摘要
目的:应用p75NTR进行人食管肿瘤干细胞的分选,并对其生物学特性进行鉴定。方法:对人食管癌标本癌细胞及食管癌细胞株TE-1、Eca109进行培养,应用流式细胞仪检测p75NTR在人食管癌细胞(esophageal cancer cells,ECCs)中的表达,应用磁珠分选(MACS)法进行p75NTR阳性细胞与阴性细胞的分选,观察p75NTR阳性细胞的增殖、分化及软琼脂克隆形成能力,并进行裸鼠接种以观察其致瘤能力;应用化疗药物作用于ECCs后检测其中p75NTR阳性细胞与阴性细胞存活率,以评价p75NTR阳性细胞对化疗药物的耐受性。结果:8个食管肿瘤细胞系(株)中,除SHEC-1、SHEC-5未检测到p75NTR阳性细胞外,其余6个细胞系(株)SHEC-4、SHEC-6、SHEC-7、SHEC-8、Eca109、TE-1中均检测到p75NTR阳性细胞,阳性细胞比例分别为2.71%、0.32%、3.35%、1.13%、2.15%、0.45%。与阴性及未分选细胞相比,MACS分选后的p75NTR阳性细胞具有较强的增殖能力,具有分化产生其他表型细胞的能力,在软琼脂中具有较强的克隆形成能力(P<0.01);行裸鼠接种时,p75NTR阳性细胞表现出较强的致瘤性,其中SHEC-7细胞只需2×103个即可致瘤,其致瘤能力是未分选细胞的50倍。化疗药物分别作用于p75NTR阳性细胞与阴性细胞48h后,p75NTR阳性细胞的存活比例明显高于阴性细胞(P<0.05)。结论:人食管肿瘤细胞中p75NTR阳性细胞具有自我更新、分化、增殖能力,对化疗药物具有较强的耐受性,并具有较强的致瘤能力,具有肿瘤干细胞的特性。
Objective： To isolate and identify cancer stem cells from esophageal cancer cells （ECCs） using cell surface marker p75^NTR. Methods： ECCs were cultured from surgically resected ECC specimens; ECC cell lines TE-1 and Eca109 were also cultured. The expression of p75^NTR in human ECCs was examined by flow cytometry, p75^NTR positive cells were isolated from ECCs using magnetic activated cell sorting （MACS） method. The proliferation,differentiation,and the ability of colony-forming in soft agar of the p75^NTR positive cells were observed. The p75^NTR positive cells were injected into BALB/c nude mice subcutaneously to observe their tumorigenesis ability. The survival rates of p75^NTRpositive and negative cells were assessed after treated with chemotherapy drugs to evaluate the resistance of p75^NTR positive cells. Results： Six out of the eight cell lines, including SHEC-4, SHEC-6, SHEC-7, SHEC-8, Eeal09, and TE-1, were positive of p75^NTR , with the positive rates being 2.71%, 0.32% , 3.35% , 1.13% , 2.15%, and 0.45%, respectively. It was showed that p75^NTR positive cells possessed higher proliferation ability compared with p75^NTR negative cells （P〈0.01）. The p75^NTR positive cells had higher colony-forming ability in soft agar compared with p75^NTR negative cells （P〈0.01）. The p75^NTR positive cells demonstrated stronger tumorigenesis ability in nude mice. As few as 2 000 SHEC-7 cells could give rise to new tumors in xenotransplantation,with a tumorigenesis ability 50 times as high as that of the p75^NTR negative cells. When treated with chemotherapy drugs for 48 h,p75^NTRpositive cells had significantly higher survival rate than p75^NTR negative cells （P〈 0. 05 ）. Conclusion： The p75^NTR positive ECCs possess self-renewal, differentiation,and proliferation abilities; they are strongly resistant to chemotherapy drugs, which gives them strong tumorigenesis ability and the characters of tumor stem cells.
出处
《第二军医大学学报》
CAS
CSCD
北大核心
2009年第5期481-486,共6页
Academic Journal of Second Military Medical University
基金
国家自然科学基金(30471718)~~
