【摘要】 目的 探究维生素D对非三阴性乳腺癌（non-triple negative breast cancer, Non-TNBC）和TNBC癌细胞增殖的影响及分子机制。方法 收集TNBC和Non-TNBC患者乳腺组织，原代培养TNBC和Non-TNBC患者乳腺细胞；免疫荧光检测乳腺细胞雌激素受体（ER）、孕激素受体（PR）和HER2蛋白表达；免疫荧光检测Non-TNBC和TNBC乳腺癌组织和乳腺细胞中VDR蛋白表达；CCK-8检测细胞活力变化；流式细胞术检测细胞增殖和细胞周期的变化；光学比色法检测细胞谷氨酰胺合成酶（glutamine synthetase, GS）活力的变化；ELISA检测细胞培养上清和胞内谷氨酰胺的水平；Western blot检测细胞GS和VDR蛋白表达的变化；CHIP-PCR检测VDR对GS的转录调控。结果 TNBC患者外周血中维生素D水平和癌组织VDR蛋白表达明显低于Non-TNBC患者（P<0.05）。相比于原代Non-TNBC乳腺癌细胞，原代TNBC患者乳腺癌细胞低表达雌激素受体（ER）、孕激素受体（PR）和HER2蛋白表达；TNBC乳腺癌细胞VDR表达水平明显低于Non-TNBC乳腺癌细胞（P<0.05）;VD(10^-7～10^-4mol/L)可明显剂量依赖性的抑制Non-TNBC乳腺癌细胞和TNBC乳腺癌细胞细胞活力（P<0.05）。相比于0mol/L VD组，10³mol/L的VD组Non-TNBC乳腺癌细胞和TNBC乳腺癌细胞细胞增殖比例明显减少（P<0.05）;相比于0mol/L VD组，10³mol/L的VD组Non-TNBC乳腺癌细胞G1/G0期细胞比例明显减少（P<0.05）;10³mol/L的VD组TNBC乳腺癌细胞G1/G0期细胞比例明显减少，S期细胞比例明显增加（P<0.05）;相比于0mol/L VD组，10³mol/L的VD组Non-TNBC和TNBC乳腺癌细胞谷氨酰胺合成酶活力、谷氨酰胺合成酶蛋白、细胞内和细胞外谷氨酰胺水平表达比例明显减少（P<0.05）;相比于0mol/L VD组，10³mol/L的VD组Non-TNBC乳腺癌细胞和TNBC乳腺癌细胞VDR蛋白表达明显增加（P<0.05）;10³mol/L的VD组TNBC乳腺癌细胞VDR可转录调控GS基因转录（P<0.05）。结论 维生素D通过下调谷氨酰胺合成酶抑制乳腺癌细胞增殖。更多还原

【Abstract】 Objective To explore the effect and molecular mechanism of vitamin D（VD） on the proliferation of non-triple negative breast cancer（Non-TNBC） and TNBC cancer cells.Methods We collected breast tissues from patients with TNBC and Non-TNBC, and cultured primary breast cells from patients with TNBC and Non-TNBC; Estrogen receptor（ER）, progesterone receptor（PR） and HER2 protein expressions in breast cells immunofluorescence was detected by immunofluorescence; VDR protein expression in Non-TNBC and TNBC breast cancer tissues and breast cells was detected by immunofluorescence; Cell viability was detected by CCK-8 method; Cell proliferation and cell cycle was detected by flow cytometry; Glutamine synthetase（GS） activity was detected by optical colorimetry method; Intracellular and extracellular glutamine were detected by ELISA; GS and VDR protein expressions were detected by western blot; The transcriptional regulation of GS by VDR was detected by CHIP-PCR.Results VD concentration in peripheral blood and the VDR protein expression in cancer tissues of patients with TNBC were significantly lower than those of patients with Non-TNBC（P <0.05）; ER, PR, HER2 and VDR protein expressions in breast cancer cells of primary TNBC breast cancer cells were lower than primary Non-TNBC breast cancer cells（P<0.05）; VD(10^-7-10^-4mol/L) could significantly inhibit the cell viability of Non-TNBC breast cancer cells and TNBC breast cancer cells in a dose-dependent manner（P<0.05）. Compared with the 0 mol/L VD group, the proliferative proportion of Non-TNBC breast cancer cells and TNBC breast cancer cell in the 10³mol/L VD group was significantly reduced（P<0.05）; Compared with the 0 mol/L VD group, the proportion of non-TNBC breast cancer cells in the G1/G0 phase of the 10³mol/L VD group was significantly reduced（P<0.05）; The proportion of TNBC breast cancer cells in the G1/G0 phase of the 10³mol/L VD group was significantly reduced, and the proportion of cells in the S phase was significantly increased（P<0.05）; Compared with 0 mol/L VD group, Non-TNBC and TNBC breast cancer cell GS activity, GS protein, intracellular and extracellular glutamine content expression ratio of the 10³mol/L VD group was significantly reduced（P<0.05）; Compared with the 0 mol/L VD group, VDR protein expression of Non-TNBC breast cancer cells and TNBC breast cancer cells in the 10³mol/L VD group was significantly increased（P<0.05）; 10³mol/L The VDR of TNBC breast cancer cells in the VD group could transcriptionally regulate the transcription of GS gene（P<0.05）.Conclusion Vitamin D inhibits breast cancer cell proliferation by down-regulating glutamine synthetase.更多还原