Comparison of Cord Blood and Bone Marrow Mononuclear Cells by Serial Analysis of Gene Expression (SAGE).

Abstract

Hematopoietic stem/progenitor cells with their dual ability for self-renewal, expansion and multilineage differentiation constitute an essential component of hematopoietic transplantation. These qualities, together with their transfection capacity make them useful for gene therapy also. Umbilical cord blood and bone marrow represent efficient sources of these cells. Cord blood stem cells are more primitive than bone marrow cells. They have a greater capacity to self-renewal, proliferation, expansion, multilineage differentiation and are better transfected. They can also be transplanted without complete HLA compatibility.Some recent studies are demonstrating that they could have a better trafficking and homing than the others, which could promote an efficient development and proliferation in the marrow microenvironment and better recovering when transplanted. However, these mechanisms of migration/cell adhesion and cell cycle are not fully understood. The aim of this study is to compare cord blood mononuclear cells with the existing bone marrow mononuclear cells library (esage database) by means of serial analysis of gene expression in order to investigate if there are any differences that could explain those findings. We obtained, by sequencing the SAGE library of cord blood cells, a total of 44.924 tags representing 15.519 unique tags. These unique tags were distribute as: known genes= 7772; sequence predicted or annotated=3935; no matches= 3812. The bone marrow library showed total tags=36577 being unique tags=13075: known genes= 6711; sequence predicted or annotated = 3371; no matches= 2993. These genes were annotated using Gene Ontology Consortium and the distribution of the categories in both libraries was similar. The different expression genes, selected using the fold difference (fold≥5 and ≤ 5), were 238. In order to validate the library, we tested 12 different expressed genes by real time PCR. The first six of them (SMARCC2, CDC25B, S100-8, alpha-globin, beta-globin, gamma-globin) confirmed the SAGE results. The more expressed cord blood genes were SMARCC2, CDC25B and gamma-globin. Beta-globin, and S100-8 were more expressed in bone marrow. The alpha- globin result demonstrated almost the same result between cord blood and bone marrow. CDC25B is a member of the CDC25, family of phosphatases, it activates cyclin dependent kinase CDC2 which is required for entry into mitoses. SMARCC2 is a member of SWI/SNF family proteins which display helicase and ATPase activities. It is responsible for regulation of transcription of certain genes by altering the chromatin structure around those genes. S100-8 is a calcium binding protein with a prominent role in leukocyte trafficking. It stimulates shedding of L-selectin, which is also involved in cell adhesion. It makes a role in inflammatory process towards neutrophil migration to inflammatory sites. The alpha, beta and gamma globin results were those expected based on the ontogeny. The differential gene expression here obtained could be an important tool to guide new functional studies regarding the molecular mechanisms of homing, cell cycle and cell adhesion of cord blood cells.