Department of Urology, Nara Medical University, 840 Shijo-cho, Nara 634-8522, Japan.
Oncol Rep. 2011 Mar;25(3):653-60. doi: 10.3892/or.2010.1125. Epub 2010 Dec 27.
Angiogenesis is necessary for the growth, invasion, and metastasis of solid tumors. Previous studies have shown that heme oxygenase-1 (HO-1) plays an important role in angiogenesis in both normal and cancerous cells, such as vascular endothelial cells and pancreatic cancer cells, respectively. In this study, we analyzed the role of HO-1 and other angiogenic factors in urothelial carcinoma of the bladder. Specifically, we used real-time reverse transcription polymerase chain reaction (PCR) and Western blotting to investigate the upregulation of 7 angiogenic factors, namely, HO-1, vascular endothelial growth factor (VEGF), hypoxia-inducible factor (HIF)-1α, HIF-2α, cyclooxygenase-2 (COX-2), interleukin-8 (IL-8), and basic fibroblast growth factor (bFGF) under hypoxic conditions in the T24 urothelial carcinoma cell line. We also used enzyme-linked immunosorbent assay (ELISA) to measure the amount of VEGF secreted into the growth media. In addition, we administered an HO-1 inhibitor, zinc protoporphyrin IX, to mice with subcutaneous T24 tumors to assess the modulation of angiogenesis in solid tumors in vivo. We also performed immunohistochemical analyses of 23 primary bladder cancer specimens with high-grade tumors infiltrating into the stroma (pT1) for expression of HO-1, VEGF, HIF-1α, HIF-2α, COX-2, and CD31. Image analysis of CD31 staining was performed to estimate microvessel density (MVD), a measure of angiogenesis. Hypoxic conditions induced upregulation of HO-1, VEGF, HIF-1α, HIF-2α, and COX-2 in T24 cells and increased VEGF secretion, which could be suppressed by zinc protoporphyrin IX. In vivo, inhibition of HO-1 decreased tumor growth and MVD by suppressing angiogenic factors, particularly VEGF and HIF-1α. In clinical specimens of bladder cancer, high expression of HO-1 was correlated with high expression of HIF-1α (P=0.027) and high MVD (P=0.005), but not with VEGF expression (P=0.19). In conclusion, since overexpression of HO-1 promotes angiogenesis in urothelial carcinoma cells, HO-1 inhibitors could be used as novel therapeutics for urothelial carcinoma of the urinary bladder.
血管生成对于实体瘤的生长、侵袭和转移是必要的。先前的研究表明，血红素加氧酶-1（HO-1）在正常细胞和癌细胞（如血管内皮细胞和胰腺癌细胞）中的血管生成中发挥重要作用。在这项研究中，我们分析了 HO-1 和其他血管生成因子在膀胱尿路上皮癌中的作用。具体来说，我们使用实时逆转录聚合酶链反应（PCR）和 Western blot 来研究在 T24 尿路上皮癌细胞系缺氧条件下 7 种血管生成因子（即 HO-1、血管内皮生长因子（VEGF）、缺氧诱导因子（HIF）-1α、HIF-2α、环氧化酶-2（COX-2）、白细胞介素-8（IL-8）和碱性成纤维细胞生长因子（bFGF））的上调情况。我们还使用酶联免疫吸附测定（ELISA）测量 VEGF 分泌到生长培养基中的量。此外，我们给皮下 T24 肿瘤的小鼠施用 HO-1 抑制剂锌原卟啉 IX，以评估体内实体瘤中血管生成的调节。我们还对 23 例浸润到基质中的高级别膀胱癌标本（pT1）进行 HO-1、VEGF、HIF-1α、HIF-2α、COX-2 和 CD31 的免疫组织化学分析。对 CD31 染色进行图像分析，以估计微血管密度（MVD），这是血管生成的一个衡量标准。缺氧条件诱导 T24 细胞中 HO-1、VEGF、HIF-1α、HIF-2α 和 COX-2 的上调，并增加 VEGF 的分泌，锌原卟啉 IX 可抑制其分泌。在体内，通过抑制血管生成因子，特别是 VEGF 和 HIF-1α，HO-1 的抑制可降低肿瘤生长和 MVD。在膀胱癌的临床标本中，HO-1 的高表达与 HIF-1α 的高表达（P=0.027）和高 MVD（P=0.005）相关，但与 VEGF 表达无关（P=0.19）。总之，由于 HO-1 的过度表达促进了尿路上皮癌细胞中的血管生成，因此 HO-1 抑制剂可作为膀胱癌的新型治疗药物。
