骨形成蛋白4在支气管肺发育不良相关肺动脉高压的肺微循环内皮细胞损伤中的机制研究  

Role of bone morphogenetic protein 4 in pulmonary microvascular endothelial cell damage in bronchopulmonary dysplasia-related pulmonary hypertension:A mechanistic study

作　　者：曾森强 刘蓉[2] 吴佩琼 谢志伟 黄建军 金颖康 Zeng Senqiang;Liu Rong;Wu Peiqiong;Xie Zhiwei;Huang Jianjun;Jin Yingkang(Department of Respiratory Diseases,Guangzhou Women and Children’s Medical Center,Guangzhou 510120;State Key Laboratory of Respiratory Diseases,First Affiliated Hospital of Guangzhou Medical University,Guangzhou)

机构地区：[1]广州市妇女儿童医疗中心呼吸科,广东广州510120 [2]广州医科大学附属第一医院,呼吸疾病国家重点实验室,广东广州510120

出　　处：《中华生物医学工程杂志》2021年第6期597-605,共9页Chinese Journal of Biomedical Engineering

基　　金：广东省自然科学基金面上项目(2018A030313965);广州市科技计划项目市校(院)联合资助项目(202102010143);广州市妇女儿童医疗中心博士启动基金(2018-2020)。

摘　　要：目的初步探讨骨形成蛋白4(BMP4)在低氧性支气管肺发育不良相关肺动脉高压(BPD-PH)的肺微循环内皮细胞(PMVEC)持续损伤中的作用机制。方法动物实验:制备低氧性BPD-PH小鼠模型,通过HE染色、免疫荧光、Western印迹比较BMP4基因半敲除(BmP4^(+/-))和野生型(wt)两组小鼠的肺组织形态学、肺微循环数量、肺血管内皮细胞形态和心脏功能;细胞实验:分离PMVEC,免疫荧光鉴定PMVEC,通过免疫印迹、细胞增殖和小管形成实验,分别以不同浓度人重组BMP4(rhBMP4)、不同程度低氧和BMP4抑制剂Noggin评价BMP4对PMVEC持续损伤的影响。结果动物实验发现与野生型相比,BmP4^(+/-)小鼠死亡率降低,体重增加,肺组织BPD样表现(辐射状肺泡计数增加和平均肺泡内衬间隔减少,P<0.05)和右心室肥厚指数均改善(P<0.05),肺微循环标记物血管内皮生长因子(VEGF)表达上调(P<0.01),肺血管内皮受损程度减轻;细胞实验发现:过高浓度的BMP4不利于PMVEC增殖和小管形成;低氧可上调PMVEC中的BMP4水平(P<0.01),2%氧浓度下PMVEC增殖受到抑制(P<0.01),小管成环减少(P<0.01),而加入BMP4抑制剂Noggin后,促进了PMVEC增殖(P<0.01),小管成环增加(P<0.05)。结论低氧引起的BMP4过度表达可以通过影响PMVEC增殖和小管形成导致肺微循环网减少,参与BPD-PH的发病机制。Objective To preliminarily explore the action mechanism of bone morphogenetic protein 4(BMP4)in sustained damage of pulmonary microvascular endothelial cells(PMVEC)in hypoxic bronchopulmonary dysplasia-related pulmonary hypertension(BPD-PH).Methods In the animal experiment,a hypoxic BPD-PH mouse model was established.Mice with BMP4 partial knockout(BmP4^(+/-))and wild-type(wt)mice were compared for lung histology,lung microcirculation,pulmonary vascular endothelial cell morphology and heart function with HE staining,immunofluorescence,and Western blotting.In the cellular experiment,PMVECs were isolated,and identified by immunofluorescence.Western-blotting,cell proliferation assay and tube formation assay were used to determine the effect of BMP4 on sustained damage of PMVECs under different concentrations of human recombinant BMP4(rhBMP4),varying degrees of hypoxia and in the presence of BMP4 inhibitor Noggin.Results Our animal experiment showed that compared with wild-type mice,BmP4^(+/-)mice had lower mortality,higher body weight,improvements in BPD-like appearance of lung tissues(increased radial alveolar count and decreased mean lining intercept,P<0.05)and right ventricular hypertrophy(P<0.05),up-regulated expression of the pulmonary microcirculation marker vascular endothelial growth factor(VEGF)(P<0.01),and milder damage to the pulmonary vascular endothelium.Our cellular experiment showed that excessively high concentrations of BMP4 were not conducive to proliferation and tube formation of PMVECs.Hypoxia can up-regulate BMP4 expression in PMVECs(P<0.01).PMVECs showed suppressed proliferation(P<0.01)and less tube formation(P<0.01)under 2%oxygen.Addition of the BMP4 inhibitor Noggin was found to promote the proliferation(P<0.01)and increased tube formation(P<0.05)of PMVECs.Conclusion Overexpression of BMP4 caused by hypoxia can reduce the pulmonary microcirculation network by affecting proliferation and tube formation of PMVECs,and thereby participates in the pathogenesis of BPD-PH.

关 键 词：支气管肺发育不良 肺动脉高压 骨形成蛋白4 低氧 肺微循环内皮细胞 

分 类 号：R544.1[医药卫生—心血管疾病]