摘要：目的探讨中国流行株HIV-1gag与hIL-2／hIL-6共表达重组核酸疫苗质粒的免疫效果。方法以核酸疫苗质粒pIRES1neo为表达载体，构建重组核酸疫苗质粒pIRES1-gag、pIRES1-gag-hIL-2、pIRES1-gag-hIL-6，通过间接免疫荧光试验、Dot-ELISA检测gag／hIL-2／hIL-6基因的表达产物。另将此重组核酸疫苗质粒免疫Balb／c小鼠，进行淋巴细胞转化试验、CD4⁺、CD8⁺T淋巴细胞数量测定、细胞毒性T淋巴细胞（CTL）特异性杀伤作用检测及血清抗体检测，结果构建的重组质粒转染BHK细胞后可表达目的基因，免疫小鼠后可有效地刺激淋巴细胞增殖、诱导特异性CTL反应，当和hIL-2／hIL-6共表达时免疫效果更加显著。讨论与Gag蛋白共表达的hIL-2／hIL-6能够进一步增强免疫鼠的细胞免疫与体液免疫水平，构建的重组质粒为HIV-1DNA疫苗的研究提供了一定实验基础。

分类号：R392.7文献标识码：A

文章编号：1000-8861（2001）01-0005-04

Construction of DNA vaccine with HIV-1 gag and hIL-2／hIL-6 and its immunological study

QIN Yun-long（Department of Microbiology, Norman Bethune University of Medical Sciences，Changchun 130021，China）

JIN Ning-yi（Viral Laboratory，the Quartermaster University of PLA，Changchun 130062，China）

LUO Kun（Department of Microbiology, Norman Bethune University of Medical Sciences，Changchun 130021，China）

LIFan（Department of Microbiology, Norman Bethune University of Medical Sciences，Changchun 130021，China）

WANG Hong-wei（Viral Laboratory，the Quartermaster University of PLA，Changchun 130062，China）

WANG Li-xin（Viral Laboratory，the Quartermaster University of PLA，Changchun 130062，China）

ZHANG Yong-sheng（Department of Microbiology, Norman Bethune University of Medical Sciences，Changchun 130021，China）

YIN Zhen（Viral Laboratory，the Quartermaster University of PLA，Changchun 130062，China）

Abstract：ObjectiveTo study the immunological effects of recombinant HIV-1 DNA vaccine plasmids in mice.MethodsTwo DNA vaccine recombinant plasmids pIRES1-gag-hIL-2，pIRES1-gag-hIL-6 were constructed by inserting the HIV-1 gag and hIL-2／hIL-6 gene into a DNA vaccine expression vector（pIRES1neo）． The expressed product of gag／hIL-2／hIL-6 gene was examined by Dot-ELISA and indirect immunofluorescent assay. Immune response of Balb／c mice immunized by the two DNA vaccine recombinant plasmids was examined by lymphocyte transformation test（LTT），cytotoxic T lymphocyte specific killer test，quantity change of T lymphocyte (CD4⁺， CD8⁺) and dynamics of specific serum IgG antibody.ResultsHigh effective expression was found in BHK cells transfected with pIRES1-gag-hIL-2，pIRES1-gag-hIL-6. Recombinant plasmids could effectively induce the proliferation of effector lymphocytes and the generation of specific lytic activity in vitro. The effects were more significant when co-expressed with human interleukins gene. ConclusionCytokine hIL-2／hIL-6 can enhance both humoral and cellular immunity in immunized mice. Constructed recombinant plasmids might provide a basis for the development of HIV-1 DNA vaccine.

Keywords：HIV-1；Gag protein；hIL-2／hIL-6基金项目：国家自然科学基金（3977066）和国家杰出青年基金（39770661）资助项目

作者简介：秦云龙（1972－），男，黑龙江双鸭山市人，助教，硕士，主要从事分子病毒学的研究。Tel：（0431）7981365；E-mail：QinYL72＠263.net参考文献：

［1］金宁一.HIV基因结构及其疫苗研究［J］. 中国病毒学， 1997，12（4）：285－294.

［2］Hanke T， Neumann VC， Blanchard TJ， et al. Effective induction of HIV-specificCTL by multiepitope using gene gun in a combined vaccination regime ［J］. Vaccine， 1999，17(6)：589－596.

［3］Kim JJ， Tsai A， Nottingham LK，et al. Intracellular adhesion molecule-1 modulates beta-chemokines and directly co-stimulates T cells in vivo ［J］. J Clin Invest， 1999，103(6)：869－877.

［4］Ayyavoo V， Nagashunmugam T，Phung MT， et al. Construction of attenuated HIV-1 accessory gene immunization cassettes ［J］. Vaccine，1999，16(19)：1872－1 879.

［5］Bagarazzi ML， Boyer JD， Ugen KE， et al. Safety and immunogenicity of HIV-1 DNA constructs in chimpanzees ［J］. Vaccine，1998，16(19)：1 836－1841.

［6］Peet NM， Mckeating JA， Souza JB， et al. The effect of low-profile serine substitutions in the V3 loop of HIV-1 gp120 IIIB／LAT on the immunogenicity of the envelope protein ［J］. Virology，1998，251(1)：59－70.

［7］Ugen KE， NylanlSB， Boyer JD， et al. DNA vaccination with HIV-1 expressing constructs elicits immune responses in human ［J］. Vaccine，1998，16(19)：1 818－1 821.

［8］Winchell JM， Routray S， Betts PW， et al. Mucosal and systemic antibody responses to a C4／V3 construct following DNA vaccination of rabbits via the Peyer’s patch ［J］. J Infect Dis，1998，178(3)：850－853.

收稿日期：2000年9月20日

修稿日期：2000年10月25日

出版日期：2001年1月15日