CircIdh2 Attenuates Endothelial-to-Mesenchymal Transition and Myocardial Fibrosis and Exhibits Translational Potential

Background

Endothelial-to-mesenchymal transition (EndMT) contributes to myocardial fibrosis after cardiac injury. Circular RNAs (circRNAs) have been implicated in the regulation of EndMT, but whether circRNAs participate in this process through translational potential remains unclear. This study investigated the role of circIdh2 in EndMT and examined whether its coding capacity may contribute to its biological effects.

Methods

Rat endothelial progenitor cells (rEPCs) were treated with TGF-β1 (5 ng/mL) for 7 days to induce EndMT. The expression of circSorbs1, circIdh2, and circSntb2 was measured by qRT-PCR. EndMT-related markers, including VE-cadherin, vimentin, and N-cadherin, were detected by western blotting. Cell migration was assessed using Transwell and scratch wound healing assays. The translational potential of circIdh2 was examined using an Idh2-IRES luciferase reporter assay, FLAG immunoprecipitation followed by western blotting, and mass spectrometry. A circIdh2 mutant construct and a rat model of acute myocardial infarction (AMI) were used to further evaluate the contribution of circIdh2 and its putative circIdh2-derived product.

Results

TGF-β1 reduced the expression of circSorbs1, circIdh2, and circSntb2 in rEPCs. CircIdh2 showed the largest decrease in the initial qRT-PCR screen and was selected before any circIdh2-specific functional experiment. The Idh2 host gene supplied metabolic and redox context for the prespecified screen but was not used as evidence of circIdh2 function. CircIdh2 overexpression increased VE-cadherin expression, decreased vimentin and N-cadherin expression, and attenuated TGF-β1–induced cell migration. In contrast, circIdh2 knockdown promoted EndMT-associated marker changes and cell migration. The Idh2-IRES reporter assay, FLAG immunoprecipitation, and mass spectrometry supported the translational potential of circIdh2 and detected a FLAG-tagged product derived from the circIdh2 expression construct. The FLAG-tagged circIdh2 expression construct modulated EndMT-related markers and reduced cell migration, whereas the coding-disrupted mutant construct did not reproduce the effects of wild-type circIdh2. In vivo, AAV-mediated delivery of circIdh2 or circIdh2-aa attenuated myocardial fibrosis and partly reversed EndMT-related marker changes after AMI.

Conclusions

CircIdh2 attenuated EndMT-associated changes in rEPCs and was associated with reduced myocardial fibrosis after AMI. These findings support the translational potential of circIdh2 and suggest that its putative circIdh2-derived product may contribute to its effects.