Journal of human virology最新文献

Objective: This study was undertaken to determine the contribution of HIV co-receptors and beta chemokine secretion to the increased susceptibility for human immunodeficiency virus (HIV) infection of peripheral blood mononuclear cells (PBMC) obtained from HIV-seronegative Ethiopian immigrants in Israel (ETH).

Study design: Immune activation markers and HIV co-receptor expression on lymphocytes and monocytes, and beta chemokine secretion by CD8+ cells, were compared between ETH and non-Ethiopian Israeli (IS) HIV-negative individuals.

Results: The percentage of lymphocytes and monocytes expressing CCR5 was 1.6 and 3.0 times higher in ETH (n = 83) than in IS (n = 45), respectively (P < .001), whereas RANTES and MIP-1alpha secretion was 0.5 and 0.7 times lower (P < .01 and P < .05). The percentage of CCR5-expressing cells and RANTES secretion were inversely correlated (r = -0.7; P < .002). No differences were found in the proportion of CXCR4-expressing cells. No correlation between CCR5 expression and cell activation profile in the whole ETH population was found. However, in highly activated individuals (HLA-DR/CD3 > 7%), a significant decrease in CCR5 expression was observed.

Conclusions: An increased proportion of CCR5-expressing cells with decreased beta chemokine secretion observed in ETH may account for the increased susceptibility to HIV infection of cells obtained from this group. These findings may partly explain the higher susceptibility for HIV infection in Africa and thus the rapid spread of acquired immunodeficiency syndrome (AIDS) in that continent.

Objective: Inheritance of a mutant allele of the SDF1 gene delays the onset of human immunodeficiency virus type 1 (HIV-1) disease. Because the mutation lies in the 3' untranslated region of the gene, it was suggested that this mutation may upregulate transcription of the gene, resulting in more abundant SDF1, which in turn inhibits T-tropic HIV-1 and delays disease onset. This implies that this segment of SDF1 gene contains a negative regulatory element. We directly tested this hypothesis in vitro.

Study design/methods: We cloned the wild-type and the mutant SDF1 gene in an HIV-2 gene transfer vector as well as in a baculovirus expression vector. We expressed the cloned genes in human and insect cells in culture and analyzed the abundance of SDF1 RNA by hybridization and protein using antiviral assays.

Results: The abundance of SDF1 RNA synthesized by the mutant clone with the mutation in the 3' untranslated region was no different from that synthesized by the wild-type clone in cultured cells. This was the case for both the HIV-2 long terminal repeat (LTR)-directed expression in human cells and baculovirus promoter-directed expression in insect cells. Both clones apparently synthesized SDF1 with equivalent biologic activity. Similar results were obtained for a mutant with the deletion of a GC-rich segment in the 5' untranslated region.

Conclusions: Mutation of the 3' untranslated exon did not affect SDF1 RNA synthesis in vitro. It also did not appear to affect translation of SDF1 RNA. A similar mutational analysis of the 5' noncoding exon suggested that this region also did not regulate SDF1 expression.

Objectives: To detect HIV-1 in cellular and acellular fractions of cervicovaginal secretions obtained by cervicovaginal lavage (CVL) and evaluate viral genotypes in the HIV-1-positive CVL samples.

Study design/methods: This study consists of 37 HIV-1-seropositive pregnant and nonpregnant women from the United States. A total of 63 paired CVL and blood samples were collected. HIV-1 DNA from cervical cells (CC) and virion RNA from cervical supernatant (CS) was detected by gag polymerase chain reaction (PCR) assays. The HIV-1 genotypes were determined by analyzing the nested PCR-amplified V3 region sequences of the HIV-1 gp120 envelope gene.

Results: Within this cohort, 95% of the women were on single or combination antiretroviral therapy. Of the pregnant women, 63% of samples had HIV-1 viral DNA in the CC, and 29% of samples were positive for viral RNA in the CS. Among nonpregnant women, 71% of samples were positive for HIV-1 DNA in CC, and 46% of samples tested positive for virion RNA in CS. Plasma viral load ranged between 10,000 and 100,000 copies/mL and showed significant correlation with the detection of HIV-1 RNA in the CVL; this relation was less apparent with viral DNA in CC. The viral blood and CVL specimens were further analyzed by evaluating the genotypes of HIV-1 variants. In most patients, a high degree of similarity was observed between the viral sequences derived from blood and CVL samples. Two patients demonstrated closely related but somewhat distinct genotypic variants in CVL and blood. One subject showed clear compartmentalization in which distinct viral genotypes were observed in CVL and blood. Based on V3 loop analyses of gp120, with one exception, the cervicovaginal secretions harbored viral populations with a macrophage (CCR5)-tropic phenotype.

Conclusions: This study demonstrates the unique characteris tics of HIV-1 strains in the genital secretions of a relatively large cohort of HIV-1-infected women in the United States. These results are important for further analysis of HIV-1 transmission and pathogenesis in vivo and for rational vaccine design.

Objective: To investigate the transactivating activity of Vpr proteins from human immunodeficiency viruses types 1 and 2 (HIV-1 and HIV-2) and simian immunodeficiency viruses (SIVs) on various primate lentivirus long terminal repeats (LTRs), and to determine whether the Vpx proteins shared by HIV-2 and SIV are able to transactivate any HIV or SIV promoter.

Study design/methods: The vpr and vpx genes of the HIVs and SIVs encode virion-associated proteins, which are implicated in viral replication and pathogenesis. HIV-1 Vpr is involved in the transport of the preintegration complex (PIC) to the nucleus, transactivates the viral LTR, and induces cell cycle arrest. HIV-2 and SIV Vpx proteins share amino acid sequence similarities with Vpr and are involved in PIC translocation into the nucleus but are unable to induce cell cycle arrest. We cloned and expressed the vpr and vpx genes from several primate lentiviruses and tested their transactivating ability on HIV-1, HIV-2, SIVmac and SIVagm LTRs cloned upstream of the CAT reporter gene.

Results: All Vpr tested had a transactivating effect on several viral LTRs; however, none of the Vpx proteins showed a detectable transactivating effect.

Conclusions: These results indicate that the transactivating properties of Vpr proteins were conserved throughout evolution in primate lentiviruses, which suggests that they have an important role in virus replication.

Objective: A novel 2-amino acid insertion between codons 69 and 70 of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) which confers multiple drug resistance has recently been reported. Independently, we have identified similar insertion mutations in Japanese hemophiliacs and attempted to analyze their emergence in conjunction with therapy regimens and their contribution to drug resistance using recombinant technology.

Methods: The plasma and peripheral blood mononuclear cells (PBMCs) of 348 HIV-1-infected hemophiliacs were screened for HIV-1 RT mutations relevant to nucleoside analogue inhibitors and isolating viruses. Contribution of each insertion to drug resistance was studied by introducing the mutations into a T-cell line-tropic NL4-3 infectious clone and testing the drug susceptibilities of the recovered virus.

Results: Insertion of the 2-amino acid residue was found in 4 of the 348 cases and was strongly associated with prolonged chemotherapy with zidovudine (AZT) and didanosine (ddI). The virus isolated from 1 of the 4 cases possessed the same insertion. Characterization of these virus and the recombinant NL4-3 with the insertion strongly suggested that the insertion caused resistance not only to AZT and ddI but also to lamivudine (3TC) and zalcitabine (ddC).

Conclusion: A 2-amino acid insertion between codons 69 and 70 of RT was detected in 4 of 348 (1.1%) Japanese hemophiliacs and was found to be associated with multiple drug resistance to nucleoside analogue RT inhibitors.

Objectives: We examined the effect and time of addition of beta-chemokines on human immunodeficiency virus type 1 (HIV-1) replication, binding, and uncoating in human macrophages and measured CCR5 receptor expression during virus binding and uncoating.

Methods: Macrophages were treated with beta-chemokines before infection, at infection, or postinfection, and virus replication was determined by p24 antigen level. Binding and uncoating of 35[S]-methionine-labeled HIV-1 was measured. CCR5 expression was determined by flow cytometry.

Results: The beta-chemokines potently inhibited virus replication. The strongest inhibition occurred when cultures were pretreated and maintained with beta-chemokines. Beta-chemokines also caused strong inhibition of viral uncoating and a considerable decrease in CCR5 expression during uncoating.

Conclusions: CCR5 receptors appear to be internalized and recycled to the cell surfaces during HIV entry. The down-regulation of CCR5 expression by beta-chemokines during virus uncoating probably accounts for the reduction in virus uncoating (entry) and hence in virus replication.

Objectives: Widespread dendritic injury may be one mechanism involved in the neurologic impairment that occurs in HIV-1 infection. The objectives of this study were to quantitate the extent of dendritic injury in a primate model of central nervous system (CNS) infection, investigate the role of nitric oxide (NO) as a mediator of neuropathologic changes, and evaluate the relation of these changes to cognitive and motor function.

Study design/methods: Cognitive and motor function was assessed in rhesus macaque monkeys infected with simian immunodeficiency virus (SIV). In situ hybridization, immunohistochemistry, and quantitative image analysis were employed to assess the relations among productive infection, NO synthase (iNOS), and dendritic injury.

Results: Productive infection of cells of the macrophage lineage in CNS is associated with inflammation, increased expression of iNOS, and dendritic injury. The tests of cognitive and motor function employed were abnormal in both animals that had evidence of productive infection and those that did not.

Conclusions: Increased NO accompanying productive infection and encephalitis may be one cause of neuronal injury in lentivirus infections of the CNS. Extension of tests of cognitive and motor function to late-stage AIDS in rhesus monkeys is needed to assess the potential role of NO-induced dendritic damage in lentiviral encephalopathy/AIDS dementia complex.

Objectives: The pathogenesis of coxsackievirus B3 (CVB3)-induced myocarditis was investigated in immunocompetent C57BL/6 and MHC class II knockout mice with identical genetic backgrounds.

Study design/methods: We analyzed the histology and immunohistology of myocardial injury, the replicating virus titer, and antibody response in the early and late phase of disease.

Results: CVB3-infected C57BL/6 mice showed acute myocarditis, with spontaneous healing, virus elimination, anti-CVB3 IgM/IgG production, and neutralizing antibody response. In contrast, MHC class II knockout mice developed less severe acute myocarditis, persistence of infiltrations and strong fibrosis, virus persistence, and weak IgG response, with absence of virus neutralizing antibodies.

Conclusions: Immunodeficient organisms are more susceptible to long-term heart muscle injuries after infection with CVB3. The presence of CD4+ T cells are necessary to prevent the development of chronic disease.

Objectives: To characterize replication patterns and cytopathic effects during human cytomegalovirus (HCMV) infection of brain cells.

Design: Primary human mixed glial/neuronal cells, as well as purified microglial, astroglial, and enriched neuronal cell cultures, were infected with HCMV strains AD169 and RC256 to determine the ability of the different brain cell types to support viral replication.

Results: Mixed glial/neuronal cell cultures were fully permissive for viral replication. Based on previous studies, we hypothesized that human microglial cells would preferentially support productive HCMV replication. However, HCMV did not replicate or display genomic expression in microglial cells. In contrast, primary astrocytes were fully permissive and displayed HCMV-induced cytopathic effects resulting in cell death. In highly enriched neuronal cultures, productive infection and viral expression occurred only in scattered astrocytes. Early in the infection, apoptotic plasma membrane changes were induced in astrocytes. However, nuclear fragmentation was not apparent until later during the course of infection.

Conclusions: These results suggest that HCMV possesses astrocytotropic properties that confer preferential expression and cytopathic replication in astrocytes over microglia or neuronal cells. Apoptotic cell death, which is a result of HCMV infection, appears to be delayed until peak viral replication has occurred.