EGF诱导结肠腺癌HCT-116细胞神经内分泌分化实验
Experimental Study on Neuroendocrine Differentiation of Colon Cancer Cell Line HCT-116 Induced by Epidermal Growth Factor
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摘要: 目的 利用表皮生长因子 (EGF) 及表皮生长因子受体 (EGFR) 抑制剂西妥昔单抗 (Cetuximab) 诱导和阻断诱导结肠腺癌HCT-116细胞神经内分泌分化 (NED) , 探讨诱导和阻断诱导后肿瘤细胞生物学变化特点及相关因素.方法 通过HE和免疫组化染色, 观察肿瘤细胞裸鼠种植成瘤后及NED诱导前后肿瘤细胞形态变化特点和免疫组化表达情况;低密度平板克隆实验检测肿瘤细胞克隆形成率;流式细胞术检测肿瘤细胞生长周期;MTT法绘制肿瘤细胞生长曲线.结果 结肠腺癌HCT-116裸鼠种植后成肿瘤细胞明显异型;经EGF及EGFR作用前后HCT-11肿瘤细胞形态无明显变化;EGF组克隆形成率增加 (P>0.05) , 肿瘤细胞Cg A、Ki-67、Bcl-2表达增强, 增殖活性增高, EGF作用后肿瘤细胞与其它各组相比差异有显著统计学意义 (P<0.01) .而CEA、CA19-9、CDX2、COX2、Syn、NSE在各组表达差异无统计学意义 (P>0.05) .结论 EGF可诱导结肠腺癌细胞HCT-116 NED;肿瘤细胞增殖活性增加、抗凋亡能力增强;EGFR可阻断EGF诱导HCT-116 NED及相应生物学特性变化;而CEA、CA19-9、CDX2、COX2对肿瘤细胞NED和增殖凋亡无明显调节作用.Abstract: Objective To induce and block to induce neuroendocrine differentiation of colon adenocarcinoma cell HCT-116 by using the epidermal growth factor (EGF) and epidermal growth factor receptor (EGFR) inhibitor Cetuximab, and further explore the intrinsical changes and association with the characteristics of colonic cancer cells HCT-116 in proliferation, apoptosis, possible mechanism, and the adjustment factors of neuroendocrine differentiation. Methods HE and immunohistochemical stains were conducted to observe morphological characters of colonic cancer cultured cells HCT-116 and tumor in the nude mouse by expression of Cg A, Ki-67, Bcl-2, Syn, CEA, CA19-9, COX2, CDX2. Cell growth curve of HCT-116 was drawn by MTT, flow cytometry technique was used to determine the cell cycle of HCT-116, and Low Density Plate Clone Test was applied to measure the cloned forming rate of HCT-116. Results Morphology of HCT-116 in the induced courses was similar. EGF acted on cultured HCT-116 cells to strengthen the expression of Cg A, Ki-67 and Bcl-2, to increase cell proliferation rate and proliferation activity in cell cycle of S and G2 phase. The colony formation rate was also increased significantly compared with the control group. Difference with cetuximab group was statistically significant (P <0.001) . Changes of proliferation index of Ki-67 in cultured HCT-116 cells was statistically significant in EGF group compared with the other groups (P<0.01) . The differential expression of NSE, CEA, CA19-9, CDX2, COX2, and Syn in each group was no statistically significant (P>0.05) . Conclusion EGF can induce neuroendocrine differentiation in cultured HCT-116 cells NED, increase cell proliferation activity, strengthen antiapoptotic ability of colonic cancer cells. EGFR can block EGF to induce HCT-116 NED, and inhibit changes of relevant biological features. However, CEA, CA19-9, COX2, and CDX2 do not regulate NED, proliferation, and apoptosis.
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