中国组织工程研究
中國組織工程研究
중국조직공정연구
Journal of Clinical Rehabilitative Tissue Engineering Research
2015年
10期
1585-1591
,共7页
薛晨晖%马迅%关晓明%张辉%张丽
薛晨暉%馬迅%關曉明%張輝%張麗
설신휘%마신%관효명%장휘%장려
干细胞%脂肪干细胞%髓核间充质干细胞%椎间盘%转化生长因子β1%诱导分化
榦細胞%脂肪榦細胞%髓覈間充質榦細胞%椎間盤%轉化生長因子β1%誘導分化
간세포%지방간세포%수핵간충질간세포%추간반%전화생장인자β1%유도분화
Adipose Tissue%Intervertebral Disk%Mesenchymal Stem Cels%Transforming Growth Factor beta1
背景：大量研究表明多种组织来源的成体干细胞在体外均可向类髓核细胞分化。椎间盘髓核组织来源间充质干细胞在转化生长因子β1诱导下能否向类髓核细胞分化？其分化能力与脂肪间充质干细胞相比是否有差异目前未见报道。目的：比较脂肪间充质干细胞与髓核间充质干细胞在转化生长因子β1诱导下向类髓核细胞诱导分化能力的差异。方法：分别取大鼠腹股沟处脂肪组织与尾段脊柱，采用机械酶消化法分离培养脂肪间充质干细胞与髓核间充质干细胞。流式细胞仪检测两种细胞 CD105、CD90、CD29、CD45、CD44、CD34、CD24的表达。脂肪间充质干细胞与髓核间充质干细胞各自分为诱导组、无因子诱导组和对照组，诱导组以转化生长因子β1标准软骨诱导液培养，无因子诱导组以不含转化生长因子β1的软骨诱导液培养，对照组以含体积分数为10%胎牛血清的DMEM/F12培养液培养。培养14 d后用RT-PCR检测各组细胞Ⅱ型胶原、蛋白多糖、SOX-9基因的表达。结果与结论：两种细胞CD105、CD90、CD29表达阳性，CD45、CD44、CD34、CD24表达阴性。向类髓核细胞诱导培养14 d后两种细胞诱导组Ⅱ型胶原、蛋白多糖、SOX-9基因表达水平较对照组均明显升高，差异有显著性意义(P <0.05)；髓核间充质干细胞诱导组3种基因的表达水平明显高于脂肪间充质干细胞诱导组，差异有显著性意义(P <0.05)。提示脂肪间充质干细胞与髓核间充质干细胞均具有向类髓核细胞分化的能力，而髓核间充质干细胞相对于脂肪间充质干细胞其软骨相关基因表达更高，可能更适合于作为组织工程髓核研究的种子细胞。
揹景：大量研究錶明多種組織來源的成體榦細胞在體外均可嚮類髓覈細胞分化。椎間盤髓覈組織來源間充質榦細胞在轉化生長因子β1誘導下能否嚮類髓覈細胞分化？其分化能力與脂肪間充質榦細胞相比是否有差異目前未見報道。目的：比較脂肪間充質榦細胞與髓覈間充質榦細胞在轉化生長因子β1誘導下嚮類髓覈細胞誘導分化能力的差異。方法：分彆取大鼠腹股溝處脂肪組織與尾段脊柱，採用機械酶消化法分離培養脂肪間充質榦細胞與髓覈間充質榦細胞。流式細胞儀檢測兩種細胞 CD105、CD90、CD29、CD45、CD44、CD34、CD24的錶達。脂肪間充質榦細胞與髓覈間充質榦細胞各自分為誘導組、無因子誘導組和對照組，誘導組以轉化生長因子β1標準軟骨誘導液培養，無因子誘導組以不含轉化生長因子β1的軟骨誘導液培養，對照組以含體積分數為10%胎牛血清的DMEM/F12培養液培養。培養14 d後用RT-PCR檢測各組細胞Ⅱ型膠原、蛋白多糖、SOX-9基因的錶達。結果與結論：兩種細胞CD105、CD90、CD29錶達暘性，CD45、CD44、CD34、CD24錶達陰性。嚮類髓覈細胞誘導培養14 d後兩種細胞誘導組Ⅱ型膠原、蛋白多糖、SOX-9基因錶達水平較對照組均明顯升高，差異有顯著性意義(P <0.05)；髓覈間充質榦細胞誘導組3種基因的錶達水平明顯高于脂肪間充質榦細胞誘導組，差異有顯著性意義(P <0.05)。提示脂肪間充質榦細胞與髓覈間充質榦細胞均具有嚮類髓覈細胞分化的能力，而髓覈間充質榦細胞相對于脂肪間充質榦細胞其軟骨相關基因錶達更高，可能更適閤于作為組織工程髓覈研究的種子細胞。
배경：대량연구표명다충조직래원적성체간세포재체외균가향류수핵세포분화。추간반수핵조직래원간충질간세포재전화생장인자β1유도하능부향류수핵세포분화？기분화능력여지방간충질간세포상비시부유차이목전미견보도。목적：비교지방간충질간세포여수핵간충질간세포재전화생장인자β1유도하향류수핵세포유도분화능력적차이。방법：분별취대서복고구처지방조직여미단척주，채용궤계매소화법분리배양지방간충질간세포여수핵간충질간세포。류식세포의검측량충세포 CD105、CD90、CD29、CD45、CD44、CD34、CD24적표체。지방간충질간세포여수핵간충질간세포각자분위유도조、무인자유도조화대조조，유도조이전화생장인자β1표준연골유도액배양，무인자유도조이불함전화생장인자β1적연골유도액배양，대조조이함체적분수위10%태우혈청적DMEM/F12배양액배양。배양14 d후용RT-PCR검측각조세포Ⅱ형효원、단백다당、SOX-9기인적표체。결과여결론：량충세포CD105、CD90、CD29표체양성，CD45、CD44、CD34、CD24표체음성。향류수핵세포유도배양14 d후량충세포유도조Ⅱ형효원、단백다당、SOX-9기인표체수평교대조조균명현승고，차이유현저성의의(P <0.05)；수핵간충질간세포유도조3충기인적표체수평명현고우지방간충질간세포유도조，차이유현저성의의(P <0.05)。제시지방간충질간세포여수핵간충질간세포균구유향류수핵세포분화적능력，이수핵간충질간세포상대우지방간충질간세포기연골상관기인표체경고，가능경괄합우작위조직공정수핵연구적충자세포。
BACKGROUND:A large number of studies have shown that adult stem cels derived from multiple tissues are available to differentiate towards nucleus pulposus-like celsin vitro. It is unclear whether mesenchymal stem cels derived from nucleus pulposus tissues have the ability to differentiate towards nucleus pulposus-like phenotypes induced by transforming growth factor-beta 1. Up to now, there are few reports on the difference between the differentiation ability of mesenchymal stem cels derived from nucleus pulposus tissues and adipose-derived mesenchymal stem cels. OBJECTIVE:To compare the ability of mesenchymal stem cels derived from nucleus pulposus tissues and adipose-derived mesenchymal stem cels differentiating into nucleus pulposus-like cels under induction of transforming growth factor-beta 1. METHODS:The groin fat tissue and the coccygeal spine of rats were taken respectively to isolate and culture mesenchymal stem cels derived from nucleus pulposus tissues and adipose-derived mesenchymal stem cels by mechanical enzyme digestion method. Flow cytometry was employed to detect the expression of CD105, CD90, CD29, CD45, CD44, CD34, and CD24 of both two kinds of stem cels. Mesenchymal stem cels derived from nucleus pulposus tissues and adipose-derived mesenchymal stem cels were divided into complete induction group (complete induction medium with transforming growth factor-beta 1), incomplete induction group (complete induction medium without transforming growth factor-beta 1) and control group(DMEM/F12 containing 10% fetal bovine serum and 100 mg/L penicilin/streptomycin), respectively. After 14 days of culture, real-time PCR was used to detect the expression of colagen type II, Aggrecan and SOX-9 in each group. RESULTS AND CONCLUSION:CD105, CD90, CD29 expressed positively and CD45, CD44, CD34, CD24 negatively in both two kinds of stem cels. After 14 days of induced differentiation, the expressions of colagen type II, Aggrecan and SOX-9 in the two kinds of cels were significantly higher in the complete induction groups than in the control groups (P < 0.05). Under the induction of transforming growth factor-beta 1, the expression of colagen type II, Aggrecan and SOX-9 in mesenchymal stem cels derived from nucleus pulposus tissues was significantly higher than that in adipose-derived mesenchymal stem cels (P < 0.05). These findings suggest that both two kinds of mesenchymal stem cels have the ability to differentiate towards nucleus pulposus-like cels induced by transforming growth factor-beta, and mesenchymal stem cels derived from nucleus pulposus tissues may be more suitable as seed cels for nucleus pulposus tissue engineering research.
