CircRNA YAP1对股骨头微血管内皮细胞血管生成的作用研究  

Effect of CircRNA YAP1 on Angiogenesis of Femoral Head Microvascular Endothelial Cells

作　　者：日夏提·帕尔哈提 翟生[1] 吕青[1] RIXIATI Paerhati;ZHAI Sheng;LV Qing(The Fifth Affiliated Hospital of Xinjiang Medical University,Xinjiang Urumqi 830000,China)

机构地区：[1]新疆医科大学第五附属医院骨科中心,新疆乌鲁木齐830000

出　　处：《河北医学》2023年第6期899-906,共8页Hebei Medicine

基　　金：新疆维吾尔自治区自然科学基金项目,(编号:2021D01C426)。

摘　　要：目的:探讨环状RNA(circRNA)YAP1对激素性股骨头坏死(SONFH)大鼠股骨头微血管内皮细胞(MVECs)血管生成的作用与机制。方法:成年雄性SD大鼠接受甲基强的松龙(MPS,20mg·kg^(-1)·d^(-1)肌肉注射3周,以诱导SONFH模型(n=30只)。Ctrl组(n=30只)注射等量生理盐水。用micro-CT扫描和HE染色评估是否造模成功。分离大鼠的股骨头MVECs。并用CD31蛋白作为标志物用免疫荧光化学法(IF)鉴定股骨头MVECs。将MVECs分为过表达circRNA YAP1组(pcDNA-YAP1组)、阴性对照组(pcDNA-NC组)、以及pcDNA-YAP1联合Wnt1/β-catenin通路拮抗剂dickkopf-1处理组(pcDNA-YAP1+dickkopf-1组)。用qPCR检测circRNA YAP1表达,Western blot法分别检测Wnt1、β-catenin、VEGF-A和vimentin的表达。用CCK-8实验检测各组细胞的增殖活性。Transwell小室检测细胞的迁移率。管形成实验检测细胞的血管生成能力。结果:HE染色结果显示Ctrl组表现为健康状态,无骨坏死,SONFH组大鼠均重度股骨头坏死,股骨头恶化。micro-CT扫描的结果显示Ctrl组表现骨小梁致密规则,骨小梁数量和形态均正常,SONFH组伴空骨陷窝,骨小梁减少,骨小梁缩短。与Ctrl组(1.00±0.08;1.00±0.12;1.00±0.15)比较,SONFH组股骨组织中circRNA YAP1(0.27±0.04)、Wnt1(0.49±0.03)、β-catenin(0.33±0.03)表达量均显著减少(t=12.158,P=0.009;t=10.596,P=0.012;t=10.080,P=0.014)。另外,成功分离Ctrl组和SONFH组的股骨头MVECs。与Ctrl组(1.00±0.00;1.00±0.02;1.00±0.01)比,SONFH组股骨头MVECs中circRNA YAP1(0.15±0.01)、Wnt1(0.28±0.05)、β-catenin(0.31±0.02)表达量都显著减少(t=17.625,P=0.004;t=16.154,P=0.005;t=13.596,P=0.007)。在MVECs中,与pcDNA-NC组比,pcDNA-YAP1组的circRNA YAP1、Wnt1、β-catenin、VEGF-A和vimentin的表达均上调,细胞增殖率、细胞迁移以及小管形成率都明显增加(均P<0.05)。与pcDNA-YAP1组比,pcDNA-YAP1+dickkopf-1组的circRNA YAP1表达变化无统计学意义(P>0.05),而Wnt1、β-catenin、VEGF-A和vimentin的表达均Objective:To explore the role and mechanism of circular RNA(circRNA)YAP1 in the regulation of microvascular endothelial cell(MVEC)angiogenesis in steroid-induced osteonecrosis of the femoral head(SONFH)rats.Methods:Adult male SD rats were injected with methylprednisolone(MPS,20 mg/kg/d)for 3 weeks to induce the SONFH model(n=30).The control group(n=30)was injected with an equal amount of saline.Micro-CT scanning and HE staining were performed to evaluate whether the model was successfully established.MVECs were isolated from the femoral heads of the rats using CD31 as a marker and identified by immunofluorescence staining.The MVECs were divided into three groups:the pcDNA-YAP1 group(overexpression of circRNA YAP1),the pcDNA-NC group(negative control),and the pcDNA-YAP1+dickkopf-1 group(treated with Wnt1/β-catenin pathway inhibitor dickkopf-1).The expression of circRNA YAP1 was detected using qPCR,while the expressions of Wnt1,β-catenin,VEGF-A,and vimentin were detected using Western blotting.Cell proliferation was detected using the CCK-8 assay,while cell migration was detected using the Transwell assay.Tube formation assays were performed to detect the angiogenic ability of the cells.Results:The HE staining results showed that the control group was healthy,with no bone necrosis,while the SONFH group showed severe femoral head necrosis and deterioration.The results of micro-CT scanning showed that the control group had dense and regular bone trabeculae,with normal numbers and morphology of bone trabeculae.The SONFH group was accompanied by an empty bone pit,a decrease in the number of bone trabeculae,and shortened bone trabeculae.Compared with the control group(1.00±0.08;1.00±0.12;1.00±0.15),the expressions of circRNA YAP1(0.27±0.04),Wnt1(0.49±0.03),andβ-catenin(0.33±0.03)in the SONFH group were significantly reduced(t=12.158,P=0.009;t=10.596,P=0.012;t=10.080,P=0.014).In addition,MVECs from the femoral heads of the Ctrl and SONFH groups were successfully isolated.Compared with the control group(1.00±0.0

关 键 词：激素性股骨头坏死 环状RNA YAP1 Wnt1/β-catenin信号通路 股骨头微血管内皮细胞 血管生成 

分 类 号：R681.8[医药卫生—骨科学]