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Mitochondrial quality control is a new target for myocardial protection. Notch signaling plays an important role in heart development, maturation, and repair. However, the role of Notch in the myocardial mitochondrial quality control remains elusive. In this study, we isolated myocardial cells from rats and established myocardial ischemia reperfusion injury (IRI) model. We modulated Notch1 expression level in myocardial cells via infection with recombinant adenoviruses Ad-N1ICD and Ad-shN1ICD. We found that IR reduced myocardial cells viability, but Notch1 overexpression increased the viability of myocardial cells exposed to IRI. In addition, Notch1 overexpression improved ATP production, increased mitochondrial fusion and decreased mitochondrial fission, and inhibited mitophagy in myocardial cells exposed to IRI. However, N1ICD knockdown led to opposite effects. The myocardial protection role of Notch1 was related to the inhibition of Pink1 expression and Mfn2 and Parkin phosphorylation. In conclusion, Notch1 exerts myocardial protection and this is correlated with the maintenance of mitochondrial quality control and the inhibition of Pink1/Mfn2/Parkin signaling.

PURPOSE During myocardial infarction (MI), cardiac fibroblasts (CFs) transform into myofibroblast (CMT). This study aimed to investigate the crosstalk of Notch1 and transforming growth factor-β1 (TGF-β1)/Smad3 signaling in the regulation of CMT and myocardial fibrosis. METHODS Primary CFs were isolated from young rats and treated with TGF-β1 or adenovirus to overexpress or knockdown Notch1 intracellular domain (N1ICD) or Smad3. RESULTS TGF-β1 decreased the expression of fibroblast markers but increased the expression of myofibroblast markers in rat CFs. TGF-β1 increased the proliferation, invasion, and adhesion, and the secretion of collagen I of CFs, and these effects were inhibited by N1ICD overexpression. Moreover, endogenous Smad3 phosphorylation in CFs was enhanced by N1ICD knockdown, whereas TGF-β1 induced Smad3 phosphorylation was antagonized by the N1ICD overexpression. Conversely, endogenous N1ICD activation in CFs was antagonized by Smad3, whereas TGF-β1 induced N1ICD inactivation was antagonized by Smad3 knockdown. Coimmunoprecipitation showed that N1ICD interacted with Smad3 and immunostaining revealed the colocalization of N1ICD and Smad3 in the nuclei of CFs. Moreover, we demonstrated the functional antagonism of N1ICD and Smad3 on the phenotypes of CFs. Finally, TGF-β1/Smad3 signaling promoted whereas Notch signaling inhibited myocardial fibrosis in rat MI model. CONCLUSION Notch signaling inhibits CMT by antagonizing TGF-β1/Smad3 signaling. Notch signaling activators and TGF-β1/Smad3 signaling inhibitors could be exploited for therapeutic intervention to inhibit myocardial fibrosis after MI.