酸性神经磷脂酶在维生素E琥珀酸酯诱导胃癌细胞凋亡过程中对内质网应激的影响

doi:10.3969/j.issn.1004-616x.2013.05.001

癌变·畸变·突变

• 论著 • 下一篇

酸性神经磷脂酶在维生素E琥珀酸酯诱导胃癌细胞凋亡过程中对内质网应激的影响

董瑞华¹,赵巍²,王立明²,张旭光^1,吴坤^1，*

( 1. 哈尔滨医科大学营养与食品卫生教研室，黑龙江哈尔滨 150081；2. 哈尔滨市第一医院，黑龙江哈尔滨 150001 )

收稿日期:2013-03-06 修回日期:2013-05-27 出版日期:2013-09-30 发布日期:2013-09-30
通讯作者: 吴坤，E-mail：wukun_15000@163.com
作者简介:董瑞华 (1988- )，女，山东省青岛市人，硕士研究生，研究方向：肿瘤的化学预防。
基金资助:
国家自然科学基金资助项目 (81172651

Effects of acid sphingomyelinase on endoplasmic reticulum stress in VES- induced apoptosis in human gastric cancer cells

DONG Rui-hua¹，ZHAO Wei²，WANG Li-ming²，ZHANG Xu-guang¹，WU Kun¹，*

(1. Department of Nutrition and Food Hygiene, Harbin Medical University, Harbin 150081; 2. The First Hospital of Harbin, Harbin 150001, Heilongjiang, China)

Received:2013-03-06 Revised:2013-05-27 Online:2013-09-30 Published:2013-09-30
Contact: WU Kun，E-mail：wukun_15000@163.com

摘要/Abstract

摘要：

目的：探讨酸性神经磷脂酶 (acid sphingomyelinase，ASMase)在维生素E琥珀酸酯 (vitamin E succinate，VES)诱导胃癌细胞凋亡过程中对内质网应激的影响。方法：常规体外培养人低分化胃癌SGC-7901细胞，将细胞分为ASMase抑制剂地昔帕明 (desipramine，DES)处理组 (12.5 μmol/L DES作用2 h)，对照组 (含2％胎牛血清的RPMI-1640培养液)，VES+DES组 (用12.5 μmol/L DES抑制ASMase活性2 h后加20 μg/mL VES)以及VES处理组 (20 μg/mL)。以ASMase活性试剂盒检测0、1.5、3、6 h时VES+DES组及VES处理组细胞中ASMase活性。MTT法分别检测12、24和48 h时各组细胞的增殖情况。再在处理细胞24 h后，采用倒置显微镜观察各组细胞形态；流式细胞仪检测细胞凋亡率；Western blot法检测细胞中内质网应激相关蛋白GRP78、GRP94、Perk、p-Perk、Chop和Caspase-4的表达。结果：VES (20 μg/mL)处理能够诱导细胞中ASMase活性增加，在3 h时细胞中ASMase活性达到最高，随后逐渐降低；VES+DES组细胞中ASMase活性始终维持在较低的水平。与对照组相比，DES组在各项检测中均无明显变化，而VES组与VES+DES组在处理细胞12、24与48 h后，细胞的增殖受到明显抑制 (P均<0.05)，并呈时间-效应趋势，且VES+DES组细胞不同时间点的相对增殖率均明显高于VES组 (P均<0.05)；细胞形态学显示20 μg/mL VES处理细胞后，镜下死细胞明显增多，而VES+DES组死亡细胞数量较VES组减少；流式细胞术检测凋亡率显示VES处理组为 39.21%±1.90%，明显高于空白对照组(3.91%±1.68%)与DES组(4.07%±1.39%) (P均<0.05)，VES+DES组 (19.47%±4.46%)较VES组明显降低 (P<0.05)；Western blot结果显示VES+DES组细胞中内质网应激相关蛋白GRP78、GRP94、p-Perk、Chop以及c-Caspase-4表达水平较单纯VES处理组降低 (P均<0.05)。结论：ASMase参与VES通过内质网应激诱导人胃癌SGC-7901细胞凋亡的过程，起促进凋亡的作用。

关键词: 酸性神经磷脂酶, 维生素E琥珀酸酯, 内质网应激, 凋亡

Abstract:

OBJECTIVE: To study the effect of acid sphingomyelinase (ASMase) on endoplasmic reticulum stress (ERS) in vitamin E succinate (VES)-induced apoptosis in human gastric cancer cells. METHODS： SGC-7901 human gastric cancer cells were cultured in vitro. The cells were divided into four groups: desipramine (DES) group (cells were treated at 12.5 μmol/L for 2 h)，control group (containing 2% FBS RPMI-1640 culture medium)，VES+ DES group (cells were treated with DES at 12.5 μmol/L for 2 h，then with VES at 20 μg/ml) and VES group (20 μg/ml). ASMase activity detection kit was used to assess the ASMase activity of VES and VES + DES groups at 0，1.5，3 and 6 h. MTT assay was used to measure the growth inhibition at 12，24，48 h. Cell morphology was examined by the inverted microscope after treated for 24 h，and flow cytometry was used to evaluate the apoptosis，then we measured the ERS-related protein expressions of GRP78，GRP94，Perk，p-Perk，Chopand Caspase-4 by Western blot. RESULTS： ASMase activity continuously increased for VES (20 μg/mL) treatment，peaked at 3 h，then gradually decreased. ASMase activity of VES+DES group remained at a low level. There was no significant change in control group and DES group throughout the tests. Cell proliferation in VES group and VES+DES group was obviously inhibited after treatment for 12，24 and 48 h (all P<0.05) and showed a time-effect trend. Cell relative growth rates at different time points in VES+DES group were significantly higher than those in VES group(all P<0.05). The number of dead cells obviously increased in VES group，and the number of dead cells in VES+DES group decreased compared with VES group. Apoptosis rate evaluated by flow cytometry revealed that VES group (39.21%±1.90%) was significantly higher than the control group(3.91%±1.68%) and the DES group(4.07%±1.39%)(P<0.05)，VES+DES group (19.47%±4.46%) was significantly lower than the VES group(P<0.05). ERS-related protein GRP78，GRP94，p-Perk，Chop and c-Caspase-4 expression levels in VES+DES group were lower than those in VES group (all P<0.05). CONCLUSION：VES may induce apoptosis of human gastric cancer cell SGC-7901 cells though ERS by ASMase regulation pathways.

Key words: acid sphingomyelinase, vitamin E succinate, endoplasmic reticulum stress, apoptosis

董瑞华,赵巍,王立明,张旭光,吴坤. 酸性神经磷脂酶在维生素E琥珀酸酯诱导胃癌细胞凋亡过程中对内质网应激的影响[J]. 癌变·畸变·突变, doi: 10.3969/j.issn.1004-616x.2013.05.001.

DONG Rui-hua，ZHAO Wei，WANG Li-ming，ZHANG Xu-guang，WU Kun. Effects of acid sphingomyelinase on endoplasmic reticulum stress in VES- induced apoptosis in human gastric cancer cells[J]. Carcinogenesis, Teratogenesis & Mutagenesis, doi: 10.3969/j.issn.1004-616x.2013.05.001.

参考文献

[1] 贾莉，王彦军，张旭光，等. 维生素E琥珀酸酯通过氧化应激诱导胃癌SGC-7901细胞凋亡的研究[J]. 癌变·畸变·突变，2010，22 (3): 186-190.

[2] 黄晓莉，赵艳，张志宏，等. 维生素E琥珀酸酯通过内质网应激诱导胃癌SGC-7901细胞凋亡的研究[J]. 癌变·畸变·突变，2012，24 (3): 195-198.

[3] Zhang X，Peng X，Yu W，et al. Alpha-tocopheryl succinate enhances doxorubicin-induced apoptosis in human gastric cancer cells via promotion of doxorubicin influx and suppression of doxorubicin efflux[J]. Cancer Lett，2011，307 (2): 174-181.

[4] Park MA，Zhang G，Martin AP，et al. Vorinostat and sorafenib increase ER stress，autophagy and apoptosis via ceramide-dependent CD95 and PERK activation[J]. Cancer Biol Ther， 2008，7 (10): 1648-1662.

[5] Li J，Yu W，Tiwary R，et al. Alpha-TEA-induced death receptor dependent apoptosis involves activation of acid sphingomyelinase and elevated ceramide-enriched cell surface membranes[J]. Cancer Cell Int，2010，10 (1): 40.

[6] Erdreich-Epstein A，Tran LB，Cox OT，et al. Endothelial apoptosis induced by inhibition of integrins alphavbeta3 and alphavbeta5 involves ceramide metabolic pathways[J]. Blood， 2005，105 (11): 4353-4361.

[7] Perrotta C，De Palma C，Falcone S，et al. Nitric oxide，ceramide and sphingomyelinase-coupled receptors: a tale of enzymes and messengers coordinating cell death，survival and differentiation [J]. Life Sci，2005，77 (14): 1732-1739.

[8] Zhu Q，Lin L，Cheng Q，et al. The role of acid sphingomyelinase and caspase 5 in hypoxia-induced HuR cleavage and subsequent apoptosis in hepatocytes[J]. Biochim Biophys Acta，2012， 1821 (12):1453-1461.

[9] Barth BM，Gustafson SJ，Hankins JL，et al. Ceramide kinase regulates TNF alpha-stimulated NADPH oxidase activity and eicosanoid biosynthesis in neuroblastoma cells[J]. Cell Signal， 2012，24 (6): 1126-1133.

[10] Lin WC，Chuang YC，Chang YS，et al. Endoplasmic reticulum stress stimulates p53 expression through NF-kappa B activation[J]. PLoS One，2012，7 (7): 39120.

[11] de Ridder G，Ray R，Misra UK，et al. Modulation of the unfolded protein response by GRP78 in prostate cancer[J]. Methods Enzymol，2011，489 (2): 245-257.

[12] Kuang E，Wan Q，Li X，et al. ER Ca2+ depletion triggers apoptotic signals for endoplasmic reticulum (ER) overload response induced by overexpressed reticulon 3 (RTN3/HAP) [J]. J Cell Physiol，2005，204 (2): 549-559.

[13] Gentile CL，Frye M，Pagliassotti MJ. Endoplasmic reticulum stress and the unfolded protein response in nonalcoholic fatty liver disease[J]. Antioxid Redox Signal，2011，15 (2): 505-521.

[14] Rojas-Rivera D，Armisén R，Colombo A，et al. TMBIM3/GRINA is a novel unfolded protein response (UPR) target gene that controls apoptosis through the modulation of ER calcium homeostasis[J]. Cell Death Differ，2012，19 (6): 1013-1026.

[15] Saito A，Ochiai K，Kondo S，et al. Endoplasmic reticulum stress response mediated by the PERK-eIF2 (alpha)-ATF4 pathway is involved in osteoblast differentiation induced by BMP2[J]. J Biol Chem，2011，286 (6): 4809-4818.

[16] Obeng EA，Boise LH. Caspase-12 and caspase-4 are not required for caspase-dependent endoplasmic reticulum stress-induced apoptosis[J]. J Biol Chem，2005，280 (33): 29578-29587.

[17] Schroder M. Endoplasmic reticulum stress responses[J]. Cell Mol Life Sci，2008，65 (6): 862-894.

[18] Oshitari T，Hata N，Yamamoto S. Endoplasmic reticulum stress and diabetic retinopathy[J]. Vasc Health Risk Manag，2008，4 (1): 115-122.

[19] Huang X，Li L，Zhang L，et al. Crosstalk between endoplasmic reticulum stress and oxidative stress in apoptosis induced by alpha-tocopheryl succinate in human gastric carcinoma cells[J]. Br J Nutr，2012，7 (1): 1-9.

酸性神经磷脂酶在维生素E琥珀酸酯诱导胃癌细胞凋亡过程中对内质网应激的影响

Effects of acid sphingomyelinase on endoplasmic reticulum stress in VES- induced apoptosis in human gastric cancer cells

PDF

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 10

编辑推荐

Metrics

本文评价

[1]	郭茜晨, 张婷婷, 王润新, 赵法明, 盛夏. 内质网应激反应IRE1α信号通路与细胞衰老在癌症中的作用及关系的研究进展[J]. 癌变·畸变·突变, 2021, 33(1): 72-76.
[2]	黄晓莉, 吴坤, 赵莎莎. 淬灭胃癌SGC-7901细胞内活性氧对维生素E琥珀酸脂诱导内质网应激的影响[J]. 癌变·畸变·突变, 2018, 30(4): 270-274.
[3]	张超, 白剑英, 王东新, 闫丹丹, 王盼, 刘艳飞. 甲醛对人肝癌细胞株HepG2内质网应激相关Bip和CANX mRNA及蛋白表达水平的影响[J]. 癌变·畸变·突变, 2017, 29(5): 340-345,351.
[4]	黄晓莉, 吴坤, 赵莎莎. 维生素E琥珀酸酯诱导人胃癌细胞发生未折叠蛋白反应的研究[J]. 癌变·畸变·突变, 2017, 29(4): 256-259,265.
[5]	张艳, 白剑英, 雷佩玉, 王幼萍, 赵春燕, 梁瑞峰. 亚硫酸钠对HL-7702细胞极低密度脂蛋白组装分泌和内质网应激的影响[J]. 癌变·畸变·突变, 2016, 28(3): 169-173,178.
[6]	宋华翠, 侯丽颖, 綦峥, 吴坤. 维生素E琥珀酸酯处理人胃癌细胞过程中自噬与内质网应激的交互作用[J]. 癌变·畸变·突变, 2015, 27(3): 161-167.
[7]	曹文波,莫赛军,柯少瑞,张宏波,杨胜利. 钙结合蛋白S100A8和S100A9在慢性炎症相关肿瘤发生中的调控作用[J]. 癌变·畸变·突变, 2013, 25(5): 403-406.
[8]	侯丽颖,孙燕佩,张旭光,赵栋,吴坤. 维生素E琥珀酸酯通过Akt/mTOR信号通路诱导人胃癌细胞SGC-7901发生保护性自噬[J]. 癌变·畸变·突变, 2013, 25(5): 343-347.
[9]	黄晓莉,赵艳,张志宏,张旭光,王晓琳,贾莉,吴坤. 维生素E琥珀酸酯通过内质网应激诱导胃癌SGC-7901细胞凋亡的研究[J]. , 2012, 24(3): 195-198.
[10]	王晓琳/黄晓莉/赵艳/张志宏/张旭光/吕和/吴坤*. 内质网应激在芹菜素诱导胃癌细胞凋亡中的作用[J]. , 2012, 24(2): 115-120.