摘要
为探讨人胚胎骨髓间充质干细胞(mesenchymal stem cells from human fetal bone marrow,hfBM-MSCs)的增殖能力及其向三个胚层起源细胞分化的"类胚胎干细胞"特性,本文利用细胞差速贴壁生长特性分离纯化hfBM-MSCs,用流式细胞仪检测hfBM-MSCs的细胞周期和MSCs特异的表面标志;用免疫细胞化学方法和RT-PCR法分别在蛋白和RNA水平检测人端粒酶逆转录酶(human telomerase reverse transcriptase,hTERT)和胚胎干细胞特异性抗原Oct4、SSEA-4的表达;用各种化学试剂诱导hfBM-MSCs向神经元、脂肪细胞和胰岛B细胞分化并加以鉴定。流式细胞仪检测结果显示,第4、5代hfBM-MSCs分别有92.3%和96.1%的细胞处于G0/G1期;这些细胞表达CD29、CD44和CD106,不表达造血细胞标志CD34和CD45,极低地表达与移植物抗宿主病(graft-versus-host disease,GVHD)相关的HLA-DR、CD40和CD80。免疫细胞化学和RT-PCR结果显示,hfBM-MSCs表达Oct4、SSEA-4等胚胎干细胞标志,hTERT也呈阳性表达。在特定诱导条件下,hfBM-MSCs可分化为神经元样细胞(外胚层)、脂肪样细胞(中胚层)和胰岛B样细胞(内胚层)。因此,人胚胎骨髓中含有丰富的MSCs,hfBM-MSCs具有胚胎干细胞的主要生物学特性,且免疫原性弱,是组织工程和细胞治疗较为理想的种子细胞。
Strong proliferative capacity and the ability to differentiate into the derivative cell types of three embryonic germ layers are the two important characteristics of embryonic stem cells. To study whether the mesenchymal stem cells from human fetal bone marrow （hfBM-MSCs） possess these embryonic stem cell-like biological characteristics, hfBM-MSCs were isolated from bone barrows and further purified according to the different adherence of different kinds of cells to the wall of culture flask. The cell cycle of hfBM-MSCs and MSC-specific surface markers such as CD29, CD44, etc were identified using flow cytometry. The expressions of human telomerase reverse transcriptase （hTERT）, the embryonic stem cell-specific antigens, such as Oct4 and SSEA-4 were detected with immunocytochemistry at the protein level and were also tested by RT-PCR at the mRNA level. Then, hfBM-MSCs were induced to differentiate toward neuron cells, adipose cells, and islet B cells under certain conditions. It was found that 92.3% passage-4 hfBM- MSCs and 96.1% passage-5 hfBM-MSCs were at G0/G1 phase respectively, hfBM-MSCs expressed CD44, CD 106 and adhesion molecule CD29, but not antigens of hematopoietic cells CD34 and CD45, and almost not antigens related to graft-versus-host disease （GVHD）, such as HLA-DR, CD40 and CD80. hfBM-MSCs expressed the embryonic stem cell-specific antigens such as Oct4, SSEA-4,and also hTERT. Exposure of these cells to various inductive agents resulted in morphological changes towards neuron-like cells, adipose-like cells, and islet B-like cells and they were tested to be positive for related characteristic markers. These results suggest that there are plenty of MSCs in human fetal bone marrow, and hfBM-MSCs possess the embryonic stem cell-like biological characteristics, moreover, they have a lower immunogenic nature. Thus, hfBM-MSCs provide an ideal source for tissue engineering and cellular therapeutics.
出处
《生理学报》
CAS
CSCD
北大核心
2008年第3期425-430,共6页
Acta Physiologica Sinica
基金
the Key Subject Construction Foundation of the State Council of China (No. 2005)
the DoctorDegree Dissertation Innovation Foundation of Jinan University (No. 52005031)
关键词
胎儿
骨髓
间充质干细胞
胚胎干细胞
fetus
bone marrow
mesenchymal stem cells
embryonic stem cell
