中华结直肠疾病电子杂志
中華結直腸疾病電子雜誌
중화결직장질병전자잡지
Chinese Journal of Colorectal Diseases
2013年
4期
170-176
,共7页
杜航向%杨治力%杨逸%盛能全%王志刚
杜航嚮%楊治力%楊逸%盛能全%王誌剛
두항향%양치력%양일%성능전%왕지강
AC133%上皮细胞黏附分子%结直肠癌干细胞
AC133%上皮細胞黏附分子%結直腸癌榦細胞
AC133%상피세포점부분자%결직장암간세포
AC 133%Epithelial cell adhesion molecule%Colorectal cancer stem cells
目的：利用AC133抗原联合EpCAM检测人结直肠癌组织中结直肠癌干细胞，并初步观察其生物学特性，为后续结直肠癌干细胞研究提供实验基础。方法利用新鲜结直肠癌组织，无血清悬浮成球培养，流式细胞检测AC133、EpCAM 的表达情况，Balb/C小鼠移植观察其成瘤情况。组间比较采用t检验，以P＜0．05为差异有统计学意义。结果从人原代结肠腺癌中分离、纯化AC133＋EpCAM ＋结直肠癌干细胞，结直肠癌原代细胞中AC133＋EpCAM ＋细胞比例为1．5％～35％（平均4．8％）。单克隆形成实验证实结直肠癌组织中存在肿瘤干细胞。其比例为2．18±0．15％，分离获得的AC133＋EpCAM ＋细胞能在无血清培养基中“成球”，在血清诱导下能贴壁分化；将AC133＋EpCAM＋细胞移植在Balb/C裸鼠体内，表现出很强的致瘤性，移植瘤中AC133＋EpCAM ＋细胞比例为4．9％～40．2％（平均17．2％）。结论人结肠腺癌组织中存在AC133＋EpCAM ＋细胞群，具有和普通干细胞相类似的无限增殖、自我更新和分化能力，可为人结直肠癌干细胞的后续研究提供实验基础。
目的：利用AC133抗原聯閤EpCAM檢測人結直腸癌組織中結直腸癌榦細胞，併初步觀察其生物學特性，為後續結直腸癌榦細胞研究提供實驗基礎。方法利用新鮮結直腸癌組織，無血清懸浮成毬培養，流式細胞檢測AC133、EpCAM 的錶達情況，Balb/C小鼠移植觀察其成瘤情況。組間比較採用t檢驗，以P＜0．05為差異有統計學意義。結果從人原代結腸腺癌中分離、純化AC133＋EpCAM ＋結直腸癌榦細胞，結直腸癌原代細胞中AC133＋EpCAM ＋細胞比例為1．5％～35％（平均4．8％）。單剋隆形成實驗證實結直腸癌組織中存在腫瘤榦細胞。其比例為2．18±0．15％，分離穫得的AC133＋EpCAM ＋細胞能在無血清培養基中“成毬”，在血清誘導下能貼壁分化；將AC133＋EpCAM＋細胞移植在Balb/C裸鼠體內，錶現齣很彊的緻瘤性，移植瘤中AC133＋EpCAM ＋細胞比例為4．9％～40．2％（平均17．2％）。結論人結腸腺癌組織中存在AC133＋EpCAM ＋細胞群，具有和普通榦細胞相類似的無限增殖、自我更新和分化能力，可為人結直腸癌榦細胞的後續研究提供實驗基礎。
목적：이용AC133항원연합EpCAM검측인결직장암조직중결직장암간세포，병초보관찰기생물학특성，위후속결직장암간세포연구제공실험기출。방법이용신선결직장암조직，무혈청현부성구배양，류식세포검측AC133、EpCAM 적표체정황，Balb/C소서이식관찰기성류정황。조간비교채용t검험，이P＜0．05위차이유통계학의의。결과종인원대결장선암중분리、순화AC133＋EpCAM ＋결직장암간세포，결직장암원대세포중AC133＋EpCAM ＋세포비례위1．5％～35％（평균4．8％）。단극륭형성실험증실결직장암조직중존재종류간세포。기비례위2．18±0．15％，분리획득적AC133＋EpCAM ＋세포능재무혈청배양기중“성구”，재혈청유도하능첩벽분화；장AC133＋EpCAM＋세포이식재Balb/C라서체내，표현출흔강적치류성，이식류중AC133＋EpCAM ＋세포비례위4．9％～40．2％（평균17．2％）。결론인결장선암조직중존재AC133＋EpCAM ＋세포군，구유화보통간세포상유사적무한증식、자아경신화분화능력，가위인결직장암간세포적후속연구제공실험기출。
Objective To isolate colorectal cancer stem cells from human fresh carcinoma samples obtained at surgical operation and to study their biological characteristics .Then to provide foundation for the follow-up experiments .Methods Colorectal cancer tissues obtained from surgically resected specimens of colorectal cancer patients were fully chopped ,trypsinized,and filtered to prepare single cell suspensions .The cells were cultured in serum-free DMEM/F12 medium,with LIF,EGF,bFGF and 2% fetal bovine serum weekly.The stem cell-markers AC133 and EpCAM on the isolated cells were detected by using flow cytometry.The tumorigenic potent of the isolated cells was evaluated via the transplantation into Balb /C nude mice.All the variables were tested with T-test.P<0.05 was considered to be statistically significant .Results AC133 +EpCAM + colorectal cancer stem cells were isolated and purified from the human primary colorectal carcinoma.The percentage of AC133 +EpCAM +colorectal cancer stem cells in primary colorectal cancer was 1.5%~35%( mean 4.8%).There were colon cancer stem cells in colon cancer by limited dilution assay.The percentage was 2.18 ±0.15%.The suspension spheres were generated by the isolated AC 133 +EpCAM +Co-CSCs in serum-free medium.Their differentiation were induced in medium containing 10%FBS.AC133 +EpCAM +CCSCs were implanted into Balb/C nude mice ,and observed stronger capacity of tumor initiation.The percentage of AC133 +EpCAM + cells in xenografts was 4.9% ~40.2%( mean 17.2%).Human CRC xenografts established in Balb/C nude mice grew as differentiated carcinomas ,which formed gland-like structures and scored positive for expression of colon-specific differentiation markers ,such as cytokeratin-20(CK20),and neutral epithelial mucins .Conclusions There are colon cancer stem cells in human colorectal carcinoma .These cells are characterized by AC 133 +EpCAM + surface marker , which possess apparent feature of stem cells and stronger capacity of initiating tumors ,moreover ,generated spheroid in serum-free medium.These findings could provide foundation for the follow-up experiments .
