Conclusion
In conclusion, miR-539-5p alleviated sepsis-induced ALI via suppressing its downstream target ROCK1, suggesting a therapeutic potential of miR-539-5p for the management of sepsis-induced ALI.
Material and methods
Adult male C57BL/6 mice were intravenously injected with or without miR-539-5p agomir or scrambled control one week before CLP operation. MPVECs were transfected with miR-539-5p mimics or control mimics, followed by lipopolysaccharide (LPS) stimulation. ROCK1 was predicted and confirmed as a direct target of miR-539-5p using dual-luciferase reporter assay. In rescue experiment, MPVECs were co-transfected with lentiviral vector expressing ROCK1 (or empty vector) and miR-539-5p mimics 24 h before LPS treatment. The transcriptional activity of caspase-3, the apoptosis ratio, the levels of miR-539-5p, interleukin-1b (IL-1b), interleukin-6 (IL-6), and ROCK1 were assessed.
Methods
Adult male C57BL/6 mice were intravenously injected with or without miR-539-5p agomir or scrambled control one week before CLP operation. MPVECs were transfected with miR-539-5p mimics or control mimics, followed by lipopolysaccharide (LPS) stimulation. ROCK1 was predicted and confirmed as a direct target of miR-539-5p using dual-luciferase reporter assay. In rescue experiment, MPVECs were co-transfected with lentiviral vector expressing ROCK1 (or empty vector) and miR-539-5p mimics 24 h before LPS treatment. The transcriptional activity of caspase-3, the apoptosis ratio, the levels of miR-539-5p, interleukin-1b (IL-1b), interleukin-6 (IL-6), and ROCK1 were assessed.
Results
Compared to sham group, mice following CLP showed pulmonary morphological abnormalities, elevated production of IL-1b and IL-6, and increased caspase-3 activity and apoptosis ratio in the lung. In MPVECs, LPS stimulation resulted in a significant induction of inflammatory cytokine levels and apoptosis compared to untreated cells. The overexpression of miR-539-5p in septic mice alleviated sepsis-induced pulmonary injury, apoptosis, and inflammation. MiR-539-5p also demonstrated anti-apoptotic and anti-inflammatory effect in LPS-treated MPVECs. The upregulation of ROCK1 in MPVECs recovered miR-539-5p-suppressed caspase-3 activity and proinflammatory cytokine production.