ABSTRACT
Essentials Obesity is a potential risk factor for development of thrombotic thrombocytopenic purpura (TTP). Obese ADAMTS-13-deficient mice were triggered with von Willebrand factor (VWF). Depletion of hepatic and splenic macrophages protects against thrombocytopenia in this model. VWF enhances phagocytosis of platelets by macrophages, dose-dependently. SUMMARY: Background Thrombotic thrombocytopenic purpura (TTP) is caused by the absence of ADAMTS-13 activity. Thrombocytopenia is presumably related to the formation of microthrombi rich in von Willebrand factor (VWF) and platelets. Obesity may be a risk factor for TTP; it is associated with abundance of macrophages that may phagocytose platelets. Objectives To evaluate the role of obesity and ADAMTS-13 deficiency in TTP, and to establish whether macrophages contribute to thrombocytopenia. Methods Lean or obese ADAMTS-13-deficient (Adamts-13-/- ) and wild-type (WT) mice were injected with 250 U kg-1 of recombinant human VWF (rVWF), and TTP characteristics were evaluated 24 h later. In separate experiments, macrophages were depleted in the liver and spleen of lean and obese WT or Adamts-13-/- mice by injection of clodronate-liposomes, 48 h before injection of rVWF. Results Obese Adamts-13-/- mice had a lower platelet count than their lean counterparts, suggesting that they might be more susceptible to TTP development. Lean Adamts-13-/- mice triggered with a threshold dose of rVWF did not develop TTP, whereas typical TTP symptoms developed in obese Adamts-13-/- mice, including severe thrombocytopenia and higher lactate dehydrogenase (LDH) levels. Removal of hepatic and splenic macrophages by clodronate injection in obese Adamts-13-/- mice before treatment with rVWF preserved the platelet counts measured 24 h after the trigger. In vitro experiments with cultured macrophages confirmed a VWF dose-dependent increase of platelet phagocytosis. Conclusions Obese Adamts-13-/- mice are more susceptible to the induction of TTP-related thrombocytopenia than lean mice. Phagocytosis of platelets by macrophages contributes to thrombocytopenia after rVWF injection in this model.
Subject(s)
ADAMTS13 Protein/deficiency , Blood Platelets/drug effects , Clodronic Acid/pharmacology , Macrophages/drug effects , Obesity/drug therapy , Phagocytosis/drug effects , Purpura, Thrombotic Thrombocytopenic/prevention & control , Spleen/drug effects , ADAMTS13 Protein/genetics , Animals , Blood Platelets/metabolism , Cells, Cultured , Disease Models, Animal , Kupffer Cells/drug effects , Kupffer Cells/metabolism , Macrophages/metabolism , Male , Mice, 129 Strain , Mice, Inbred C57BL , Mice, Knockout , Obesity/blood , Obesity/complications , Purpura, Thrombotic Thrombocytopenic/blood , Purpura, Thrombotic Thrombocytopenic/etiology , Spleen/metabolism , Time Factors , von Willebrand FactorABSTRACT
BACKGROUND: Inhibition of angiogenesis may impair adipose tissue development. METHODS: The effect of fumagillin (a methionine aminopeptidase-2 inhibitor) on adipocyte differentiation and de novo adipogenesis was investigated in murine model systems. RESULTS: During in vitro differentiation of murine 3T3-F442A preadipocytes, administration of fumagillin (>/=1 muM) resulted in reduced expression of methionine aminopeptidase-2, and in enhanced differentiation rate. In vivo, de novo development of adipose tissue following injection of preadipocytes in nude mice kept on high fat diet was somewhat, but not significantly (p=0.06), reduced by administration of fumagillin (1 mg/kg/day during 4 weeks by oral gavage). This was not associated with effects on blood vessel size or density, whereas blood vessel density normalized to adipocyte density was enhanced upon fumagillin treatment. In vivo BrdU incorporation experiments did not reveal effects of fumagillin on cell proliferation in adipose tissues, and cellular apoptosis was also not affected. Treatment with fumagillin enhances in vitro differentiation of preadipocytes, but has only a minor effect on in vivo adipogenesis. GENERAL SIGNIFICANCE: These studies on in vitro and in vivo preadipcoyte differentiation thus do not support an anti-obesity effect of fumagillin as a result of effects on adipocyte differentiation.
Subject(s)
Adipocytes/cytology , Adipogenesis/drug effects , Cell Differentiation/drug effects , Cyclohexanes/pharmacology , Fatty Acids, Unsaturated/pharmacology , Sesquiterpenes/pharmacology , 3T3 Cells , Adipocytes/drug effects , Adipocytes/physiology , Animals , Body Weight/drug effects , Cell Division/drug effects , Energy Intake/drug effects , Humans , Male , Mice , Mice, Inbred BALB C , Mice, Nude , Obesity/prevention & controlABSTRACT
BACKGROUND: A functional role for several components of the fibrinolytic (plasminogen/plasmin) system in development of adipose tissue has been demonstrated. No information is available, however, on a potential role of plasminogen activator inhibitor-2 (PAI-2) in obesity. METHODS: In vitro, 3T3-F442A murine pre-adipocytes were differentiated into mature adipocytes. In vivo, 5-week-old male PAI-2-deficient (PAI-2(-/-)) mice and wild-type (WT) controls of the same genetic background (C57Bl/6) were kept on a high fat diet (HFD, caloric value of 20.1 kJ g(-1)) for 15 weeks. RESULTS: Semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) revealed expression of PAI-2 mRNA during in vitro differentiation of pre-adipocytes and in vivo in s.c. and gonadal (GON) adipose tissues of WT mice, where it was localized both in the stromal/vascular cell fraction and in adipocytes. During HFD feeding, food intake and body weight gain were comparable for WT and PAI-2(-/-) mice. Subcutaneous plus GON fat mass was, however, significantly lower in PAI-2(-/-) mice (3.15 +/- 0.21 vs. 3.91 +/- 0.18 g; P < 0.05). Immunohistochemical analysis of adipose tissues revealed significant adipocyte hypotrophy in s.c. fat of PAI-2(-/-) mice (about 25% reduction in size; P < 0.01). Blood vessel density, normalized to adipocyte number, was comparable in s.c. fat, but was lower (P < 0.05) in GON fat of PAI-2(-/-) mice. Adipose tissue-associated fibrinolytic activity was not affected by PAI-2 deficiency. CONCLUSION: PAI-2 promotes adipose tissue development in mice via a mechanism independent of its antifibrinolytic effect.
Subject(s)
Adipogenesis , Adipose Tissue/growth & development , Plasminogen Activator Inhibitor 2/deficiency , Plasminogen Activator Inhibitor 2/physiology , 3T3 Cells , Animals , Diet , Energy Intake , Fibrinolysis , Mice , Mice, Inbred C57BL , Mice, Knockout , Plasminogen Activator Inhibitor 2/analysisABSTRACT
OBJECTIVE: To substantiate a potential role of plasminogen activator inhibitor-1 (PAI-1) in adipogenesis, we have studied its effects on in vitro adipocyte differentiation and on in vivo adipose tissue formation. RESULTS: Our in vitro data do not support a functional role of PAI-1, as substantiated by our findings that: (i) inhibition of PAI-1 with a neutralizing antibody did not affect differentiation of 3T3-F442A preadipocytes; (ii) overexpression of murine PAI-1 in 3T3-F442A cells had no effect on differentiation; and (iii) differentiation of PAI-1-deficient murine embryonic fibroblasts into mature adipocytes was comparable to wild-type (WT) cells. Furthermore, our in vivo studies did not reveal an important role for PAI-1, as suggested by our findings that: (i) de novo fat pad formation in NUDE mice following injection of 3T3-F442A cells was not affected by a PAI-1 neutralizing antibody; and (ii) adipose tissue formation following combined injection of Matrigel and basic fibroblast growth factor was comparable in WT and in PAI-1 deficient mice. CONCLUSION: Taken together, these in vitro and in vivo studies in murine model systems do not support an important functional role of PAI-1 in adipogenesis.
