Biomedical subjects

Yinming Chen

Publications and source records attributed to Yinming Chen.

3 recordsLinked to original sources

PubMed2003

Customized rapid subtraction hybridization (RaSH) gene microarrays identify overlapping expression changes in human fetal astrocytes resulting from human immunodeficiency virus-1 infection or tumor necrosis factor-alpha treatment.

Genes displaying altered expression as a function of human immunodeficiency virus (HIV)-1 infection of cultured primary human fetal astrocytes (PHFA) were previously identified using a rapid subtraction hybridization (RaSH) method. This scheme identified both known and novel genes displaying elevated expression, astrocyte elevated genes (AEG), and decreased expression, astrocyte suppressed genes (ASG), in PHFA as a consequence of infection with HIV-1 or treatment with HIV-1 envelope glycoprotein (gp120). RaSH also identified both known and novel genes displaying enhanced (HR) or reduced (HS) expression in HIV-1 resistant versus HIV-1 susceptible human T-cell clones. In the present study, a customized microarray approach employing these RaSH-derived genes was used to distinguish overlapping gene expression changes occurring in PHFA as a function of treatment with HIV-1 and the neurotoxic agent tumor necrosis factor (TNF)-alpha. RaSH cDNAs were spotted (microarrayed) on nylon membranes and probed with temporally isolated reverse transcribed cDNAs from HIV-1-infected and TNF-alpha-treated PHFA. This strategy identified genes displaying parallel changes after TNF-alpha treatment as observed following HIV-1 infection. Confirmation of genuine differential expression was achieved by Northern blotting. These studies document that TNF-alpha can induce a set of corresponding changes in specific AEGs and ASGs as does HIV-1 infection in PHFA. Furthermore, this customized microarray approach with RaSH-derived clones represents an efficient and sensitive methodology for elucidating molecular changes in PHFA occurring as a consequence of treatment with pharmacological agents affecting astrocyte physiology.

Astrocytes↗

PubMed2003

Identification of gene products suppressed by human immunodeficiency virus type 1 infection or gp120 exposure of primary human astrocytes by rapid subtraction hybridization.

Neurodegeneration and human immunodeficiency virus type 1 (HIV-1)-associated dementia (HAD) are the major disease manifestations of HIV-1 colonization of the central nervous system (CNS). In the brain, HIV-1 replicates in microglial cells and infiltrating macrophages and it persists in a low-productive, noncytolytic state in astrocytes. Astrocytes play critical roles in the maintenance of the brain microenvironment, responses to injury, and in neuronal signal transmission, and disruption of these functions by HIV-1 could contribute to HAD. To better understand the potential effects of HIV-1 on astrocyte biology, the authors investigated changes in gene expression using an efficient and sensitive rapid subtraction hybridization approach, RaSH. Primary human astrocytes were isolated from abortus brain tissue, low-passage cells were infected with HIV-1 or mock infected, and total cellular RNAs were isolated at multiple time points over a period of 1 week. This approach is designed to identify gene products modulated early and late after HIV-1 infection and limits the cloning of genes displaying normal cell-cycle fluctuations in astrocytes. By subtracting temporal cDNAs derived from HIV-1-infected astrocytes from temporal cDNAs made from uninfected cells, 10 genes displaying reduced expression in infected cells, termed astrocyte suppressed genes (ASGs), were identified and their suppression was confirmed by Northern blot hybridization. Both known and novel ASGs, not reported in current DNA databases, that are down-regulated by HIV-1 infection are described. Northern blotting confirms suppression of the same panel of ASGs by treatment of astrocytes with recombinant HIV-1 envelope glycoprotein, gp120. These results extend our previous analysis of astrocyte genes induced or enhanced by HIV-1 infection and together they suggest that HIV-1 and viral proteins have profound effects on astrocyte physiology, which may influence their function in the CNS.

AIDS Dementia Complex↗

PubMed2002

Identification and cloning of human astrocyte genes displaying elevated expression after infection with HIV-1 or exposure to HIV-1 envelope glycoprotein by rapid subtraction hybridization, RaSH.

Neurodegeneration and dementia are common complications of AIDS caused by human immunodeficiency virus type 1 (HIV-1) infection of the central nervous system. HIV-1 target cells in the brain include microglia, infiltrating macrophages and astrocytes, but rarely neurons. Astrocytes play an important role in the maintenance of the synaptic micro-environment and in neuronal signal transmission. To investigate potential changes in cellular gene expression associated with HIV-1 infection of astrocytes, we employed an efficient and sensitive rapid subtraction hybridization approach, RaSH. Primary human astrocytes were isolated from abortus brain tissue and low-passage cells were infected with HIV-1. To identify genes that display both early and late expression modifications after HIV-1 infection and to avoid cloning genes displaying normal cell cycle fluctuations in astrocytes, RNAs were isolated and pooled from 6, 12, 24 h and 3 and 7 day uninfected and infected cells and used for RaSH. Temporal cDNA libraries were prepared from double-stranded cDNAs that were enzymatically digested into small fragments, ligated to adapters, PCR amplified, and hybridized by incubation of tester and driver PCR fragments. By subtracting temporal cDNAs derived from uninfected astrocytes from temporal cDNAs made from HIV-1 infected cells, genes displaying elevated expression in virus infected cells, termed astrocyte elevated genes (AEGs), were identified. Both known and novel AEGs, not reported in current DNA databases, are described that display early or late expression kinetics following HIV-1 infection or treatment with recombinant HIV-1 envelope glycoprotein (gp120). For selected AEGs, expression of their protein products was also tested by Western blotting and found to display elevated expression following HIV-1 infection. The comparable pattern of regulation of the AEGs following HIV-1 infection or gp120 treatment suggest that HIV-1 exposure of astrocytes, even in the absence of productive infection, can induce changes in cellular gene expression.

Actinin↗