The molecular basis for the cytokine-induced defect in homing and engraftment of hematopoietic stem cells

Abstract

Objective: Hematopoietic stem cell homing and engraftment is dramatically altered by cytokine exposure. These studies address the molecular mechanisms responsible for the observed changes in transplantation biology.

Methods: Primitive murine hematopoietic stem cells were isolated by fluorescence-activated cell sorting of lineage depleted (Lin(-)) cells exhibiting low staining of Hoechst 33342 and rhodamine 123 dyes or Lin(-) cells bearing Sca. Adhesion receptor expression was examined by immunofluorescence and reverse transcriptase polymerase chain reaction. In vitro adhesion assays were employed to define binding interactions between stem cells and stroma or extracellular matrix proteins.

Results: Adhesion of Lin(-)Sca+ cells to Dexter stroma could be blocked by about 90% with antibodies to PECAM-1, alphaa(4), or beta(1), and partially blocked by antibodies to alpha(5), CD44, or L-selectin. By immunofluorescence, about 30% of purified Lin(-)Ho(lo)Rho(lo) cells expressed alpha(4), alpha(5), beta(1), and L-selectin, about 15% expressed alpha(L) and alpha(6), half expressed PECAM-1, and none expressed alpha(1) or alpha(2). After 48 hours in expansion cytokines, only 9% of the cells expressed alpha(4) and none expressed beta(1), whereas alpha(L) expression was fully restored, PECAM-1 and L-selectin partially restored, CD44 expression was newly induced, and adhesion to both fibronectin and laminin was reduced. Adhesion to purified collagen, fibronectin, or laminin enhanced expression of beta(1) integrins.

Conclusion: Expansion cytokines that move quiescent primitive hematopoietic stem cells into S phase markedly altered adhesion receptor expression and reduced their functional binding to extracellular matrix, which could reduce engraftment after transplant.