徐润强　刘海盛

【摘要】 目的：探究靶向血管内皮生长因子A（vascularendothelial growth factor A， VEGFA）调控促分裂原活化的蛋白激酶/细胞外调解蛋白激酶（mitogen-activated protein kinase/extracellular regulated protein kinase， MAPK/ERK）和磷脂酰肌醇3-激酶/蛋白激酶B（phosphatidylinositol 3- kinase/protein kinase B， PI3K/Akt）信号通路在结直肠癌血管生成中的作用及机制。方法：收集本院2015年1月-2019年12月82例结直肠癌患者癌组织及癌旁正常组织，使用qRT-PCR检测患者癌组织、癌旁正常组织中VEGFA水平，分析VEGFA水平与患者临床病理特征的关系。将SW480细胞分为五组，分别为对照组（正常人结肠细胞），空白组（无转染质粒），pMIR-miR-29b组（转染miR-29b），MAPK/ERK组（转染miR-29b后添加MAPK/ERK信号通路抑制剂PD98059），PI3K/Akt组（转染miR-29b后添加PI3K/Akt信号通路抑制剂wortmanin）。检测各组细胞中VEGFA、MAPK/ERK及PI3K/Akt信号通路靶蛋白ERK、Akt、mTOR、Bcl-2的表达水平。将各组细胞移植至裸鼠，建立结直肠癌裸鼠转移瘤模型，观察各组裸鼠移植瘤生长情况，检测各组裸鼠肿瘤微血管密度变化情况。结果：癌组织VEGFA表达水平高于癌旁正常组织（P<0.05）;癌组织内Ⅲ、Ⅳ期患者VEGFA mRNA水平高于Ⅰ、Ⅱ期患者（P<0.05），淋巴结转移患者VEGFA表达水平高于无淋巴结转移患者（P<0.05）。与對照组相比，空白组、pMIR-miR-29b组、MAPK/ERK组、PI3K/Akt组VEGFA、ERK、Akt、mTOR、Bcl-2 mRNA及蛋白表达水平均上升（P<0.05）。MAPK/ERK组与PI3K/Akt组肿瘤重量、微血管密度均低于空白组（P<0.05），pMIR-miR-29b组裸鼠微血管密度、肿瘤重量均明显低于MAPK/ERK组与PI3K/Akt组（P<0.05）。结论：VEGFA可有效下调MAPK/ERK及PI3K/Akt信号通路，从而抑制结肠癌血管新生，为临床治疗结直肠癌提供了新的临床依据。

【关键词】 VEGFA MAPK/ERK PI3K/Akt 结直肠癌 血管生成

[Abstract] Objective： To investigate the effect of vascular endothelial growth factor A （VEGFA） on mitogen activated protein kinase/extracellular regulated protein kinase （MAPK/ERK） and the role and mechanism of phosphatidylinositol 3-kinase/protein kinase B （PI3K/Akt） signaling pathways in colorectal cancer angiogenesis. Method： A total of 82 patients with colorectal cancer from January of 2015 to December of 2019 in our hospital were collected. The levels of VEGFA in cancer tissue and normal tissue were detected by qRT PCR， and the relationship between VEGFA level and clinicopathological characteristics was analyzed. SW480 cells were divided into five groups： control group （normal colon cells）， blank group （without transfection plasmid）， pMIR-miR-29b group （transfected with miR-29b）， MAPK/ERK group （adding MAPK/ERK signal pathway inhibitor PD98059） and PI3K/Akt group （adding PI3K/Akt signal pathway inhibitor wortmanin after miR-29b transfection）. The expression levels of VEGFA， MAPK/ERK， ERK， Akt， mTOR and Bcl-2 were detected. The cells were transplanted into nude mice to establish colorectal cancer metastasis model in nude mice. The growth of transplanted tumor and the change of tumor microvessel density were observed. Result： The expression level of VEGFA in cancer tissue was higher than that in the normal tissues （P<0.05）， the level of VEGFA mRNA in stage Ⅲ and Ⅳ patients was higher than that in stage Ⅰ and Ⅱ patients （P<0.05）， and that of patients with lymph node metastasis was higher than that in patients without lymph node metastasis （P<0.05）. Compared with the control group， the expression levels of VEGFA， ERK， Akt， mTOR， Bcl-2 mRNA and protein in blank group， pMIR-miR-29b group， MAPK/ERK group， PI3K/Akt group were all increased （P<0.05）. The tumor weight and microvessel density of MAPK/ERK group and PI3K/Akt group were lower than that in blank group （P<0.05）， and that of pMIR-miR-29b group was significantly lower than those of

MAPK/ERK group and PI3K/Akt group （P<0.05）. Conclusion： VEGFA can effectively down regulate MAPK/ERK and PI3K/Akt signaling pathways， thus inhibiting angiogenesis of colon cancer， which provides a new clinical basis for clinical treatment of colorectal cancer.

[Key words] VEGFA MAPK/ERK PI3K/Akt Colorectal cancer Angiogenesis

First-authors address： Zengcheng District Peoples Hospital， Guangzhou 511300， China

doi：10.3969/j.issn.1674-4985.2021.09.008

在肿瘤进展过程中，病理性血管生长处于激活状态，从而促进肿瘤组织发生远处转移。肿瘤在生长及转移过程中，血管内皮生长因子A（vascularendothelial growth factor A， VEGFA）可有效促进细胞分裂增殖。有临床研究显示，结直肠癌患者血清中VEGFA表達水平与其血管新生状态密切相关，是患者预后的独立预测因子[1]。促分裂原活化的蛋白激酶/细胞外调解蛋白激酶（mitogen-activated protein kinase/extracellular regulated protein kinase，MAPK/ERK）和磷脂酰肌醇3-激酶/蛋白激酶B（phosphatidylinositol 3- kinase/protein kinase B，PI3K/Akt）信号通路与肿瘤细胞血管内皮细胞生长及血管新生有重要作用[2-3]。有研究表明，VEGFA下降可导致MAPK/ERK、PI3K/Akt信号通路活化抑制，从而发挥抗血管生成作用[4]。但结直肠癌肿瘤发生发展的过程中血管生成的机制尚不清楚，本研究解释了VEGFA在结直肠癌发生、血管生成中的作用，为直肠癌靶向治疗提供依据，现报道如下。……