Cardiac Circular RNAs CDR1as, Circ-RCAN2, Circ-C12orf29 Show Cell-Specific Hypoxia-Induced Dysregulation and Distinct...

Area of Science:

• Cardiovascular Biology
• Molecular Biology
• Epigenetics

Background:

• Circular RNAs (circRNAs) are emerging as key regulators in cardiovascular diseases, including myocardial infarction (MI).
• Understanding the specific roles and regulation of novel circRNAs in cardiac cells during ischemic conditions is crucial for developing therapeutic strategies.
• CDR1as, circ-RCAN2, and circ-C12orf29 have been implicated in MI, but their circularity, cell-specific responses to hypoxia, and functional effects require detailed investigation.

Purpose of the Study:

• To confirm the circular nature of CDR1as, circ-RCAN2, and circ-C12orf29.
• To investigate the cell-specific expression patterns of these circRNAs in porcine cardiac progenitor cells (pCPCs), bone marrow mesenchymal stem cells (pMSCs), and cardiac fibroblasts (pCFs) under hypoxic conditions.
• To assess the functional impact of circ-RCAN2 and circ-C12orf29 on cell viability and their regulation of downstream microRNA (miRNA) targets.

Main Methods:

• Circularity confirmation using RNase R treatment, qPCR, gel electrophoresis, and Sanger sequencing.
• Hypoxia and serum deprivation to mimic ischemia in pCPCs, pMSCs, and pCFs.
• Assessment of cell viability via MTT assay and miRNA target analysis using siRNA-mediated knockdown of circ-RCAN2 and circ-C12orf29.

Main Results:

• RNase R treatment and subsequent analyses confirmed the circularity of CDR1as, circ-RCAN2, and circ-C12orf29.
• Hypoxia induced cell-specific circRNA expression: pCPCs showed upregulation of all candidates, pMSCs showed upregulation of circ-RCAN2 and circ-C12orf29 with prolonged hypoxia, while pCFs exhibited no significant changes.
• Knockdown of circ-RCAN2 and circ-C12orf29 demonstrated differential effects on cell viability (e.g., circ-RCAN2 knockdown reduced pCF viability, increased pMSC viability) and circ-C12orf29 knockdown upregulated specific miRNAs (ssc-miR-21-5p, ssc-miR-181c) in pCPCs.

Conclusions:

• CDR1as, circ-RCAN2, and circ-C12orf29 are confirmed as circular RNAs with dysregulation patterns that are time- and cell-type-specific following hypoxic insult.
• circ-RCAN2 and circ-C12orf29 play distinct roles in regulating cell viability across different cardiac cell types.
• circ-C12orf29 influences downstream miRNA targets, highlighting its complex regulatory network in cardiac cells during ischemia.