Effect of macrophages on the expression of vascular cell adhesion molecule 1 in ovarian carcinoma cells and its mechanism
10.3760/cma.j.issn.1006-9801.2018.10.001
- VernacularTitle:巨噬细胞对卵巢癌细胞血管细胞黏附分子1表达的影响及其机制研究
- Author:
Ru ZHOU
1
;
Qian FENG
;
Shentong YU
;
Tong YANG
;
Shuhong YU
;
Shuang LIU
;
Zhuqing CUI
;
Jing ZHANG
Author Information
1. 空军军医大学基础医学院病理学教研室暨西京医院病理科
- Keywords:
Ovarian neoplasms;
Macrophages;
Cytokines;
Vascular cell adhesion molecule 1
- From:
Cancer Research and Clinic
2018;30(10):649-654
- CountryChina
- Language:Chinese
-
Abstract:
Objective To explore the effect of macrophages on the expression of vascular cell adhesion molecule 1 (VCAM1) in ovarian carcinoma cells and its mechanism. Methods Phorbol ester and lipopolysaccharide were used to activate the monocyte THP-1 that would become macrophages . Enzyme linked immunosorbent assay (ELISA) was used to detect the cytokines level in the supernatant of macrophages. The effect of macrophages ' supernatant on VCAM1 mRNA expression of ovarian HEY and IGROV1 carcinoma cells was detected by using quantitative real-time and polymerase chain reaction (qRT-PCR). Western blot was used to detect the effect of macrophages ' supernatant on VCAM1 protein expression of ovarian carcinoma cells with VCAM1 over expression (HEY-VCAM1 and IGROV1-VCAM1). Dual-luciferase report gene assay was used to detect the effect of macrophages' supernatant and the cytokines on promoter transcriptional activity in different truncations of human embryonic kidney cells HEK293T VCAM1 gene. Results Compared with the supernatant of THP-1 cells, the release number of tumor necrosis factor α (TNF-α), interleukin (IL)-6 and IL-12 in supernatant from macrophages was increased (all P< 0.05), and IL-10 was decreased (t=3.841, P=0.019). The levels of VCAM1 mRNA in HEY and IGROV1 cells were upregulated by macrophages' supernatant and 1 ng/ml TNF-α, and macrophages' supernatant could promote the expression levels of VCAM1 protein in HEY-VCAM1 cells and IGROV1-VCAM1 cells. Compared with the empty vector (pGV354) control group [(8.6 ±0.2) ×10-3 relative light unit (RLU)], the reporter gene luciferase activity of human embryonic kidney cells HEK293T VCAM1 gene promoter region were upregulated by supernatant from macrophages located at -1641 bp to +12 bp including the transcription binding site of AP-1 [(109.4±3.4)×10-3 RLU], and there was a significant difference (t=29.42, P<0.001). Compared with the negative control group untreated by cytokine [(21.0 ±0.5) ×10-3 RLU], 100 ng/ml TNF-α could promote the transcriptional activity of -1641 bp to +12 bp promoter of VCAM1 in HEK293T cells [(23.4±0.4)×10-3 RLU;t=4.134, P=0.001]. 150 ng/ml IL-6 had no effect on the transcriptional activity of the promoter [(21.4±1.0)× 10-3 RLU; t= 0.328, P= 0.708]. 5 ng/ml IL-12 inhibited the transcriptional activity of the promoter [(14.3 ± 1.0)×10-3 RLU;t= 6.390, P< 0.001]. Conclusion Macrophages can promote VCAM1 expression in ovarian carcinoma cells by secreting inflammatory factors like TNF-α to affect VCAM1 promoter region containing AP1 transcription binding site and can promote VCAM1 mRNA expression in ovarian cancer cells.
