PF-477736 modulates vascular smooth muscle cells phenotypic transition through Chk1/p53/CD44 pathway

The phenotypic transition of vascular smooth muscle cells (VSMCs) from a quiescent, contractile type to a secretory phenotype with high proliferation and mobility is a key event in vascular remodeling. PF-477736 is an ATP-competitive inhibitor of Chk1 which induces the accumulation of DNA damage by increasing the level of replicative stress, and ultimately inhibiting cell proliferation or causing cell death. Although this compound has been utilized as an anti-tumor drug, its role in vascular remodeling remains unclear.

In vitro, Human aortic smooth muscle cell line (HAVSMC) and primary rat aortic smooth muscle cells were used to establish phenotype transformation model with PDGF-bb; Western blot was used to detect the expression of VSMCs phenotype marker α-SMA, Vimentin; MTT and EdU assays were used to evaluate the proliferation ability of VSMCs; wound healing assay was used to evaluate the migration ability of VSMCs. In vivo, we established ballon injury of carotid artery in rats, and the function of the PF-477736 was evaluated by several histological stainings.

The results exhibit that PF-477736 effectively inhibited VSMCs phenotypic transition, resulting in G1/S phase arrest and decreased proliferation and migration ability of VSMCs. Furthermore, while PDGF-bb down-regulated p53 protein and up-regulated CD44 expression, PF-477736 significantly countered these effects. Pretreatment of VSMCs with p53 siRNA blocked the effect of PF-477736, up-regulated the expression of CD44, and promoted VSMCs' proliferation and migration. Conversely, CD44 silencing through siRNA mitigated the phenotypic transition of VSMCs. In addition, the H&E, Masson' staining and the immunohistochemistry of PCNA, p53 and CD44 showed that PF-477736 substantially inhibits vascular remodeling in the balloon injury model.

Our findings show that PF-477736 exerts anti-vascular remodeling effect by inhibiting phenotypic transition through the Chk1/p53/CD44 pathway in VSMCs, providing novel therapeutic strategies for preventing and treating vascular remodeling.