热休克蛋白70与汉坦病毒持续性感染乳大鼠大脑皮层星形胶质细胞

doi:10.16098/j.issn.0529-1356.2018.01.002

摘要/Abstract

摘要：

目的探讨热休克蛋白70（HSP70）过表达可能有助于汉坦病毒（HTNV）持续感染。方法在HTNV持续性感染过程中，通过Westen blotting及RT-PCR检测大脑皮层星形胶质细胞中HSP70的表达情况，并通过免疫荧光显微技术、AnnexinV 及RT-PCR检测细胞凋亡。结果与对照组相比，感染HTNV 76-118或 SEOV L99 的星形胶质细胞HSP70蛋白的表达增加，且在HTNV 76-118感染后3d或SEOV L99感染后2d HSP70蛋白表达达到峰值（n=15，P＜0.01，P＜0.05）；感染HTNV 76-118及SEOV L99上调星形胶质细胞HSP70基因的表达，且在HTNV 76-118感染后2~5d或SEOV L99感染后2~4d HSP70基因表达达到峰值（n=15，P＜0.01，P＜0.05）。HTNV76-118或SEOV L99感染过程中，星形胶质细胞无细胞病变效应，并且病毒复制未诱导细胞凋亡。结论 HTNV感染星形胶质细胞激活了HSP70，这种改变可能抑制细胞凋亡，促进病毒持续性感染。

关键词: 汉坦病毒, 大脑皮层, 星形胶质细胞, 热休克蛋白70, 免疫印迹法, 反转录-聚合酶链反应, 大鼠

Abstract:

Objective To define that heat shock protein 70（HSP70）overpression might contributed to the persistent infection of Hantaviruses （HTNV）. Methods The expression of HSP70 in cortical astrocytes was examined by Western blotting and RT-PCR during Hantaviruses persistent infection. The following apoptosis was detected by immunofluorescence microscopy, AnnexinV assays and RT-PCR assay. Results Compared to control group ,the expression of HSP70 protein in astrocytes infected with HTNV 76-118 or SEOV L99 increased, peak expression of HSP70 at day 3 following HTNV 76-118 infection or at day 2 following SEOV infection (n=15, P<0.01, P<0.05), and the expression of HSP70 gene in astrocytes infected with HTNV 76-118 or SEOV L99 up-regulated, peak expression of HSP70 at day 2-5 post HTNV76-118 infection and at day 2-4 post SEOV L99 infection (n=15, P<0.01, P<0.05). In the course of HTNV76-118 or SEOV L99 infection, any cytopathic effect was not shown in astrocytes and cell apoptosis was not induced by viral replication. Conclusion HSP70 induced by Hantaviruses infection might inhibit apoptosis and contribute to the viral persistent infection.

Key words: Hantavirus, Cerebral cortex, Astrocyte, Heat shock protein 70, Western blotting, RT-PCR, Rat

中图分类号:

Ｒ373. 3 ⁺ 2

贺帅杨守京. 热休克蛋白70与汉坦病毒持续性感染乳大鼠大脑皮层星形胶质细胞[J]. 解剖学报, 2018, 49(1): 7-13.

HE Shuai YANG Shou-jing. Heat shock protein 70 and Hantavirus persistent infection of cortical astrocytes from newborn rats[J]. Acta Anatomica Sinica, 2018, 49(1): 7-13.

参考文献

［1］Nichol ST, Spiropoulou CF, Morzunov S, et al. Genetic identification of a hantavirus associated with an outbreak of acute respiratory illness［J］.Science,1993,262(5135):914-917.［2］Linderholm M, Elgh F. Clinical characteristics of hantavirus infections on the Eurasian continent［J］. CurrTop Microbiol Immunol, 2001, 256:135-151.

［3］Pensiero MN, Sharefkin JB, Dieffenbach CW, et al. Hantaan virus infection of human endothelial cells［J］.J Virol, 1992,66(10):5929-5936.

［4］Temonen M, Vapalahti O, Holthofer H, et al. Susceptibility of human cells to Puumala virus infection［J］. J Gen Virol, 1993,74 (Pt 3):515-518.

［5］Zaki SR, Greer PW, Coffield LM, et al. Hantavirus pulmonary syndrome. Pathogenesis of an emerging infectious disease［J］. Am J Pathol, 1995,146(3):552-579.

［6］Liu JS, Kuo SR, Makhov AM, et al. Human Hsp70 and Hsp40 chaperone proteins facilitate human papillomavirus-11 E1 protein binding to the origin and stimulate cell-free DNA replication［J］. J Biol Chem,1998, 273(46):30704-30712.

［7］Wang RY, Stork J, Pogany J, et al. A temperature sensitive mutant of heat shock protein 70 reveals an essential role during the early steps of tombusvirus replication［J］. Virology, 2009, 394(1):28-38.

［8］Chen YJ, Chen YH, Chow LP, et al. Heat shock protein 72 is associated with the hepatitis C virus replicase complex and enhances viral RNA replication［J］. J Biol Chem, 2010,285(36):28183-28190.

［9］Livingston CM, DeLuca NA, Wilkinson DE, et al. Oligomerization of ICP4 and rearrangement of heat shock proteins may be important for herpes simplex virus type 1 prereplicative site formation［J］. J Virol, 2008, 82 (13): 6324-6336.

［10］Conti C, De Marco A, Mastromarino P, et al. Antiviral effect of hyperthermic treatment in rhinovirus infection［J］. Antimicrob Agents Chemother, 1999, 43(4):822-829.

［11］Pica F, Palamara AT, Rossi A, et al. Delta(12)-prostaglandin J(2) is a potent inhibitor of influenza A virus replication［J］. Antimicrob Agents Chemother, 2000, 44(1):200-204.

［12］Rozera C, Carattoli A, De Marco A, et al. Inhibition of HⅣ-1 replication by cyclopentenone prostaglandins in acutely infected human cells. Evidence for a transcriptional block［J］. J Clin Invest, 1996, 97(8):1795-1803.

［13］Kang JI, Park SH, Lee PW, et al. Apoptosis is induced by hantaviruses in cultured cells［J］. Virology,1999, 264(1):99-105.

［14］Li XD, Kukkonen S, Vapalahti O, et al. Tula hantavirus infection of Vero E6 cells induces apoptosis involving caspase 8 activation［J］. J Gen Virol, 2004, 85(Pt 11):3261-3268.

［15］Kurata T, Tsai TF, Bauer SP, et al. Immunofluorescence studies of disseminated Hantaan virus infection of suckling mice［J］. Infect Immun, 1983, 41(1):391-398.

［16］Akhmatova NK, Yusupova RS, Khaiboullina SF, et al. Lymphocyte apoptosis during hemorragic fever with renal syndrome［J］. Russ J Immunol, 2003, 8(1):37-46.

［17］Bouillet P, O’Reilly LA. CD95, BIM and T cell homeostasis［J］. Nat Rev Immunol, 2009, 9(7):514-519.

［18］McCarthy KD, de Vellis J. Preparation of separate astroglial and oligodendroglial cell cultures from rat cerebral tissue［J］. J Cell Biol, 1980, 85(3):890-902.

［19］Stoltz M, Klingstrom J. Alpha/beta interferon (IFN-alpha/beta)-independent induction of IFN-lambda1 (interleukin-29) in response to Hantaan virus infection［J］. J Virol, 2010, 84(18):9140-9148.

［20］Rang A, Gunther S, Will H. Effect of interferon alpha on hepatitis B virus replication and gene expression in transiently transfected human hepatoma cells［J］. J Hepatol,1999, 31(5):791-799.

［21］Shin OS, Yanagihara R, Song JW. Distinct innate immune responses in human macrophages and endothelial cells infected with shrew-borne hantaviruses［J］. Virology, 2012,434:43-49.

［22］Conti C, De Marco A, Mastromarino P, et al. Antiviral effect of hyperthermic treatment in rhinovirus infection［J］. Antimicrob Agents Chemother,1999, 43(4):822-829.

［23］Boatright KM, Salvesen GS. Mechanisms of caspase activation［J］. Curr Opin Cell Biol, 2003, 15 (6): 725-731.

［24］Stennicke HR, Jurgensmeier JM, Shin H,et al. Pro-caspase-3 is a major physiologic target of caspase-8［J］. J Biol Chem,1998, 273(42):27084-27090.

［25］Sch ?nrich G, Rang A, Lütteke N, et al. Hantavirus-0induced immunity in rodent reservoirs and humans［J］. Immunol Rev,2008, 225: 163-189.

［26］Vapalahti O, Mustonen J, Lundkvist ?, et al. Hantavirus infections in Europe［J］. Lancet Infect Dis, 2003,3(10): 653-661.

［27］Mackow ER, Gavrilovskaya IN.Hantavirus regulation of endothelial cell functions［J］. Thromb Haemost, 2009, 102(6): 1030-1041.

［28］Gao W,Wang XY, Liu W, et al. Study on human embryonic kidney 293 cell infected by hantavirus and its mechanism inducing apoptosis［J］. Chinese Journal of Nosocomiology, 2011,21(22): 4671-4673.(in Chinese)

高巍，王晓燕，刘伟，等.汉坦病毒感染人胚肾细胞诱导凋亡的研究［J］.中华医院感染学杂志, 2011,21(22): 4671-4673.

［29］Li XD, Kukkonen S, Vapalahti O, et al.Tula hantavirus infection of Vero E6 cells induces apoptosis involving caspase 8 activation［J］. J Gen Virol, 2004, 85(11):3261-3268.

［30］Gupta S, Braun M, Tischler ND, et al. Hantavirus-infection confers resistance to cytotoxic lymphocyte-mediated apoptosis ［J］. PLoS Pathog, 2013, 9(3): e1003272.

［31］Parcellier A, Gurbuxani S, Schmitt E, et al. Heat shock proteins, cellular chaperones that modulate mitochondrial cell death pathways［J］. Biochem Biophys Res Commun, 2003, 304(3):505-512.

［32］Jiang B, Liang P, Deng G, et al. Increased stability of Bcl-2 in HSP70-mediated protection against apoptosis induced by oxidative stress［J］. Cell Stress Chaperones, 2011, 16(2): 143-152.

［33］Chuquet J, Quilichini P, Nimchinsky EA, et al.Predominant enhancement of glucose uptake in astrocytes versus neurons during activation of the somatosensory cortex［J］. J Neurosci,2010, 30(45):15298-15303.

［34］Rouach N, Koulakoff A, Abudara V, et al. Astroglial metabolic networks sustain hippocampal synaptic transmission［J］. Science,2008, 322(5907):1551-1555.

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