Effect of miR-330-3p on hepatic ischemia-reperfusion injury in mice
10.3760/cma.j.cn113884-20200915-00491
- VernacularTitle:微小RNA-330-3p对小鼠肝脏缺血再灌注损伤影响的研究
- Author:
Shipeng LI
;
Zhijun ZHU
;
Liying SUN
;
Haiming ZHANG
;
Guangpeng ZHOU
;
Jie SUN
;
Bin CUI
- From:
Chinese Journal of Hepatobiliary Surgery
2021;27(5):371-376
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To study the effect of microRNA (miR)-330-3p on hepatic ischemia-reperfusion injury (IRI) in mice, meanwhile, and to determine potential molecular mechanism.Methods:Eighty male C57BL/6 mice, aged 7-8 weeks, 23-25 g, specific pathogen free, were randomly divided into 8 groups (10 mice in each group) using random number table: reperfusion 2 h group, 6 h group, 12 h group, 24 h group, sham group, miR-330-3p agomir group (preoperative injection of agonist), miR-330-3p antagomir group (preoperative injection of inhibitor) and miRNA-NC group. Except for the sham group, the hepatic IRI model were established in mice. Polymerase chain reaction (PCR), Western blot and immunohistochemistry were used to detect the expression of miR-330-3p and phosphoglycerate mutase family member 5 (PGAM5), cleave caspase-1 and GSDMD. Luciferase reporter assay was performed to investigate whether miR-330-3p directly targets PGAM5. At the same time, AML12 cells were also treated with miR-330-3p mimics/inhibitor or PGAM5 siRNA, then the expression of PGAM5, NLRP3, cleave caspase-1 and GSDMD were detected by Western blot analysis.Results:Level of miR-330-3p gradually decreased after reperfusion, however, mRNA level of PGAM5 was increased thereafter ( P<0.05) as compared with the sham group. Serum level of AST and ALT were decreased in miR-330-3p agomir group while that of were increased in miR-330-3p antagomir group as a function of time following reperfusion, and the differences were statistically significant (all P<0.05). Cleave caspase-1 expression was decreased in miR-330-3p agomir group but was increased in miR-330-5p antagomir group ( P<0.05). Luciferase reporter assay was performed to determine PGAM5 was a target gene of miR-330-3p. SiRNA-mediated knockdown of PGAM5 decreased level of GAM5 (0.24±0.09), NLRP3(0.12±0.07), cleave caspase-1 (0.15±0.07) and GSDMD (1.08±0.08) as compared with the siRNA-NC group (1.17±0.14), (0.36±0.09), (0.68±0.09), (1.36±0.08), and the differences were statistically significant (all P<0.05). Conclusion:MiR-330-3p can alleviate hepatic IRI in mice, which may be related to inhibition of PGAM5-induced pyroptosis.
