CTRP3 ameliorates cerulein-induced severe acute pancreatitis in mice via SIRT1/NF-κB/p53 axis

Figures

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  Figure 1. CTRP3 down-regulation is observed in the pancreas of cerulein-induced SAP mice (A) CTRP3 mRNA expression levels were determined by qPCR analysis in the pancreas of mice from normal control group (n=5) and SAP group (n=5). (B) CTRP3 protein expression levels were examined by Western blotting analysis in the pancreas of mice from normal control group and SAP group. (C) Serum amylase and lipase activities of mice from normal control group and SAP group were measured using amylase assay kit and lipase assay kit, respectively. (D) The levels of inflammatory cytokines (TNFα, IL-1β, IL-6 and IL-10) were determined using ELISA in the serum and pancreas of mice from normal control group and SAP group. (E) NO concentration and NOS activity in the pancreas and serum of mice from normal control group and SAP group were detected via NO assay kit and NOS assay kit, respectively. (F) The protein expression levels of SIRT1, acetylated p53 and phosphorylated NF-κB p65 were examined by Western blotting analysis in the pancreas of mice from normal control group and SAP group. **P<0.01.

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  Figure 2. CTRP3 overexpression alleviates pancreatic injury in cerulein-induced SAP mice (A) CTRP3 protein expression was examined by Western blotting analysis in the pancreas of mice from different treatment groups after tail intravenous injection of AAV-NC or AAV-CTRP3. (B) Representative images of H&E staining of pancreatic tissues of mice from different treatment groups (magnification, 100×). (C) Serum amylase and lipase activities of mice from different treatment groups were determined by amylase assay kit and lipase assay kit, respectively. **P<0.01.

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  Figure 3. CTRP3 overexpression reduces the production of pro-inflammatory mediators in SAP mice (A) The levels of inflammatory cytokines (TNFα, IL-1β, IL-6 and IL-10) were determined using ELISA in the serum and pancreas of mice from different treatment groups after tail intravenous injection of AAV-NC or AAV-CTRP3. (B) NO concentration and NOS activity in the serum and pancreas of mice from different treatment groups were detected via NO assay kit and NOS assay kit, respectively, after tail intravenous injection of AAV-NC or AAV-CTRP3. **P<0.01.

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  Figure 4. CTRP3 overexpression decreases MPO activity and increases SOD activity in SAP mice (A) MPO protein expression in the pancreas of mice from different treatment groups was visualized by immunohistochemical staining after tail intravenous injection of AAV-NC or AAV-CTRP3. (B) MPO activity in the pancreas of mice from different treatment groups was analyzed by MPO assay kit after tail intravenous injection of AAV-NC or AAV-CTRP3. (C) SOD activity in the pancreas of mice from different treatment groups was evaluated by SOD assay kit after tail intravenous injection of AAV-NC or AAV-CTRP3. **P<0.01.

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  Figure 5. CTRP3 regulates p53 and NF-κB signaling pathways via SIRT1 in primary pancreatic acinar cells of mice (A) The protein expression levels of CTRP3, SIRT1, acetylated p53 and phosphorylated NF-κB p65 were examined by Western blotting analysis after transfection of SIRT1 specific inhibitor EX527 in primary pancreatic acinar cells treated with CTRP3 expression vectors. (B) The protein expression levels of CTRP3, SIRT1, acetylated p53 and phosphorylated NF-κB p65 were examined by Western blotting analysis after transfection of SIRT1-specific activator SRT1720 in shCTRP3-treated primary pancreatic acinar cells. **P<0.01; ns, not significant.

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  Figure 6. CTRP3 modulates p53 and NF-κB signaling pathways via SIRT1 in vivo (A) Immunohistochemical staining was used to visualize the protein expression of phosphorylated NF-κB p65 in the pancreas of mice from different treatment groups after tail intravenous injection of AAV-NC or AAV-CTRP3. (B) The protein expression levels of CTRP3, SIRT1, acetylated p53 and phosphorylated NF-κB p65 in the pancreas of mice from different treatment gruops were examined by Western blotting analysis after tail intravenous injection of AAV-NC or AAV-CTRP3. **P<0.01.