Thrombopoietin (TPO) is a cytokine that is involved in the regulation of platelet production. The receptor for TPO is c-Mpl. To further investigate the role and specificity of this receptor in regulating megakaryocytopoiesis, c-mpl-deficient mice were generated by gene targeting. The c-mpl-/- mice had an 85 percent decrease in their number of platelets and megakaryocytes but had normal amounts of other hematopoietic cell types. These mice also had increased concentrations of circulating TPO. These results show that c-mpl specifically regulates megakaryocytopoiesis and thrombopoiesis through activation by its ligand TPO.

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The effects of recombinantly produced ob protein were compared to those of food restriction in normal lean and genetically obese mice. Ob protein infusion into ob/ob mice resulted in large decreases in body and fat-depot weight and food intake that persisted throughout the study. Smaller decreases in body and fat-depot weights were observed in vehicle-treated ob/ob mice that were fed the same amount of food as that consumed by ob protein-treated ob/ob mice (pair feeding). In lean mice, ob protein infusion significantly decreased body and fat-depot weights, while decreasing food intake to a much lesser extent than in ob/ob mice. Pair feeding of lean vehicle-treated mice to the intake of ob protein-treated mice did not reduce body fat-depot weights. The potent weight-, adipose-, and appetite-reducing effects exerted by the ob protein in ob protein-deficient mice (ob/ob) confirm hypotheses generated from early parabiotic studies that suggested the existence of a circulating satiety factor of adipose origin. Pair-feeding studies provide compelling evidence that the ob protein exerts adipose-reducing effects in excess of those induced by reductions in food intake.Recent epidemiologic studies have reported that more than one-third of U.S. adults 20 yr of age or older are overweight and that this prevalence increased by 8% over a 15-yr period (1). Obesity is associated with increased risk for several co-morbid conditions and diseases, including insulin resistance, non-insulin-dependent diabetes mellitus, cardiovascular disease, hypertension, hypertriglyceridemia, dyslipoproteinemia, and some forms of cancer (2, 3). The recent cloning and sequencing of the mouse ob gene and its human homologue (4) represent a significant step toward a better understanding of a possible biochemical cause of obesity.Parabiosis experiments performed >20 yr ago predicted that the genetically obese (ob/ob) mouse does not produce a satiety factor that regulates its food intake, whereas the diabetic (db/db) mouse produces, but does not respond to, a satiety factor (5, 6). Recent reports have demonstrated that daily injections of recombinant ob protein profoundly inhibit food intake and reduce body weight and fat in ob/ob but not in db/db mice (7-9), suggesting that the ob protein is such a satiety factor, as proposed in early cross-circulation studies. Although modest effects of daily injections of the ob protein on food intake and body weight were reported in lean mice, there was a significant reduction in body fat as assessed by carcass composition in one (8) but not in another (7) of these reports, despite equivalent decreases in body weight. To elucidate the activity of the ob protein, a comprehensive analysis of the effects of low-dose continuous infusions of the ob protein in lean and obese mice on body weight, food intake, and adipose-depot mass is presented here, including a comparison of the effects of ob protein treatment to those of pair feeding in lean and obese mice. The data suggest that aThe publication costs of this ar...

The involvement of platelets and the c-mpl receptor in the regulation of thrombopoietin (TPO) plasma concentrations and tissue mRNA levels was investigated in both normal mice and mice defective in c-mpl (c-mpl- /-). Although c-mpl-/- mice have fewer platelets and higher plasma TPO activity than normal mice, there was no increase in TPO mRNA levels as measured by an S1 nuclease protection assay. After the intravenous injection of 125I-TPO, specific uptake of radioactivity by the spleen and blood cells was present in the normal mice, but absent in the c-mpl- /- mice. Platelet-rich plasma (PRP) from normal mice was able to bind and internalize 125I-TPO, whereas PRP from c-mpl-/- mice lacked this ability. Analysis of 125I-TPO binding to normal PRP indicated that binding was specific and saturable, with an approximate affinity of 560 pmol/L and 220 receptors per platelet. PRP from normal mice was also able to degrade 125I-TPO into lower molecular weight fragments. After the intravenous injections, c-mpl-/- mice cleared a dose of 125I-TPO at a much slower rate than did normal mice. Injection of washed platelets from normal mice into c-mpl-/- mice resulted in a dramatic, but transient, decrease in plasma TPO levels. These data provide evidence that platelets regulate plasma TPO levels via binding to the c-mpl receptor on circulating platelets.

Recently, the ligand for c-mpl has been identified and cloned. Initial studies of this molecule indicate that it is the platelet regulatory factor, thrombopoietin (TPO). Previous work has indicated that c-mpl is expressed in very immature hematopoietic precursors and thus raised the possibility that TPO may act directly on the hematopoietic stem cell. Therefore, in these studies, we investigate the effects of TPO on hematopoietic stem cell populations isolated from the murine fetal liver and bone marrow. Cocultivation of stem cells with fetal liver stroma give rise to multilineage expansion of the stem cells but with little or no megakaryocytopoiesis. Addition of TPO to these cocultures gives significant megakaryocyte production. This production is enhanced in combination with Kit ligand or interleukin-3. The addition of TPO to stem cell suspension cultures produces a dynamic thrombopoietic system in which stem cells undergo differentiation to produce megakaryocytes and proplatelets. These experiments show that the megakaryocytopoietic and thrombopoietic activities of TPO are initiated at the level of an early progenitor cell or upon the hematopoietic stem cell.