中华烧伤杂志
中華燒傷雜誌
중화소상잡지
16
2012年
4期
274-277
,共4页
李勇铁%刘德伍%刘德明%毛远桂%彭燕%宁璞%胡翔%邹萍%邹永红%余群洪
李勇鐵%劉德伍%劉德明%毛遠桂%彭燕%寧璞%鬍翔%鄒萍%鄒永紅%餘群洪
리용철%류덕오%류덕명%모원계%팽연%저박%호상%추평%추영홍%여군홍
诱导多功能干细胞%细胞分化%表皮样干细胞
誘導多功能榦細胞%細胞分化%錶皮樣榦細胞
유도다공능간세포%세포분화%표피양간세포
Induced pluripotent stem cells%Cell differentiation%Epidermal-like stem cells
目的 探讨人诱导性多潜能干细胞( iPSC)向表皮样干细胞分化的可能性.方法 (1)以经过灭活处理的小鼠胚胎Fb株作为滋养层,将人iPSC株接种于其上,加入胚胎干细胞完全培养液培养,Ⅳ型胶原酶消化法传代,倒置相差显微镜下观察人iPSC形态及生长状况并行碱性磷酸酶(AKP)染色.以胚胎干细胞不完全培养液悬浮培养iPSC,观察拟胚体形成能力.(2)将人iPSC接种于铺有人羊膜的6孔培养板培养,设为诱导组;另于未铺羊膜的6孔培养板上培养iPSC,作为对照组.观察2组iPSC形态,免疫细胞化学染色法检测整合素β1和细胞角蛋白19( CK19)表达.结果 (1)人iPSC在胚胎干细胞完全培养液中呈典型的干细胞克隆状生长,边界清楚,增殖旺盛;AKP染色阳性.iPSC在无滋养层条件下悬浮培养可形成拟胚体.(2)诱导组iPSC经培养4d后形成干细胞克隆,部分细胞整合素β1和CK19表达阳性.对照组细胞大量死亡,未见整合素β1和CK19表达.结论 人iPSC在羊膜诱导下可定向分化为表皮样干细胞,有望成为皮肤组织工程新的种子细胞.
目的 探討人誘導性多潛能榦細胞( iPSC)嚮錶皮樣榦細胞分化的可能性.方法 (1)以經過滅活處理的小鼠胚胎Fb株作為滋養層,將人iPSC株接種于其上,加入胚胎榦細胞完全培養液培養,Ⅳ型膠原酶消化法傳代,倒置相差顯微鏡下觀察人iPSC形態及生長狀況併行堿性燐痠酶(AKP)染色.以胚胎榦細胞不完全培養液懸浮培養iPSC,觀察擬胚體形成能力.(2)將人iPSC接種于鋪有人羊膜的6孔培養闆培養,設為誘導組;另于未鋪羊膜的6孔培養闆上培養iPSC,作為對照組.觀察2組iPSC形態,免疫細胞化學染色法檢測整閤素β1和細胞角蛋白19( CK19)錶達.結果 (1)人iPSC在胚胎榦細胞完全培養液中呈典型的榦細胞剋隆狀生長,邊界清楚,增殖旺盛;AKP染色暘性.iPSC在無滋養層條件下懸浮培養可形成擬胚體.(2)誘導組iPSC經培養4d後形成榦細胞剋隆,部分細胞整閤素β1和CK19錶達暘性.對照組細胞大量死亡,未見整閤素β1和CK19錶達.結論 人iPSC在羊膜誘導下可定嚮分化為錶皮樣榦細胞,有望成為皮膚組織工程新的種子細胞.
목적 탐토인유도성다잠능간세포( iPSC)향표피양간세포분화적가능성.방법 (1)이경과멸활처리적소서배태Fb주작위자양층,장인iPSC주접충우기상,가입배태간세포완전배양액배양,Ⅳ형효원매소화법전대,도치상차현미경하관찰인iPSC형태급생장상황병행감성린산매(AKP)염색.이배태간세포불완전배양액현부배양iPSC,관찰의배체형성능력.(2)장인iPSC접충우포유인양막적6공배양판배양,설위유도조;령우미포양막적6공배양판상배양iPSC,작위대조조.관찰2조iPSC형태,면역세포화학염색법검측정합소β1화세포각단백19( CK19)표체.결과 (1)인iPSC재배태간세포완전배양액중정전형적간세포극륭상생장,변계청초,증식왕성;AKP염색양성.iPSC재무자양층조건하현부배양가형성의배체.(2)유도조iPSC경배양4d후형성간세포극륭,부분세포정합소β1화CK19표체양성.대조조세포대량사망,미견정합소β1화CK19표체.결론 인iPSC재양막유도하가정향분화위표피양간세포,유망성위피부조직공정신적충자세포.
Objective To investigate the feasibility of differentiation of human induced pluripotent stem cells (iPSCs) into epidermal-like stem cells. Methods ( 1 ) Human strain of iPSCs were plated onto trophoblast of inactivated Fb strain of mouse embryos and cultured in complete medium of embryonic stem cells.iPSCs were subcultured by collagenase Ⅳ digestion method.The morphology and growth of iPSCs were observed under inverted phase contrast microscope,and the cells were stained with alkaline phosphatase (AKP).iPSCs were cultured in incomplete medium of embryonic stem cells to observe the ability of embryoid body formation.(2) Human iPSCs were inoculated onto 6-well plate covered with human amniotic membrane to culture as induction group.Other iPSCs were cultured on 6-well plate without human amniotic membrane as control group.Morphological changes in iPSCs in two groups were observed.Expressions of integrin β1 and CK19 of iPSCs in two groups were determined by immunocytochemical staining. Results Human iPSCs showed a typical stem cell clone-like growth with a clear boundary,and they proliferated vigorously in complete medium of embryonic stem cells.These cells were AKP-positve.iPSCs formed embryoid body in trophoblast-free and suspension culture conditions.After 4 days of co-culture,stem cell clones were formed on the surface of amniotic membrane in induction group,and part of the cells were integrin β1 and CK19positive.Most of the cells died,and no integrin β1 and CK19 positive cells were found in control group.Conclusions Human iPSCs can be differentiated into epidermal-like stem cells by amniotic membrane induction,and it lays an experimental basis for providing new source of seed cells of skin tissue engineering.
