MTORC1 is a key regulator that mediates OGD- and TGFβ1-induced myofibroblast transformation and chondroitin-4-sulfate expression in cardiac fibroblasts.

Abstract

Ischemia-reperfusion infarct-derived chondroitin sulfate proteoglycans (CSPGs) are important for sustaining denervation of the infarct. Sympathetic denervation within the heart after myocardial infarction (MI) predicts the probability of a higher risk for serious ventricular arrhythmias. Chondroitin-4-sulfate (C4S) is the predominant chondroitin sulfate component in the heart. However, the mechanisms that induce CSPG expression in fibroblasts following MI remain to be elucidated. The present study found that oxygen-glucose deprivation (OGD) and TGFβ1 stimulation induced myofibroblast transformation and C4S synthesis in vitro by using reverse transcription-quantitative PCR, western blotting and immunofluorescence. MTT assay was used to detect cell viability following OGD or OGD + TGF lotreatment. Using the PI3K inhibitor ZSTK474, the Akt inhibitor MK2206, or the mTOR inhibitor AZD8055, it was observed that OGD and TGFβ1 stimulation induced myofibroblast transformation and that C4S synthesis was mTOR-dependent, whereas the upstream canonical PI3K/Akt axis was dispensable by using western blotting and immunofluorescence. siRNA knockdown of Smad3, Raptor, or Rictor, indicated that mTORC1 was critical for promoting OGD- and TGFβ1-induced myofibroblast transformation and C4S synthesis by using western blotting and immunofluorescence. This response, may be mediated via cooperation between canonical Smad3 and mTORC1 signaling. These data suggested that inhibiting myofibroblast transformation may reduce C4S synthesis. Target mTORC1 may provide additional insight into the regeneration of sympathetic nerves and the reduction of fibrosis after MI at the cellular level. These findings may contribute to the understanding of the mechanism by which C4S overproduction in the hearts of patients with MI is associated with myocardial fibrosis.