Abstract
It has been recognized that dysfunction of CB immunity is in part due to the immaturity of the neonatal immune system (Cairo, Blood, 1997). However, biological pathways and molecular mechanisms associated with the immaturity of CB immunity are still poorly understood. Recently we have utilized oligonucleotide microarray to examine gene expression profiling of CB versus APB Mo and have demonstrated significant differential gene expression patterns, including surface molecules, cytokines, signaling molecules, transcription factors and apoptotic genes (Jiang/Cairo, et al, J. of Immunol., 2004). We sought to examine whether there are differential expressed genes occurred in Mo-derived CB versus APB DC and their impact on DC mediated T cell activity. Briefly, Mo were purified from fresh CB or APB and cultured for 8 days with GM-CSF and IL-4 (immature DC (iDC)) and LPS for mature DC (mDC). mRNA was isolated and oligonucleotide microarray was carried out (Affymetrix, U133A). Data was analyzed by Microarray Suite Version 5.0 (Affymetrix) and GeneSpring 5.0 software (Silicon Genetics). Selected genes were analyzed by RT-PCR (SuperScript, Invetrogen). We identified gene expression patterns that were significantly lower in CB versus APB DC including surface molecules HLA-DQA1 (4F), HLA-DRB3 (5F), HLA-DRB4 (5.5F), CD80 (3F), CD38 (3.8F); cytokine/chemokine genes IL-1b (2.5F), IL6 (2.9F), IL12B (3.5F), CXCL10 (6.6F); immunoregulatory genes ISG20 (11F), IFI27 (7.6F), TNFSF10 (4.5F), SOCS3 (2.5F). Moreover, several transcription factor genes whose proteins may involve in the activation of expression of these immune regulator genes were also differentially expressed (IRF-5 (3F), IRF7 (3F), MAD (6.3F)). We therefore compared CB versus APB DC antigen presentation activity to APB CD8 T cells by ELISPOT assay for interferon-r (IFNr) production (BD Pharmagen). Briefly, the purified CD8 T cells (MHC HLA A2) were incubated with CB or APB DC that were loaded without or with influenza peptide onto ELISPOT plate (Larsson, et al, J. of Immunol., 2000). The ELISPOT plates were developed, scanned and quantitated by an ELISPOT reader (C.T.L. Technology). Our results demonstrated that, although CB or APB mDC had allogeneic effects, influenza peptide loaded CB mDC was not able to induce CD8 T cells to produce IFNr while APB mDC loaded with influenza peptide strongly induced CD8 T cells to produce IFNr. This stimulatory effect of APB mDC on CD8 T cells to produce IFNr was 3.5 fold greater than that of CB mDC. We further examined DC antigen presentation activity to CD4 T cells and observed that APB-DC had stronger effects on CD4 T cell proliferation (3 fold for mDC vs. iDC) compared with CB-DC (only 1.5 fold for mDC vs. iDC) by CFSE assay (Molecular Probe). We postulate that decreased expression of specific surface molecules and other genes resulting in lower surface protein expression in CB DC may in part be responsible for the lack of initiation of signaling events from cell surface to trigger CB-DC to stimulate activation of CD8 and CD4 T cells. The decreased expression of transcription factor genes may also in part be responsible for the lower expressed surface molecule genes. Furthermore, these decreased expressed genes in other molecular categories in LPS-CB vs. APB DC may also partially be responsible for differential innate and adaptive immune function and properties of CB vs. APB.
Metallothionein and bZIP Transcription Factor Genes from Velvetleaf and Their Differential Expression Following Colletotrichum coccodes Infection
