Sodium butyrate affected the proliferation and apoptosis of steatosis HepG2 cells via G protein-coupled receptor 43/109a-phosphorylated protein kinase B-mammalian target of rapamycin signaling pathway
10.3760/cma.j.cn311367-20201111-00654
- VernacularTitle:丁酸钠通过G蛋白偶联受体43/109a-磷酸化蛋白激酶B-哺乳动物雷帕霉素靶蛋白通路影响脂肪变性HepG2细胞的增殖与凋亡
- Author:
Yu LI
1
;
Mengting LI
;
Taotao LIU
;
Da ZHOU
Author Information
1. 复旦大学附属中山医院消化科，上海 200032
- Keywords:
Sodium butyrate;
Steatosis;
Hepatocytes;
Cell proliferation;
Apoptosis
- From:
Chinese Journal of Digestion
2021;41(7):471-477
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the effects and mechanism of gut metabolite sodium butyrate on the proliferation and apoptosis of steatosis HepG2 cells in vitro. Methods:The in vitro steatosis hepatocyte model was established with human liver cell line HepG2 and free fatty acid (FFA; the concentration ratio of oleic acid to palmitic acid was 2∶1). Normal control group, model group and intervention groups with different concentration (1, 2, 5, 10, 20 and 50 mmol/L) of sodium butyrate were set up. The inhibition of sodium butyrate on the proliferation of steatosis HepG2 cells was detected by cell counting kit (CCK-8). The proportion of apoptotic cells of normal control group, model group and sodium butyrate 5 mmol/L (sodium butyrate intervention) group was detected by flow cytometry. Normal control group, model group, intervention group with different concentration (1, 2, 5 and 10 mmol/L) of sodium butyrate, negative small interfering RNA (siRNA) control group, G protein-coupled receptor (GPR) 43-siRNA group, GPR109a-siRNA group, GPR43+ GPR109 a double knockout group were set up. The change of the levels of phosphorylated protein kinase B (p-AKT) and phosphorylated mammalian target of rapamycin (p-mTOR) before and after transfection were detected by Western blotting. One-way analysis of varivance, SNK- q test and logistic regression analysis were used for statistical analysis. Results:The results of CCK-8 test indicated that sodium butyrate inhibited the proliferation of steatosis HepG2 cells in a dose-dependent and time-dependent manner. The results of flow cytometry showed that the proportion of apoptotic cells of the sodium butyrate intervention group was higher than that of the model group and normal control group ((3.400±0.100)% vs. (1.800±0.400)% and(1.067±0.451)%), and the differences were statistically significant ( t=6.721 and 8.705, both P<0.01). There was no significant difference in the proportion of apoptotic cells between the model group and the normal control group ( P>0.05). Before transfection, the expressions of p-AKT and p-mTOR at protein level of the model group were both higher than those of the normal control group (2.300±0.058 vs. 1.000±0.012, 2.160±0.125 vs. 1.000±0.052), and the differences were statistically significant ( t=22.080 and 8.575, both P<0.05). The expressions of p-AKT and p-mTOR at protein level of sodium butyrate intervention groups at 1, 2, 5 and 10 mmol/L were all lower than those of the model group (1.530±0.085, 1.407±0.096, 1.032±0.035 and 1.036±0.099 vs. 2.300±0.058; 1.483±0.073, 1.297±0.048, 1.067±0.035 and 0.970±0.072 vs. 2.160±0.125), and the differences were statistically significant ( t=7.491, 7.997, 19.790, and 11.020; 4.683, 6.445, 8.424, and 8.245; all P<0.05). After transfection, the expressions of p-AKT and p-mTOR at protein level of GPR43-siRNA group, GPR109a-siRNA group and GPR43/ GPR109 a double knockout group were all higher than those of the negative siRNA control group and 5 mmol/L sodium butyrate group (1.474±0.045, 1.471±0.058 and 2.067±0.120 vs. 1.158±0.030 and 1.139±0.031; 1.850±0.082, 1.683±0.058 and 2.160±0.091 vs. 1.469±0.037 and 1.490±0.116), and the differences were statistically significant ( tp-AKT=5.807, 4.816, 7.322, 6.109, 5.080 and 7.463; tp-mTOR=4.235, 3.113, 7.044, 2.542, 1.497 and 4.562; all P<0.05). Conclusions:The effect of sodium butyrate on the proliferation and apoptosis of steatosis HepG2 cells is associated with the GPR43/GPR109a-pAKT-mTOR signaling pathway.
