Cytokine-Dependent Proliferation of Human CD34+ Progenitor Cells in the Absence of Serum is Suppressed by the Production of Serine Proteinases by their Progeny

¹ Laboratory of Experimental Hematology, Department of Hematology, Leiden University Medical Center, Leiden, The Netherlands
² Department of Pulmonology, Leiden University Medical Center, Leiden, The Netherlands

^* To whom correspondence should be addressed. E-mail: H.M.Goselink{at}lumc.nl.

	Abstract

In this study, we demonstrate that the synthesis and release of serine proteinases by hematopoietic cells affects the in vitro proliferation of hematopoietic progenitor cells (HPC) in response to proteins including hematopoietic growth factors (HGF), transferrin, insulin and albumin in serum-free cultures. In serum-free cultures, bone marrow mononuclear cells (BMMNC) or the CD34-negative progeny of the CD34⁺ cells were shown to release the serine proteinases human neutrophil elastase (HNE), cathepsin G (Cath G) and proteinase 3 (Pr3). In the absence of serum, we showed that HNE, Cath G and Pr3 rapidly and dose-dependently degraded HGF and other proteins present in the medium resulting in decreased proliferation of the HPC. Addition of the serine proteinase inhibitors 1-Proteinase inhibitor (1-PI) or the Secretory Leukocyte Proteinase Inhibitor (SLPI), but not leupeptin, aprotinin or 4-(2-aminoethyl)-benzenesulfonylfluoride hydrochloride (AEBSF), could completely prevent the degradation of proteins relevant for the growth of hematopoietic cells. Thus, the addition of serine proteinase inhibitors like 1-PI or SLPI may be critical for the expansion of CD34⁺ cells or gene transfer into CD34⁺ cells or other hematopoietic cells in vitro using serum-free media under good manufacturing practice (GMP) conditions.