Molecular Analysis Of Human Naive And Memory T Cells

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Summary: Identification of a set of cDNA clones that are differentially expressed in memory CD4+ T cells. In an attempt to determine the molecular features of memory CD4+ T cells, we have utilized cDNA microarrays to measure gene expression of freshly isolated human memory and naive CD4+ T cells from peripheral blood over 100 donors. cDNA microarray method allows us to simultaneously assess genes at genome scales, providing a powerful tool for comparative analysis of gene expression profiles and for identifying differentially expressed genes in certain cell populations. After a sequential analyses of cDNA microarray filters from commercial source (consisting of over 50,000 cDNA clones) and custom-made selected clones from the commercial source and this laboratory (~2,800 unique clones), we provide the first glimpse into gene expression patterns of memory and naive CD4+ T cells at the genome-scale. We found that freshly isolated memory and naive CD4+ T cells expressed similar numbers of genes, and that 14 cDNA clones expressed higher levels of transcripts in memory cells than in naive cells. Identification of activation induced down- and up-regulated genes in memory CD4+ T cells. We have identified 135 (130 known genes and 5 ESTs) up-regulated and 68 (42 known genes and 26 ESTs) down-regulated cDNA clones in memory CD4+ T after 16 hours of in vitro stimulation with anti-CD3 plus anti-CD28. Interestingly, the increase in mRNA levels of up-regulated genes was greater in memory than in naive CD4+ T cells after in vitro stimulation, and was higher with anti-CD3 plus anti-CD28 than with anti-CD3 alone in both memory and naive CD4+ T cells. We then confirmed the changes in expression of actin and cytokine genes identified by cDNA microarrays by Northern analysis. Furthermore, we extended mRNA analysis to protein expression and found that the levels of mRNA and protein of cytokines and actins were correlated. Together, we have identified approximately 200 cDNA clones whose expression levels changed after activation, and suggest that the level of expression of up-regulated genes is a molecular mechanism that differentiates the response of memory from naive CD4+ T cells.