Regulatory effect of Rac1 protein on vascular endothelial growth factor secreted in breast cancer MDAMB-231 cells

doi:10.3877/cma.j.issn.1674-0807.2016.01.003

Abstract

Abstract:

Objective

To investigate the regulatory effect of Rac1, one member of Rho protein family, on vascular endothelial growth factor (VEGF) in breast cancer MDA-MB-231 cells.

Methods

MDAMB-231 cells were transfected by Rac1 constitutively-active mutant plasmid pCEFL-GST-V12Rac1 (V12Rac1 group), blank plasmid pCEFL-GST-neo (blank control group), small interference Rac1 RNA (siRac1 group)and small interference RNA of control (si-control group) respectively. VEGF expression levels in MDA-MB-231 cells were determined by ELISA assay at different time points(24, 48, 72 h after transfection). Western blot assay was performed to detect the relative expressions of angiogenesis-related factors p53 and VEGF.VEGF expression levels were compared by repeated measurement analysis of variance and t test was used to compare the quantitative data of Western blot.

Results

ELISA assays showed that after up-regulating Rac1 expression in MDA-MB-231 cells, VEGF level was gradually increased with time, indicating a significant difference compared with blank control group (group comparison: F=837.122, P ＜0.001; comparison of different time points:F=57 806.374,P＜0.001;interaction of grouping and time:F=7 663.095,P＜0.001).After down-regulating Rac1,VEGF level was significantly lower than that in si-control group (group comparison:F=511.891,P＜0.001; comparison of different time points: F=268.078,P＜0.001; interaction of grouping and time: F=120.708,P=0.001). Western blot assay showed that in MDA-MB-231 cells, the increased Rac1 expression could inhibit p53 expression (t=-6.392, P=0.003) and enhance VEGF expression (t=7.497, P=0.002) compared with blank control group, while decreased Rac1 expression could elevate p53 expression (t=5.307, P=0.006) and inhibit VEGF expression (t=-7.395, P=0.002) compared with sicontrol group.

Conclusion

Rac1 expression could affect VEGF expression in breast cancer MDA-MB-231 cells and Rac1 may promote VEGF expression through inhibiting p53 expression.

Key words: Breast neoplasms, Vascular endothelial growth factors, Tumor suppressor protein p53, RAC1 protein, human

Ji Ma, Qingli Zhao, Yi Zhao, Ying Liu. Regulatory effect of Rac1 protein on vascular endothelial growth factor secreted in breast cancer MDAMB-231 cells[J]. Chinese Journal of Breast Disease(Electronic Edition), 2016, 10(01): 10-13.

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Figures/Tables 5

组别	实验次数	VEGF蛋白表达水平(pg/ml, $\overset{ˉ}{x}$ ±s)
组别	实验次数	24 h	48 h	72 h
空白对照组	3	102.21±2.01	135.31±3.60	168.09±2.62
V12Rac1组	3	113.54±4.53	173.11±3.55	255.02±4.55

组别	实验次数	VEGF蛋白表达水平(pg/ml, $\overset{ˉ}{x}$ ±s)
组别	实验次数	24 h	48 h	72 h
空白对照组	3	102.21±2.01	135.31±3.60	168.09±2.62
V12Rac1组	3	113.54±4.53	173.11±3.55	255.02±4.55

组别	实验次数	VEGF蛋白表达水平(pg/ml, $\overset{ˉ}{x}$ ±s)
组别	实验次数	24 h	48 h	72 h
沉默对照组	3	102.04±3.11	136.55±3.72	168.62±4.35
siRac1组	3	93.11±3.20	104.23±4.00	107.15±3.83

组别	实验次数	VEGF蛋白表达水平(pg/ml, $\overset{ˉ}{x}$ ±s)
组别	实验次数	24 h	48 h	72 h
沉默对照组	3	102.04±3.11	136.55±3.72	168.62±4.35
siRac1组	3	93.11±3.20	104.23±4.00	107.15±3.83

组别	实验次数	p53^a	VEGF^a
沉默对照组	3	1.407±0.077	1.163±0.140
siRac1组	3	2.623±0.389	0.483±0.075
t值		5.307	-7.395
P值		0.006	0.002

组别	实验次数	p53^a	VEGF^a
空白对照组	3	2.043±1.114	0.643±0.130
V12Rac1组	3	1.425±0.123	1.323±0.089
t值		-6.392	7.497
P值		0.003	0.002

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