胎羊体外循环通过microRNAs导致胎心功能失调

Differential expression of myocardial microRNAs result in cardiac dysfunction after cardiopulmonary bypass in fetal goats

目的本文重点研究胎羊体外循环影响心肌microRNAs的表达情况，探索microRNAs导致胎羊体外循环（CPB）后胎心功能失调的可能机制。方法孕120—140d母羊6只，其中两只孕羊有双胎，共孕8只胎羊，随机分为对照组（n=4）和胎羊CPB组（n=4）。对照组胎羊开胸，不建立CPB。CPB组建立胎羊心脏转流模型，转流30rain。分别于转流前和转流30min结束后测胎羊心率和血肌钙蛋白I（TnI）的变化，B超检测左、右心室Tei指数。取胎羊心尖部心肌组织，采用microRNA芯片和荧光定量聚合酶链式反应（PCR）技术检测两组心肌组织的microRNAs的表达水平。使用生物信息学方法预测microR—NAs的靶基因，并进行蛋白印迹实验（WB）验证。结果两组胎羊心率变化无统计学差异（P〉0．05）。CPB组胎羊左、右心室Tei指数、血TnI含量均显著大于对照组（P〈0．05）。两组的microRNAs表达谱显著不同，有19个microRNAs显著差异性表达，其中11个microRNAs（miR-125b＊，-151，-127，-144＊，-141，-199b，-224，-301a，-30e＊，-374a和-424＊）在组织中被上调，而8个microRNAs（miR-99a，-151＊，-21，-223，-320，-129，let-7a和-7e）被下调。肌浆内质网Ca^2＋转运蛋白2（SERCA2）是miR-151的靶基因，并在CPB组的心肌组织中被明显下调。结论MicroRNAs的差异性表达与胎羊体外循环后胎心功能失调密切相关。可能是通过miR-151／SERCA2通路来实现。

Objective This study focuses on evaluating the different expression of myocardial microRNAs of fetal lamb, and exploring the possible mechanism of fetal cardiac dysfunction after cardiopulmonary bypass. Methods Six ewes at gestation of 120 to 140 days have eight fetuses which were randomly divided into control group （n= 4） and fetal cardiopulmonary bypass group （n= 4）. Control group underwent sham procedure that fetal stemotomy was performed. Bypass group underwent fetal cardiac bypass with centrifugal pump and placenta for 30 minute. Fetal heart rate and plasma troponin I were recorded before bypass and 30 minutes during bypass. Tei index of two ventricles were also recorded with uhrasonography. Expression levels of microRNAs were obtained from examination of apical myocardial tissues through mieroRNA microarray and quantitative real time polymerase chain reaction （qRT-PCR）. By using bioinformatics methods to predict the target genes of microRNAs, and then verify these results by Western blot experiments （WB）. Results There was no statistical difference between two groups in heart rate. Tie index of two ventricles and plasma troponin I of the bypass group increased remarkably compared with the control group. In the bypass group, microRNAs expression profiling were significantly different by more than two-fold compared to the control group. There are 19 microRNAs significantly differentially expressed, and 11 mieroRNAs were up-regnlated （miR-125b ＊ , -151, -127, -144 ＊ , -141, -199b, -224, -301a, -30e ＊ , -374a and -424 ＊ ）, whereas 8 were down-regulated （miR-99a, -151 ＊ , -21, -223, -320, -129, let-7a and -7e）. By Western blot analysis, it was found that the ATPase sarcoplasmic/endoplasmic reticulum Ca^2＋ transporting 2 （SERCA2） was significantly downregulated in apical myo- cardial tissues of the bypass group, and miR-151 may be involved in this process. Conclusion We demonstrated an association between microRNAs and fetal cardiac dysfunction, consistent with its known mechanism of the miR-151/SERCA2 pathway. Because the management of fetal cardiac dysfunction is often difficult, we suggest that microRNAs should be further studied as a potential treatment.