吡格列酮抗高糖诱导H9c2心肌细胞炎性损伤的机制研究

目的 探讨PPAR-γ激动剂吡格列酮（pioglitazone, PGZ）通过抑制p38MAPK/NF-κB信号通路减轻高糖诱导的H9c2大鼠心肌细胞炎性损伤。方法 离体培养的H9c2细胞随机分6组,分别为低糖组（5.5 mmol/L葡萄糖,NG组）、高糖组（35 mmol/L葡萄糖,HG组）、高糖+不同浓度PGZ组（HG+5 μmol/L PGZ 、HG+10 μmol/L PGZ、HG+15 μmol/L PGZ、HG+20 μmol/L PGZ）,采用CCK8法检测各组细胞干预24、48 h后细胞活性变化;据CCK8结果选取NG组、HG组、HG+10 μmol/L PGZ （Low组）、HG+20 μmol/L PGZ （High组） 4组细胞干预48 h,ELISA检测各组细胞炎性因子TNF-α、IL-1β的含量,Western-blot检测各组细胞p-p38、p-p65蛋白的表达。结果 CCK8检测结果显示,与NG组比较,24、 48 h后HG组明显活力下降（P<0.01）,表明高糖可诱导心肌细胞损伤;与HG组比较,48 h后HG+10 μmol/L PGZ、HG+ 15 μmol/L PGZ两组细胞活力均升高（P<0.05）,HG+20 μmol/L PGZ组细胞活力明显升高（P<0.01）,提示PGZ可对高糖环境下心肌细胞起保护作用。干预48 h后ELISA结果显示,HG组TNF-α、IL-1β含量与NG组比明显升高（P<0.01）,提示高糖环境可致心肌细胞发生炎性损伤;与HG组比较,Low组IL-1β含量降低（P<0.05）,High组TNF-α含量降低（P<0.05）,IL-1β含量明显降低（P<0.01）。提示PGZ可减轻高糖诱导心肌细胞炎性损伤。干预48 h后Western-blot结果显示,与NG组比较,HG组p-p65和p-p38蛋白表达水平均明显升高（P<0.01）;与HG组比较,Low组p-p65和p-p38蛋白表达均降低（P<0.05）,High组p-p65和p-p38蛋白表达水平均明显降低（P<0.01）。提示PGZ可下调高糖诱导心肌细胞NF-κB/p38MAPK通路的表达。结论 PPAR-γ激动剂吡格列酮可以减轻高糖诱导H9c2大鼠心肌细胞的炎性损伤,其机制可能与抑制p38MAPK/NF-κB信号转导通路有关。

Mechanism of pioglitazone against high glucose-induced inflammatory injury in H9c2 cardiomyocytes

Objective To investigate PPAR-γ agonist pioglitazone alleviates high glucose-induced inflammatory damage of H9c2 rat cardiomyocytes through inhibiting P38MAPK/NF-κB signaling pathway. Methods The in vitro cultured H9c2 cells were randomized into 6 groups: low-glucose group (5.5 mmol/L glucose, NG group), high glucose group (35 mmol/L glucose, HG group), high glucose + different concentration pioglitazone (PGZ) Group (Hg+PGZ 5, 10, 15, 20 μmol/L). CCK8 method was used to detect cell activity change after 24 h, 48 h of cell intervention; according to CCK8 results, NG group, HG group, HG+10 μmol/L PGZ (Low group), HG+20 μmol/L PGZ (High group) of cells were selected for 48 h of cellular intervention, and the content of cell inflammatory factors TNF-α, IL-1β in each group of cells were detected by ELISA, and the expression of p-p38 and p-p65 proteins in each group of cells were detected by Western-blot assays. Results CCK8 results: Compared with NG group, the activity of HG group decreased significantly after 24 h and 48 h (P<0.01), indicating that high glucose can induce myocardial cell injury. Compared with HG group, the cell viability of HG+10 μmol/L PGZ and HG+15 μmol/L PGZ groups increased after 48 h (P<0.05), and the cell viability of HG+20 μmol/L PGZ group increased significantly (P<0.01), suggesting that PGZ can protect cardiomyocytes under high glucose environment. ELISA results after 48 h intervention: Compared with NG group, the contents of TNF-α and IL-1β in HG group were significantly increased (P<0.01), suggesting that high glucose environment can cause inflammatory damage to cardiomyocytes; Compared with HG group, the content of IL-1β in low group was decreased (P<0.05), the content of TNF-α in high group was decreased (P<0.05), and the content of IL-1β was significantly decreased (P<0.01). It is suggested that PGZ can reduce the inflammatory damage of cardiomyocytes induced by high glucose. Western-blot results after 48 h intervention: Compared with NG group, the protein expression levels of p-p65 and p-p38 in HG group were significantly increased (P<0.01); the protein expressions of p-p65 and p-p38 in low group were decreased (P<0.05), and the protein expressions of p-p65 and p-p38 in high group were significantly decreased (P<0.01). These results suggest that PGZ can down-regulate the expression of NF-κB /p38MAPK pathway in cardiomyocytes induced by high glucose. Conclusion PPAR-γ agonist pioglitazone can reduce inflammatory damage of H9c2 rat cardiomyocytes in high glucose, which may be related to inhibiting P38MAPK/NF-κB signal transduction pathways.