Immune Netw.  2001 Apr;1(1):87-94. 10.4110/in.2001.1.1.87.

Analysis of stromal Cells Developed from Cord Blood CD34+ Cells

• Ryu KH
• Park SJ
• Kim KH
• Seoh JY
• Khan M
• Shin HY
• Ahn HS

• KMID: 2150561
• DOI: http://doi.org/10.4110/in.2001.1.1.87

Abstract

BACKGROUND: Cytokine-mediated ex vivo expansion has been proposed as a means of increasing the number of cord blood (CB) hematopoietic stem cells for transplantation. As well as stem cell number, stromal cells are necessary for functional maturation of hematopoiesis. The purpose of this study was to analyze the development of stromal cells during ex vivo expansion of CB CD34+ cells.
METHODS
CD34+ cells were purified from CB by magnetic bead selection. The levels of of interleukin-3, interleukin-1 beta, interleukin-6, granulocyte macrophage-colony stimulating factor and tumor necrosis factor-alpha were measured in culture supernatants on 0, 1, 2, and 3 weeks, using ELISA techniques. CB CD34+ cells were expanded in Iscoves modified Dulbeccos medium in the presence of several cytokines. The expression of E-selectin, vascular cell adhesion molecule- 1, intercellular adhesion molecule- 1, platelet/endothelial cell adhesion molecule-1, von Willebrand factor, vimentin, and CD14 in newly developed stromal cells was examined by immunocytochemical method. Relevant extracellular matrix (ECM) proteins and proper cytokines were also assayed for the most suitable condition for expansion of stromal cells.
RESULTS
Several cytokines were found to have been produced by CB CD34+ cells as well as bone marrow-derived CD34+ cells. During ex vivo expansion of CB CD34+ cells, stromal cells appeared in the culture by day 4 and expanded over the following 7- 10 days before being confluent by day 2 1. These cells expressed surface markers characteristic of cells of endothelial lineage. Furthermore, these stroaml cells also expanded effectively when treated with thrombopoietin+flt-3 ligand+stem cell factor+leukemia inhibitory factor or 0. 1% poly-L-lysine-coated wells.
CONCLUSION
Stromal cells were developed during ex vivo expansion of CB CD34+ cells and that this development could be enhanced further by treating the stromal cells with cytokines or ECM.

Keyword

cord blood; exvivo expansion; stromal cells; cytokines; extracellular matrix

MeSH Terms

Cell Adhesion
Cytokines
E-Selectin
Enzyme-Linked Immunosorbent Assay
Extracellular Matrix
Fetal Blood*
Granulocytes
Hematopoiesis
Hematopoietic Stem Cells
Interleukin-1beta
Interleukin-3
Interleukin-6
Stem Cells
Stromal Cells*
Tumor Necrosis Factor-alpha
Vimentin
von Willebrand Factor
Cytokines
E-Selectin
Interleukin-1beta
Interleukin-3
Interleukin-6
Tumor Necrosis Factor-alpha
Vimentin
von Willebrand Factor