摘要
背景:以往研究脂肪间充质干细胞培养方式多为单层培养,诱导方式主要集中于细胞因子的方法,而椎间盘体内微环境,除细胞因子对细胞影响外,在其三维环境中细胞间的相互作用还值得进一步研究。目的:在体外分别通过髓核细胞和转化生长因子β1/胰岛素样生长因子1两种条件诱导,观察脂肪间充质干细胞向髓核细胞分化的差异。方法:分别单层培养兔的脂肪间充质干细胞与髓核细胞细胞,脂肪间充质干细胞培养至3代经鉴定后,按5×106个细胞制成微球,置于Transwell培养板中培养,分别通过髓核细胞微球及转化生长因子β1/胰岛素样生长因子1诱导;于诱导前、诱导7,14d,观察细胞形态变化,并通过RT-PCR对Ⅱ型胶原、蛋白多糖水平进行测定。结果与结论:在体外诱导7,14d,两组脂肪间充质干细胞微团体积、形态无明显区别。RT-PCR检测结果显示7d两组与髓核诱导组均有Ⅱ型胶原及蛋白多糖mRNA表达,但转化生长因子β1/胰岛素样生长因子1组表达更强;诱导14d髓核诱导组Ⅱ型胶原及蛋白多糖mRNA表达明显增高,优于转化生长因子β1/胰岛素样生长因子1组。结果表明,在体外髓核细胞和转化生长因子β1/胰岛素样生长因子1对脂肪间充质干细胞进行诱导均有促进其向髓核细胞分化作用,而转化生长因子β1、胰岛素样生长因子1为正常髓核细胞分泌重要因子,说明髓核细胞对脂肪间充质干细胞的诱导之间除细胞因子作用外,尚存在相互促进增殖分化作用。
BACKGROUND：Previous studies have addressed adipose-derived mesenchymal stem cells （ADSCs） culture way for monolayer culture;induction way was mainly cytokine method,and under intervertebral discs micro-environment,cell interactions also deserved further study in its three-dimensional environment,in addition to the effects on cell cytokines.OBJECTIVE：To observe differences in differentiation of ADSCs into NPCs under two induced conditions：nucleus pulposus cells （NPCs） and transforming growth factor β1 （TGF-β1）/insulin-like growth factor （IGF-1） in vitro.METHODS：Rabbit ADSCs monolayer culture and NPCs were cultured respectively,3 generations of ADSCs were qualified;5×106 cells were made into microspheres and placed in Transwell culture plates for culture under TGF-β 1/IGF-1 induction.Cell shape changes were observed before,7,14 days after induction.Reverse transcription-polymerase chain reaction （RT-PCR） was used to determine levels of type II collagen and proteoglycan.RESULTS AND CONCLUSION：At 7 and 14 days,no significant difference was detected in size and shape of ADSCs microspheres in both groups.RT-PCR detection results displayed that type II collagen and proteoglycan mRNA expression was determined in 7-day TGF-β1/IGF-1 group and NPCs-induced group,but expression was stronger in the TGF-β1/IGF-1 group.Type II collagen and proteoglycan mRNA expression was obviously increased in 14-day NPCs-induced group,superior to TGF-β1/IGF-1 group.Results have suggested that in vitro NPCs and TGF-β1/IGF-1 lead to the ADSCs,which had promoting effects on NPCs differentiation.TGF-β 1 and IGF-1 are normal NPCs-secreted important factors.The promoting effects on proliferation and differentiation exist,with the exception of cytokine induction effect,in differentiation of ADSCs into NPCs.
出处
《中国组织工程研究与临床康复》
CAS
CSCD
北大核心
2010年第23期4217-4221,共5页
Journal of Clinical Rehabilitative Tissue Engineering Research
基金
北京市自然科学基金资助项目(5062039)
项目名称:组织工程化髓核结合等离子消融修复退变椎间盘实验研究~~
