with p38 inhibitor (SB203580). Transfection of siRNA for ERK or JNK significantly diminished the protein secretions of IL-8 and MCP-1 from SW480 cells, but not for p38. (C) Mean numbers of migrated THP-1 cells at x200 magnification. Migration of THP-1 cells was not inhibited by MAPK inhibitors when cultured without SW480 cells (no cells). Incubation with ERK inhibitor (PD98059) or JNK inhibitor (SP600125) significantly diminished the migration of THP-1 cells. In contrast, p38 inhibitor (SB203580) did not affect the chemotaxis enhanced by CHI3L1 (left panel). Transfection of siRNA for ERK or JNK in SW480 cells significantly diminished the migration of THP-1 cells, but siRNA for p38 had no effect (right panel). (D) Mean numbers of migrated HUVECs at x200 magnification. Migration of HUVECs was not inhibited by MAPK inhibitors when cultured without SW480 cells (no cells). Incubation with ERK inhibitor (PD98059) or JNK inhibitor (SP600125) significantly diminished the migration of HUVECs. In contrast, p38 inhibitor (SB203580) did not affect the chemotaxis enhanced by CHI3L1 (left panel). Transfection of siRNA for ERK or JNK in SW480 cells significantly diminished the migration of HUVECs, but siRNA for p38 had no effect (right panel). (E) Mean numbers of tube-forming structures at x100 magnification. Tube formation was not inhibited by MAPK inhibitors when cultured in basal medium. Addition of ERK inhibitor (PD98059) or JNK inhibitor (SP600125) during preparation of conditioned medium significantly inhibited the tube formation. In contrast, p38 inhibitor (SB203580) did not affect the tube formation enhanced by CHI3L1 (left panel). Transfection of siRNA for ERK or JNK in SW480 cells significantly diminished the tube formation, but siRNA for p38 had no effect (right panel). All results (B-E) are the mean (± SEM) of more than three separate experiments. * P
Figure Legend Snippet: Enhanced chemotaxis of macrophages and angiogenesis through ERK1/2 and JNK signaling but not p38 in colon cancer cells (A) Western blot analysis of phospho-ERK, -JNK, -p38, and total-ERK, -JNK, -p38 expression in SW480 cells. SW480 cells were transfected with empty- (EV), CHI3L1 expression- (CH), negative control miRNA- (NC), or two kinds of CHI3L1 specific miRNAs (miRNA 1, miRNA 2)- vector for 48 hrs. CHI3L1 overexpression in SW480 enhanced ERK and JNK phosphorylation but not p38. CHI3L1 knockdown diminished ERK and JNK phosphorylation but not p38. The fold expression of phospho-ERK/JNK/p38 normalized by total ERK/JNK/p38 was shown relative to the control (EV or NC). (B) The protein secretions of IL-8 and MCP-1 from SW480 cells assessed by ELISA. Incubation with ERK inhibitor (PD98059) or JNK inhibitor (SP600125) significantly diminished the protein secretions of IL-8 and MCP-1 from SW480 cells, but not with p38 inhibitor (SB203580). Transfection of siRNA for ERK or JNK significantly diminished the protein secretions of IL-8 and MCP-1 from SW480 cells, but not for p38. (C) Mean numbers of migrated THP-1 cells at x200 magnification. Migration of THP-1 cells was not inhibited by MAPK inhibitors when cultured without SW480 cells (no cells). Incubation with ERK inhibitor (PD98059) or JNK inhibitor (SP600125) significantly diminished the migration of THP-1 cells. In contrast, p38 inhibitor (SB203580) did not affect the chemotaxis enhanced by CHI3L1 (left panel). Transfection of siRNA for ERK or JNK in SW480 cells significantly diminished the migration of THP-1 cells, but siRNA for p38 had no effect (right panel). (D) Mean numbers of migrated HUVECs at x200 magnification. Migration of HUVECs was not inhibited by MAPK inhibitors when cultured without SW480 cells (no cells). Incubation with ERK inhibitor (PD98059) or JNK inhibitor (SP600125) significantly diminished the migration of HUVECs. In contrast, p38 inhibitor (SB203580) did not affect the chemotaxis enhanced by CHI3L1 (left panel). Transfection of siRNA for ERK or JNK in SW480 cells significantly diminished the migration of HUVECs, but siRNA for p38 had no effect (right panel). (E) Mean numbers of tube-forming structures at x100 magnification. Tube formation was not inhibited by MAPK inhibitors when cultured in basal medium. Addition of ERK inhibitor (PD98059) or JNK inhibitor (SP600125) during preparation of conditioned medium significantly inhibited the tube formation. In contrast, p38 inhibitor (SB203580) did not affect the tube formation enhanced by CHI3L1 (left panel). Transfection of siRNA for ERK or JNK in SW480 cells significantly diminished the tube formation, but siRNA for p38 had no effect (right panel). All results (B-E) are the mean (± SEM) of more than three separate experiments. * P

Techniques Used: Chemotaxis Assay, Western Blot, Expressing, Transfection, Negative Control, Plasmid Preparation, Over Expression, Enzyme-linked Immunosorbent Assay, Incubation, Migration, Cell Culture

Enhanced chemotaxis of macrophages and angiogenesis by CHI3L1-induced IL-8 and MCP-1 protein secretions (A) The protein secretions of IL-8 and MCP-1 in the supernatant assessed by ELISA. CHI3L1 transfection (CH) in SW480 cells for 48 hrs significantly and dose-dependently increased the protein secretion of IL-8 and MCP-1 as compared with empty vector (EV) transfection. Stimulation with CHI3L1 (80 ng/ml) in SW480 cells significantly increased the protein secretion of IL-8 and MCP-1. (B) The mRNA expressions of IL-8 and MCP-1 determined by quantitative RT-PCR. The mRNA levels of IL-8/GAPDH and MCP-1/GAPDH were significantly increased in CHI3L1 overexpression (CH) as compared with those in empty vector (EV) transfection. (C) The protein secretions of IL-8 and MCP-1 in the supernatant assessed by ELISA. Two kinds of CHI3L1 miRNAs (miRNA 1, miRNA 2) transfection in SW480 cells for 48 hrs significantly decreased the protein secretions of IL-8 and MCP-1 as compared with negative control miRNA (NC) transfection. (D) Mean numbers of migrated THP-1 cells at x200 magnification. Incubation with IL-8 or MCP-1 neutralizing Ab significantly diminished the migration of THP-1 cells induced by CHI3L1 in SW480 cells as compared with control goat IgG. (E) Mean numbers of migrated HUVECs at x200 magnification. Incubation with IL-8 or MCP-1 neutralizing Ab significantly diminished the migration of HUVECs induced by CHI3L1 in SW480 cells as compared with control goat IgG. (F) Mean numbers of tube-forming structures at x100 magnification. Addition of IL-8 or MCP-1neutralizing Ab during preparation of conditioned medium from empty vector-transfected or CHI3L1-overexpressed SW480 cells significantly inhibited the tube formation of HUVECs as compared with control goat IgG. All results (A-F) are the mean (± SEM) of more than three separate experiments. * P
Figure Legend Snippet: Enhanced chemotaxis of macrophages and angiogenesis by CHI3L1-induced IL-8 and MCP-1 protein secretions (A) The protein secretions of IL-8 and MCP-1 in the supernatant assessed by ELISA. CHI3L1 transfection (CH) in SW480 cells for 48 hrs significantly and dose-dependently increased the protein secretion of IL-8 and MCP-1 as compared with empty vector (EV) transfection. Stimulation with CHI3L1 (80 ng/ml) in SW480 cells significantly increased the protein secretion of IL-8 and MCP-1. (B) The mRNA expressions of IL-8 and MCP-1 determined by quantitative RT-PCR. The mRNA levels of IL-8/GAPDH and MCP-1/GAPDH were significantly increased in CHI3L1 overexpression (CH) as compared with those in empty vector (EV) transfection. (C) The protein secretions of IL-8 and MCP-1 in the supernatant assessed by ELISA. Two kinds of CHI3L1 miRNAs (miRNA 1, miRNA 2) transfection in SW480 cells for 48 hrs significantly decreased the protein secretions of IL-8 and MCP-1 as compared with negative control miRNA (NC) transfection. (D) Mean numbers of migrated THP-1 cells at x200 magnification. Incubation with IL-8 or MCP-1 neutralizing Ab significantly diminished the migration of THP-1 cells induced by CHI3L1 in SW480 cells as compared with control goat IgG. (E) Mean numbers of migrated HUVECs at x200 magnification. Incubation with IL-8 or MCP-1 neutralizing Ab significantly diminished the migration of HUVECs induced by CHI3L1 in SW480 cells as compared with control goat IgG. (F) Mean numbers of tube-forming structures at x100 magnification. Addition of IL-8 or MCP-1neutralizing Ab during preparation of conditioned medium from empty vector-transfected or CHI3L1-overexpressed SW480 cells significantly inhibited the tube formation of HUVECs as compared with control goat IgG. All results (A-F) are the mean (± SEM) of more than three separate experiments. * P

Techniques Used: Chemotaxis Assay, Enzyme-linked Immunosorbent Assay, Transfection, Plasmid Preparation, Quantitative RT-PCR, Over Expression, Negative Control, Incubation, Migration

8) Product Images from "The Transcription Factor C/EBP delta Has Anti-Apoptotic and Anti-Inflammatory Roles in Pancreatic Beta Cells"

Article Title: The Transcription Factor C/EBP delta Has Anti-Apoptotic and Anti-Inflammatory Roles in Pancreatic Beta Cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0031062

C/EBPδ silencing increases cytokine-induced chemokine production by enhancing STAT1 activation. INS-1E cells were transfected with either siCtrl (white dots/bars), siC/EBPδ #1 (black triangles/bars) or siC/EBPδ #2 (grey squares/bars) and subsequently left untreated, or treated with IL-1β+IFN-γ for the indicated time points; (A–E) C/EBPδ, CXCL1, 9, 10 CCL20 mRNA expressions were assayed by RT-PCR and normalized for the housekeeping gene GAPDH; (F–G) CXCL1 and CXCL9 protein secretions were evaluated by ELISA. (H) C/EBPδ, phospho-STAT1, total STAT1, IRF-1 and α-tubulin expressions were evaluated by Western blot. (I) 24 h after siRNA transfection, cells were transfected with a STAT1 luciferase reporter+pRL-CMV and subsequently left untreated or exposed to cytokines for 16 h as indicated. Results are mean Relative Luciferase Unit (R.L.U.) ± SEM. (J–L) Mean optical density measurements of IRF1, phospho-STAT1 and total STAT1 Western blots corrected for α-tubulin (representative figure in H). (G) SOCS-1 mRNA expression was assayed by RT-PCR and normalized for the housekeeping gene GAPDH. Results are mean ± SEM of 4–6 experiments; *: p
Figure Legend Snippet: C/EBPδ silencing increases cytokine-induced chemokine production by enhancing STAT1 activation. INS-1E cells were transfected with either siCtrl (white dots/bars), siC/EBPδ #1 (black triangles/bars) or siC/EBPδ #2 (grey squares/bars) and subsequently left untreated, or treated with IL-1β+IFN-γ for the indicated time points; (A–E) C/EBPδ, CXCL1, 9, 10 CCL20 mRNA expressions were assayed by RT-PCR and normalized for the housekeeping gene GAPDH; (F–G) CXCL1 and CXCL9 protein secretions were evaluated by ELISA. (H) C/EBPδ, phospho-STAT1, total STAT1, IRF-1 and α-tubulin expressions were evaluated by Western blot. (I) 24 h after siRNA transfection, cells were transfected with a STAT1 luciferase reporter+pRL-CMV and subsequently left untreated or exposed to cytokines for 16 h as indicated. Results are mean Relative Luciferase Unit (R.L.U.) ± SEM. (J–L) Mean optical density measurements of IRF1, phospho-STAT1 and total STAT1 Western blots corrected for α-tubulin (representative figure in H). (G) SOCS-1 mRNA expression was assayed by RT-PCR and normalized for the housekeeping gene GAPDH. Results are mean ± SEM of 4–6 experiments; *: p

Techniques Used: Activation Assay, Transfection, Reverse Transcription Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Western Blot, Luciferase, Expressing

9) Product Images from "A TSPO ligand is protective in a mouse model of multiple sclerosis"

Article Title: A TSPO ligand is protective in a mouse model of multiple sclerosis

Journal: EMBO Molecular Medicine

doi: 10.1002/emmm.201202124

Etifoxine modulation of T-cell activity during EAE Spinal cords of vehicle- or etifoxine-treated mice were either used for flow cytometry analysis or ELISA analysis. Cells stained for CD4, CD8, IL-17 and IFN-γ were gated according to fluorescent intensity. Cells were then measured as a percentage of the total cell population. There was a decrease in the percentage of CD4 + (* p = 0.04), CD4 + /IL-17 + (* p = 0.04), CD8 + (* p = 0.01) and CD8 + /IFNγ + (* p = 0.03) cells in the drug-treated group Protein levels of the inflammatory cytokines IL-1β (* p = 0.001), IL-17 (* p = 0.02) and IFN-γ (* p = 0.02) were also measured, and there was a significant decrease in all cytokines in response to etifoxine treatment The p -values were calculated using t -test, n = 8/group.
Figure Legend Snippet: Etifoxine modulation of T-cell activity during EAE Spinal cords of vehicle- or etifoxine-treated mice were either used for flow cytometry analysis or ELISA analysis. Cells stained for CD4, CD8, IL-17 and IFN-γ were gated according to fluorescent intensity. Cells were then measured as a percentage of the total cell population. There was a decrease in the percentage of CD4 + (* p = 0.04), CD4 + /IL-17 + (* p = 0.04), CD8 + (* p = 0.01) and CD8 + /IFNγ + (* p = 0.03) cells in the drug-treated group Protein levels of the inflammatory cytokines IL-1β (* p = 0.001), IL-17 (* p = 0.02) and IFN-γ (* p = 0.02) were also measured, and there was a significant decrease in all cytokines in response to etifoxine treatment The p -values were calculated using t -test, n = 8/group.

Techniques Used: Activity Assay, Mouse Assay, Flow Cytometry, Cytometry, Enzyme-linked Immunosorbent Assay, Staining

10) Product Images from "IL-4 Deficiency Decreases Mortality but Increases Severity of Arthritis in Experimental Group B Streptococcus Infection"

Article Title: IL-4 Deficiency Decreases Mortality but Increases Severity of Arthritis in Experimental Group B Streptococcus Infection

Journal: Mediators of Inflammation

doi: 10.1155/2009/394021

Chemokine production in joints . IL-4-deficient or wt mice infected with 1 × 10 7 GBS/mouse. Supernatants from joint homogenates were collected at the indicated times after infection and assayed for Mip-1 α and MIP-2 by ELISA as detailed in materials and methods (see Section 2 ). Values are the mean ± SD of three separate experiments each consisting of 20 mice per experimental group. Three mice per group were sacrificed at each time point. Statistical differences between the experimental groups were marked with asterisk ( P
Figure Legend Snippet: Chemokine production in joints . IL-4-deficient or wt mice infected with 1 × 10 7 GBS/mouse. Supernatants from joint homogenates were collected at the indicated times after infection and assayed for Mip-1 α and MIP-2 by ELISA as detailed in materials and methods (see Section 2 ). Values are the mean ± SD of three separate experiments each consisting of 20 mice per experimental group. Three mice per group were sacrificed at each time point. Statistical differences between the experimental groups were marked with asterisk ( P

Techniques Used: Mouse Assay, Infection, Enzyme-linked Immunosorbent Assay

11) Product Images from "The Inflammatory Response to Enterotoxigenic E. coli and Probiotic E. faecium in a Coculture Model of Porcine Intestinal Epithelial and Dendritic Cells"

Article Title: The Inflammatory Response to Enterotoxigenic E. coli and Probiotic E. faecium in a Coculture Model of Porcine Intestinal Epithelial and Dendritic Cells

Journal: Mediators of Inflammation

doi: 10.1155/2018/9368295

Protein expression (in pg/ml) of (a) IL-8 and (b) TSLP detected by ELISA in supernatants of IPEC-J2 cells after stimulation with either E. faecium ( Ecf ) or ETEC. In IPEC-J2/MoDC cocultures, Ecf or ETEC was added either to the apical side of IPEC-J2 cells or to the MoDC compartment. In IPEC-J2 monocultures, the bacteria were added to the apical compartment. Samples were taken at 6 h after addition of bacteria (means ± SEM). N = 3 independent experiments per bar. Results of the ANOVA are indicated below each graph. Results of post hoc tests are presented in Supplementary Table 4 .
Figure Legend Snippet: Protein expression (in pg/ml) of (a) IL-8 and (b) TSLP detected by ELISA in supernatants of IPEC-J2 cells after stimulation with either E. faecium ( Ecf ) or ETEC. In IPEC-J2/MoDC cocultures, Ecf or ETEC was added either to the apical side of IPEC-J2 cells or to the MoDC compartment. In IPEC-J2 monocultures, the bacteria were added to the apical compartment. Samples were taken at 6 h after addition of bacteria (means ± SEM). N = 3 independent experiments per bar. Results of the ANOVA are indicated below each graph. Results of post hoc tests are presented in Supplementary Table 4 .

Techniques Used: Expressing, Enzyme-linked Immunosorbent Assay

Protein expression (in pg/ml) of (a) IL-1 β , (b) IL-8, (c) TGF- β , and (d) TSLP detected by ELISA in supernatants of porcine MoDC after stimulation with either E. faecium ( Ecf ) or ETEC. In IPEC-J2/MoDC cocultures, Ecf or ETEC was added either to the apical side of IPEC-J2 cells or to the MoDC compartment. In MoDC monocultures, the bacteria were added to the basolateral compartment. Samples were taken at 6 h after addition of bacteria (means ± SEM). N = 3 − 4 independent experiments per bar. Results of the ANOVA are indicated below each graph. Results of post hoc tests are presented in Supplementary Table 5 .
Figure Legend Snippet: Protein expression (in pg/ml) of (a) IL-1 β , (b) IL-8, (c) TGF- β , and (d) TSLP detected by ELISA in supernatants of porcine MoDC after stimulation with either E. faecium ( Ecf ) or ETEC. In IPEC-J2/MoDC cocultures, Ecf or ETEC was added either to the apical side of IPEC-J2 cells or to the MoDC compartment. In MoDC monocultures, the bacteria were added to the basolateral compartment. Samples were taken at 6 h after addition of bacteria (means ± SEM). N = 3 − 4 independent experiments per bar. Results of the ANOVA are indicated below each graph. Results of post hoc tests are presented in Supplementary Table 5 .

Techniques Used: Expressing, Enzyme-linked Immunosorbent Assay

12) Product Images from "The protective effect of sinapic acid in osteoarthritis: In vitro and in vivo studies, et al. The protective effect of sinapic acid in osteoarthritis: In vitro and in vivo studies"

Article Title: The protective effect of sinapic acid in osteoarthritis: In vitro and in vivo studies, et al. The protective effect of sinapic acid in osteoarthritis: In vitro and in vivo studies

Journal: Journal of Cellular and Molecular Medicine

doi: 10.1111/jcmm.14096

Effect of SA on Nrf2/HO‐1 pathway. Chondrocytes were pre‐treated for 1 hour with various concentrations of SA (40, 80, and 160 μmol/L), followed by stimulation with or without IL‐1β (10 ng/mL) for 2 hours. Nrf2 activation (in nucleus) and HO‐1 (in cytoplasm) were determined by Western blot (A) and quantification analysis (B). After Nrf2 knock down, the protein expressions of Nrf2 and p65 in nuclear and HO‐1 in cytoplasm in chondrocytes treated as above were visualized by Western blot (C), and quantified in (D). The production of PGE2, MMP‐13 and collagen‐ll was assessed by ELISA, and the expression of NO was assessed using Griess reagent (E). The values are mean ± SD of three independent experiments. ## P
Figure Legend Snippet: Effect of SA on Nrf2/HO‐1 pathway. Chondrocytes were pre‐treated for 1 hour with various concentrations of SA (40, 80, and 160 μmol/L), followed by stimulation with or without IL‐1β (10 ng/mL) for 2 hours. Nrf2 activation (in nucleus) and HO‐1 (in cytoplasm) were determined by Western blot (A) and quantification analysis (B). After Nrf2 knock down, the protein expressions of Nrf2 and p65 in nuclear and HO‐1 in cytoplasm in chondrocytes treated as above were visualized by Western blot (C), and quantified in (D). The production of PGE2, MMP‐13 and collagen‐ll was assessed by ELISA, and the expression of NO was assessed using Griess reagent (E). The values are mean ± SD of three independent experiments. ## P

Techniques Used: Activation Assay, Western Blot, Enzyme-linked Immunosorbent Assay, Expressing

Effect of SA on extracellular matrix degradation from IL‐1β‐induced human OA chondrocytes. Chondrocytes were pre‐treated for 1 hour with various concentrations of SA (40, 80, and 160 μmol/L), followed by stimulation with or without IL‐1β (10 ng/mL) for 24 hours. The mRNA expression levels of MMP‐13, ADAMTS‐5 (A), aggrecan (E) and collagen‐II (F) were assayed by qRT‐PCR. The protein expression levels of MMP‐13, ADAMTS‐5 were determined by ELISA (B). The protein expression levels of collagen‐II (C) and MMP‐13 (D) were determined by immunofluorescence. The values are mean ± SD of three independent experiments. ## P
Figure Legend Snippet: Effect of SA on extracellular matrix degradation from IL‐1β‐induced human OA chondrocytes. Chondrocytes were pre‐treated for 1 hour with various concentrations of SA (40, 80, and 160 μmol/L), followed by stimulation with or without IL‐1β (10 ng/mL) for 24 hours. The mRNA expression levels of MMP‐13, ADAMTS‐5 (A), aggrecan (E) and collagen‐II (F) were assayed by qRT‐PCR. The protein expression levels of MMP‐13, ADAMTS‐5 were determined by ELISA (B). The protein expression levels of collagen‐II (C) and MMP‐13 (D) were determined by immunofluorescence. The values are mean ± SD of three independent experiments. ## P

Techniques Used: Expressing, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Immunofluorescence

13) Product Images from "Sex-related survival differences in murine cardiomyopathy are associated with differences in TNF-receptor expression"

Article Title: Sex-related survival differences in murine cardiomyopathy are associated with differences in TNF-receptor expression

Journal: Journal of Clinical Investigation

doi:

Cytokine expression in the myocardium. ( a – e ) Representative images ( a ) and quantitative results of RNase protection assays for TNF-α ( b ), IL-1β ( c ), MCP-1 ( d ), and TGF-β1 ( e ). Quantitative results are reported as the ratio of the pixel intensity of each protected probe normalized to that of the GAPDH probe included in each template set as an internal control, as described in Methods. Although TNF1.6 mice expressed significant amounts of TNF-α and other cytokines, the transcript levels of TNF-α and other downstream cytokines were not significantly different between male and female mice. Y -axis values correspond to the signal intensity of the hybridized probe in pixels normalized to that of GAPDH. ( f – h ) Results of ELISA for TNF-α ( f ), IL-1β ( g ), and MCP-1 ( h ) showed significantly elevated levels in TNF1.6 versus wild-type mice, but no difference between sexes of the same genotype. Values are mean ± SD ( n = 6). TG, TNF1.6 mice; WT, wild-type mice; M, male; F, female. A P
Figure Legend Snippet: Cytokine expression in the myocardium. ( a – e ) Representative images ( a ) and quantitative results of RNase protection assays for TNF-α ( b ), IL-1β ( c ), MCP-1 ( d ), and TGF-β1 ( e ). Quantitative results are reported as the ratio of the pixel intensity of each protected probe normalized to that of the GAPDH probe included in each template set as an internal control, as described in Methods. Although TNF1.6 mice expressed significant amounts of TNF-α and other cytokines, the transcript levels of TNF-α and other downstream cytokines were not significantly different between male and female mice. Y -axis values correspond to the signal intensity of the hybridized probe in pixels normalized to that of GAPDH. ( f – h ) Results of ELISA for TNF-α ( f ), IL-1β ( g ), and MCP-1 ( h ) showed significantly elevated levels in TNF1.6 versus wild-type mice, but no difference between sexes of the same genotype. Values are mean ± SD ( n = 6). TG, TNF1.6 mice; WT, wild-type mice; M, male; F, female. A P

Techniques Used: Expressing, Mouse Assay, Enzyme-linked Immunosorbent Assay

14) Product Images from "NGF Activation of TrkA Induces Vascular Endothelial Growth Factor Expression via induction of Hypoxia-Inducible Factor-1?"

Article Title: NGF Activation of TrkA Induces Vascular Endothelial Growth Factor Expression via induction of Hypoxia-Inducible Factor-1?

Journal: Molecular and cellular neurosciences

doi: 10.1016/j.mcn.2010.12.002

NGF regulates HIF-1 α and VEGF levels via PI3K/mTOR and MAPK pathways Panel A. Western analysis of 30 μg of HIF-1α levels in protein lysates from the serum-deprived (6h) high TrkA-expressing TA25 cells (TET−) pretreated with control solvent or indicated inhibitor for 1 h followed by stimulation with NGF for 8 h. HIF-1β is shown as a loading control. Panel B. Conditioned media from the high TrkA-expressing TA25 cells (TET−) (1 × 10 6 ) incubated with serum-free media for 6 h following a 1 h pre-incubation with control solvent or indicated inhibitor was analyzed by ELISA for VEGF expression. Results represent mean + standard deviation of triplicate values for each condition (*: p
Figure Legend Snippet: NGF regulates HIF-1 α and VEGF levels via PI3K/mTOR and MAPK pathways Panel A. Western analysis of 30 μg of HIF-1α levels in protein lysates from the serum-deprived (6h) high TrkA-expressing TA25 cells (TET−) pretreated with control solvent or indicated inhibitor for 1 h followed by stimulation with NGF for 8 h. HIF-1β is shown as a loading control. Panel B. Conditioned media from the high TrkA-expressing TA25 cells (TET−) (1 × 10 6 ) incubated with serum-free media for 6 h following a 1 h pre-incubation with control solvent or indicated inhibitor was analyzed by ELISA for VEGF expression. Results represent mean + standard deviation of triplicate values for each condition (*: p

Techniques Used: Western Blot, Expressing, Incubation, Enzyme-linked Immunosorbent Assay, Standard Deviation

15) Product Images from "Recombinant human IL-26 facilitates the innate immune response to endotoxin in the bronchoalveolar space of mice in vivo"

Article Title: Recombinant human IL-26 facilitates the innate immune response to endotoxin in the bronchoalveolar space of mice in vivo

Journal: PLoS ONE

doi: 10.1371/journal.pone.0188909

Effects of rhIL-26 on lung tissue inflammation and MPO protein content. Mice received intranasal instillation of recombinant human (rh) IL-26 protein or its vechicle (PBS), with or without prior instillation of endotoxin (LPS) or its vehicle (PBS). Lung tissue samples were harvested 6 h (n = 8), 24 h (n = 10) and 72 h (n = 8) instillations. Histological evaluations were performed on lung tissue sections stained with hematoxylin and eosin (H E) and quantified with a scoring system. Representative H E-staining pictures (magnification: 10×, scale bar: 50 μm) are shown (A) . The inflammation in four tissue compartments (the interstitial area, alveolar space, peribronchial area and perivascular area) on lung tissue sections was evaluated and scored (see Methods section) at (B) 6 h, (C) 24 h, and (D) 72 h after the intranasal instillation, respectively. The MPO protein in lung tissue was analyzed using immunoblot. The bands of each mouse and the densitometry ratio of MPO to actin are shown in (E) . Each panel relates to a time-point and each lane of the bands represents a separate blot. (F) The MPO content in cell-free BAL fluid samples was quantified using ELISA. Data are presented as mean ± SEM.
Figure Legend Snippet: Effects of rhIL-26 on lung tissue inflammation and MPO protein content. Mice received intranasal instillation of recombinant human (rh) IL-26 protein or its vechicle (PBS), with or without prior instillation of endotoxin (LPS) or its vehicle (PBS). Lung tissue samples were harvested 6 h (n = 8), 24 h (n = 10) and 72 h (n = 8) instillations. Histological evaluations were performed on lung tissue sections stained with hematoxylin and eosin (H E) and quantified with a scoring system. Representative H E-staining pictures (magnification: 10×, scale bar: 50 μm) are shown (A) . The inflammation in four tissue compartments (the interstitial area, alveolar space, peribronchial area and perivascular area) on lung tissue sections was evaluated and scored (see Methods section) at (B) 6 h, (C) 24 h, and (D) 72 h after the intranasal instillation, respectively. The MPO protein in lung tissue was analyzed using immunoblot. The bands of each mouse and the densitometry ratio of MPO to actin are shown in (E) . Each panel relates to a time-point and each lane of the bands represents a separate blot. (F) The MPO content in cell-free BAL fluid samples was quantified using ELISA. Data are presented as mean ± SEM.

Techniques Used: Mouse Assay, Recombinant, Staining, Enzyme-linked Immunosorbent Assay

Effects of rhIL-26 on the activity and the quantity of proteinases in BAL samples. Mice received intranasal instillation of recombinant human (rh) IL-26 protein, or its vehicle (PBS), with or without prior instillation of endotoxin (LPS). Bronchoalveolar lavage (BAL) samples were harvested 6 h (n = 8), 24 h (n = 10) and 72 h (n = 8) after instillations. The total activity of gelatinase (A) was measured with zymography in BAL samples that were pooled for each study group: the left panel presents the bands for MMP-9 and MMP-2 detected at all three time-points and the right panel shows the bands densitometry data of MMP-2 (upwards) and MMP-9 (downwards) (A) . The respective net activity of (B) gelatinase and (F) elastase in the same BAL samples was measured utilizing a substrate-based method. The extracellular concentrations of gelatinases including (C) MMP-2, (D) MMP-9, (E) pro- MMP-9, and serine proteinases including (G) NE and (H) Cathepsin G (only in mice receiving instillation of LPS) were quantified in BAL samples using ELISA. Data are presented as mean.
Figure Legend Snippet: Effects of rhIL-26 on the activity and the quantity of proteinases in BAL samples. Mice received intranasal instillation of recombinant human (rh) IL-26 protein, or its vehicle (PBS), with or without prior instillation of endotoxin (LPS). Bronchoalveolar lavage (BAL) samples were harvested 6 h (n = 8), 24 h (n = 10) and 72 h (n = 8) after instillations. The total activity of gelatinase (A) was measured with zymography in BAL samples that were pooled for each study group: the left panel presents the bands for MMP-9 and MMP-2 detected at all three time-points and the right panel shows the bands densitometry data of MMP-2 (upwards) and MMP-9 (downwards) (A) . The respective net activity of (B) gelatinase and (F) elastase in the same BAL samples was measured utilizing a substrate-based method. The extracellular concentrations of gelatinases including (C) MMP-2, (D) MMP-9, (E) pro- MMP-9, and serine proteinases including (G) NE and (H) Cathepsin G (only in mice receiving instillation of LPS) were quantified in BAL samples using ELISA. Data are presented as mean.

Techniques Used: Activity Assay, Mouse Assay, Recombinant, Zymography, Enzyme-linked Immunosorbent Assay

Effects of rhIL-26 on extracellular and intracellular proteinases in relation to innate effector cells in BAL samples. Mice received prior intranasal instillation of endotoxin (LPS) with or without the subsequent addition of instillation of rhIL-26 protein or its vehicle (PBS) and bronchoalveolar lavage (BAL) samples were harvested. The average contents of extracellular proteinases per leukocyte in BAL samples were calculated to evaluate the activity of innate inflammatory cells. The intracellular contents of proteinases were measured using ELISA in whole cell lysates of BAL samples that were pooled for each study group. The panels show (A) MMP-2 per macrophage, (B) MMP-9 per leukocyte, (C) MPO per leukocyte, (D) NE per neutrophil, (E) intracellular MMP-2 per macrophage, (F) intracellular MMP-9 per leukocyte, (G) intracellular MPO per leukocyte and (H) intracellular NE per neutrophil. Data are presented as mean ± SEM.
Figure Legend Snippet: Effects of rhIL-26 on extracellular and intracellular proteinases in relation to innate effector cells in BAL samples. Mice received prior intranasal instillation of endotoxin (LPS) with or without the subsequent addition of instillation of rhIL-26 protein or its vehicle (PBS) and bronchoalveolar lavage (BAL) samples were harvested. The average contents of extracellular proteinases per leukocyte in BAL samples were calculated to evaluate the activity of innate inflammatory cells. The intracellular contents of proteinases were measured using ELISA in whole cell lysates of BAL samples that were pooled for each study group. The panels show (A) MMP-2 per macrophage, (B) MMP-9 per leukocyte, (C) MPO per leukocyte, (D) NE per neutrophil, (E) intracellular MMP-2 per macrophage, (F) intracellular MMP-9 per leukocyte, (G) intracellular MPO per leukocyte and (H) intracellular NE per neutrophil. Data are presented as mean ± SEM.

Techniques Used: Mouse Assay, Activity Assay, Enzyme-linked Immunosorbent Assay

16) Product Images from "Changes in the Anti-Allergic Activities of Sesame by Bioconversion"

Article Title: Changes in the Anti-Allergic Activities of Sesame by Bioconversion

Journal: Nutrients

doi: 10.3390/nu10020210

Effects of NS and FS extracts on TNF-α/IFN-γ-induced IL-1β, IL-6, TARC, and MDC production in HaCaT cells. The production of IL-1β ( a ); IL-6 ( b ); TARC ( c ); MDC ( d ) protein was measured by ELISA. Each bar represents the mean ± SD of triplicate determinations, n = 3; * p
Figure Legend Snippet: Effects of NS and FS extracts on TNF-α/IFN-γ-induced IL-1β, IL-6, TARC, and MDC production in HaCaT cells. The production of IL-1β ( a ); IL-6 ( b ); TARC ( c ); MDC ( d ) protein was measured by ELISA. Each bar represents the mean ± SD of triplicate determinations, n = 3; * p

Techniques Used: Enzyme-linked Immunosorbent Assay

17) Product Images from "Simvastatin treatment boosts benefits of apoptotic cell infusion in murine lung fibrosis"

Article Title: Simvastatin treatment boosts benefits of apoptotic cell infusion in murine lung fibrosis

Journal: Cell Death & Disease

doi: 10.1038/cddis.2017.260

Expression of pro-resolving cytokines after apoptotic cell instillation with or without simvastatin. Apoptotic Jurkat cells (Apo) were once instillated on day 2 or twice on days 2 and 7 after bleomycin (BLM) treatment. Simvastatin (Simv; 20 mg/kg/d, i.p) or its vehicle (Veh; 2% DMSO in saline) was administered with or without second apoptotic cell instillation and every day thereafter. Mice were killed on days 7 (2 h after second apoptotic cell or simvastatin treatment) and 14 following BLM treatment. ( a ) HGF, ( c ) IL-10, and ( e ) TGF- β 1 mRNA levels in alveolar macrophages were analyzed by quantitative real-time PCR. The levels of ( b ) HGF, ( d ) IL-10, and ( f ) TGF- β 1 in BAL fluid were quantified by ELISA. Values represent the mean±S.E.M. of results from five mice per group. * P
Figure Legend Snippet: Expression of pro-resolving cytokines after apoptotic cell instillation with or without simvastatin. Apoptotic Jurkat cells (Apo) were once instillated on day 2 or twice on days 2 and 7 after bleomycin (BLM) treatment. Simvastatin (Simv; 20 mg/kg/d, i.p) or its vehicle (Veh; 2% DMSO in saline) was administered with or without second apoptotic cell instillation and every day thereafter. Mice were killed on days 7 (2 h after second apoptotic cell or simvastatin treatment) and 14 following BLM treatment. ( a ) HGF, ( c ) IL-10, and ( e ) TGF- β 1 mRNA levels in alveolar macrophages were analyzed by quantitative real-time PCR. The levels of ( b ) HGF, ( d ) IL-10, and ( f ) TGF- β 1 in BAL fluid were quantified by ELISA. Values represent the mean±S.E.M. of results from five mice per group. * P

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

18) Product Images from "Pinus densiflora needle supercritical fluid extract suppresses the expression of pro-inflammatory mediators iNOS, IL-6 and IL-1β, and activation of inflammatory STAT1 and STAT3 signaling proteins in bacterial lipopolysaccharide-challenged murine macrophages"

Article Title: Pinus densiflora needle supercritical fluid extract suppresses the expression of pro-inflammatory mediators iNOS, IL-6 and IL-1β, and activation of inflammatory STAT1 and STAT3 signaling proteins in bacterial lipopolysaccharide-challenged murine macrophages

Journal: DARU Journal of Pharmaceutical Sciences

doi: 10.1186/s40199-017-0184-y

Effect of PDN-SCFE on LPS-induced cytokine ( a ) mRNA and ( b ) protein expression in RAW 264.7 cells. The cells were pre-treated with indicated concentrations of PDN-SCFE for 2 h, followed by LPS for 6 h (a, mRNA expression) or 18 h (b, protein expression) The cell-free supernatant was subjected to quantify TNFα, IL-6 and IL-1β levels, using ELISA kits, according to manufacturer’s protocol. Total RNA was extracted with Trizol reagent and subjected to qRT-PCR according to the manufacturer’s instructions. The graphical figures represent the change in mRNA expression normalized to actin. Data represent mean ± SD from three separate experiments. *** P
Figure Legend Snippet: Effect of PDN-SCFE on LPS-induced cytokine ( a ) mRNA and ( b ) protein expression in RAW 264.7 cells. The cells were pre-treated with indicated concentrations of PDN-SCFE for 2 h, followed by LPS for 6 h (a, mRNA expression) or 18 h (b, protein expression) The cell-free supernatant was subjected to quantify TNFα, IL-6 and IL-1β levels, using ELISA kits, according to manufacturer’s protocol. Total RNA was extracted with Trizol reagent and subjected to qRT-PCR according to the manufacturer’s instructions. The graphical figures represent the change in mRNA expression normalized to actin. Data represent mean ± SD from three separate experiments. *** P

Techniques Used: Expressing, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR

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Article Title: Inhibiting nucleolin reduces inflammation induced by mitochondrial DNA in cardiomyocytes exposed to hypoxia and reoxygenation, et al. Inhibiting nucleolin reduce inflammation induced by mitochondrial DNA in cardiomyocytes exposed to hypoxia and reoxygenation
Article Snippet: Reversible Ponceau staining was used as a loading control. .. 2.12 ELISA elisa kits were purchased from R & D systems (R & D Systems, Inc., Minneapolis, USA), and the protocol was conducted according to the manufacturer. ..

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    R&D Systems sandwich elisa kit
    Flow reduction is associated with cytokine production within the vessel wall. (a) Transcripts for <t>IL-1β,</t> IL-6, IP-10, Mig, TNF-α, and iNOS were quantified by qRT-PCR and normalized to GAPDH from right (R) and left (L) common carotid artery lysates of WT versus MyD88 −/− mice at 6 h ( n = 10), 1 d ( n = 8), 3 d ( n = 9), 7 d ( n = 11), and 14 d ( n = 6) after left external carotid artery ligation. (b) IL-1β protein was measured by <t>ELISA</t> in common carotid artery lysates and plasma at 24 h after sham operations or left external carotid artery ligation ( n = 6). Data are means ± SE, *P
    Sandwich Elisa Kit, supplied by R&D Systems, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    R&D Systems enzyme linked immuno sorbent assay elisa kit
    NE-induced PlGF expression and secretion are mediated by Egr-1. (A-C) The mRNA level of PlGF was determined by reverse-transcriptional polymerase chain reaction (RT-PCR) with primer sets for PlGF and GAPDH cDNA. (D-F) Cellular lysates were also subjected to Western blot analysis with antibodies for PlGF and β-actin. (G-I) The PlGF in the culture medium was detected by enzyme-linked <t>immuno-sorbent</t> assay <t>(ELISA).</t> Data are presented as mean ± SEM. * p
    Enzyme Linked Immuno Sorbent Assay Elisa Kit, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/enzyme linked immuno sorbent assay elisa kit/product/R&D Systems
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    Flow reduction is associated with cytokine production within the vessel wall. (a) Transcripts for IL-1β, IL-6, IP-10, Mig, TNF-α, and iNOS were quantified by qRT-PCR and normalized to GAPDH from right (R) and left (L) common carotid artery lysates of WT versus MyD88 −/− mice at 6 h ( n = 10), 1 d ( n = 8), 3 d ( n = 9), 7 d ( n = 11), and 14 d ( n = 6) after left external carotid artery ligation. (b) IL-1β protein was measured by ELISA in common carotid artery lysates and plasma at 24 h after sham operations or left external carotid artery ligation ( n = 6). Data are means ± SE, *P

    Journal: The Journal of Experimental Medicine

    Article Title: MyD88-dependent, superoxide-initiated inflammation is necessary for flow-mediated inward remodeling of conduit arteries

    doi: 10.1084/jem.20081298

    Figure Lengend Snippet: Flow reduction is associated with cytokine production within the vessel wall. (a) Transcripts for IL-1β, IL-6, IP-10, Mig, TNF-α, and iNOS were quantified by qRT-PCR and normalized to GAPDH from right (R) and left (L) common carotid artery lysates of WT versus MyD88 −/− mice at 6 h ( n = 10), 1 d ( n = 8), 3 d ( n = 9), 7 d ( n = 11), and 14 d ( n = 6) after left external carotid artery ligation. (b) IL-1β protein was measured by ELISA in common carotid artery lysates and plasma at 24 h after sham operations or left external carotid artery ligation ( n = 6). Data are means ± SE, *P

    Article Snippet: Plasma and carotid lysate levels of murine IL-1β were determined using a sandwich ELISA kit (R & D Systems).

    Techniques: Flow Cytometry, Quantitative RT-PCR, Mouse Assay, Ligation, Enzyme-linked Immunosorbent Assay

    IR induces HIF-1a and its two target genes BNIP3 and NIX in rat brain (A-C) The levels of HIF-1a (by ELISA, A), BNIP3 and NIX (by Western blot, B-C) were examined in rat brain tissue (Sham group, IR+HC group and IR+HC-HIF-1a group) after IR. N=20. * p

    Journal: Oncotarget

    Article Title: Role of HIF-1a in regulating autophagic cell survival during cerebral ischemia reperfusion in rats

    doi: 10.18632/oncotarget.21445

    Figure Lengend Snippet: IR induces HIF-1a and its two target genes BNIP3 and NIX in rat brain (A-C) The levels of HIF-1a (by ELISA, A), BNIP3 and NIX (by Western blot, B-C) were examined in rat brain tissue (Sham group, IR+HC group and IR+HC-HIF-1a group) after IR. N=20. * p

    Article Snippet: HIF-1a levels were determined by an ELISA kit (R & D System, Los Angeles, CA, USA).

    Techniques: Enzyme-linked Immunosorbent Assay, Western Blot

    Beneficial effects of HIF-1a during IR (A) Rat hematopoietic cells (HCs) were transduced them with lentivirus carrying either null (as a control) or recombinant HIF-1a under a CMV promoter. HIF-1a levels in the transduced HCs were determined by HIF-1a ELISA. (B-C) For HC reinfusion, 10 7 donor null/HIF-1a-transduced HCs were given to the receipt rats 3 days before IR. The rats were randomly divided into 3 groups of 20 each: sham-treated (no IR; Sham); IR in rats that received 10 7 donor null-transduced HCs 3 days before IR (IR-HC); IR in rats that received 10 7 donor HIF-1a-transduced HCs 3 days before IR (IR-HC-HIF-1a). Histopathologic score (HPS) was done, shown by quantification (B), and by representative images (C). (D) The neurologic scores in 3 groups. (E) The brain water content in 3 groups. (F) The Brain-Blood Barrier (BBB) permeability was evaluated, using EB extravasation. N=20. * p

    Journal: Oncotarget

    Article Title: Role of HIF-1a in regulating autophagic cell survival during cerebral ischemia reperfusion in rats

    doi: 10.18632/oncotarget.21445

    Figure Lengend Snippet: Beneficial effects of HIF-1a during IR (A) Rat hematopoietic cells (HCs) were transduced them with lentivirus carrying either null (as a control) or recombinant HIF-1a under a CMV promoter. HIF-1a levels in the transduced HCs were determined by HIF-1a ELISA. (B-C) For HC reinfusion, 10 7 donor null/HIF-1a-transduced HCs were given to the receipt rats 3 days before IR. The rats were randomly divided into 3 groups of 20 each: sham-treated (no IR; Sham); IR in rats that received 10 7 donor null-transduced HCs 3 days before IR (IR-HC); IR in rats that received 10 7 donor HIF-1a-transduced HCs 3 days before IR (IR-HC-HIF-1a). Histopathologic score (HPS) was done, shown by quantification (B), and by representative images (C). (D) The neurologic scores in 3 groups. (E) The brain water content in 3 groups. (F) The Brain-Blood Barrier (BBB) permeability was evaluated, using EB extravasation. N=20. * p

    Article Snippet: HIF-1a levels were determined by an ELISA kit (R & D System, Los Angeles, CA, USA).

    Techniques: Recombinant, Enzyme-linked Immunosorbent Assay, Permeability

    Effects of TLR agonists on TNFα ( A ) and VEGF ( B ) protein production by macrophages from C57BL/10ScN (TLR4-deficient) mice. Macrophages were treated with TLR agonists and controls at the indicated concentrations. NECA (an A 2 R agonist) and polymyxin B were added to the cells at the same time as the TLR agonists. Conditioned media were harvested 24 hours after addition of test agents and the TNFα and VEGF levels in the media were determined by ELISA. At least two replicates were studied for each condition, and each sample was analyzed for TNFα and VEGF in duplicate. Results are presented as means ± SD. The x-axis labels refer to both TNFα ( A ) and VEGF ( B ). The particular TLR studied in each group is indicated beneath the x-axis labels. An asterisk (*) indicates undetectable levels.

    Journal: The American Journal of Pathology

    Article Title: An Angiogenic Switch in Macrophages Involving Synergy between Toll-Like Receptors 2, 4, 7, and 9 and Adenosine A2A Receptors

    doi:

    Figure Lengend Snippet: Effects of TLR agonists on TNFα ( A ) and VEGF ( B ) protein production by macrophages from C57BL/10ScN (TLR4-deficient) mice. Macrophages were treated with TLR agonists and controls at the indicated concentrations. NECA (an A 2 R agonist) and polymyxin B were added to the cells at the same time as the TLR agonists. Conditioned media were harvested 24 hours after addition of test agents and the TNFα and VEGF levels in the media were determined by ELISA. At least two replicates were studied for each condition, and each sample was analyzed for TNFα and VEGF in duplicate. Results are presented as means ± SD. The x-axis labels refer to both TNFα ( A ) and VEGF ( B ). The particular TLR studied in each group is indicated beneath the x-axis labels. An asterisk (*) indicates undetectable levels.

    Article Snippet: TNFα levels in macrophage conditioned media were determined using a sandwich ELISA kit (R & D Systems), as described by the manufacturer.

    Techniques: Mouse Assay, Enzyme-linked Immunosorbent Assay

    Effects of TLR agonists on TNFα ( A ) and VEGF ( B ) production by macrophages from C57BL/10ScSn (TLR4+/+) mice. Macrophages were treated with TLR agonists and controls at the indicated concentrations. NECA (an A 2 R agonist) and polymyxin B were added to the cells at the same time as the TLR agonists. Conditioned media were harvested 24 hours after addition of test agents and the TNFα and VEGF levels in the media were determined by ELISA. At least two replicates were studied for each condition, and each sample was analyzed for TNFα and VEGF in duplicate. Results are presented as means ± SD. The x-axis labels refer to both TNFα ( A ) and VEGF ( B ). The particular TLR studied in each group is indicated beneath the x-axis labels. An asterisk (*) indicates undetectable levels.

    Journal: The American Journal of Pathology

    Article Title: An Angiogenic Switch in Macrophages Involving Synergy between Toll-Like Receptors 2, 4, 7, and 9 and Adenosine A2A Receptors

    doi:

    Figure Lengend Snippet: Effects of TLR agonists on TNFα ( A ) and VEGF ( B ) production by macrophages from C57BL/10ScSn (TLR4+/+) mice. Macrophages were treated with TLR agonists and controls at the indicated concentrations. NECA (an A 2 R agonist) and polymyxin B were added to the cells at the same time as the TLR agonists. Conditioned media were harvested 24 hours after addition of test agents and the TNFα and VEGF levels in the media were determined by ELISA. At least two replicates were studied for each condition, and each sample was analyzed for TNFα and VEGF in duplicate. Results are presented as means ± SD. The x-axis labels refer to both TNFα ( A ) and VEGF ( B ). The particular TLR studied in each group is indicated beneath the x-axis labels. An asterisk (*) indicates undetectable levels.

    Article Snippet: TNFα levels in macrophage conditioned media were determined using a sandwich ELISA kit (R & D Systems), as described by the manufacturer.

    Techniques: Mouse Assay, Enzyme-linked Immunosorbent Assay

    Effects of adenosine and an adenosine deaminase inhibitor (EHNA) on production of VEGF ( A ) and TNFα ( B ) by macrophages from C57Bl/10ScSn mice. Macrophages were treated with adenosine (A) (100 μmol/L), NECA (N) (1 μmol/L), LPS (L) (100 ng/ml), or EHNA at the indicated concentrations, either alone or in combination. Conditioned media were harvested 24 hours after addition of test agents and the TNFα and VEGF levels in the media were determined by ELISA. At least two replicates were studied for each condition, and each sample was analyzed for TNFα and VEGF in duplicate. Results are presented as means ± SD. An asterisk (*) indicates undetectable levels.

    Journal: The American Journal of Pathology

    Article Title: An Angiogenic Switch in Macrophages Involving Synergy between Toll-Like Receptors 2, 4, 7, and 9 and Adenosine A2A Receptors

    doi:

    Figure Lengend Snippet: Effects of adenosine and an adenosine deaminase inhibitor (EHNA) on production of VEGF ( A ) and TNFα ( B ) by macrophages from C57Bl/10ScSn mice. Macrophages were treated with adenosine (A) (100 μmol/L), NECA (N) (1 μmol/L), LPS (L) (100 ng/ml), or EHNA at the indicated concentrations, either alone or in combination. Conditioned media were harvested 24 hours after addition of test agents and the TNFα and VEGF levels in the media were determined by ELISA. At least two replicates were studied for each condition, and each sample was analyzed for TNFα and VEGF in duplicate. Results are presented as means ± SD. An asterisk (*) indicates undetectable levels.

    Article Snippet: TNFα levels in macrophage conditioned media were determined using a sandwich ELISA kit (R & D Systems), as described by the manufacturer.

    Techniques: Mouse Assay, Enzyme-linked Immunosorbent Assay

    NE-induced PlGF expression and secretion are mediated by Egr-1. (A-C) The mRNA level of PlGF was determined by reverse-transcriptional polymerase chain reaction (RT-PCR) with primer sets for PlGF and GAPDH cDNA. (D-F) Cellular lysates were also subjected to Western blot analysis with antibodies for PlGF and β-actin. (G-I) The PlGF in the culture medium was detected by enzyme-linked immuno-sorbent assay (ELISA). Data are presented as mean ± SEM. * p

    Journal: Respiratory Research

    Article Title: PlGF mediates neutrophil elastase-induced airway epithelial cell apoptosis and emphysema

    doi: 10.1186/s12931-014-0106-1

    Figure Lengend Snippet: NE-induced PlGF expression and secretion are mediated by Egr-1. (A-C) The mRNA level of PlGF was determined by reverse-transcriptional polymerase chain reaction (RT-PCR) with primer sets for PlGF and GAPDH cDNA. (D-F) Cellular lysates were also subjected to Western blot analysis with antibodies for PlGF and β-actin. (G-I) The PlGF in the culture medium was detected by enzyme-linked immuno-sorbent assay (ELISA). Data are presented as mean ± SEM. * p

    Article Snippet: Human and mouse recombinant PlGF protein and an enzyme-linked immuno-sorbent assay (ELISA) kit were obtained from R and D Systems (Minneapolis, MN, USA).

    Techniques: Expressing, Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction, Western Blot, Enzyme-linked Immunosorbent Assay

    NE increases expression of PlGF and activation of JNK and PKCδ signaling. (A and B) Paraffin-embedded lung tissue sections were used for immuno-histochemistry (IHC) analysis and incubated with antibodies of PlGF, p-JNK and p-PKCδ. Arrow heads indicated positive stain of PlGF, p-JNK and p-PKCδ in LE cells. (C) Mice broncho-alveolar lavage fluid was analyzed for the PlGF level by ELISA. Scale bar = 200 μm. Data are presented as mean ± SEM. * p

    Journal: Respiratory Research

    Article Title: PlGF mediates neutrophil elastase-induced airway epithelial cell apoptosis and emphysema

    doi: 10.1186/s12931-014-0106-1

    Figure Lengend Snippet: NE increases expression of PlGF and activation of JNK and PKCδ signaling. (A and B) Paraffin-embedded lung tissue sections were used for immuno-histochemistry (IHC) analysis and incubated with antibodies of PlGF, p-JNK and p-PKCδ. Arrow heads indicated positive stain of PlGF, p-JNK and p-PKCδ in LE cells. (C) Mice broncho-alveolar lavage fluid was analyzed for the PlGF level by ELISA. Scale bar = 200 μm. Data are presented as mean ± SEM. * p

    Article Snippet: Human and mouse recombinant PlGF protein and an enzyme-linked immuno-sorbent assay (ELISA) kit were obtained from R and D Systems (Minneapolis, MN, USA).

    Techniques: Expressing, Activation Assay, Immunohistochemistry, Incubation, Staining, Mouse Assay, Enzyme-linked Immunosorbent Assay