维生素D对大鼠肾上腺嗜铬细胞瘤PC12细胞中神经酰胺水平的影响

doi:10.6040/j.issn.1671-7554.0.2017.163

摘要/Abstract

摘要： 目的研究维生素D(VD)对大鼠肾上腺嗜铬细胞瘤细胞系PC12细胞中神经酰胺(Cer)水平的影响。方法采用C2-Cer(20 μmol/L)刺激PC12细胞,MTT法检测Cer对PC12细胞增殖的影响;高效液相色谱串联质谱法(HPLC-MS)测定VD(50或0 nmol/L)处理的PC12细胞中C16:0-Cer的含量。qRT-PCR法测定VD(50或0 nmol/L)处理的PC12细胞中Cer代谢相关酶mRNA的表达;Western blotting法检测VD(200、100、50、25、12.5和0 nmol/L)处理后,PC12细胞中SPTLC1和Cers6蛋白的相对表达量。结果 C2-Cer能抑制PC12细胞的活性,IC₅₀为2.800 μmol/L。与0 nmol/L VD相比,50 nmol/L VD降低了PC12细胞中C16:0-Cer的含量［(19.37±0.61)ng/mL vs(16.50±0.10)ng/mL,P=0.010］,升高了SPTLC1 mRNA的表达水平(P=0.001)和Cers6 mRNA的表达水平(P<0.001),降低了SPTLC1蛋白的相对表达量(P=0.032),升高了Cers6蛋白的相对表达量(P=0.020)。结论 VD 降低PC12中Cer的含量,可能与降低SPTLC1蛋白表达有关。

关键词: 神经酰胺, PC12细胞, 维生素D, 高效液相色谱-质谱

Abstract: Objective To invastigate the effect of vitamin D(VD)on the content of ceramide(Cer)in rat adrenal pheochromocytoma PC12 cells. Methods MTT was performed to verify the effect of short-chain C2-Cer(20 μmol/L)on cell proliferation. High performance liquid chromatography tandem mass spectrometry(HPLC-MS)was performed to detect the concentration of C16: 0-Cer in PC12 cells stimulated with VD(50 or 0 nmol/L). qRT-PCR was performed to measure the mRNA expression levels of enzymes which were related to the metabolism of Cer in PC12 cells after being stimulated with VD(50 or 0 nmol/L). Western blotting was performed to measure the protein levels of SPTLC1 and Cers6 in PC12 cells that were stimulated with VD(200, 100, 50, 25, 12.5 and 0 nmol/L). Results Activity of PC12 cells was inhibited by C2-Cer with IC50 being 2.800 μmol/L. Compared with VD at 0 nmol/L, the content of C16: 0-Cer in PC12 cells was decreased by VD at 50 nmol/L ［(19.37±0.61)ng/mL vs(16.50±0.10)ng/mL, P=0.010］, the expressions of SPTLC1 mRNA(P=0.001)and Cers6 mRNA(P<0.001)in PC12 cells were increased, the protein expression of SPTLC1 in PC12 cells was reduced(P=0.032)and the protein expression of Cers6 was increased(P=0.020). Conclusion The concentration of C16:0-Cer is decreased in PC12 cells that were treated with VD, and it may be related to the decrease of protein expression of SPTLC1.

Key words: Ceramide, PC12 cell, Vitamin D, High pressure liquid chrometography /mass spectroscopy

中图分类号:

R744.5+1

关新媛,高计芳,郭娟娟,刘师莲. 维生素D对大鼠肾上腺嗜铬细胞瘤PC12细胞中神经酰胺水平的影响[J]. 山东大学学报 (医学版), 2017, 55(12): 13-17.

GUAN Xinyuan, GAO Jifang, GUO Juanjuan, LIU Shilian. Effects of Vitamin D on the content of Ceramide in rat adrenal pheochromocytoma PC12 Cells[J]. Journal of Shandong University (Health Sciences), 2017, 55(12): 13-17.

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