NORMAL HUMAN-SERUM CONTAINS A FACTOR(S) CAPABLE OF INHIBITING MEGAKARYOCYTE COLONY FORMATION

  • 1 January 1982
    • journal article
    • research article
    • Vol. 10  (8) , 650-660
Abstract
Growth of megakaryocyte colonies from human bone marrow progenitors was achieved in plasma clot culture. Megakaryocyte [MK] colonies were identified either by cytological examination or by immunofluorescent labeling using a monoclonal antiplatelet antibody (J 15). No significant differences were observed in the quantitation of the colonies by these 2 methods. In the absence of a stimulating factor, MK colonies were detectable when high cellular concentrations were seeded. Among the numerous conditioned media tested for their ability to stimulate MK colony formation, conditioned medium from leukocytes stimulated by PHA (PHA-LCM [phytohemagglutinin/leukocyte-condionted medium]) was the most effective. Under standard conditions of culture corresponding to 20% normal human sera, colony formation was not related linearly to seeding density, even when cultures were stimulated by PHA-LCM. Upon reduction of the concentration of serum (2.5-5%) in the culture medium, colony formation displayed a linear relationship with seeding density only when PHA-LCM was used as the stimulating factor. At the same time, the size of the colonies increased. Such inhibition was observed with all the human sera tested, but it varied in extent from 1 batch to another. Replacement of serum by albumin, Fe-saturated transferrin, .alpha.-thioglycerol and low density lipoproteins at physiological concentration, but in the presence of bovine plasma, provided a culture system whose ability to support colony formation equaled that of low concentrations of whole serum; spontaneous MK colony formation still occurred. Evidence for the presence of an inhibitor(s) of MK colony formation in normal human sera and the role of cellular factors in stimulating MK colony formation were demonstrated.