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目的:研究喉癌细胞系Hep-2中CD133的表达;比较CD133~+细胞、未分选细胞、CD133~-细胞的体外增殖、克隆形成能力及其在裸鼠体内的成瘤能力;探讨喉癌肝细胞对化疗药物顺铂(cisplatin,DDP)的抵抗作用。方法:采用流式细胞仪检测CD133在Hep-2细胞系中的表达;免疫磁珠分选技术纯化CD133阳性肿瘤细胞;使用四甲基偶氮唑蓝(MTT)法和平板克隆形成实验检测分选所得各细胞亚群细胞以及未分选细胞的体外增殖能力和克隆形成能力;将CD133阳性肿瘤细胞和CD133阴性肿瘤细胞以一定的数量级注入重症联合免疫缺陷小鼠腹部皮下,比较其成瘤差异性;此外,使用DDP干预分选所得各细胞亚群细胞,检测比较CD133阳性肿瘤细胞和CD133阴性肿瘤细胞的体外增殖能力与体内成瘤能力。结果:流式细胞仪示CD133在Hep-2细胞系中呈微量恒定表达,表达概率为40.12±1.32%;CD133阳性肿瘤细胞的体外增殖能力显著强于CD133阴性肿瘤细胞的增殖能力(P<0.05),且其克隆形成能力也强于CD133阴性肿瘤细胞;体内成瘤实验结果显示CD133阳性肿瘤细胞较CD133阴性细胞、未分选细胞在重症联合免疫缺陷小鼠体内具有更强的成瘤性(P<0.05);在DDP的干预下,相对于CD133阴性肿瘤细胞,CD133阳性肿瘤细胞表现出更强的抵抗力。结论:喉癌Hep-2细胞系中,CD133阳性癌细胞具有强的体外增殖能力、体内成瘤能力且对化疗药物具有较强的抵抗性,可作为喉癌肿瘤干细胞的标志之一。
OBJECTIVE: To study the expression of CD133 in Hep-2 cell line of laryngeal cancer. To compare the proliferation, clonality and CD133 ~ - cell proliferation ability of CD133 ~ + cells, unsorted cells and CD133 ~ - cells in nude mice. Hepatocellular carcinoma cells resist chemotherapeutic drug cisplatin (DDP). Methods: The expression of CD133 in Hep-2 cell line was detected by flow cytometry. The CD133 positive tumor cells were purified by magnetic beads sorting technique. MTT assay and plate clone formation assay Selected from each cell subpopulation cells and unsorted cells in vitro proliferation and clonality; CD133-positive tumor cells and CD133-negative tumor cells by an order of magnitude into the severe combined immunodeficiency mice abdominal subcutaneous comparison of tumor formation difference In addition, DDP was used to intervene in each cell subpopulation of sorted cells to detect the in vitro proliferation ability and in vivo tumorigenicity of CD133-positive tumor cells and CD133-negative tumor cells. Results: Flow cytometry showed that the expression of CD133 in Hep-2 cell line was constant and the expression rate was 40.12 ± 1.32%. The proliferation ability of CD133 positive cells was significantly stronger than that of CD133 negative cells (P <0.05) ), And its clonogenic capacity is also stronger than that of CD133 negative tumor cells. In vivo tumorigenicity test results show that CD133 positive tumor cells have stronger tumorigenicity than CD133 negative cells and unsorted cells in severe combined immunodeficient mice P <0.05). CD133-positive tumor cells showed more resistance than CD133-negative tumor cells under the intervention of DDP. Conclusion: Among laryngeal cancer Hep-2 cell lines, CD133-positive cancer cells have strong ability of proliferation in vitro, tumorigenic ability in vivo and strong resistance to chemotherapeutic drugs, which can be used as one of the markers of cancer stem cells in laryngeal cancer.