Analysis of glycosyltransferase gene expression in human monocytes, macrophages and DC

ABSTRACT: So far, although differentiation and activation of Ag presenting cells, including monocytes, dendritic cells (DCs) and macrophages are accompanied by programmed remodelling of cell surface glycosylated molecules with potentially biologically important consequences, no global analysis of the expression of genes involved in glycan biosynthesis (essentially glycosyltransferases (GTs) and modification (mainly sulfotransferases) has been reported in these cells. Our project aims to obtain global information relating to the expression of genes encoding GTs and sulfotransferases in human monocytes, DCs and macrophages isolated or differentiated from the same donor. We also aimed to compare the expression profile of these genes in immature versus mature (immunostimulatory phenotype) DCs and macrophages (LPS stimulation, 4 and 18 hrs). To this end we have performed three independent experiments (n = 7 x 3 samples) using the dedicated gene microarray (glycogene-chip v3). Although, globally the data are interesting(please see the summary below), they lack significance and are not yet exploitable. The relatively high variability between samples probably results from the human donor sources given their heterogeneous genetic makeup and natural history. Thus, to reach significance, we believe that additional donors need to be analyzed. Of note, the time point “4hrs” can be omitted since it does not appear to be indicative. Three x 5 samples would be thus necessary. Utilizing a focused gene microarray, we intended to compare the glycosyltransferase (GT) and sulfotransferase gene expression profile of human monocytes relative to immature dendritic cells (DCs) and to macrophages and to characterize variations in DCs and macrophages undergoing maturation. Micro-array analysis indicated that monocytes express transcripts for a full set of enzymes involved in the biosynthesis of N- and O-glycans potentially elongated by poly-LacNAc chains with type II terminal sequences. Monocytes also express, at a very good level, genes encoding enzymes involved in glycosaminoglycan biosynthesis, but seem to have a limited capacity for glycolipid biosynthesis. Immature DCs and macrophages exhibit similar pattern of GT and sulfotransferase expression with only a limited number of cell type specific gene expression. Approximately 15 % of GT and sulfotransferase transcripts varied in both DCs and macrophages, the majority of them being up-regulated compared to monocytes. Strikingly, stimulation of DCs and Ms with lipopolysaccharide caused a general alteration in gene expression, in particular in DCs, mainly affecting genes found to be positively modulated during the differentiation steps. LPS treatment also caused a relatively minor increase in gene expression in macrophages whereas in DCs, the expression levels of a significant number of genes were enhanced. Validation of this analysis was partially provided by quantitative RT-PCR and flow cytometry of cell surface glycan epitopes. Collectively, this preliminary study implies an important modification of the pattern of glycosylation in differentiated and activated APCs with potential biological consequences. Our project aims to obtain global information relating to the expression of genes encoding GTs and sulfotransferases in human monocytes, DCs and macrophages isolated or differentiated from the same donor. We also aimed to compare the expression profile of these genes in immature versus mature (immunostimulatory phenotype) DCs and macrophages (LPS stimulation, 4 and 18 hrs).

ORGANISM(S): Homo sapiens

SUBMITTER: Steven Head 

PROVIDER: E-GEOD-29937 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress