miR-424在人骨髓间充质干细胞分化过程中的表达  

Expressions of miR-424 during differentiation of human bone marrow-derived mesenchymal stem cells

作　　者：高杰[1] 韩建伟[1] 朱洪繁[1] 杨彤涛[1] 范清宇[1] 马保安[1] 

机构地区：[1]第四军医大学附属唐都医院骨科,西安710038

出　　处：《中华创伤杂志》2010年第8期752-756,共5页Chinese Journal of Trauma

基　　金：国家自然科学基金面上资助项目（30672143）

摘　　要：目的 筛选可作为人骨髓间充质干细胞（MSCs）分子标记物的microRNA（miRNA）.方法 以从骨髓中分离培养的MSCs及其诱导分化的成骨细胞和软骨细胞为实验对象,利用基因芯片检测miRNA的表达情况,由芯片显著性分析（SAM）得到MSCs较其诱导分化细胞高表达的miRNA,再由实时定量PCR进行验证.结果 成功分离培养出MSCs,并分别诱导其分化为成骨细胞和软骨细胞;MSCs及由其诱导分化的成骨细胞和软骨细胞经miRNA芯片检测及SAM分析得到8个MSCs较由其诱导分化的成骨细胞高表达miRNA（has-miR-424、has-miR-34a、has-miR-593、has-miR-10a、has-miR-148a、has-miR-602、mmu-miR-709、mmu-miR-665）,3个MSCs较由其诱导分化的软骨细胞高表达miRNA（has-miR-424、PREDICTED_MIR189、mmu-miR-665）.其中MSCs较成骨细胞和软骨细胞均高表达的人源性miRNA为has-miR-424,差异倍数分别为6.6倍和4.4倍;在原样本中对进行实时定量PCR的验证,结果提示与诱导分化的成骨细胞相比,MSCs中的has-miR-424表达升高约3.6倍,而与诱导分化的软骨细胞相比,MSCs中的has-miR-424表达升高约3.1倍,结果与芯片结果相符合.结论 MSCs较其诱导分化细胞高表达的miRNA有望成为人骨髓间充质干细胞的一种特异性的分子标记物,这些miRNA对MSCs自我更新和未分化状态起着重要的维持作用.Objective To identify the specific microRNA （miRNA） that can be taken as a molecular marker for human bone marrow-derived mesenchymal stem cells （MSCs）. Methods MSCs were isolated and cultured from bone marrow through density centrifugation and then were induced to differentiate into osteoblasts and chondrocytes. Samples of MSCs, osteoblasts and chondrocytes were detected by miRNA microarrays single channel fluorescence chip to determine the expression levels of miRNAs. Significance Analysis of Microarrays （ SAM, version 2.1 ） software was used to analyze the raw data to determine the miRNAs overexpressed in MSCs, which was validated in the same sample using real time reserve transcriptase polymerase chain reaction （RT-PCR）. Results MSCs were successfully isolated from bone marrow and induced to differentiate into osteoblasts and chondrocytes in vitro. Microarrays showed that eight miRNAs （has-miR-424, has-miR-34a, has-miR-593, has-miR-10a, has-miR-148a,has-miR-602, mmu-miR-709 and mmu-miR-665） were overexpressed in MSCs but underexpressed in osteoblasts. Three miRNAs including has-miR-424, PREDICTED_MIR189 and mmu-miR-665, were overexpressed in MSCs but underexpressed in chondrocytes. The has-miR-424 expression in MSCs was 6.6times higher than in osteoblasts and 4.4 times higher than in chondrocytes. The results of real time RTPCR showed that the miR-424 was overexpressed in MSCs, 3.6 times higher than that in osteoblasts and 3.1 times higher than that in chondrocytes, which was coincident with the results of microarray. Conclusions The screened MSCs express more miRNAs in comparison with osteoblasts and chondrocytes,play important roles in maintaining self renewal and undifferentiation of MSCs and is a promising specific molecule marker for MSCs.

关 键 词：骨髓 间充质干细胞 MICRORNA 

分 类 号：R686[医药卫生—骨科学]