Zhang Xu-Dong, Wu Qiang, Yang Shu-Hua
Dr. Xu-dong Zhang, Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
Prof. Qiang Wu, Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
Pak J Med Sci. 2017 Mar-Apr;33(2):341-346. doi: 10.12669/pjms.332.12587.
To explore angiogenesis in osteosarcoma under the condition of hypoxia-inducible factor (HIF)-1α gene silenced by small interference RNA (siRNA).
The SaOS-2 osteosarcoma cells, transfected with the recombinant plasmid pSilencer2.1-HIF-1α or pSilencer2.1-SCR, were classified as HIF-1α/siRNA group or SCR/siRNA group, respectively. In which, vascular endothelial growth factor (VEGF) immunohistochemistry were performed. HIF-1α and VEGF protein contents were detected by western blot. Gene expressions of HIF-1α and VEGF were quantified by qPCR. Then the transfected SaOS-2 cells were inoculated in nude mice and transplantation tumor were checked via HE staining, VEGF and CD34 immunohistochemistry, and calculation of microvascular density (MVD).
In vitro, VEGF immunohistochemistry stains, HIF-1α and VEGF protein contents, and the relative expressions of HIF-1α mRNA and VEGF mRNA in HIF-1α/siRNA group were obviously reduced. In vivo, morphological observation illustrated that heteromorphism were not obvious in the cells of HIF-1α/siRNA group and vascular systems were sparse in its transplantation tumor tissue, and immunohistochemistry revealed that both VEGF and CD34 stains were significantly decreased in HIF-1α/siRNA group, and MVD in HIF-1α/siRNA group (7.3±1.1) were obviously less than that in SCR/siRNA group (17.2±3.2) (<0.05).
Angiogenesis in osteosarcoma can be inhibited by siRNA-mediated HIF-1α gene silencing, which is expected to provide a novel and attractive target of therapeutic strategies of osteosarcoma.
探讨小干扰RNA（siRNA）沉默缺氧诱导因子（HIF）-1α基因条件下骨肉瘤的血管生成情况。
将重组质粒pSilencer2.1-HIF-1α或pSilencer2.1-SCR转染人骨肉瘤SaOS-2细胞，分别分为HIF-1α/siRNA组和SCR/siRNA组，进行血管内皮生长因子（VEGF）免疫组化检测；采用蛋白质印迹法检测HIF-1α和VEGF蛋白含量；运用实时荧光定量PCR（qPCR）检测HIF-1α和VEGF基因表达。将转染后的SaOS-2细胞接种于裸鼠，通过苏木精-伊红（HE）染色、VEGF和CD34免疫组化检测及微血管密度（MVD）计算观察移植瘤情况。
体外实验中，HIF-1α/siRNA组VEGF免疫组化染色、HIF-1α和VEGF蛋白含量以及HIF-1α mRNA和VEGF mRNA相对表达均明显降低。体内实验中，形态学观察显示HIF-1α/siRNA组细胞异型性不明显，其移植瘤组织血管系统稀疏；免疫组化结果显示，HIF-1α/siRNA组VEGF和CD34染色均显著降低，HIF-1α/siRNA组MVD为（7.3±1.1），明显低于SCR/siRNA组的（17.2±3.2）（P<0.05）。
siRNA介导的HIF-1α基因沉默可抑制骨肉瘤血管生成，有望为骨肉瘤治疗策略提供新的、有吸引力的靶点。
