ABSTRACT
Myocardial ischemiaâreperfusion injury (MI/RI) is closely related to pyroptosis. alkB homolog 5 (ALKBH5) is abnormally expressed in the MI/RI models. However, the detailed molecular mechanism of ALKBH5 in MI/RI has not been elucidated. In this study, rats and H9C2 cells served as experimental subjects and received MI/R induction and H/R induction, respectively. The abundance of the targeted molecules was evaluated using RT-qPCR, Western blotting, immunohistochemistry, immunofluorescence, and enzyme-linked immunosorbent assay. The heart functions of the rats were evaluated using echocardiography, and heart injury was evaluated. Cell viability and pyroptosis were determined using cell counting Kit-8 and flow cytometry, respectively. Total m6A modification was measured using a commercial kit, and pri-miR-199a-5p m6A modification was detected by Me-RNA immunoprecipitation (RIP) assay. The interactions among the molecules were validated using RIP and luciferase experiments. ALKBH5 was abnormally highly expressed in H/R-induced H9C2 cells and MI/RI rats. ALKBH5 silencing improved injury and inhibited pyroptosis. ALKBH5 reduced pri-miR-199a-5p m6A methylation to block miR-199a-5p maturation and inhibit its expression. TNF receptor-associated Factor 3 (TRAF3) is a downstream gene of miR-199a-5p. Furthermore, in H/R-induced H9C2 cells, the miR-199a-5p inhibitor-mediated promotion of pyroptosis was reversed by ALKBH5 silencing, and the TRAF3 overexpression-mediated promotion of pyroptosis was offset by miR-199a-5p upregulation. ALKBH5 silencing inhibited pri-miR-199a-5p expression and enhanced pri-miR-199a-5p m6A modification to promote miR-199a-5p maturation and enhance its expression, thereby suppressing pyroptosis to alleviate MI/RI through decreasing TRAF3 expression.
Subject(s)
AlkB Homolog 5, RNA Demethylase , MicroRNAs , Myocardial Reperfusion Injury , Pyroptosis , Animals , Rats , Adenine , AlkB Homolog 5, RNA Demethylase/genetics , AlkB Homolog 5, RNA Demethylase/metabolism , Demethylation , MicroRNAs/metabolism , Myocardial Reperfusion Injury/genetics , Myocardial Reperfusion Injury/metabolism , TNF Receptor-Associated Factor 3/genetics , TNF Receptor-Associated Factor 3/metabolismABSTRACT
BACKGROUND: Polyphenols in extra virgin olive oil (EVOO) have been found to reduce the expression of PPARγ2, inhibit adipocyte differentiation, and enhance the formation of osteoblasts from bone marrow stem cells. However, the underlying mechanisms of their action remain unknown. PURPOSE: To determine the sequential effects of EVOO on marrow fat expansion induced by estrogen deprivation using 3.0-T proton magnetic resonance (MR) spectroscopy in an ovariectomy (OVX) rabbit model of postmenopausal bone loss over a six-month period. MATERIAL AND METHODS: A total of 45 female New Zealand rabbits were equally divided into sham-operation, OVX controls, and OVX treated with EVOO for six months. Marrow fat fraction was measured by MR spectroscopy at baseline conditions, and three and six months postoperatively, respectively. Serum bone biomarkers, lumbar and femoral bone mineral density, microtomographic parameters, biomechanical properties, and quantitative parameters of marrow adipocytes were studied. RESULTS: OVX was associated with marrow adiposity in a time-dependent manner, accompanied with increased bone turnover and impaired bone mass and trabecular microarchitecture. In OVX rabbits, EVOO markedly alleviated trabecular bone loss and reduced the accumulation of lipid droplets including adipocyte size, density, and areas of fat deposits in the bone marrow. EVOO prevented such changes in terms of both marrow adiposity and bone remodeling. CONCLUSION: Early EVOO treatment may exert beneficial effects on bone by modulating marrow adiposity, which would support their protective effect against bone pathologies.
