The influence of INK4 proteins on growth and self-renewal kinetics of hematopoietic progenitor cells

This study investigated the influence of expression of proteins of the INK4 family, particularly p16, on the growth and self-renewal kinetics of hemat opoietic cells. First, retrovirus-mediated gene transfer (RMGT) was used to restore p16(INK4a) expression in the p16(INK4a)-deficient lymphoid and mye loid cell lines BV173 and K562, and it was confirmed that this inhibited th eir growth. Second, to sequester p16(INK4a) and related INK4 proteins, cycl in-dependent kinase 4 (CDK4) was retrovirally transduced into normal human CD34(+) bone marrow cells and then cultured in myeloid colony-forming cell( CFC) assays. The growth of CDK4-transduced colonies was more rapid; the cel l-doubling time was reduced; and, upon replating, the colonies produced gre ater yields of secondary colonies than mock-untransduced controls. Third, c olony formation was compared by marrow cells from p16(INK4a-/-) mice and wi ld-type mice. The results from p16(INK4a-/-) marrow were similar to those f rom CDK4-transduced human CFCs, in terms of growth rate and replating abili ty, and were partially reversed by RMGT of p16(INK4a). Lines of immature gr anulocytic cells were raised from 15 individual colonies grown from the mar row of p16(INK4a-/-) mice. These had a high colony-forming ability (15%) an d replating efficiency (96.7%). The p16(INK4a-/-) cell lines readily became growth factor-independent upon cytokine deprivation, Taken together, these results demonstrate that loss of INK4 proteins, in particular p16(INK4a), increases the growth rate of myeloid colonies in vitro and, more importantl y, confers an increased ability for clonal expansion on hematopoietic proge nitor cells. (Blood. 2001;97:2604-2610) (C) 2001 by The American Society of Hematology.