脐血间充质干细胞对淋巴细胞增殖的作用

Effects of umbilical cord blood mesenchymal stem cells on lymphocyte proliferation

目的:骨髓间充质干细胞抑制混合淋巴细胞反应及丝裂原刺激的T细胞增殖,是通过自分泌转化生长因子发挥其免疫调节作用,而关于脐血间充质干细胞对淋巴细胞增殖作用的影响及其机制尚不清楚。实验拟观察验证脐血间充质干细胞对植物血凝素诱导的淋巴细胞增殖的影响。方法:实验于2005-07/2006-03在南昌大学第二附属医院血液病研究所、江西省医学分子重点实验室完成。①实验材料:健康产妇分娩后的脐带血及正常志愿者外周血各9份,由南昌大学第二附属医院妇产科提供,产妇及家属对实验知情同意,并经医院伦理委员会批准。②实验方法:采脐血进行间充质干细胞的培养扩增,并经形态学、细胞表面分子测定及定向分化功能加以鉴定。将1.0×105,0.75×105,0.5×105,0.25×105,0.1×105,0.05×105,0.02×105,0.01×105的间充质干细胞加入到经植物血凝素诱导的外周血淋巴细胞增殖体系中,应用MTT法测定细胞增殖率,观察脐血间充质干细胞对淋巴细胞增殖的影响。应用ELISA法测定脐血间充质干细胞培养上清液中转化生长因子β1水平。结果:①不同数量脐血间充质干细胞对植物血凝素诱导的淋巴细胞增殖均有抑制作用,抑制作用在一定范围内呈细胞剂量依赖性,以淋巴细胞:间充质干细胞为1:0.75时抑制作用最强。②转化生长因子β1水平与脐血间充质干细胞数量有关,随着间充质干细胞数量增多,转化生长因子β1水平渐增高。③将脐带血间充质干细胞对淋巴细胞增殖的抑制率与转化生长因子β1水平进行相关性分析,两者间存在明显相关性(r=0.85)。结论:①脐血间充质干细胞对植物血凝素诱导的淋巴细胞增殖有抑制作用,在一定范围内呈细胞剂量依赖性。②脐血间充质干细胞所分泌的转化生长因子β1含量与脐带血间充质干细胞数量呈正相关性。

AIM： Mesenchymal stem cells showed inhibitory effect on mixed lymphocyte reaction and phytohemagglutinin-induced lymphocyte proliferation by secreting transforming growth factor-β1, while the effect of umbilical cord blood mesenchymal stem cells on lymphocyte proliferation and the possible mechanism are unknown. The study observed effects of umbilical cord blood mesenchymal stem cells on phytohemagglutinin induced lymphocytes proliferation. METHODS： Experiments were conducted from July 2005 to March 2006 at Jiangxi Key Laboratory of Molecular Medicine, Institute of Hematopathy, Second Affiliated Hospital, Nanchang University. （1）Nine umbilical cord blood samples from healthy puerperants and nine peripheral blood samples from normal volunteers were provided by Department of Gynecology and Obstetrics of Second Affiliated Hospital, Nanchang University. Peurperants and theirs family members signed the informed consent. The experiment was approved by Hospital Ethics Committee. （2）Umbilical cord blood was collected for mesenchymal stem cell culture. Umbilical cord blood mesenchymal stem cells were identified by morphology, cell surface molecule and differentiation. Mesenchymal stem cells were added to phytohemagglutinin induced proliferation with various numbers as follows 1.0 ×10^5, 0.75 ×10^5, 0.5 ×10^5, 0.25 ×10^5, 0.1 ×10^5, 0.05 ×10^5, 0.02 ×10^5, 0.01 ×10^5. The proliferation of mesenchymal stem cells was measured by MTT method. Effects of umbilical cord blood mesenchymal stem cells on the proliferation of lymphocytes were observed. Transforming growth factor-β 1 levels in supernatant of umbilical cord blood mesenchymal stem cells were measured by ELISA. RESULTS： （1）Umbilical cord blood mesenchymal stem cells of different numbers had a inhibitory effect on phytohemagglutinin induced proliferation of lymphocytes, with dose-dependent effects. The inhibitory effect was the strongest at 1：0.75 of lymphocytes and mesenchymal stem cells. （2）Transforming growth factor-β1 levels were positively associated with the amount of umbilical cord blood mesenchymal stem cells. （3）The inhibitory effect of umbilical cord blood mesenchymal stem cells on the proliferation of lymphocytes was significantly associated with the transforming growth factor-β 1 levels （r=0.85） . CONCLUSION： （1）Umbilical cord blood mesenchymal stem cells show a inhibitory effect on phytohemagglutinin-induced lymphocyte proliferation, with a dose dependent effect. （2）Transforming growth factor- β1 levels of umbilical cord blood mesenchymal stem cells are positively associated with the amount of umbilical cord blood mesenchymal stem cells.