circ_TGFBR2 Inhibits Vascular Smooth Muscle Cells Phenotypic Switch and Suppresses Aortic Dissection Progression by Sponging miR-29a.
circ_TGFBR2 Inhibits Vascular Smooth Muscle Cells Phenotypic Switch and Suppresses Aortic Dissection Progression by Sponging miR-29a.
复制标题
Circ_TGFBR2 通过海绵 miR-29a 抑制血管平滑肌细胞表型转换并抑制主动脉夹层进展
DOI：
10.2147/jir.s336094
复制
发表时间：
2021
影响因子：
4.5
通讯作者：
Pan J
中科院分区：
文献类型：
--
作者：
Xu Z;Zhong K;Guo G;Xu C;Song Z;Wang D;Pan J
主动脉夹层（Aortic夹层，AD）是一种具有高致死率且治疗手段有限的致命性血管病。越来越多的证据表明，环状rna在调节各种心血管疾病中起着至关重要的作用。然而，circRNAs在AD中的生物学功能和分子机制尚不清楚。本研究的目的是阐明hsa_circ_TGFBR2在体外和体内的潜在功能作用和机制。从正常主动脉和AD组织中分离血管平滑肌细胞（VSMCs）和AD-VSMCs。采用实时聚合酶链反应（RT-PCR）和荧光原位杂交（FISH）检测circ_TGFBR2、miR-29a和KLF4的表达。采用CCK-8法、菌落形成法和EDU法检测细胞增殖情况。通过transwell实验评估细胞迁移。通过双荧光素酶报告基因测定和RNA下拉来鉴定circ_TGFBR2与miR-29a或miR-29a与KLF4之间的相互作用。将circ_TGFBR2或KLF4野生型序列克隆到荧光素酶报告基因质粒中，采用双荧光素酶报告基因检测系统测定其活性。对于RNA下拉，使用RT-PCR确认circ_TGFBR2和miR-29a的相对RNA富集。Western Blot检测表型开关相关蛋白的表达。采用β-单马来酸氨基丙腈（BAPN）诱导AD大鼠模型验证circ_TGFBR2的作用和机制。Circ_TGFBR2抑制AD-VSMCs细胞的增殖和迁移。过表达circ_TGFBR2可促进AD-VSMCs细胞中收缩性标记物（α-SMA、SM22α）的表达，抑制合成标记物（MGP、OPN）的表达。Circ_TGFBR2作为靶向KLF4的miR-29a的海绵。MiR-29a模拟物挽救了circ_TGFBR2过表达诱导的生物学作用。体内实验显示，过表达TGFBR2抑制AD的进展，增加收缩标记物的表达，抑制合成标记物的表达。我们的研究发现circ_TGFBR2调节VSMCs表型开关，抑制AD的进展。
Aortic dissection (AD) is a threatening and catastrophic vascular disease with high mortality rate and limited therapeutic strategies. There is emerging evidence showing that circular RNAs play crucial role in regulating various cardiovascular diseases. However, the biological functions and molecular mechanisms of circRNAs in AD still remains elusive. The purpose of this study was to illustrate the potential functional roles and mechanisms of hsa_circ_TGFBR2 in vitro and in vivo. The vascular smooth muscle cells (VSMCs) and AD-VSMCs were isolated from normal aorta and AD tissues. The expression of circ_TGFBR2, miR-29a and KLF4 were detected by realtime polymerase chain reaction (RT-PCR) and fluorescence in situ hybridization (FISH). Cell proliferation was assessed by CCK-8 assay, colony formation and EDU assay. Cell migration was evaluated through transwell assay. Dual-luciferase reporter assay and RNA pulldown were performed to identify the interaction between circ_TGFBR2 and miR-29a or between miR-29a and KLF4. The wild-type sequence of circ_TGFBR2 or KLF4 were cloned into the luciferase reporter plasmid, and the activity was measured using dual-luciferase reporter assay system. And for RNA pulldown, the relative RNA enrichment of circ_TGFBR2 and miR-29a were confirmed using RT-PCR. Western Blot measured the expression of phenotype switch-related proteins. AD rat model induced by β-aminopropionitrile monofumarate (BAPN) was used to verify the role and mechanism of circ_TGFBR2. Circ_TGFBR2 inhibited cell proliferation and migration of AD-VSMCs cells. Overexpression of circ_TGFBR2 promoted the expression of contractile markers (α-SMA, SM22α) and inhibited the expression of synthetic markers (MGP, OPN) in AD-VSMCs cells. Circ_TGFBR2 served as a sponge for miR-29a targeting KLF4. MiR-29a mimics rescued biological roles induced by circ_TGFBR2 overexpression. The in vivo experiments revealed that overexpression of TGFBR2 suppressed the progression of AD and increased the expression of contractile markers while inhibited the expression of synthetic markers. Our study revealed that circ_TGFBR2 regulated VSMCs phenotype switch and suppressed the progression of AD.