Acquired Immunodeficiency Syndrome: Tobiume M

Iwata N, Yoshida H, Tobiume M, Ono F, Shimazaki T, Sata T, Nakajima N. · Department of Pathology, National Institute of Infectious Disease, Tokyo, Japan. · J Neurovirol. · Pubmed #17454444 No free full text.

Abstract: The pathogenesis of neurologic dysfunctions caused by human immunodeficiency virus type 1 (HIV-1) infection is not yet well understood. Simian immunodeficiency virus (SIV) infection of macaques is an important animal model for HIV-1 infection. This is the first report to characterize brain progenitor cells (BPCs) isolated from embryonic brain of cynomolgus monkeys (Macaca fascicularis) by neurosphere assay and utilize BPC-derived cell culture for studying SIV infection. The self-renewal and multilineage differentiation properties of BPCs are convenient for planning viral infection experiments. The BPC-derived culture does not contain macrophage/microglial cells, fibroblasts, or endothelial cells. Thus, this culture is appropriate for studying direct relation between SIV infection and neuronal and glial cells. First, the authors characterized undifferentiated and differentiated simian BPCs by immunocytochemistry, flow cytometry analysis, real-time polymerase chain reaction (PCR), and reverse transcriptase (RT)-PCR. The BPCs induced to differentiate by the addition of 1% fetal bovine serum (FBS) were composed of heterogeneous cells expressing nestin, glial fibrillary acidic protein (GFAP), and/or tubulin beta III isoform (Tuj). None of them expressed the monocyte/macrophage/microglial marker. mRNA expression of CD4, CXCR4, CCR5, GPR1, STRL33, and APJ in both undifferentiated and differentiated BPCs were shown by RT-PCR method, suggesting that SIV would infect and replicate in this culture system. Then, it was confirmed that the neurotropic SIV strain, SIV17/E-Fr, replicated productively in BPC-derived cells. The SIV/17E-FrDelta nefGFP was inoculated to identify the infected cells and immunocytochemistry analysis revealed that green fluorescent protein (GFP)-expressing cells were mostly GFAP positive and coexpressed with SIV p27 antigen. Thus, BPC-derived cell culture system is applicable for studying SIV infection in glial and neuronal cells.

Murakami Y, Fukazawa H, Kobatake T, Yamagoe S, Takebe Y, Tobiume M, Matsuda M, Uehara Y. · Department of Bioactive Molecules, National Institute of Infectious Diseases, 1-23-1, Toyama, Shinjuku-ku, 162-8640, Tokyo, Japan. · Antiviral Res. · Pubmed #12076760 No free full text.

Abstract: In the scope of the search for new anti-HIV agents interacting with a new target, we developed a high-throughput screening system to detect the interactions between Nef and Hck. Nef is an accessory protein of HIV, which is involved in the pathogenicity of the acquired immunodeficiency syndrome (AIDS). Nef is also a signaling molecule because it binds to many host molecules. It has especially high affinity to Hck, a member of src family tyrosine kinase. Using a mammalian two-hybrid system, the interaction between Nef and the SH3 domain of Hck induced luciferase activity with high sensitivity and a Nef-PXXP peptide inhibited this interaction; and so did the anticancer drug adriamycin. We also developed another assay system by coexpression of full-length Hck and Nef, and found that Hck kinase was activated depending on the dose of Nef plasmid. Using the second system, we found that adriamycin interfered with the Nef-Hck interaction by reducing the amount of the Hck protein. The mammalian two-hybrid system may show utility in screening inhibitors of Nef-Hck interaction.