敲减免疫调节蛋白B7-H3基因对膀胱癌细胞恶性增殖、侵袭和干细胞样特性的影响

doi:10.3969/j.issn.1674-8115.2021.11.008

上海交通大学学报(医学版) ›› 2021, Vol. 41 ›› Issue (11): 1454-1460.doi: 10.3969/j.issn.1674-8115.2021.11.008

敲减免疫调节蛋白B7-H3基因对膀胱癌细胞恶性增殖、侵袭和干细胞样特性的影响

王晓寒¹(), 叶云林², 肖康华², 赵志鑫³, 吴静雅⁴

^1.河南护理职业学院外科教研室，安阳 455000
^2.中山大学附属肿瘤医院泌尿肿瘤科，广州 510030
^3.河南护理职业学院药学教研室，安阳 455000
^4.河南省安阳地区医院中医科，安阳 455000

出版日期:2021-11-28 发布日期:2021-12-03
通讯作者: 王晓寒 E-mail:wang136473@163.com
作者简介:王晓寒（1983—），女，副教授，硕士；电子信箱：wang136473@163.com。
基金资助:
2018年度河南省医学科技攻关计划项目(2018020950)

Effects of knockdown of immunoregulatory protein B7-H3 gene on malignant proliferation, invasion and stem cell-like characteristics of bladder cancer cells

Xiao-han WANG¹(), Yun-lin YE², Kang-hua XIAO², Zhi-xin ZHAO³, Jing-ya WU⁴

^1.Department of surgery, Henan Vocational College of nursing, Anyang 455000, China
^2.Department of Urology and oncology, Cancer Hospital Affiliated to Zhongshan University, Guangzhou 510030, China
^3.Pharmacy teaching and Research Office, Henan Vocational College of nursing, Anyang 455000, China
^4.Department of traditional Chinese medicine, Henan Province Anyang District Hospital, Anyang 455000, China

Online:2021-11-28 Published:2021-12-03
Contact: Xiao-han WANG E-mail:wang136473@163.com
Supported by:
2018 Henan Province Medical Science and Technology Tackling Project(2018020950)

摘要/Abstract

摘要：

目的·探讨干扰免疫调节蛋白B7-H3基因对膀胱癌J82细胞的恶性增殖和干细胞样特性的影响。方法·shRNA干扰效率的检测采用反转录聚合酶链式反应（reverse transcription-polymerase chain reaction，RT-PCR）。J82细胞增殖采用克隆形成实验检测。细胞凋亡采用流式细胞术检测。细胞侵袭采用Transwell检测。肿瘤克隆能力采用肿瘤成球实验观察。细胞凋亡、侵袭和干细胞样相关蛋白表达采用蛋白质印迹法（Western blotting）检测。结果·B7-H3在膀胱癌组织及膀胱癌细胞系中高表达（P=0.000）。确定shRNA2为设计的3组shRNAs中干扰B7-H3基因效率最高（P=0.000）。B7-H3干扰抑制了J82细胞的增殖、侵袭、肿瘤克隆能力及干细胞样特性（P=0.000）。并且，B7-H3干扰促进了J82细胞的凋亡（P=0.000）。结论·免疫调节蛋白B7-H3基因干扰可对膀胱癌J82细胞的恶性增殖、侵袭及干细胞样特性产生抑制作用。

关键词: B7-H3, 膀胱癌, J82细胞, 干细胞样特性

Abstract:

Objective·To investigate the effect of gene interfering with immune regulatory protein B7-H3 on malignant proliferation and stemness characteristics of bladder cancer J82 cells.

Methods·Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect shRNA interference efficiency. The proliferation of J82 cells was detected by clone formation assay. Cell apoptosis was detected by flow cytometry. Cell invasion was detected using Transwell asssay. The tumor-cloning ability was observed using tumor sphere formation experiment. Apoptosis, invasion, and expression and stemness-related proteins were detected by Western blotting.

Results·B7-H3 is highly expressed in bladder cancer tissues and bladder cancer cell lines (P=0.000). shRNA2 was identified as the most efficient interference with B7-H3 gene in three shRNAs (P=0.000). The interference of B7-H3 inhibited the proliferation, invasion, tumor cloning ability and stem cell-like characteristics of J82 cells (P=0.000). Furthermore, B7-H3 interference promoted the apoptosis of J82 cells (P=0.000).

Conclusion·The gene interference of immunoregulatory protein B7-H3 can inhibit the malignant proliferation, invasion and stemness characteristics of bladder cancer J82 cells, providing a new possibility and effective evidence for the treatment of bladder cancer.

Key words: B7-H3, Bladder cancer, J82 cells, Stemness characteristics

中图分类号:

R737.14

王晓寒, 叶云林, 肖康华, 赵志鑫, 吴静雅. 敲减免疫调节蛋白B7-H3基因对膀胱癌细胞恶性增殖、侵袭和干细胞样特性的影响[J]. 上海交通大学学报(医学版), 2021, 41(11): 1454-1460.

Xiao-han WANG, Yun-lin YE, Kang-hua XIAO, Zhi-xin ZHAO, Jing-ya WU. Effects of knockdown of immunoregulatory protein B7-H3 gene on malignant proliferation, invasion and stem cell-like characteristics of bladder cancer cells[J]. JOURNAL OF SHANGHAI JIAOTONG UNIVERSITY (MEDICAL SCIENCE), 2021, 41(11): 1454-1460.

图/表 6

图1 B7-H3在膀胱癌组织和膀胱癌细胞系中的表达Note： A. Expression of B7-H3 in bladder cancer tissues. B. Expression of B7-H3 in bladder cancer cell lines. ①P=0.000, compared with the adjacent group; ②P=0.000, compared with the normal group.

Fig 1 Expression of B7-H3 in bladder cancer tissues and bladder cancer cell lines

图2 膀胱癌细胞J82转染3种shRNAs后对B7-H3干扰效率的检测Note： A. Expression level of B7-H3 mRNA in shRNA of the three groups. B. B7-H3 protein expression in shRNA of the three groups. C. Histogram of B7-H3 protein expression in the three shRNA groups. ①P=0.000 compared with the control group.

Fig 2 Detection of B7-H3 interference efficiency of bladder cancer cells J82 transfected with the three shRNAs

图3 J82细胞增殖情况的检测Note： ①P =0.000, compared with the control group.

Fig 3 Detection of J82 cells proliferation

图4 J82细胞的凋亡情况的检测Note： A. Apoptosis rate of J82 cells. B. Expression of apoptosis-related proteins Bax and Bcl-2. ①P=0.000, compared with the control group.

Fig 4 Detection of apoptosis of J82 cells

图5 J82细胞侵袭情况Note： A. Invading J82 cells. B. Western blotting. ①P=0.000, compared with the control group.

Fig 5 Invasion of J82 cells

图6 J82细胞的干细胞样特性Note： A.Tumor into a ball experiment. B. Diameter of the tumor spheres. C. Number of clones of tumor spheres. D. Expression of stem cell markers SOX2 and OCT4. ①P =0.000, compared with the control group.

Fig 6 Stemness characteristics of J82 cells

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敲减免疫调节蛋白B7-H3基因对膀胱癌细胞恶性增殖、侵袭和干细胞样特性的影响

Effects of knockdown of immunoregulatory protein B7-H3 gene on malignant proliferation, invasion and stem cell-like characteristics of bladder cancer cells

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