Role of mitophagy in hydrogen-induced reduction of LPS-caused mitochondrial injury to macrophages of mice / 中华麻醉学杂志

ABSTRACT

Objective To evaluate the role of mitophagy in hydrogen-induced reduction of lipopolysaccharide (LPS)-caused mitochondrial injury to macrophages of mice.Methods Macrophage line RAW264.7 cells of mice were routinely cultured and divided into 4 groups (n =6 each) using a random number table method:control group (Con group),LPS group,LPS plus hydrogen group (LPS + H2 group) and LPS plus hydrogen plus mitophagy inhibitor 3-methyladenine (3-MA) group (LPS+H2+3-MA group).Cells were incubated for 6 h with LPS at the concentration of 1 μg/ml in LPS group.Cells were incubated for 6 h with LPS 1 μg/ml and hydrogen-rich medium 0.6 mmol/L.Cells were incubated for 1 h with 2 mmol/L 3-MA and then incubated for 6 h with LPS 1 μg/ml and hydrogen-rich medium 0.6 mmol/L in LPS+H2+3-MA group.Mitochondrial respiratory control ratio (RCR) was measured using a Clark-type electrode.Mitochondrial membrane potential (MMP) was determined by JC-1 staining.Autophagosomes were counted with a transmission electron microscope.The expression of PTEN-induced putative kinase 1 (PINK1),E3 ubiquitin ligase (Parkin),microtubule-associated protein 1 light chain 3 Ⅱ (LC3 Ⅱ)and Beclin-1 was determined by Western blot.Results Compared with Con group,RCR and MMP were significantly decreased,the expression of PINK1,Parkin,LC3 Ⅱ and Beclin-1 was up-regulated,and the number of autophagosomes was increased in LPS group (P＜0.05).Compared with LPS group,RCR and MMP were significantly increased,the expression of PINK1,Parkin,LC3 Ⅱ and Beclin-1 was up-regulated,and the number of autophagosomes was increased in LPS+H2group (P＜0.05).Compared with LPS+H2 group,RCR and MMP were significantly decreased,the expression of PINK1,Parkin,LC3 Ⅱ and Beclin-1 was down-regulated,and the number of autophagosomes was decreased in LPS + H2 + 3-MA group (P＜0.05).Conclusion Enhanced mitophagy is involved in hydrogen-induced reduction of LPS-caused mitochondirial injury to macrophages of mice.