摘要
目的通过观察转化生长因子-β(TGF-β)3种不同亚型(β1、β2和β3)对前软骨干细胞(PSCs)的增殖分化及细胞外基质合成的影响,为构建组织工程学软骨组织寻找理想的细胞因子。方法通过免疫磁珠分选法得到经过筛选和纯化的PSCs细胞,分别用含有浓度为10ng/mL的3种亚型的重组TGF-β培养液对PSCs进行培养。PSCs培养后第0、3、6、9天,采用MTT法与BrdU/PI双参法测细胞增殖及DNA合成水平,采用RT-PCR法测定ColⅡ、Aggrecan基因的表达。采用免疫组化法测定培养2周后各组细胞SOX9蛋白表达情况。结果培养3d后,TGF-β1组PSCs的增殖活性明显较对照组下降(P<0.05),而TGF-β2、TGF-β3组增殖活性与对照组相比差异无统计学意义(P>0.05)。6d后,与对照组相比,TGF-β1组细胞增殖活性明显降低(P<0.05);而TGF-β2、TGF-β3组则显示了相反的结果,增殖活性较对照组明显增强,且TGF-β3组高于TGF-β2组(P<0.05)。DNA合成呈同样趋势。RT-PCR分析显示,随着时间的延长,与对照组相比,TGF-β1、TGF-β2、TGF-β3组ColⅡ基因表达均呈上升趋势;TGF-β1与TGF-β3组Aggrecan的表达亦较对照组增加,但TGF-β2组Aggrecan的基因表达无明显改变。免疫组化检测结果显示,TGF-β1、TGF-β3组的SOX9蛋白合成高于对照组(P<0.05),而TGF-β2组与对照组相比差异无统计学意义(P>0.05)。结论 TGF-β1能抑制PSCs增殖活性,而TGF-β2和TGF-β3具有促进PSCs增殖的作用,TGF-β1与TGF-β3均能促进PSCs细胞分化,而TGF-β2对于分化的诱导作用不明显。
Objective To examine the effect of different TGF-βisoforms（TGF-β1,TGF-β2and TGF-β3）on the proliferation and differentiation of precartilaginous stem cells（PSCs）in order to seek an ideal cytokine for tissue engineering cartilage construction.Methods PSCs were obtained after isolation and purification by means of immun-magnetic activated cell sorting.The cells were then cultured in the media containing TGF-β1,TGF-β2 or TGF-β3at 10ng/mL.At 0,3,6,9days after culture,the proliferation and DNA synthesis of PSCs in response to TGF-βisoforms were examined by MTT and BrdU/PI double staining,and the mRNA expression of ColⅡand Aggrecan was detected by RT-PCR.The expression of SOX9was immunohistochemically determined at 2 weeks.Results Three days after culture,the proliferation potential of PSCs was significantly decreased in the TGF-β1group relative to the control group（P0.05）,but it had no significant changes in both TGF-β2and TGFβ3groups（P0.05）.Six days later,the PSCs proliferation continued to decline in the TGF-β1group but increase in the TGF-β2 and TGF-β3 groups with difference being significant when compared with the control group（P0.05）.RT-PCR showed the mRNA expression of ColⅡ was increased with time in all the three groups.The expression of Aggrecan in the three groups except the TGF-β2group was much stronger than in the control group.Immunohistochemistry showed the synthesis of SOX9 protein was increased in both TGF-β1and TGF-β3groups when compared with the control group（P0.05）.Conclusion TGF-β1 can inhibit the proliferation of PSCs,while TGF-β2and TGF-β3can promote it.Both TGF-β1and TGF-β3in stead of TGF-β2can promote the differentiation of PSCs.
出处
《华中科技大学学报（医学版）》
CAS
CSCD
北大核心
2013年第6期659-664,共6页
Acta Medicinae Universitatis Scientiae et Technologiae Huazhong
关键词
前软骨干细胞
转化生长因子-Β
增殖
分化
precartilaginous stem cells（PSCs）
transforming growth factor-β
proliferation
differentiation
