[1]程剑△,沈晓洁,刘艳.携带人IL10基因的腺病毒载体的构建及其对大鼠肝细胞损伤的保护作用*[J].郑州大学学报(医学版),2013,(04):477.
 CHENG Jian,SHEN Xiaojie,LIU Yan.Construction of adenovirus vector carrying human interleukin10 gene and its protection effects on rat hepatocytes during liver injury[J].JOURNAL OF ZHENGZHOU UNIVERSITY(MEDICAL SCIENCES),2013,(04):477.
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携带人IL10基因的腺病毒载体的构建及其对大鼠肝细胞损伤的保护作用*
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《郑州大学学报(医学版)》[ISSN:1671-6825/CN:41-1340/R]

卷:
期数:
2013年04期
页码:
477
栏目:
论著
出版日期:
2013-07-20

文章信息/Info

Title:
Construction of adenovirus vector carrying human interleukin10 gene and its protection effects on rat hepatocytes during liver injury
作者:
程剑沈晓洁刘艳
无锡卫生高等职业技术学校病理学教研室 无锡 214028
Author(s):
CHENG Jian SHEN Xiaojie LIU Yan
Department of Pathology, Wuxi Higher Health Vocational Technology School, Wuxi 214028
关键词:
白细胞介素10腺病毒载体大鼠肝细胞保护作用
Keywords:
interleukin10 adenovirus vector rat hepatocyte protection effect
分类号:
R657.3;R575
摘要:
目的:构建携带人白细胞介素10(hIL10)基因的腺病毒载体,并验证其对大鼠肝细胞损伤的保护作用。方法:以pORFhIL10质粒为模板,PCR扩增hIL10,插入腺病毒质粒pDC315,构建pDC315hIL10。将pDC315hIL10质粒与腺病毒包装质粒共转染293细胞,产生复制缺陷型重组腺病毒Ad5hIL10;感染大鼠肝细胞,免疫学方法鉴定目的基因的表达;建立CCL4诱导的大鼠肝细胞损伤模型,观察Ad5hIL10不同感染强度(MOI)预处理对损伤肝细胞存活率的影响。结果:经PCR扩增,证明已正确构建重组腺病毒Ad5hIL10。大鼠肝细胞感染Ad5hIL10后,随着病毒MOI的升高,hIL10表达上升(F=52.667,P<0.001)。不同感染强度(MOI=1、5、10和50)Ad5hIL10预处理组大鼠肝细胞存活率较损伤对照组升高,差异有统计学意义(F=23.645、22.533、30.697、35.559,P均<0.001)。结论:成功构建了重组腺病毒Ad5hIL10,其能提高肝细胞的存活能力,对肝细胞有保护作用。
Abstract:
Aim: To construct the adenovirus vector carrying human interleukin10 (hIL10) gene, and then identify the expression of hIL10 and detect its protection effects on rat hepatocytes during liver injury. Methods:The hIL10 gene was amplified by PCR from template plasmid pORFhIL10, then digested with endonuclease and inserted into pDC315 to construct pDC315hIL10. The plasmid pDC315hIL10 and the adenovirus vector were cotransfected to 293 cells togenerate the replicationdeficient recombinant adenovirus Ad5hIL10. Ad5hIL10 was used to infect rat hepatocytes and the expression of hIL10 was identified by immunology method. Rat hepatocyte injury model was constructed, and the protection effects of Ad5hIL10 was investigated in rat model at different multiplicities of infection (MOI). Results: The recombi ant adenovirus Ad5hIL10 was recombined successfully. Rat hepatocytes expressed hIL10 after infection of Ad5hIL10. The expression level of hIL10 was increased along with the increase of MOI(F=52.667,P<0.001). Cell viability of rat hepatocytes in Ad5hIL10treated group(MOI=1,5,10 and 50) was higher than that in the control group,the difference has a statistical significance(F=23.645,22.533,30.697,35.559,P<0.001). Conclusion: The recombinant adenovirus Ad5hIL10 has been successfully constructed, which could improve hepatocytes viability and protect hepatocytes.

参考文献/References:

[1]Si ZZ, Li JQ, Qi HZ, et al. Recombinant adenovirus vector AdhIL10 protects grafts from cold ischemiareperfusion injury following orthotopic liver transplantation in rats [J]. Hepatobiliary Pancreat Dis Int, 2010, 9(2):144
[2]Donckier V, Loi P, Closset J, et al. Preconditioning of donors with interleukin10 reduces hepatic ischemiareperfusion injury after liver transplantation in pigs [J].Transplantation,2003,75(6):902
[3]Douglas DB, Beiting DP, Loftus JP, et al. Combinatorial effects of interleukin 10 and interleukin 4 determine the progression of hepatic inflammation following murine enteric parasitic infection [J]. Hepatology, 2010, 51(6):2162
[4]Papeleu P, Vanhaecke T, Henkens T, et al. Isolation of rat hepatocytes [J]. Methods Mol Biol, 2006,320:229
[5]Kiyota T, Ingraham KL, Swan RJ, et al. AAV serotype 2/1mediated gene delivery of antiinflammatory interleukin10 enhances neurogenesis and cognitive function in APP+PS1 mice [J]. Gene Ther, 2012,19(7):724
[6]Andreoli CG, Genro VK, Souza CA, et al. T helper (Th)1, Th2, and Th17 interleukin pathways in infertile patients with minimal/mild endometriosis [J]. Fertil Steril, 2011,95(8):2477
[7]Kosaka S, Tamauchi H, Terashima M, et al. IL10 controls Th2type cytokine production and eosinophil infiltration in a mouse model of allergic airway inflammation [J]. Immunobiology, 2011, 216(7):811
[8]Cui HD, Qi ZM, Yang LL, et al. Interleukin10 receptor expression and signalling were downregulated in CD4+ T cells of lupus nephritis patients [J]. Clin Exp Immunol, 2011, 165(2):163
[9]Bouaziz JD, Calbo S, MahoVaillant M, et al. IL10 produced by activated human B cells regulates CD4(+) Tcell activation in vitro [J]. Eur J Immunol, 2010, 40(10):2686
[10]ChávezSánchez L, ChávezRueda K, LegorretaHaquet MV, et al. The activation of CD14, TLR4, and TLR2 by mmLDL induces IL1β, IL6, and IL10 secretion in human monocytes and macrophages [J]. Lipids Health Dis, 2010,9:117

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备注/Memo

备注/Memo:
*江苏省卫生职业技术教育研究基金资助项目J201017 女,1967年10月生,硕士,副教授,研究方向:病毒与基因治疗,Email:251810058@qq.com
更新日期/Last Update: 2013-07-25