Mace K, Seif I, Anjard C, De Maeyer-Guignard J, Dodon M D, Gazzolo L, De Maeyer E
UMR 30 CNRS-UCBL, Faculté de Médecine Alexis Carrel, Lyon, France.
J Immunol. 1991 Nov 15;147(10):3553-9.
Cells of the monocyte lineage act as a major reservoir for HIV, and ways of enhancing the resistance of mononuclear phagocytes to HIV replication would be useful for delaying the onset of AIDS in infected individuals. Seif et al. (J. Virol. 65:664, 1991) have recently shown the possibility of obtaining stable antiviral expression (SAVE), directed against three nonretroviral RNA viruses, and normal cell viability in a significant percentage of murine BALB/c 3T3 cells transformed with an IFN-beta expression plasmid under the control of the 0.6-kb XhoII-NruI promoter region of the murine H-2Kb MHC gene. In the present paper, we show that it is possible to establish SAVE in human promonocytic cells. Cells of the human promonocytic U937 line were stably transfected with a human IFN-beta expression plasmid carrying the neo- and human IFN-beta-coding sequences under the control of the H-2Kb promoter fragment previously used in murine cells. After selection with G418, two transformed clones were isolated that released small amounts of human IFN-beta into the culture medium, without affecting the expression of CD4 and leucocyte function-associated Ag-1 differentiation Ag. The presence of construct-derived IFN-beta mRNA was demonstrated by polymerase chain reaction amplification of cDNA, and the level of 2-5A synthetase, one of the major IFN-induced antiviral proteins, was shown to be constitutively increased. These clones were less permissive for HIV-1 than control clones transformed with the neo gene only. The antiviral state could be modulated by anti-IFN-beta antibodies, in that the continuous presence of antibodies in the culture medium abolished the enhanced resistance to HIV-1 replication, whereas the withdrawal of the antiserum restored the antiviral state, indicating that it did indeed result from the constitutive synthesis of human IFN-beta. These results demonstrate the possibility of restricting HIV-1 replication in human promonocytic cells by establishing SAVE. Further exploration of this method as a possible approach to somatic cell gene therapy of HIV infection appears worthwhile.
单核细胞系的细胞是HIV的主要储存库，增强单核吞噬细胞对HIV复制的抵抗力的方法对于延缓感染个体中艾滋病的发病将是有用的。Seif等人（《病毒学杂志》65:664，1991）最近表明，在用鼠H-2Kb MHC基因的0.6-kb XhoII-NruI启动子区域控制下的IFN-β表达质粒转化的相当比例的鼠BALB/c 3T3细胞中，有可能获得针对三种非逆转录RNA病毒的稳定抗病毒表达（SAVE）以及正常的细胞活力。在本文中，我们表明在人原单核细胞中建立SAVE是可能的。用人IFN-β表达质粒稳定转染人原单核细胞U937系细胞，该质粒携带在鼠细胞中先前使用的H-2Kb启动子片段控制下的新霉素和人IFN-β编码序列。用G418筛选后，分离出两个转化克隆，它们向培养基中释放少量人IFN-β，而不影响CD4和白细胞功能相关抗原-1分化抗原的表达。通过cDNA的聚合酶链反应扩增证明了构建体衍生的IFN-β mRNA的存在，并且显示主要的IFN诱导抗病毒蛋白之一的2-5A合成酶的水平持续增加。这些克隆对HIV-1的易感性低于仅用新霉素基因转化的对照克隆。抗病毒状态可以通过抗IFN-β抗体进行调节，因为培养基中抗体的持续存在消除了对HIV-1复制的增强抵抗力，而抗血清的撤除恢复了抗病毒状态，表明它确实是由人IFN-β的组成性合成导致的。这些结果证明了通过建立SAVE来限制人原单核细胞中HIV-1复制的可能性。进一步探索这种方法作为HIV感染体细胞基因治疗的一种可能途径似乎是值得的。
