Mechanism of CircANKRD36 regulating cell heterogeneity and endothelial mesenchymal transition in aortic valve stromal cells by regulating miR-599 and TGF-β signaling pathway

Objective: To explore the mechanism of CircANKRD36 regulating cell heterogeneity and endothelial mesenchymal transition in aortic valve stromal cells by regulating miR-599 and TGF-beta signaling pathway. Methods: Human tissue specimens were divided into Control group (n = 25) and CAVD group (n = 25). The mRNA expressions of CircANKRD36 and miR-599 in tissue samples were analyzed by qRT-PCR. Western blot was used to analyze the protein expression of osteogenic differentiation related factors induced by OM.The expressions of ALP, osteocalcin, osteopontin, Runx2 and Cadherin11 were detected by Western blot. Results: The expression of CircANKRD36mRNA in CAVD tissue was lower than that in Control tissue (P < 0.05), and the expression of miR-599mRNA in CAVD tissue was higher than that in Control tissue (P < 0.05). CircANKRD36 was negatively correlated with ALP, osteocalcin, osteopontin, Runx2, Cadherin11 expression level after OM induced osteogenic differentiation. The expression level of miR-599 was positively correlated with ALP, osteocalcin, osteopontin, Runx2 and Cadherin11 after OM induced osteogenic differentiation. The expression of ALP, osteocalcin, osteopontin, Runx2 and Cadherin11 protein in circ+miR-599 group was lower than that in circ+miR-NC group (P < 0.05). Compared with Vector+miR-NC group, the protein expressions of TGF-beta 1, TGF beta 2 and SMAD4 in circ+miR-NC group decreased (P < 0.05), while the protein expressions of TGF-beta 1, TGF-beta 2 and SMAD4 in circ+miR-599 group increased (P < 0.05). Conclusion: CircANKRD36 can inhibit the expression of miR-599 and the activation of TGF-beta signaling pathway, thus inhibiting the expression of differentiation-related factors of VIC osteogenesis and the formation of calcified nodules. Therefore, circANKRD36-miR-599-TGF-beta axis can be a new theoretical basis for treating CAVD.

Automated summary

In this study, it was found that CircANKRD36 can inhibit the expression of miR-599 and the activation of the TGF-beta signaling pathway, thereby affecting cell heterogeneity and endothelial mesenchymal transition in aortic valve stromal cells. This provides a new theoretical basis for treating calcific aortic valve disease (CAVD).