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印记基因TSSC3对骨肉瘤血管生成的影响
Effect of overexpression of imprinted gene TSSC3 on angiogenesis in human osteosarcoma
- 第三军医大学学报 2018年40卷第10期 页码：855-861
- 作者机构：
1.400037 重庆，陆军军医大学(第三军医大学)第二附属医院病理科
2.400016 重庆，重庆医科大学第一附属医院骨科
- 作者简介：
[ 郭乔楠, E-mail: qiaonan85@263.net GUO Qiaonan, E-mail: qiaonan85@263.net
- 基金信息：国家自然科学基金面上项目(81602591);重庆市基础科学与前沿技术研究重点项目(CSTC2015jcyjBX0067)
DOI：10.16016/j.1000-5404.201801125
中图分类号： R394.3;R364.3;R738.1收稿：2018-01-15，
修回：2018-02-10，
纸质出版：2018-05-30
- 稿件说明：
移动端阅览
高自然, 赵郭盛, 彭动斌, 等. 印记基因TSSC3对骨肉瘤血管生成的影响[J]. 第三军医大学学报, 2018,40(10):855-861.
Ziran GAO, Guosheng ZHAO, Dongbin PENG, et al. Effect of overexpression of imprinted gene TSSC3 on angiogenesis in human osteosarcoma[J]. Journal of Third Military Medical University, 2018, 40(10): 855-861.
高自然, 赵郭盛, 彭动斌, 等. 印记基因TSSC3对骨肉瘤血管生成的影响[J]. 第三军医大学学报, 2018,40(10):855-861. DOI： 10.16016/j.1000-5404.201801125.
Ziran GAO, Guosheng ZHAO, Dongbin PENG, et al. Effect of overexpression of imprinted gene TSSC3 on angiogenesis in human osteosarcoma[J]. Journal of Third Military Medical University, 2018, 40(10): 855-861. DOI： 10.16016/j.1000-5404.201801125.
摘要
目的
2
通过体内外实验观察印记基因TSSC3对骨肉瘤血管生成的影响。
方法
2
利用慢病毒建立稳定过表达TSSC3的骨肉瘤MTH细胞株，收集骨肉瘤细胞上清制备条件培养基(conditional medium，CM)；通过划痕实验、Transwell实验及小管形成实验观察CM对人脐静脉内皮细胞(human umbilical vein endothelial cells，HUVEC)的迁移和小管形成能力的影响；RT-PCR、Western blot及酶联免疫吸附试验(ELISA)检测骨肉瘤细胞中血管内皮生长因子A(vascular endothelial growth factor A，VEGFA)mRNA及蛋白水平变化；建立裸鼠移植瘤模型，免疫组化检测肿瘤组织中VEGFA蛋白及微血管密度(microvessel density，MVD)的表达情况。
结果
2
过表达TSSC3(overTSSC3)组CM抑制内皮细胞迁移和小管形成，与对照组比较差异有统计学意义(
P
<
0.01)；RT-PCR、Western blot及ELISA检测结果显示，overTSSC3组细胞较对照组VEGFA mRNA和蛋白表达水平降低，差异有统计学意义(
P
<
0.01)；裸鼠移植瘤免疫组化检测结果显示，overTSSC3组肿瘤中VEGFA蛋白表达量及MVD明显低于对照组。
结论
2
印记基因TSSC3可能通过下调VEGFA的表达进而抑制骨肉瘤血管生成。
Abstract
Objective
2
To observe the effect of imprinted gene TSSC3 overexpression on angiogenesis in human osteosarcoma.
Methods
2
TSSC3 gene was transduced in human osteosarcoma MTH cells
via
lentiviral vector
and the culture supernatant was collected from the cells with and without TSSC3 gene transduction to prepare the conditional medium (CM) of osteosarcoma cells. Wound healing assay
Transwell migration assay and tube formation assay were used to assess the effect of CM on the migration and tube formation ability of human umbilical vein endothelial cells (HUVEC)
in vitro
. Real-time fluorescence quantitative PCR (RT-PCR) and Western blotting were used to detect the expression of vascular endothelial growth factor A (VEGFA) mRNA and protein in MTH cells
and VEGFA concentration in the CM was detected using enzyme-linked immunosorbent assay (ELISA). The expression of VEGFA and CD34 in MTH cell xenograft in a nude mice model was determined using immunohistochemistry.
Results
2
The CM from MTH cells overexpressing TSSC3 significantly suppressed the migration and tube formation of HUVECs. The results of RT-PCR
Western blotting and ELISA showed that TSSC3 overexpression caused significantly lowered expression of VEGFA at both the mRNA and protein levels in MTH cells (
P
<
0.01). In the tumor-bearing nude mice
VEGFA expression and microvessel density was significantly lowered in TSSC3-overexpressing MTH cell xenograft as compared with those in the control group.
Conclusion
2
TSSC3 suppresses angiogenesis in osteosarcoma possibly by down-regulating the expression of VEGFA.
关键词
Keywords
references
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