南方医科大学学报

南方医科大学学报 ›› 2022, Vol. 42 ›› Issue (7): 1100-1108.doi: 10.12122/j.issn.1673-4254.2022.07.21

• • 上一篇    

JAG1影响血管生成并促进三阴性乳腺癌细胞的迁移、侵袭和粘附

刘俊平，石宇彤，吴敏敏，徐梦岐，张凤梅，何志强，唐 敏   

  1. 重庆医科大学检验医学院，临床检验诊断学教育部重点实验室，重庆 400016
  • 出版日期:2022-07-20 发布日期:2022-07-15

JAG1 promotes migration, invasion, and adhesion of triple-negative breast cancer cells by promoting angiogenesis

LIU Junping, SHI Yutong, WU Minmin, XU Mengqi, ZHANG Fengmei, HE Zhiqiang, TANG Min   

  1. Key Laboratory of Clinical Laboratory and Diagnostics of Ministry of Education, College of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, China
  • Online:2022-07-20 Published:2022-07-15

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摘要： 目的 探究JAG1对三阴性乳腺癌（TNBC）恶性表型的影响，并研究乳腺癌微环境血管生成的机制。方法 体外培养人TNBC细胞231和231B，Q-PCR实验检测Notch相关分子的表达水平。动物实验将雌性裸鼠分为231组和231B组，5只/组，乳腺脂肪垫分别注射231和231B细胞1×106/只。4~6周取肿瘤组织进行免疫组化和免疫荧光实验。使用JAG1重组蛋白处理231细胞和DAPT处理231B细胞，实验分4组：231空白对照组；rJAG1处理231组；231B空白对照组；DAPT处理231B组。用CCK-8法、Hoechst 33258染色实验、划痕愈合实验、Transwell小室实验和内皮细胞粘附实验检测TNBC的生物学特性变化。使用Western blot检测231和231B生物学特性相关蛋白的水平。接下来体外培养血管内皮细胞HUVEC，TNBC条件培养基处理HUVEC，实验分5组：（1）HUVEC阴性对照组；（2）231空白条件培养基处理组；（3）rJAG1处理231的条件培养基处理组；（4）231B空白条件培养基处理组；（5）DAPT处理231B的条件培养基处理组。CCK-8实验和基质胶成管实验分别检测HUVEC的增殖与成管能力。结果 与231细胞相比，231B表达更高的JAG1（P<0.05）。231B肿瘤表达更高的VEGFA和CD31。与231空白组相比，rJAG1处理组中231细胞的迁移、侵袭和粘附能力明显受到促进（P<0.05）。Twist1和Snail的蛋白水平均增加（均P<0.01），抗凋亡蛋白Bcl-2表达量增加（P<0.05），而DAPT明显抑制231B的相关现象和指标。JAG1过表达的条件培养基增加了HUVEC的细胞数以及成管数量（P<0.05）。结论 JAG1可能影响TNBC的恶性表型，并促进微环境的血管生成。

关键词: JAG1；三阴性乳腺癌；血管生成；血管内皮细胞生长因子；肿瘤微环境

Abstract: Objective To investigate the effect of JAG1 on the malignant phenotype of triple-negative breast cancer (TNBC) and its role in angiogenesis in breast cancer microenvironment. Methods The expressions of Notch molecules were detected in human TNBC 231 and 231B cells using RT-qPCR. Five female nude mice were inoculated with 231 cells and another 5 with 231B cells into the mammary fat pads, and 4-6 weeks later, the tumors were collected for immunohistochemical and immunofluorescence tests. 231 cells and 231B cells were treated with recombinant JAG (rJAG) protein and DAPT, respectively, and changes in their malignant phenotypes were assessed using CCK-8 assay, Hoechst 33258 staining, wound healing assay, Transwell chamber assay and endothelial cell adhesion assay. Western blotting was used to detect the changes in the expressions of proteins related with the malignant phenotypes of 231 and 231B cells. The effects of conditioned medium (CM) derived from untreated 231 and 231 B cells, rJAG1-treated 231 cells and DAPT-treated 231B cells on proliferation and tube formation ability of cultured human umbilical vein endothelial cells (HUVECs) were evaluated using CCK-8 assay and tube-forming assay. Results The expression of JAG1 was higher in 231B cells than in 231 cells (P<0.05). Tumor 231B showed higher expression of VEGFA and CD31. Compared with 231-Blank group, the migration, invasion and adhesion of 231 cells in 231-rJAG1 were significantly enhanced (P<0.05). Protein levels of Twist1 and Snail increased (P<0.01), anti-apoptotic protein Bcl-2 increased (P<0.05), while DAPT inhibited the related phenomena and indicators of 231B. The 231-rJAG1-CM increased the cell number and tubule number of HUVEC (P<0.05). Conclusion JAG1 may affect the malignant phenotype of TNBC and promote angiogenesis in the tumor microenvironment.

Key words: JAG1; triple-negative breast cancer; angiogenesis; vascular endothelial cell growth factor-A; tumor microenvironment