SIRT1/NF-κB通路在人参皂苷Rb1对高糖处理的H9C2细胞凋亡与炎症反应作用的研究

Role of SIRT1/NF-κB pathway in the effect of ginsenoside Rb1 on apoptosis and inflammation of H9C2 cells treated with high glucose

目的探讨沉默信息调节因子2相关酶1(SIRT1)/核因子-κB(NF-κB)通路在人参皂苷Rb1对高糖处理的H9C2细胞的凋亡与炎症反应中的作用。方法采用不同浓度的人参皂苷Rb1处理高糖环境(33μmol/L)下的H9C2细胞,CCK-8法检测细胞活性,ELISA检测细胞上清液TNF-α、IL-6的含量,确定人参皂苷Rb1对高糖环境下H9C2细胞的最佳处理浓度。用SIRT1的siRNA转染沉默SIRT1基因,并通过蛋白免疫印迹法检测细胞的Bax、Bcl-2、cleaved caspase-3、SIRT1、NF-κB等蛋白含量,ELISA检测细胞上清液TNF-α、IL-6的含量来探索人参皂苷Rb1是否通过SIRT1/NF-κB通路减轻H9C2细胞凋亡与炎症反应。结果 CCK8法示33μmol/L糖浓度下H9C2细胞活性即出现下降(P<0.05),33μmol/L糖浓度中人参皂苷Rb1在50μmol/L时细胞活性最高(P <0.05),ELISA法示TNF-α与IL-6在50μmol/L的人参皂苷Rb1处理时与其他浓度相比均最低(P均<0.05)。Rb1处理组与高糖组NF-κB的乙酰化水平比较差异有统计学意义(P <0.05)。SIRT1的siRNA转染后的人参皂苷Rb1处理组与单纯转染SIRT1的siRNA高糖组相比,Bax/Bcl-2与Acetyl-NF-κB/NF-κB表达量差异均无统计学意义(P均<0.05),TNF-α与IL-6的含量比较差异均无统计学意义(P均<0.05)。结论人参皂苷Rb1可通过SIRT1/NF-κB通路对高糖处理的H9C2细胞的凋亡与炎症反应产生保护作用。

Objective To investigate the role of silent information regulation 2 homolog 1(SIRT1)/NF-κB pathway in the effect of ginsenoside Rb1 upon the apoptosis and inflammation of H9 C2 cells treated with high glucose. Methods The H9 C2 cells were cultured in high glucose medium(33 μmol/L) and cotreated with different concentrations of ginsenoside Rb1. The cell viability was detected by CCK-8 assay. The concentrations of TNF-α and IL-6 in the cellular supernatant were measured by ELISA. The optimal treatment concentration of ginsenoside Rb1 for the high glucose-treated H9 C2 cells was determined. SIRT1 gene was silenced by siRNA transfection. The expression levels of Bax, bcl-2, cleaved caspase-3, SIRT1 and NF-κB proteins were quantitatively detected by Western blot. The content of TNF-α and IL-6 in the cellular supernatant was measured by ELISA to determine whether ginsenoside Rb1 could alleviate the apoptosis and inflammation of H9 C2 cells through SIRT1/NF-κB pathway. Results CCK8 assay demonstrated that the viability of H9 C2 cells began to significantly decline when treated with 33 μmol/L glucose(P < 0.05). The viability of H9 C2 cells co-treated with 33 μmol/L glucose and 50μmol/L ginsenoside Rb1 had the highest cell viability(P < 0.05). ELISA revealed that the concentrations of TNF-α(P < 0.05) and IL-6(P < 0.05) were the lowest when the cells were co-treated with 50 μmol/L ginsenoside Rb1. The acetylation level of NF-κB significantly differed between the ginsenoside Rb1 and high glucose treatment groups(P < 0.05). The expression levels of Bax/Bcl-2 and Acetyl-NF-κB/NF-κB did not significantly differ between the siRNA transfection plus ginsenoside Rb1 treatment group and siRNA transfection plus high glucose treatment groups(both P > 0.05). The content of TNF-α and IL-6 did not significantly differ(both P > 0.05). Conclusion Ginsenoside Rb1 can protect the H9 C2 cells with high glucose treatment from apoptosis and inflammation via regulating the Sirt1/NF-κB pathway.