Effect of miR-181a on acute lung injury induced by sepsis in mice by targeting SIRT1 and its mechanism / 西安交通大学学报(医学版)

ABSTRACT

Objective To explore the effect and mechanism of miR-181a on sepsis-induced acute lung injury in mice by targeting silent information regulator 1 (SIRT1). Methods Lipopolysaccharide was used to induce the A549 cell model of acute lung injury. miR-181 inhibitor and inhibitor negative control (inhibitor NC) were transfected in the cells. Cecal ligation and puncture (CLP) was used to establish the mouse model of sepsis-induced acute lung injury. The mice were intravenously infused with miR-181a antagomir and antagomir NC. Then survival rate and wet-to-dry ratio of the lungs were examined, the targeting relationship between miR-181a and SIRT1 was tested by luciferase reporter assay, RT-PCR was used to detect the gene expression of miR-181a and SIRT1 in vitro and vivo. Western blotting was used to detect the protein level of SIRT1 in vitro and vivo, as well as the levels of apoptosis-related protein B-cell lymphoma-2 (Bcl-2), Bcl-2 associated X protein (Bax), and cleave caspase-3 (cl-CASP3). Cell apoptosis was quantified by flow cytometry, cell viability was measured by CCK-8, CASP3 activity was detected by kit. HE staining was used to observe the histopathological changes in the lungs; TUNNEL staining was used to detected the cell apoptosis in the lungs. The concentrations of inflammatory cytokines were measured by ELISA. Results In cellular experiments, compared with those in control group, the expression of miR-181a was increased and the expression of SIRT1 was decreased in LPS group (P<0.01). Meanwhile the cell apoptosis rate was increased, CASP3 activity was increased, and cell viability was decreased. Treatment by miR-181a inhibitor could reverse the changes of the above indicators (P<0.01). In animal experiments, compared with those in sham group, in CLP group the mice's survival rate was decreased, wet-to-dry ratio of the lungs was increased, significant pathological changes and cell apoptosis in the lungs could be observed. Meanwhile, the concentrations of inflammatory cytokines were increased, the protein levels of miR-181a, Bax and cl-CASP3 were increased, and the protein levels of SIRT1 and Bcl-2 were decreased (P<0.01). Treatment by miR-181a antagomir could reverse the changes of the above indicators (P<0.05, P<0.01). Conclusion miR-181a can be targeted to SIRT1, and inhibition of miR-181a expression can increase SIRT1 expression, there by protecting against acute lung injury induced by sepsis in mice.