CREB/ATF3 signaling mediates indoxyl sulfate-induced vascular smooth muscle cell proliferation and neointimal formation in uremia

Abstract Background and aims Uremic patients are characterized by an increased risk of atherosclerotic cardiovascular diseases. Vascular smooth muscle cell (VSMC) proliferation contributes to neointimal formation, a main pathological feature in atherosclerosis. Activation of CREB/ATF3 signaling is pivotal in VSMC proliferation, yet its role in uremic atherosclerosis is unknown. This study aimed to explore whether CREB/ATF3 signaling is involved in the molecular mechanism underlying neointimal formation in uremia. Methods and Results Treatment of VSMCs with uremic toxin (indoxyl sulfate [IS]) activated cAMP/CREB/ATF3/cyclin D signaling, which was reflected by increased VSMC proliferation. Blocking cAMP/PKA/CREB/ATF3 signaling attenuated the promoting effect of IS on cyclin D1 expression and VSMC proliferation. Loss-of-function and time-dependent experiments showed that ATF3 lies downstream of the CREB signaling. Mutational analysis of cyclin D1 promoter along with chromatin immunoprecipitation assays showed that CREB/ATF3 signaling participated in IS-induced cyclin D transcription. In vivo, phosphorylated CREB (an active form of CREB) and ATF3 were prominently upregulated in the neointima of experimental uremic rats, the atherosclerotic plaques of uremic ApoE−/− mice, and the iliac arteries of uremic patients. Notably, the use of lentivirus to knock down ATF3 in the neointima of balloon-injured arteries could suppress the effect of uremia in vivo, including neointimal formation and cyclin D expression. Conclusions In this study, we demonstrated that CREB/ATF3-related signaling may be involved in IS-induced VSMC proliferation and the pathogenesis of neointimal formation during uremia.