T103A变异MxA蛋白抑制水泡性口膜炎病毒复制的体外研究

目的研究T103A变异MxA蛋白抑制水疱性口膜炎病毒（VSV）复制活性。方法将野生型、T103A变异MxA蛋白表达载体和对照质粒分别瞬时转染Wish细胞,24h后VSV感染细胞,48h后用MTr法检测各组细胞增殖;另取Wish细胞转染上述3种质粒,转染24h加入VSV感染细胞,24h后收集细胞采用RT-PCR检测VSVmRNA水平;Western blot检测各组MxA蛋白表达。结果野生型、T103A变异MxA蛋白均在Wish细胞有较好表达;MTT检测结果提示T103A变异组细胞增殖数显著低于野生型组（P〈0．01）;RT—PCR结果显示T103A变异组VSVmRNA水平显著高于野生型组（P〈0．01）,但与对照组比较差异无统计学意义（P〉0．05）。结论T103A变异MxA蛋白失去了抑制VSV复制活性。     

Adenovirus Susceptibility to Human Interferon During One-Step Replication

Susceptibility of adenovirus types 2, 7, and 12 to human interferon was measured in three human diploid cell strains during a single-cycle infection. Although the relative susceptibility of adenovirus to interferon varied in these cell strains, the final yield of each type in each cell strain decreased as the interferon dose increased. On the other hand, wide difference in interferon susceptibility of adenoviruses and vesicular stomatitis virus (VSV) was noted, as interferon doses above 100 units profoundly inhibited VSV but not the adenoviruses.     

Inhibition of influenza C viruses by human MxA protein

Human MxA protein was analyzed for its ability to inhibit the replication of different influenza C viruses. Three laboratory derivatives of viral strain C/Ann Arbor/1/50 were investigated, namely the parental wild-type virus C/AA-wt, the persistent variant C/AA-pi and the highly cytopathogenic variant C/AA-cyt. In addition, strain C/Paris/214/91 isolated from an influenza patient was used. Multiplication of all four viruses was suppressed in MxA-expressing Vero cells, as indicated by a decrease in viral RNA synthesis, viral protein synthesis, virion production and induction of a cytopathic effect. Inhibition correlated with the level of MxA expression. Furthermore, inhibition was independent of cell clone-specific differences in expression of virus receptors, as demonstrated by receptor reconstitution experiments. Thus, human MxA protein has antiviral activity against influenza C viruses.     

Inhibition of Dugbe nairovirus replication by human MxA protein

Sensitivity to the interferon-induced protein, MxA, has previously been demonstrated for viruses belonging to the Orthobunyavirus, Hantavirus and Phlebovirus genera of the Bunyaviridae family. We have extended these findings to a member of the fourth and remaining genus containing viruses that infect man and other animals, the nairovirus Dugbe virus (DUGV). Indirect immunofluorescence experiments using VA9 cells (Vero cells permanently transfected with MxA cDNA) revealed strongly reduced DUGV antigen expression, suggesting that MxA inhibited DUGV replication. Western and Northern blot analyses showed significantly lower DUGV nucleocapsid (N) protein expression and DUGV genomic RNA, respectively, in the presence of MxA. Viral titres were also reduced by more than two orders of magnitude in VA9 cells compared with control VN36 cells. This finding may have application to nairovirus therapeutics.     

Interferon Production by Lymphocytes in Human Milk

Despite their limited ability to synthesize immunoglobulins (only IgA and not IgM or IgG) lymphocytes of human colostrum and human breast milk can be stimulated by Newcastle disease virus to produce interferon in the same amount as do blood lymphocytes. The maximum interferon production by milk lymphocytes was found on the 4th and the 5th day postpartum.     

Pathogenesis of Human Parainfluenza Type 3 Virus Infection in Hamster Tracheal Organ Culture

Hamster tracheal organ culture was employed as a model for the study of the pathogenesis of human parainfluenza type 3 virus infection. It clearly supported replication of the virus over a 2-week period of time. Infected tracheal explants were examined with light, electron, and immunofluorescence microscopy. They exhibited specific cytopathologic alterations including nuclear swelling and chromatin margination, multinucleated syncytia and binucleated epithelial cells, and fibroblasts and chondrocytes. Focal destruction and denudation of the respiratory epithelium occurred in later stages of infection. Virus was detected in close association with cilia and was observed budding off the unit membrane of epithelial cells.     

Complete Inhibition of SIVmac Replication by its Capsid Mutants

Mutations were introduced into a genomic region encoding the C-terminal portion of Gag capsid protein of pathogenic simian immunodeficiency virus (SIVmac239). All the mutants generated were defective for virion production and were non-infectious for monkey cells. They all efficiently suppressed the replication of wild type SIVmac in monkey cells. These results were in good agreement with those obtained for human immunodeficiency virus type 1, showing the importance of SIV/monkey model system for studies on Gag. This revised version was published online in July 2006 with corrections to the Cover Date.     

Reversal of Human Immunodeficiency Virus Type 1 Protein-Induced Inhibition of Natural Killer Cell Activity by Alpha Interferon and Interleukin-2

A recombinant fusion peptide, Env-Gag, derived from the human immunodeficiency virus type 1 (HIV-1) genome corresponding to a defined portion of the envelope (Env) and internal core (Gag) proteins was examined for immunoregulatory effects on the cytotoxic activity of natural killer (NK) cell-enriched, large granular lymphocytes (LGL) from healthy donors. Percoll-separated, NK cell-enriched LGL precultured for 24 h with Env-Gag at 10- and 50-ng/ml concentrations, which significantly stimulated lymphocyte proliferation, caused significant suppression of NK cell activity. Denatured Env-Gag did not cause any effect on the NK cell activity of LGL. Two other control peptides, one derived from the Escherichia coli vector used to clone the HIV Env-Gag fusion peptide and the other derived from a non-HIV-1 viral antigen (rubeola virus), did not produce any observable effect on the NK cell activity of LGL, demonstrating the specificity of the effect produced by Env-Gag. Subsequent treatment of LGL with alpha interferon (IFN-α) or interleukin 2 (IL-2) alone partially reversed the Env-Gag-induced suppression of NK cell activity. However, LGL treated with both IFN-α and IL-2 completely reversed the suppression of NK cell cytotoxicity by Env-Gag. The combined effect of IFN-α and IL-2 in enhancing NK cell activity may provide a novel therapeutic approach to the restoration of depressed NK cell activity observed in HIV-infected patients.     

RNA干扰技术抑制病毒复制的研究进展

RNA干扰是指dsRNA抑制细胞内同源基因表达的现象。dsRNA不仅参与内源基因表达调控 ,而且能够抑制宿主内病原微生物基因的表达 ,参与构筑生物体的防御机制。近年来 ,运用RNAi技术在哺乳动物中的研究不断深入 ,尤其是抑制病毒复制的研究成果令人欣喜 ,这为人类抗病毒治疗提供了新的思路。     

HTLV-1 (Human T-Cell Leukemia Virus-1) and Inflammation

Inflammation Research -     

Production of Human Lymphoblastoid Interferon

The interferon response of 21 lines of human lymphoblasts varied greatly. Interferon from the best producer (11,000 U/ml) resembled human leukocyte interferon.     

MXA蛋白抑制HBV复制的体外研究

目的研究MXA蛋白抑制HBV复制的活性。方法将pcDNA3．1-MXA重组质粒和PU19-1．24HBV重组质粒分别按1：1、2：1共转染HepG2细胞（MXA组），对照组使用空pcDNA3．1、Salon DNA和PU19-1．24HBV重组质粒共转染，3d后Western blot检测MXA蛋白表达，Abbott法检测细胞上清HBeAg和HBsAg分泌量，定量PCR检测上清和胞内HBV DNA水平，统计学分析结果。pcDNA3．1-MXA与PU19-1．24HBV重组质粒共转染HepG2细胞，对照组为pcDNA3．1-MXA重组质粒、PU19空质粒和Salon DNA共转染．3d后裂解细胞Western blot检测MXA蛋白表达。结果Western blot显示MXA组有MXA蛋白表达：与对照组相比，pcDNA3．1-MXA和PU19—1．24HBV重组质粒按1：1转染时，MXA组HBeAg下降27％．上清HBV DNA和细胞内HBV DNA分别下降1个log值和0．6个log值；按2：1比例转染时MXA组HBeAg较对照组下降66％，上清HBV DNA和细胞内HBV DNA水平分别下降1．9个和1．7个log值，差异均具有统计学意义（P〈0．05）。Western blot检测显示MXA蛋白抑制HBV组与对照组MXA蛋白表达没有明显差别。结论MXA蛋白在HepG2细胞具有抑制HBV复制活性，抑制活性与蛋白的表达量相关；在抑制HBV复制过程中MXA蛋白自身可能不发生降解。     

Inhibition of HIV/SIV Replication by Dominant Negative Gag Mutants

There are several major strategies against HIV/AIDS. Of these, the gene therapy is a novel, challenging, and promising one. The target genes, which have been extensively studied for the potential gene therapy of HIV/AIDS, include those of cellular and viral origins. Especially, trans-dominant negative Tat, Rev, Env, Pol, and Gag mutants of HIV have currently attracted considerable attention. In this brief review, we summarize the nature of the HIV/SIV mutants of this category and discuss their future use for gene therapy with special reference to the dominant negative Gag mutants of HIV-1.     

腺病毒3型诱导MxA蛋白产生及其抗病毒作用的研究

目的:观察腺病毒3型对MxA蛋白的诱导作用以及MxA蛋白的体外抗病毒作用。方法:采用流式细胞仪分析不同浓度的腺病毒3型（Ad3）诱导健康人外周血单个核细胞（PBMC）胞浆MxA蛋白的产生;采用微量细胞病变抑制法观察重组MxA蛋白对Hela细胞内腺病毒3型的抗病毒作用。结果:不同浓度的腺病毒诱异PBMC产生MxA蛋白的含量均显著高于对照组。10 ng/ml MxA蛋白质抗Hela细胞内腺病毒3型感染的效价为20TCID20。结论:腺病毒3型体外可诱导PBMC产生MxA蛋白;重组MxA蛋白具有抗腺病毒3型作用。     

Selective Enhancement of Human Mononuclear Leucocyte Cytotoxic Function by Interferon

The influence of type I interferon (IFN) on leucocyte-mediated antibody-dependent cellular cytotoxicity (ADCC) has been examined in a system in which differential alloantibody sensitization of human erythrocyte target cells allows discrimination between lymphocyte and monocyte effector function. ADCC mediated by unfractionated mononuclear leucocyte populations was regularly enhanced by interferon pretreatment over a range of concentrations of the sensitizing antibody. The capacity of IFN-augmented reactivity was, however, removed with adherent cell depletion, suggesting that lymphocyte (K-cell) effectors are not modulated by IFN, even though such populations demonstrate IFN-potentiated natural killer (NK) reactivity against K562. These results suggest that monocyte and NK cell function but not K-cell activity was influenced by interferon.