Department of Biochemistry and Molecular Biology, The Key Laboratory of Neural and Vascular Biology, China Administration of Education, Hebei Medical University, Shijiazhuang, China.
Department of Biochemistry and Molecular Biology, Hebei University of Chinese Medicine, Shijiazhuang, China.
Theranostics. 2020 Jun 19;10(17):7787-7811. doi: 10.7150/thno.46911. eCollection 2020.
Abnormal proliferation and migration of vascular smooth muscle cells VSMCs) are essential for vascular remodeling. Natural compounds with diterpene chinone or phenolic acid structure from an eminent medicinal herb widely used to treat cardiovascular diseases in China, can effectively attenuate vascular remodeling induced by vascular injury. However, it remains unknown whether -derived miRNAs can protect VSMCs from injury by environmental stimuli. Here, we explored the role and underlying mechanisms of -derived Sal-miR-1 and 3 in the regulation of VSMC migration and monocyte adhesion to VSMCs induced by thrombin. A mouse model for intimal hyperplasia was established by the ligation of carotid artery and the injured carotid arteries were in situ-transfected with Sal-miR-1 and 3 using F-127 pluronic gel. The vascular protective effects of Sal-miR-1 and 3 were assessed via analysis of intimal hyperplasia with pathological morphology. VSMC migration and adhesion were analyzed by the wound healing, transwell membrane assays, and time-lapse imaging experiment. Using loss- and gain-of-function approaches, Sal-miR-1 and 3 regulation of OTUD7B/KLF4/NMHC IIA axis was investigated by using luciferase assay, co-immunoprecipitation, chromatin immunoprecipitation, western blotting, etc. derived Sal-miR-1 and 3 can enter the mouse body after intragastric administration, and significantly suppress intimal hyperplasia induced by carotid artery ligation. In cultured VSMCs, these two miRNAs inhibit thrombin-induced the migration of VSMCs and monocyte adhesion to VSMCs. Mechanistically, Sal-miR-1 and 3 abrogate OTUD7B upregulation by thrombin via binding to the different sites of the OTUD7B 3'UTR. Most importantly, OTUD7B downregulation by Sal-miR-1 and 3 attenuates KLF4 protein levels via decreasing its deubiquitylation, whereas decreased KLF4 relieves its repression of transcription of NMHC IIA gene and thus increases NMHC IIA expression levels. Further, increased NMHC IIA represses VSMC migration and monocyte adhesion to VSMCs via maintaining the contractile phenotype of VSMCs. Our studies not only found the novel bioactive components from but also clarified the molecular mechanism underlying Sal-miR-1 and 3 inhibition of VSMC migration and monocyte adhesion to VSMCs. These results add important knowledge to the pharmacological actions and bioactive components of . Sal-miR-1 and 3-regulated OTUD7B/KLF4/NMHC IIA axis may represent a therapeutic target for vascular remodeling.
异常的血管平滑肌细胞（VSMCs）增殖和迁移是血管重构的关键。天然化合物中二萜醌或酚酸结构，来自中国一种著名的药用草药，广泛用于治疗心血管疾病，可有效减轻血管损伤引起的血管重构。然而，尚不清楚 - 衍生的 miRNA 是否可以保护 VSMCs 免受环境刺激的损伤。在这里，我们研究了 - 衍生的 Sal-miR-1 和 3 在调节凝血酶诱导的 VSMC 迁移和单核细胞与 VSMCs 黏附中的作用及其潜在机制。通过颈总动脉结扎建立内膜增生的小鼠模型，并使用 F-127 pluronic 凝胶原位转染 Sal-miR-1 和 3。通过病理形态学分析评估 Sal-miR-1 和 3 的血管保护作用。通过划痕愈合、Transwell 膜测定和延时成像实验分析 VSMC 迁移和黏附。通过荧光素酶测定、共免疫沉淀、染色质免疫沉淀、Western blot 等方法，研究了 Sal-miR-1 和 3 对 OTUD7B/KLF4/NMHC IIA 轴的调控作用。结果表明，Sal-miR-1 和 3 可以通过灌胃进入小鼠体内，并显著抑制颈总动脉结扎引起的内膜增生。在培养的 VSMCs 中，这两种 miRNA 抑制凝血酶诱导的 VSMC 迁移和单核细胞与 VSMCs 的黏附。机制上，Sal-miR-1 和 3 通过结合 OTUD7B 3'UTR 的不同位点来拮抗凝血酶诱导的 OTUD7B 上调。最重要的是，Sal-miR-1 和 3 通过减少其去泛素化来降低 KLF4 蛋白水平，而降低的 KLF4 则解除其对 NMHC IIA 基因转录的抑制，从而增加 NMHC IIA 表达水平。进一步研究表明，增加的 NMHC IIA 通过维持 VSMCs 的收缩表型来抑制 VSMC 迁移和单核细胞与 VSMCs 的黏附。我们的研究不仅发现了 中的新型生物活性成分，还阐明了 Sal-miR-1 和 3 抑制 VSMC 迁移和单核细胞与 VSMCs 黏附的分子机制。这些结果为 的药理学作用和生物活性成分增添了重要知识。Sal-miR-1 和 3 调节的 OTUD7B/KLF4/NMHC IIA 轴可能代表血管重构的治疗靶点。
