粉防己碱对先天性膈疝大鼠模型胎仔肺Rho/Rho激酶表达的影响和意义

目的 探讨中药粉防己碱(TET)产前干预对先天性膈疝(CDH)大鼠模型胎仔肺内RhoA蛋白和Rho激酶表达的影响和意义.方法 将孕9.5d的SD雌鼠随机分为对照组、膈疝组和TET组,给予膈疝组和TET组除草醚灌胃建立CDH动物模型.孕16.5d给予TET进行干预.孕21.5d剖宫产取出胎肺,测肺质量/体质量比(Lw/Bw),苏木素-伊红(HE)染色光镜下观察肺发育及肺血管情况,免疫组化和Western blot法检测Rho蛋白A及Rho激酶ROCK1的表达情况.结果 膈疝组胎肺明显发育不良,TET组胎肺发育接近对照组.膈疝组肺发育指标Lw/Bw、肺泡面积比(PAA％)及肺血管重构指标管腔面积与血管总面积比值(LA％)明显低于对照组(2.11±0.36 vs.4.24±0.31;33.60±3.12 vs.58.81±2.92;38.58±2.15 vs.61.20±3.23,均P＜0.05).TET干预后Lw/Bw、PAA％、LA％指标较膈疝组明显改善(3.61±0.24 vs.2.11±0.36;42.46±3.68 vs.33.60±3.12;56.07±3.32 vs.38.58±2.15,均P＜0.05);膈疝组肺小动脉管壁厚度占血管外径百分比(WT％)和肺小动脉中膜厚度百分比(MT％)均明显高于对照组(26.64±2.41 vs.13.50±1.45;25.98±2.79 vs.16.47±2.07,均P＜0.05),TET组WT％、MT％显著低于膈疝组(16.02±2.35 vs.26.64±2.41和17.96±1.95 vs.25.98±2.79,均P＜0.05).免疫组化及West-ern blot检测提示,RhoA、ROCK1表达由低到高为对照组、TET组、膈疝组(P=0.000).结论 先天性膈疝胎仔肺内存在肺发育不良和肺血管重构,Rho/Rho激酶信号通路参与此过程,产前给予TET可能通过调节Rho/Rho激酶信号通路发挥肺保护作用.

Effect and significance of tetrandrine on expression of Rho/Rho kinase in fetal lung of congenital diaphragmatic hernia rats

Objective To investigate the effect and significance of the traditional Chinese medicine tetrandrine(TET) prenatal intervention on the expression of RhoA protein and Rho kinase ROCK1 in the fetal lung of congenital diaphragmatic hernia (CDH) rat model.Methods SD female rats with 9.5 d of gestation were randomly divided into the control group,CDH group and TET group.The CDH group and TET intervention group were administered with nitrofen by gavage for establishing CDH model.The TET intervention was given on 16.5 d of gestation.The fetal rat lungs were taken by cesarean section on 21.5 d of gestation and the lung weight/body ratio(Lw/Bw) was measured.The lung development and small pulmonary arterial morphologic changes in HE staining in all groups were observed with microscopy.The protein expression of RhoA and Rho kinase ROCK1were respectively examined by immunohistochemistry and Western blot.Results In the CDH group,the lungs had obvious maldevelopment and the fetal lung development in the TET group was close to that in the control group.The lung development indicators of Lw/Bw,PAA％,and lung vascular remodeling indicators of lumen area and vascular total area ratio(LA％) in the CDH group were significantly lower than those in the control group(2.11±0.36 vs.4.24±0.31;33.60±3.12 vs.58.81 ±2.92;38.58±2.15 vs.61.20±3.23,P＜0.05),the indicators of Lw/Bw.PAA％ and LA％ after TET intervention were significantly improved compared with the CDH group(3.61±0.24 vs.2.11±0.36;42.46±3.68 vs.33.60±3.12;56.07±3.32 vs.38.58±2.15,all P ＜0.05);the ratio of small pulmonary artery wall thickness to vascular external diameter (WT％) and the medium thickness percentage (MT％) in CDH group were significantly higher than those in the control group(26.64±2.41 vs.13.50±1.45 and 25.98±2.79 vs.16.47±2.07,P＜0.05),WT％ and MT％ in the TET group were obviously lower than those in the CDH group (16.02±2.35 vs.26.64± 2.41 and 17.96 ± 1.95 vs.25.98 ± 2.79,P＜0.05).The immunohistochemistry and Western blot detection indicated that the expressions of RhoA and ROCK1 from low to high were the control group ＜TET group ＜ CDH group.Conclusion Pulmonary hypoplasia and lung vascular remodeling exist in fetal rats with CDH and Rho/Rho kinase signaling pathway may be involved in the process.Prenatally giving TET may play the lung protective effect by regulating the Rho/Rho signal pathway.