FTO represses NLRP3‐mediated pyroptosis and alleviates myocardial ischemia–reperfusion injury via inhibiting CBL‐mediated ubiquitination and degradation of β‐catenin

Abstract

AbstractCardiac ischemia/reperfusion (I/R) injury is a complicated pathological event, which has close association with pyroptosis. This study uncovered the regulatory mechanisms of fat mass and obesity‐associated protein (FTO) in NLRP3‐mediated pyroptosis during cardiac I/R injury. H9c2 cells were stimulated with oxygen–glucose deprivation/reoxygenation (OGD/R). Cell viability and pyroptosis were detected by CCK‐8 and flow cytometry. Western blotting or RT‐qPCR was performed to analyze target molecule expression. NLRP3 and Caspase‐1 expression was observed by immunofluorescence staining. IL‐18 and IL‐1β production was detected by ELISA. The total m6A and m6A level of CBL was determined by dot blot assay and methylated RNA immunoprecipitation‐qPCR, respectively. The interaction between IGF2BP3 and CBL mRNA was confirmed by RNA pull‐down and RIP assays. The protein interaction between CBL and β‐catenin and β‐catenin ubiquitination were evaluated by Co‐IP. Myocardial I/R model was established in rats. We determined infarct size by TTC staining and pathological changes by H&E staining. LDH, CK‐MB, LVFS, and LVEF were also assessed. FTO and β‐catenin were down‐regulated, while CBL was up‐regulated by OGD/R stimulation. FTO/β‐catenin overexpression or CBL silencing restrained OGD/R‐induced NLRP3 inflammasome‐mediated pyroptosis. CBL repressed β‐catenin expression via ubiquitination and degradation. FTO reduced the mRNA stability of CBL by inhibiting m6A modification. CBL‐mediated ubiquitination and degradation of β‐catenin were involved in FTO‐induced pyroptosis inhibition during myocardial I/R injury. FTO inhibits NLRP3‐mediated pyroptosis to attenuate myocardial I/R injury via repressing CBL‐induced ubiquitination degradation of β‐catenin.