HGF/c-Met轴介导EPCs治疗大鼠HPAH的初步实验研究

Effects of HGF/c-Met-mediated endothelial progenitor cells on pulmonary artery pressure and pulmonary vascular remodeling in rats with hypoxic pulmonary hypertension: A preliminary experimental study

摘要:

目的：探讨肝细胞生长因子（hepatocyte growth factor，HGF）/c-Met轴介导内皮祖细胞（endothelial progenitor cells，EPCs）移植入低氧性肺动脉高压（hypoxic pulmonary artery hypertension，HPAH）大鼠对大鼠的肺动脉压力和肺血管重构的影响。方法：构建HPAH模型大鼠；大鼠骨髓原代EPCs提取、培养和鉴定；构建c-Met腺病毒并转染EPCs；将大鼠分为c-Met-EPCs细胞组、EPCs细胞组、肺动脉高压组和正常组，每组10只。前2组大鼠通过尾静脉分别输注c-Met-EPCs细胞和EPCs细胞，后2组大鼠输注生理盐水；饲养一周后检测平均肺动脉压力（mean pulmonary arterial pressure，mPAP）和右心室肥大指数（right ventricle hypertrophy index，RVHI）；透射电镜检测肺动脉超微结构改变，肺组织病理切片观察肺小动脉改变；ELISA法测定血清内皮素-1（endothelin-1，ET-1）含量，Griess法测定血清一氧化氮（nitric oxide，NO）含量。结果：c-Met-EPCs细胞组、EPCs细胞组、肺动脉高压组和正常组4组大鼠的mPAP值分别为（23.17±3.07）、（30.85±3.15）、（33.55±5.47）、（20.30±1.99） mmHg；RVHI值分别为（32.48±2.38）%、（37.54±4.42）%、（38.53±2.81）%、（26.05±3.05）%；血清ET-1值分别为（69.45±6.32）、（95.76±7.31）、（99.14±8.39）、（62.35±6.06） ng/L；血清NO值分别为（99.63±11.40）、（56.07±3.32）、（48.83±6.56）、（125.50±12.26） μmol/L。EPCs细胞组、肺动脉高压组大鼠的mPAP、RVHI和血清ET-1含量明显高于正常组（P<0.05）；c-Met-EPCs细胞组较EPCs细胞组、肺动脉高压组明显降低（P<0.05）。EPCs细胞组、肺动脉高压组大鼠的血清NO含量明显低于正常组（P<0.05）；c-Met-EPCs细胞组较EPCs细胞组、肺动脉高压组明显升高（P<0.05）。透射电镜显示EPCs细胞组、肺动脉高压组肺动脉中膜平滑肌细胞线粒体和内质网肿胀、内皮细胞中断不连续，而c-Met-EPCs细胞肺动脉中膜平滑肌细胞线粒体和内质网肿胀不显著。肺组织病理切片显示EPCs细胞组、肺动脉高压组肺血管重构明显，而c-Met-EPCs细胞组肺血管重构减轻。结论：HGF/c-Met轴介导内皮祖细胞移植入低氧性肺动脉高压大鼠后可明显降低肺动脉压力和和改善肺血管重构，其机制可能与抑制ET-1释放和促进NO产生有关。

Abstract:

Objective：To investigate the effects of hepatocyte growth factor（HGF）/c-Met-mediated endothelial progenitor cells（EPCs） on pulmonary artery pressure（PAP） and pulmonary vascular remodeling in rats with hypoxic pulmonary hypertension（HPH）. Methods：A rat model of HPH was established. Primary EPCs were iso－lated from the bone marrow of rats，cultured，and identified. The recombinant adenovirus plasmid carrying the c-Met gene was constructed and transfected into EPCs. The rats were divided into c-Met-EPCs group，EPCs group，pulmonary hypertension group，and normal group，with 10 rats in each group. c-Met-EPCs and EPCs were infused into the tail vein of rats in the c-Met-EPCs group and EPCs group，respectively，while normal saline was in－fused into the tail vein of rats in the pulmonary hypertension group and normal group. After normal feeding for a week，the mean PAP（mPAP） and right ventricular hypertrophy index（RVHI） were determined. The ultrastructural pathology of the pulmonary artery was observed under a transmission electron microscope，and the pathological sections of lung tissues were prepared to observe the histopathological changes of pulmonary arterioles. ELISA was used to determine the serum level of endothelin-1（ET-1），and the Griess method was used to determine the serum level of nitric oxide（NO）. Results：The measured values of mPAP for the c-Met-EPCs group，EPCs group，pulmonary hypertension group，and normal group were （23.17±3.07） mmHg，（30.85±3.15） mmHg，（33.55±5.47） mmHg，and （20.30±1.99） mmHg，respectively；the measured values of RVHI for the four groups were （32.48±2.38）%，（37.54±4.42）%，（38.53±2.81）%，and （26.05±3.05）%，respectively；the serum level of ET-1 for the four groups were （69.45±6.32） ng/L，（95.76±7.31） ng/L，（99.14±8.39） ng/L，and （62.35±6.06） ng/L，respectively；the serum level of NO for the four groups were （99.63±11.40） μmol/L，（56.07±3.32） μmol/L，（48.83±6.56） μmol/L，and （125.50±12.26） μmol/L，respectively. The mPAP，RVHI，and serum ET-1 level in the EPCs group and pulmonary hypertension group were significantly higher than those in the normal group（P<0.05），and the above-mentioned indices were significantly lower in the c-Met-EPCs group than in the EPCs group and pulmonary hypertension group（P<0.05）. The serum NO level in the EPCs group and pul－monary hypertension group was significantly lower than that in the normal group（P<0.05），and the serum NO level in the c-Met-EPCs group was significantly higher than that in the EPCs group and pulmonary hypertension group（P<0.05）. Transmission electron mi－croscopy showed obvious swelling of mitochondria and endoplasmic reticulum in pulmonary artery smooth muscle cells and dis－continuous distribution of endothelial cells in the EPCs group and pulmonary hypertension group，while swelling of mitochondria and endoplasmic reticulum in pulmonary artery smooth muscle cells was not obvious in the c-Met-EPCs group. The pathological sections of lung tissues showed obvious pulmonary vascular remodeling in the EPCs group and pulmonary hypertension group；however，the remodeling was reduced in the c-Met-EPCs group. Conclusion：HGF/c-Met-mediated EPCs can decrease PAP and improve pul－monary vascular remodeling in rats with HPH，possibly by inhibiting ET-1 release and promoting NO production.