Wu Lun, Zhang You-Shun, Ye Meng-Liang, Shen Feng, Liu Wei, Hu Hong-Sheng, Li Sheng-Wei, Wu Hong-Wei, Chen Qin-Hua, Zhou Wen-Bo
Liver Surgery Institute of Experiment Center of Medicine, Department of Hepatobiliary Surgery, Dongfeng Hospital, Hubei University of Medicine, Shiyan, Hubei 442001, P.R. China.
Department of Biostatistics, College of Public Health and Management, Chongqing Medical University, Chongqing 400016, P.R. China.
Exp Ther Med. 2017 Jun;13(6):3529-3534. doi: 10.3892/etm.2017.4428. Epub 2017 May 4.
Rapid growth of residual tumors can occur as a result of their recurrence and progression. The present study aimed to investigate the expression of hypoxia inducible factor-2 subunit α (HIF-2α), vascular endothelial growth factor A (VEGFA), erythropoietin-producing hepatocellular A2 (EphA2) and angiogenesis in residual hepatocellular carcinoma (HCC), following treatment with high-intensity focused ultrasound (HIFU) ablation, in order to investigate the association between protein expression and tumor recurrence and growth. Athymic BALB/c (nu/nu) mice were subcutaneously inoculated with the HCC cell line HepG2, in order to create xenograft tumors. Approximately 30 days post-inoculation, eight mice were treated with HIFU, whereas eight mice received no treatment and acted as the control group. Residual tumor tissues were obtained from the experimental groups after one month. Levels of HIF-2α, VEGFA, EphA2 and cluster of differentiation 31 (CD31) expression was measured by immunohistochemical staining. CD31-positive vascular endothelial cells were counted to calculate microvascular density (MVD), and western blot analysis was performed to determine levels of HIF-2α, VEGFA, and EphA2 protein. It was found that the expression levels of HIF-2α, VEGFA, EphA2, and MVD proteins in residual HCC tissues were significantly higher than in the control group tissues (P<0.05). Tumor MVD was strongly correlated with VEGFA (R=0.957, P<0.01) and EphA2 (R=0.993, P<0.01) protein expression levels. Furthermore, there was a significant positive correlation between HIF-2α and EphA2 expression (R=0.991, P<0.01). The correlation between VEGFA and EphA2 expression was also positive (R=0.985, P<0.01). These data suggest that overexpression of HIF-2α, VEGFA and EphA2 is related to angiogenesis in residual HCC following HIFU ablation, potentially via their association with key mediators of recurrence.
残余肿瘤的快速生长可能是其复发和进展的结果。本研究旨在调查高强度聚焦超声（HIFU）消融治疗后，残余肝细胞癌（HCC）中缺氧诱导因子-2亚基α（HIF-2α）、血管内皮生长因子A（VEGFA）、促红细胞生成素产生肝细胞A2（EphA2）的表达及血管生成情况，以研究蛋白表达与肿瘤复发和生长之间的关联。将无胸腺BALB/c（nu/nu）小鼠皮下接种HCC细胞系HepG2，以建立异种移植肿瘤。接种后约30天，8只小鼠接受HIFU治疗，而8只小鼠未接受治疗，作为对照组。1个月后从实验组获取残余肿瘤组织。通过免疫组织化学染色测量HIF-2α、VEGFA、EphA2和分化簇31（CD31）的表达水平。对CD31阳性血管内皮细胞进行计数以计算微血管密度（MVD），并进行蛋白质印迹分析以确定HIF-2α、VEGFA和EphA2蛋白的水平。结果发现，残余HCC组织中HIF-2α、VEGFA、EphA2和MVD蛋白的表达水平显著高于对照组组织（P<0.05）。肿瘤MVD与VEGFA（R=0.957，P<0.01）和EphA2（R=0.993，P<0.01）蛋白表达水平密切相关。此外，HIF-2α与EphA2表达之间存在显著正相关（R=0.991，P<0.01）。VEGFA与EphA2表达之间的相关性也为正（R=0.985，P<0.01）。这些数据表明，HIF-2α、VEGFA和EphA2的过表达与HIFU消融后残余HCC中的血管生成有关，可能是通过它们与复发关键介质的关联。
