低氧对人结肠癌细胞HT-29增殖、血管生成能力的影响及其机制探讨

Moderate hypoxia up-regulates expression of NF-κB and augments proliferation and angiogenesis in colon carcinoma cell line HT-29

目的观察HT-29细胞在低氧下的增殖、血管生成能力,NF-κB的表达水平,初步探讨低氧与肿瘤恶性演进的关系。方法参照体内肿瘤的氧分压,建立低氧模型。MTT检测低氧对HT-29细胞增殖的影响,描绘细胞生长曲线;内皮细胞小管形成试验检测体外促进新生血管形成的能力。将HT-29细胞低氧处理不同时段,提取细胞核蛋白,Western blot检测细胞核内NF-κB的表达。结果低氧组HT-29细胞的形态与对照组相似,而A570值则显著高于对照组。低氧组内皮细胞形成的管状结构数为(28.5±3.6),明显多于对照组(12.4±2.8),其差异具有统计学意义(P<0.01)。低氧12 h,HT-29细胞胞核内NF-κB的表达逐渐上调,24 h达顶峰。结论在适度低氧下,HT-29细胞增殖加快,并诱导促进新生血管形成,促进其向恶性表型转化,可能与细胞核内NF-κB的表达上调有关。

【Objective】To investigate the correlation between moderate hypoxia and tumor proliferation and angiogenesis in colon carcinoma cell line HT-29, and to detect the expression of NF-κB/p65 protein in nuclei so as to explore its latent mechanism of malignant transformation.【Methods】The moderate hypoxia model was established in accordance with the PO2 of tumor in vivo. The proliferation of HT-29 cells was detected using MTT chromometry in order to draw a growth curve. Tube formation of human umbilical vein endothelial cells（HUVECs） was adopted to evaluate the ability of tumor angiogenesis in vitro. After different period in moderate hypoxia, NF-κB/p65 levels in nuclei were assessed by Western blot after extraction of nucleic protein.【Results】The morphology of HT-29 cells in moderate hypoxia was similar to that in normoxia, the absorbance values of the moderate hypoxia group were higher than those of the normoxia group with a statistic significance. The numbers of tubes formed by HUVECs were（12.4±2.8） in the control group and（28.5±3.6） in the moderate hypoxia group with a significant difference（P〈0.01）. The expression of NF-κB/p65 protein in nuclei under moderate hypoxia gradually up-regulated at 12 h and reached the peak at 24 h.【Conclusions】Moderate hypoxia could induce malignant transformation in HT-29 cells through upregulating tumor proliferation and angiogenesis, in which NF-κB could be involved in this malignant phenotype.