microRNA-379在胚胎心脏发育及P19细胞分化过程中的表达规律  

Expression pattern of microRNA-379 during the embryo heart development and differentiation of P19 cells

作　　者：王丽华[1] 宋桂仙[2] 朱金改[3] 余章斌[3] 刘明[2] 陈斌[2] 周晓玉[1] 

机构地区：[1]南京医科大学附属南京儿童医院新生儿科,210008 [2]南京医科大学第一附属医院心脏科 [3]南京市妇幼保健院儿科

出　　处：《中华实用儿科临床杂志》2014年第24期1886-1888,共3页Chinese Journal of Applied Clinical Pediatrics

基　　金：国家自然科学基金（81270725）;江苏省自然科学基金（SBK201340683）

摘　　要：目的 观察microRNA （miR）-379在正常小鼠胚胎心脏发育过程中表达水平的变化,结合其在P19细胞向心肌细胞分化过程中的表达情况,探讨其在心脏发育中可能存在的作用.方法 取孕第8.5、11.5、14.5、18.5天小鼠胚胎的心脏组织,采用实时荧光定量PCR （RT-PCR）技术检测基因miR-379的表达变化.应用10 mL/L二甲基亚砜诱导P19细胞向心肌细胞方向分化,细菌培养皿中悬浮诱导至第4天,形成细胞聚集体后转移至6孑板培养,采用RT-PCR技术分别检测诱导第0、4、6、10天P19细胞中miR-379基因的表达水平.结果 miR-379在小鼠胚胎心脏发育过程中表达量逐渐减低,各时间点之间比较差异有统计学意义（F=21.13,P＜0.05）.另一方面,在P19细胞诱导分化过程中心肌标志物肌钙蛋白T呈递增趋势,证明P19细胞被成功诱导为心肌样细胞.在P19细胞向心肌细胞分化的第0天表达量较低,第4天表达量最高,随后表达量又逐渐减低.结论 miR-379参与心脏发育过程,且可能与先天性心脏病的发生发展有关,但是具体机制还需要进一步的研究。Objective To observe the expression changes in microRNA （miR）-379 in the developmental process of the mouse heart and during the differentiation of P19 cells into cardiac myocytes,and to explore the possible relationship between miR-379 and the differentiation of cardiacmyocytes.Methods Heart tissues were collected from fetal mice in pregnant ones at their gestational age （8.5,11.5,14.5 and 18.5 days） respectively.Heart tissue sections of the fatal mice were obtained to observe the heart development process.Then total RNA was isolated from heart tissues by using the TRIzol method.Complementary DNA was synthesized from 1 μg total RNA by using a Reverse Transcriptase Kit.Finally,real-time PCR （RT-PCR） was employed to detect the expression of miR-379.At the same time,P19 cells were cultured with 10 mL/L Dimethyl sulfoxide in suspension for 4 days to form cell aggregation,and these aggregations were transferred into 6-wells plate for culturing by adherence.Beating cells were detected with microscopy on the 10th day after induction.Afterwards,total RNA was extracted from cultured P19 cells at different time points.Reverse transcription was executed to get DNA.At last,RT-PCR was used to explore the expression of miR-379 on 0,4,6,10 days after aggregation.Results The expression level of miR-379 was down-regulated gradually in the developing heart （at gestational age of 8.5,11.5,14.5,16.5 days,respectively）,and there were significant differences on the different days （F =21.13,P ＜ 0.05）.On the other hand,myocardial markers of troponin T represented an increasing trend during the process of P19 cells induction,which demonstrated that P19 cells were successfully induced into cardiomyocyte-like cells.Meanwhile,miR-379 showed a low expression on day 0 of P19 cells aggregation.On day 4,miR-379 demonstrated a higher level.Afterwards,miR-379 proved to be down-regulated gradually.Conclusions miR-379 plays a role in the process of the heart development,but the specific mechanisms need further research.

关 键 词：microRNA-379 心脏发育 心肌组织 P19细胞 

分 类 号：R541.1[医药卫生—心血管疾病]