Down Regulation of Let-7 Triggers a Critical Double-Negative Feedback Loop to Determinate the Malignant VSMCs Fate During Restenosis

Background: The excessive proliferation and migration of VSMCs is the main cause of restenosis (RS). Our previous results suggested that overexpression of Lin28a is responsible for this “malignant” behaver of VSMCs. However, the mechanisms driving the specific VSMCs behavior in RS are far less understood. Methods and Findings: Using an atherosclerosis followed by PTA treatment rats as RS model, we discovered a decreased expression of let-7c, let-7g, let-7i, miR98 and miR202, that associated with an increased expression of Lin28a in RS plaques compared to those of atherosclerosis. Combined with in vitro results, we demonstrated that let-7c, let-7g, and miR98 were Lin28a downstream targets in RS. More importantly, the decreased expression of let-7c, let-7g, in turn, will enhance the expression of Lin28a, indicating a double-negative feedback loop between Lin28a and let-7c/let-7g, which may be responsible for the vicious behavior of the VSMCs in RS. Transfection of let-7c and let-7g in vivo by AAV9 attenuated the aggregation of VSMCs and decreased the RS plaque size. In addition, unlike the other member of let-7, let-7d downregulates Lin28a and plays a restrictive regulator on the Lin28a/let-7 loop, that may bias VSMCs fate decision. Conclusion: In this study, by using a “two-step lesion” protocol to create RS models, we discovered that the double negative feedback system of Lin28a/let-7 axis act as a regulation center in vicious behavior of VSMCs in RS. These discoveries give us a better understanding of RS in lower extremity arterial disease and expected to present opportunities for discovering new therapeutic approaches. Funding: This work was supported by the National Natural Science Foundation of China Grants (No. 81670757, 81770822, 81800732, 81900685), Shandong Provincial Natural Science Foundation of China Grants (No. ZR2017LH025, ZR2020MH101), Shandong Provincial Medicine and Health Science and Technology Development Program (No. 2017WS461), China Postdoctoral Science Foundation (No. 2020M682193). Declaration of Interest: The authors confirm that there are no conflicts of interest. Ethical Approval: The protocol was authorized by the Ethics Committee of Qilu Hospital, Shandong University.