The growth of large, compact megakaryocyte colonies in cultures of human bone marrow is promoted by fresh human plasma and medium conditioned by phyto-hernagglutinin stimulated leukocytes (PHA-LCM). These colonies are typically composed of large cells with translucent cytoplasma, surrounded by a highly refractile border. In addition, they may also contain smaller cells of similar morphology. Independent of their size, all cells react positively with antibodies directed against human factor Vlll antigen. The frequency of megakaryocyte colonies may vary for different individuals from 1-35 colonies per 105 mononuclear bone marrow cells. The observed linear relationships between the number of cultured cells and the frequency of colonies suggests a single cell origin. The described culture conditions also support the development of a larger megakaryocyte component within multilineage mixed colonies, so that it will now be feasible to investigate the mechanisms involved in directing pluripotent cells towards megakaryocytopoiesis.

Bestatin, an inhibitor of leucine aminopeptidase (LAPase), significantly decreased HIV infection as reflected by a reduced number of positive immunofluorescent cells, p24 levels, reverse transcriptase activity and the number of proviral copies found in Bestatin-treated cells. Cellular and extracellular LAPase activity in infected cells was higher than the LAPase activity found in uninfected cells. However, cellular and extracellular LAPase activity as well as total protein kinase C activity was lower in Bestatin-treated cells. Conversely, the incubation of human lymphocytic HUT78 cells with LAPase promotes HIV infectivity. The possible role of LAPase in the pathophysiology of HIV was assessed by determining LAPase serum levels in HIV infected patients. LAPase activity levels were three orders of magnitude greater in sera obtained from HIV patients than those detected in sera of uninfected individuals. Although Bestatin reduced HIV infection, a moderate decrease in the reverse transcriptase activity of chronically-infected H9 human T-lymphocytic cells was observed. Based on the higher levels of LAPase present in the serum of HIV patients and on the combined inhibitory effect of Bestatin on LAPase and on protein kinase C activities, we suggest that LAPase may play an important role in the early events of HIV infection such as viral entry.

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