白细胞介素1β对大鼠髓核间质干细胞生物学特性的影响  

Interleukin-1 beta affects the biological properties of rat nucleus pulposus-derived mesenchymal stem cells

作　　者：张宁[1] 关晓明[1] 马迅 张丽 张辉[1] 宋亮[1] 

机构地区：[1]山西医科大学,山西省太原市030001 [2]山西大医院骨科,山西省太原市030032

出　　处：《中国组织工程研究》2014年第28期4437-4443,共7页Chinese Journal of Tissue Engineering Research

基　　金：山西省自然科学基金资助项目(2011011042-3);太原市大学生创新创业专题(120164064);山西省研究生优秀创新项目(20113069)~~

摘　　要：背景:内源性干细胞并未在椎间盘退变的进程发挥修复作用,作者推测可能与相关致病因素的异常作用有关,导致对髓核间质干细胞功能产生一定的抑制效应。目的:观察炎症因子白细胞介素1β对大鼠髓核间质干细胞生物学特性的影响。方法:取3月龄雄性SD大鼠脊柱腰段椎间盘髓核组织,用胶原酶,胰酶序贯消化法分离并培养髓核间质干细胞。采用流式细胞术检测细胞免疫表型CD24,CD34,CD45,CD90,CD105表达;利用RT-PCR检测干细胞基因SOX2,Nanog的表达。观察髓核间质干细胞成脂、成骨、成软骨分化能力。利用流式细胞术检测白细胞介素1β作用于髓核间质干细胞后的凋亡率,荧光定量PCR检测白细胞介素1β作用于髓核间质干细胞后SOX9,蛋白多糖,Ⅱ型胶原酶,caspase-3基因的表达变化。结果与结论:体外分离培养出的细胞表现为纺锤样长梭形;CD90、CD105表达阳性,CD45、CD34、CD24表达阴性,表达干细胞基因SOX2,Nanog,具有成骨和成软骨分化能力,无成脂分化能力。CCK-8检测白细胞介素1β可以抑制髓核间质干细胞的增殖活性,与作用时间、作用浓度存在一定的依赖关系,最佳作用时间为48 h,最佳作用浓度为50μg/L。流式细胞仪检测白细胞介素1β可以使髓核间质干细胞轻度凋亡;荧光定量PCR结果显示:SOX9,蛋白多糖,Ⅱ型胶原的mRNA表达下调,caspase-3表达上调。结果可见白细胞介素1β具有抑制髓核间质干细胞增殖活性并诱导其凋亡的作用,是导致椎间盘自身干细胞无法发挥修复作用的原因之一。BACKGROUND：Endogenous stem cells have no repair effects on the process of disc degeneration. Authors assumed that this maybe associate with abnormal effects of related etiological factor, resulting in an inhibitory effect on the function of nucleus pulposus-derived mesenchymal stem cells. OBJECTIVE：To investigate the effects of inflammatory cytokine interleukin-1βon biological characteristics of nucleus pulposus-derived mesenchymal stem cells of rats. METHODS：Lumbar spinal nucleus pulposus was obtained from 3-month-old male Sprague-Dawley rats. Nucleus pulposus-derived mesenchymal stem cells were isolated and cultured with col agenase and sequential trypsin digestion. The expression of CD24, CD34, CD45, CD90 and CD105 was detected using flow cytometry. Stem cellgene SOX2 and Nanog expression was measured using RT-PCR. Adipogenic, osteogenic and chondrogenic abilities of nucleus pulposus-derived mesenchymal stem cells were observed. The apoptotic rate of interleukin-1β-treated nucleus pulposus-derived mesenchymal stem cells was detected using flow cytometry. Fluorescent quantitative PCR was used to measure the expression of SOX9, proteoglycan, type II col agenase and caspase-3 gene after nucleus pulposus-derived mesenchymal stem cells were treated with interleukin-1β.

关 键 词：干细胞 骨髓干细胞 髓核间质干细胞 增殖活性 凋亡 大鼠 山西省自然科学基金 

分 类 号：R394.2[医药卫生—医学遗传学]