Tissue Uptake of Circulating Thrombopoietin Is Increased in Immune-Mediated Compared With Irradiated Thrombocytopenic Mice

Abstract

We have previously demonstrated a significant inverse correlation between circulating thrombopoietin (TPO) levels and peripheral platelet (PLT) counts in patients with thrombocytopenia secondary to megakaryocytic hypoplasia but not in patients with immune thrombocytopenic purpura (ITP; Chang et al, Blood 88:3354, 1996). To test the hypothesis that the differences in the circulating TPO levels in these two types of thrombocytopenia are caused by differences in the total capacity of Mpl receptor-mediated TPO clearance, thrombocytopenia was induced in female CD-1 mice either by sublethal irradiation (irradiated) or rabbit antimouse PLT serum (RAMPS) for 1 day (1 d RAMPS) and 5 days (5 d RAMPS). A well-characterized murine model of autoimmune thrombocytopenic purpura, male (NZW x BXSB) F1 mice (W/B F1), was also included in this study. All thrombocytopenic mice and their controls received trace amounts of 125I-recombinant murine TPO (125I-rmTPO) intravenously and were killed 3 hours postinjection. Blood cell-associated radioactivity was significantly decreased in all 4 groups of thrombocytopenic mice. Significantly increased plasma and decreased whole spleen-associated radioactivity was observed in the irradiated group compared with controls (P <.05). While a lesser but still significant increase in plasma and decrease in whole spleen-associated radioactivity was observed in the 1 d RAMPS mice (P <.05), there were no significant differences between the 5 d RAMPS nor the W/B F1 male mice compared with controls, although whole spleen-associated radioactivity was higher in the W/B F1 male. A significant inverse correlation of plasma and whole spleen-associated radioactivity was demonstrated in W/B F1 male mice (r = -.91, n = 6, P <.05). There was also a decrease in bone (femur)/blood-associated radioactivity in the irradiated group compared with controls (P <.05), but a significant increase in 1 d and 5 d RAMPS mice (P <.01). Furthermore, the 125I-rmTPO uptake capacity within the spleen and marrow of immune thrombocytopenic mice appeared to be associated with a higher megakaryocytic mass when tissue samples were examined by light microscopy. Internalization of 125I-rmTPO by megakaryocytes and PLTs in the spleens and marrows of ITP mice was also demonstrated directly using electron microscopic autoradiography. Labeled PLTs were also found within splenic macrophages. Additionally, the mean PLT volumes of RAMPS mice were significantly higher than those of the control and irradiated mice (P <.05), as was the bound 125I-rmTPO (cpm) per million PLT (P <.05). Finally, significantly decreased 125I-rmTPO degradation products were only found in the plasma of the irradiated mice compared with control animals (P <.05). These data suggest that the lack of Mpl+ cells in the mice with thrombocytopenia secondary to megakaryocytic hypoplasia (irradiated) results in decreased uptake and degradation of TPO and higher circulating TPO levels. Furthermore, these data also suggest that, after a brief TPO surge in response to immune thrombocytopenia (1 d RAMPS), the lack of an inverse correlation of circulating TPO with PLT counts during steady-state immune thrombocytopenic mice (5 d RAMPS + W/B F1 male) is due, at least in part, to its uptake and degradation by the high PLT turnover and increased mass of megakaryocytes.