Total Panax notoginseng saponin inhibits vascular smooth muscle cell proliferation and migration and intimal hyperplasia by regulating WTAP/p16 signals via m6A modulation

Abstract

Intimal hyperplasia, the key event of arterial restenosis, is a result of vascular smooth muscle cell (VSMC) proliferation and migration. Previous studies have demonstrated that total Panax notoginseng saponin (TPNS) represses intimal hyperplasia and inhibits the proliferation of VSMCs following balloon injury. However, the underlying roles of TPNS in intimal hyperplasia remain unclear. In this study, we first found that TPNS inhibited the intimal hyperplasia and reversed the reduced m⁶A quantity in balloon catheter-injured rat carotid artery. Then, we measured the expression profiles of m⁶A "writers" (i.e., methyltransferase like 3 (METTL3), methyltransferase like 14 (METTL14), and WT1 associated protein (WTAP)) and "erasers" (i.e., FTO alpha-ketoglutarate dependent dioxygenase (FTO) and alkB homolog 5, RNA demethylase (ALKBH5)) in vivo and found that TPNS up-regulated the reduced the WTAP expression in balloon catheter-injured rat carotid artery. Furthermore, we illustrated that TPNS inhibited the viability, proliferation, and migration potential of VSMCs via promotion of WTAP expression and suppression of WTAP restored the TPNS-induced inhibition of cell viability, proliferation and migration potential of VSMCs. In addition, we found that p16 was up-regulated in VSMCs treated with TPNS and repression of p16 restored the TPNS-induced inhibition of cell viability, proliferation and migration potential of VSMCs. Finally, we elucidated that, mechanistically, WTAP exerted its role by regulating p16 via m⁶A modification. Collectively, our results reveal the WTAP-p16 signaling axis and highlight the critical roles of m6A modification in intimal hyperplasia. Thus, this study provided a potential biomarker for the assessment of intimal hyperplasia risk following angioplasty as well as a novel therapeutic target for this disease.

Keywords: Rat carotid balloon injury; TPNS; VSMCs; WTAP; m(6)A; p16.