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

doi:10.3969/j.issn.1004-616x.2017.05.004

癌变·畸变·突变 ›› 2017, Vol. 29 ›› Issue (5): 340-345,351.doi: 10.3969/j.issn.1004-616x.2017.05.004

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

张超¹, 白剑英¹, 王东新², 闫丹丹¹, 王盼¹, 刘艳飞¹

1. 山西医科大学公共卫生学院环境卫生教研室, 山西太原 030001;
2. 山西医科大学公共卫生学院卫生检验教研室, 山西太原 030001

收稿日期:2017-04-10 修回日期:2017-06-08 出版日期:2017-09-30 发布日期:2017-09-30
通讯作者: 白剑英,E-mail:jybai66@aliyun.com E-mail:jybai66@aliyun.com
作者简介:张超,E-mail:13261519824@163.com。
基金资助:
山西省回国留学人员科研资助项目（2014-034）；山西省留学回国人员科技活动择优资助项目（晋人社厅函[2014]376号）

Effects of formaldehyde on expression of endoplasmic reticulum stress-related Bip and CANX mRNA and proteins of HepG2 cell line

ZHANG Chao¹, BAI Jianying¹, WANG Dongxin², YAN Dandan¹, WANG Pan¹, LIU Yanfei¹

1. Department of Environmental Hygiene, School of Public Health, Shanxi Medical University, Taiyuan 030001;
2. Department of Health Laboratory, School of Public Health, Shanxi Medical University, Taiyuan 030001, Shanxi, China

Received:2017-04-10 Revised:2017-06-08 Online:2017-09-30 Published:2017-09-30

摘要/Abstract

摘要： 目的：研究甲醛（FA）染毒是否会诱导人肝癌细胞株HepG2内质网应激相关基因Bip和CANX mRNA和蛋白表达水平的改变，引起内质网应激反应。方法：用Hoechst 33258进行核染色，以观察不同浓度（0、0.02、0.1、0.5、2.5、12.5 mmol/L）FA分别染毒肝细胞株24和48 h 后的细胞核形态学变化。再分别用0.004、0.02、0.1 mmol/L的FA处理人肝癌细胞株HepG2细胞24 h和48 h后，采用荧光定量PCR（qPCR）和Western blot检测内质网应激相关基因免疫球蛋白重链结合蛋白（Bip）及钙连蛋白（CANX） mRNA和蛋白的表达水平。结果：Hoechst 33258染色结果表明，染毒24和48 h后，0.5、2.5、12.5 mmol/L FA 组出现大量核呈固缩状态或颗粒状荧光死亡细胞，阴性对照组和0.1 mmol/L及以下剂量组细胞核荧光染色较浅，细胞核未固缩，故后续试验采用0.1 mmol/L及以下剂量。经0.004、0.02和0.1 mmol/L FA处理细胞24和48 h后，qPCR检测结果显示，甲醛各剂量染毒组Bip和CANX的mRNA表达量与阴性对照组比较，差异均无统计学意义（P>0.05）。Western blot结果表明，染毒24 h，不同剂量甲醛处理组与阴性对照组相比，Bip和CANX蛋白表达量均无显著差异（P>0.05）；染毒48 h，与阴性对照组相比，各FA处理组Bip蛋白表达量升高，0.02和0.1 mmol/L FA处理组CANX蛋白表达明显升高，差异均有统计学意义（P < 0.05）。结论：甲醛染毒可诱导HepG2细胞内质网应激相关蛋白Bip和CANX表达水平升高，引起肝细胞发生内质网应激反应。

关键词: 甲醛, 内质网应激, 肝细胞, 免疫球蛋白重链结合蛋白, 钙连蛋白

Abstract: OBJECTIVE: To investigate whether formaldehyde can induce endoplasmic reticulum stress in hepatocytes. METHODS: Hoechst 33258 was used for nuclear staining to observe morphological changes of hepatocytes in different formaldehyde concentration groups. The human hepatocellular carcinoma cell line HepG2 was treated with formaldehyde (FA) solution at 0.004 mmol/L,0.02 mmol/L and 0.1 mmol/L for 24 h and 48 h respectively. Expression of Bip and CANX mRNA in stress-related genes was detected by fluorescence quantitative PCR (qPCR). Western blot was used to detect the expression of stress-related immunoglobulin heavy chain binding protein (Bip) and connexin (CANX) in endoplasmic reticulum. RESULTS: Cells in the negative control group and the low dose group were significantly higher with Hoechst 33258 staining than in the control group (P < 0.05). Results of qPCR show that expression of Bip and CANX mRNA in each dose group was not significantly different from that in the negative control group. Results of Western blot show that there was no significant difference in the expression of Bip and CANX proteins among the different doses of FA group and the negative control group (P>0.05). Compared with the negative control group (P < 0.05), expression of Bip protein was increased in 0.02 mmol/L and 0.1 mmol/L FA group,the difference was statistically significant (P < 0.05). CONCLUSION: Formaldehyde exposure may induce endoplasmic reticulum stress of hepatocytes.

Key words: formaldehyde, endoplasmic reticulum stress, hepatocytes, Bip, CANX

中图分类号:

R994.6

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.

ZHANG Chao, BAI Jianying, WANG Dongxin, YAN Dandan, WANG Pan, LIU Yanfei. Effects of formaldehyde on expression of endoplasmic reticulum stress-related Bip and CANX mRNA and proteins of HepG2 cell line[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2017, 29(5): 340-345,351.

参考文献

[1] 陈霜玲,李玉光,付慧群,等. 甲醛遗体防腐剂改良研究[J]. 中国临床解剖学杂志,2016,34(2):227-231.
[2] QUE Z,FURUNO T. Formaldehyde emission from wood products:relationship between the values by the Chamber method and those by the Desiccator test[J]. Wood Sci Technol 2007,41(3):267-279.
[3] 王小玲. 甲醛对小鼠中枢神经系统毒性的研究[D]. 太原:山西医科大学,2008,10(8):67-72.
[4] KIM J Y,JEONG M S,PARK K Y,et al. Aggravation of atopic dermatitis-like symptoms by consecutive low concen-tration of formaldehyde exposure in NC/Nga mice[J]. Exp Dermato,2013,22(3):219-221.
[5] 李亚琳. 液态甲醛对雄性小鼠生殖毒性的研究[J]. 动物医学进展,2012,33(12):71-75.
[6] SWENBERG J A,MOELLER B C,LU K,et al. Formaldehyde carcinogenicity research:30 years and counting for mode of action,epidemiology,and cancer risk assessment[J]. Toxicol Pathol,2013,41(2):181-189.
[7] CIKMAZ S,KUTOGLU T,KANTER M,et al. Effect of formaldehyde inhalation on rat livers:a light and electron microscopic study[J]. Toxicol Ind Health,2010,26(2):113-118.
[8] 张杰,武晶晶,赵飞飞,等. 不同温度和不同放置时间对总RNA提取浓度和纯度的影响[J]. 实验技术与管理,2013,10(11):79-82.
[9] 梁瑞峰,原福胜,白剑英,等. 甲醛吸入对小鼠的氧化损伤作用[J]. 山西医科大学学报,2005,36(1):58-60.
[10] CANCER WHOIAFRO. IARC monographs on the evaluation of carcinogenic risks to humans:Formaldehyde,2-butoxyethanol and 1-tert-butoxypropan-2-ol[J]. Med Hypothese,2004,33(4):343-358.
[11] SCHMID O,SPEIT G. Genotoxic effects induced by formaldehyde in human blood and implications for the interpretation of biomonitoring studies[J]. Mutagenesis,2007,22(1):69-75.
[12] SABNIS R W. Hoechst 33258[J]. Circ Res,2015,167(7):239-242.
[13] REIS P A B,SOARES-RAMOS J R L,FONTES E P B,et al. Endoplasmic reticulum stress response[S]. In Tech,2011:249-253.
[14] HETZ C. The unfolded protein response:controlling cell fate decisions under ER stress and beyond[J]. Nat Rev Mol Cell Biol,2012,13(2):89-94.
[15] YAN M,SHEN J,PERSON M D,et al. Endoplasmic reticulum stress and unfolded protein response in ATM-deficient thymocytes and thymic lymphoma cells are attributable to oxidative stress[J]. Neoplasia,2008,10(2):160-167.
[16] 王洪岩,刘晓珺,杜雅菊,等. 内质网应激与肝脏疾病研究进展[J]. 世界华人消化杂志,2012,6:451-459.
[17] ZHOU A X,WANG X,LIN C S,et al. C/EBP-homologous protein (CHOP) in vascular smooth muscle cells regulates their proliferation in aortic explants and atherosclerotic lesions[J]. Circ Res,2015,116(11):1736-1740.
[18] SUYAMA K,WATANABE M,SAKABE K,et al. Overexpression of GRP78 protects glial cells from endoplasmic reticulum stress[J]. Neurosci Lett,2011,504(3):271-276.
[19] XU D,PEREZ R E,REZAIEKHALIGH M H,et al. Knockdown of ERp57 increases BiP/GRP78 induction and protects against hyperoxia and tunicamycin-induced apoptosis[J]. Am J Physiol-lung C,2009,297(1):144-148.
[20] GUÉRIN R,TURCOTTE C,LEROUX A,et al. The epigenetic calnexin-independent state is induced in response to environmental changes[J]. Fems Yeast Res,2009,9(8):1250-1259.
[21] RUTKOWSKI D T,WU J,BACK S H,et al. UPR pathways combine to prevent hepatic steatosis caused by ER stress-mediated suppression of transcriptional master regulators[J]. Dev Cell,2008,15(6):824-828.
[22] 高虹,张向颖,岳莉英,等. 内质网应激调控糖原合成酶激酶3β活性在小鼠急性肝衰竭致病中的作用[J]. 中华医学杂志,2016,96(34):2734-2738.

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

Effects of formaldehyde on expression of endoplasmic reticulum stress-related Bip and CANX mRNA and proteins of HepG2 cell line

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