ABSTRACT
<p><b>OBJECTIVE</b>To explore the role of HIV-1 tat gene variations in AIDS dementia complex (ADC) pathogenesis.</p><p><b>METHODS</b>HIV-1 tat genes derived from peripheral spleen and central basal ganglia of an AIDS patient with ADC and an AIDS patient without ADC were cloned for sequence analysis. HIV-1 tat gene sequence alignment was performed by using CLUSTAL W and the phylogentic analysis was conducted by using Neighbor-joining with MEGA4 software. All tat genes were used to construct recombinant retroviral expressing vector MSCV-IRES-GFP/tat. The MSCV-IRES-GFP/tat was cotransfected into 293T cells with pCMV-VSV-G and pUMVC vectors to assemble the recombinant retrovirus. After infection of gliomas U87 cells with equal amount of the recombinant retrovirus, TNF-α, and IL-1β concentrations in the supernatant of U87 cells were determined with ELISA.</p><p><b>RESULTS</b>HIV-1 tat genes derived from peripheral spleen and central basal ganglia of the AIDS patient with ADC and the other one without ADC exhibited genetic variations. Tat variations and amino acid mutation sites existed mainly at Tat protein core functional area (38-47aa). All Tat proteins could induce U87 cells to produce TNF-α and IL-1β, but the level of IL-1β production was different among Tat proteins derived from the ADC patient's spleen, basal ganglia, and the non-ADC patient's spleen. The level of Tat proteins derived from the ADC patient's spleen, basal ganglia, and the non-ADC patient's spleen were obviously higher than that from the non-ADC patient's basal ganglia.</p><p><b>CONCLUSION</b>Tat protein core functional area (38-47aa) may serve as the key area of enhancing the secretion of IL-1β. This may be related with the neurotoxicity of HIV-1 Tat.</p>
Subject(s)
Adult , Humans , Middle Aged , AIDS Dementia Complex , Metabolism , Pathology , Virology , Amino Acid Sequence , Basal Ganglia , Virology , Cell Line, Tumor , Gene Expression Regulation, Viral , Genes, tat , HIV-1 , Genetics , Virulence , Interleukin-1beta , Genetics , Bodily Secretions , Molecular Sequence Data , Neuroglia , Pathology , Bodily Secretions , Spleen , Virology , Tumor Necrosis Factor-alpha , Genetics , Bodily Secretions , tat Gene Products, Human Immunodeficiency Virus , Genetics , PhysiologyABSTRACT
<p><b>OBJECTIVE</b>To study the genetic diversity of HIV-1 nef genes from a patient with AIDS dementia complex(ADC) , so as to research the amino acid variability and the pathogenesis of ADC.</p><p><b>METHODS</b>The nef gene was amplified with PCR from genomic DNA which was extracted from spleen and different brain tissues(basal ganglia, frontal gray matter, meninges, temporal lobe)of a patient who died of ADC. PCR products were cloned into the pMD19-T vector, after transformation and selection by ampicillin and blue/white spotting. Five of positive clones were sequenced and confirmed with BLAST. HIV-1 nef sequences were processed with BioEdit and MEGA4 to do Neighbor-Joining tree, p-Distances, and values of ds/dn.</p><p><b>RESULTS</b>The samples were all identified as HIV-1 B and genetic variation exists in HIV-1 nef gene isolated from different tissues compared with HXB2. In addition,part of the changes were different between periphery and brain.</p><p><b>CONCLUSION</b>Variations exist in the HIV-1 nef gene extracted from the ADC patient and the variations from peripheral and central nerve tissues were different,these variations may change the function of Nef,and it needs more research.</p>