本文選題：細(xì)胞分化 + 羊膜��； 參考：《中國組織工程研究》2017年33期

【摘要】：背景:Scleraxis作為肌腱細(xì)胞特異性表達(dá)分子,不僅參與肌腱祖細(xì)胞的聚集及分化,還影響肌腱細(xì)胞外基質(zhì)的形成。人羊膜間充質(zhì)干細(xì)胞具有多向分化潛能,在體外不同誘導(dǎo)條件下可分化為骨、軟骨及其他結(jié)締組織。目的:探討Scleraxis慢病毒基因感染人羊膜間充質(zhì)干細(xì)胞能否向肌腱細(xì)胞定向分化并觀察其分化效果。方法:取足月產(chǎn)胎盤羊膜組織,兩步酶消化法分離人羊膜間充質(zhì)干細(xì)胞并采用倒置相差顯微鏡觀察和流式細(xì)胞鑒定。取第3代細(xì)胞分3組進(jìn)行培養(yǎng),單純?nèi)搜蚰らg充質(zhì)干細(xì)胞培養(yǎng)組為空白組,人羊膜間充質(zhì)干細(xì)胞經(jīng)Slclerxis基因慢病毒感染后為過表達(dá)組,人羊膜間充質(zhì)干細(xì)胞經(jīng)不攜帶Scleraxis基因慢病毒感染后為空質(zhì)粒組。細(xì)胞培養(yǎng)7 d內(nèi)CCK-8法檢測(cè)各組細(xì)胞增殖能力細(xì)胞。細(xì)胞培養(yǎng)后3 d和7 d,分別進(jìn)行實(shí)時(shí)熒光定量PCR和Western Blot檢測(cè)評(píng)價(jià)各組細(xì)胞向肌腱細(xì)胞定向分化的效果。結(jié)果與結(jié)論:(1)CCK-8檢測(cè)顯示:培養(yǎng)7 d內(nèi),過表達(dá)組、空質(zhì)粒組與空白組細(xì)胞在增殖能力上無明顯差異(P0.05);(2)Westen blot檢測(cè)顯示:過表達(dá)組Scleraxis蛋白表達(dá)水平明顯高于空質(zhì)粒組和空白組(P0.05);(3)實(shí)時(shí)熒光定量PCR顯示:3 d時(shí),過表達(dá)組Ⅰ型膠原、Ⅲ型膠原、纖連蛋白及肌腱蛋白C m RNA表達(dá)水平明顯高于空質(zhì)粒組(P0.05),而腱調(diào)蛋白的表達(dá)與空質(zhì)粒組無明顯差異(P0.05);7 d時(shí),Ⅰ型膠原、Ⅲ型膠原、纖連蛋白、肌腱蛋白C及腱調(diào)蛋白的表達(dá)水平明顯高于空質(zhì)粒組(P0.05);(4)結(jié)果提示:人羊膜間充質(zhì)干細(xì)胞經(jīng)Scleraxis慢病毒基因感染后可向肌腱細(xì)胞定向分化。
[Abstract]:Background as a specific expression molecule of tendon cells, Scleraxis not only participates in the aggregation and differentiation of tendon progenitor cells, but also affects the formation of extracellular matrix of tendon cells. Human amniotic mesenchymal stem cells have the potential to differentiate into bone, cartilage and other connective tissues under different induction conditions in vitro. Aim: to investigate whether human amniotic mesenchymal stem cells (MMSCs) infected with Scleraxis lentivirus gene can differentiate into tendon cells and observe its differentiation effect. Methods: human amniotic mesenchymal stem cells were isolated by two-step enzyme digestion and observed by inverted phase contrast microscope and identified by flow cytometry. The third passage of human amniotic mesenchymal stem cells were divided into three groups. The human amniotic mesenchymal stem cells were cultured as blank group and human amniotic membrane mesenchymal stem cells infected by Slclerxis gene lentivirus as overexpression group. Human amniotic mesenchymal stem cells were infected without Scleraxis gene lentivirus. The proliferative ability of cells in each group was detected by CCK-8 assay within 7 days of cell culture. After 3 days and 7 days of culture, real-time fluorescence quantitative PCR and Western Blot were used to evaluate the differentiation of the cells into tendon cells in each group. Results and conclusion the results of CCK-8 test showed that: within 7 days of culture, the expression of CCK-8 in the overexpressed group was higher than that in the control group. There was no significant difference in proliferative ability between empty plasmid group and blank group. The expression level of Scleraxis protein in overexpression group was significantly higher than that in blank plasmid group and blank group. The expression of Scleraxis protein in overexpression group was significantly higher than that in blank plasmid group and blank group. The C m RNA expression level of type 鈪，

本文編號(hào)：1883228