Rute Moura

Thrombospondin-1 Deficiency Accelerates Atherosclerotic Plaque Maturation in ApoE-/- Mice

Thrombospondin (TSP)1 is implicated in various inflammatory processes, but its role in atherosclerotic plaque formation and progression is unclear. Therefore, the development of atherosclerosis was compared in ApoE−/− and Tsp1−/−ApoE−/− mice kept on a normocholesterolemic diet. At 6 months, morphometric analysis of the aortic root of both mouse genotypes showed comparable lesion areas. Even when plaque burden increased ≈5-fold in ApoE−/− and 10-fold in Tsp1−/−ApoE−/− mice, during the subsequent 3 months, total plaque areas were comparable at 9 months. In contrast, plaque composition differed substantially between genotypes: smooth muscle cell areas, mostly located in the fibrous cap of ApoE−/− plaques, both at 6 and 9 months, were 3-fold smaller in Tsp1−/−ApoE−/− plaques, which, in addition, were also more fibrotic. Moreover, inflammation by macrophages was twice as high in Tsp1−/−ApoE−/− plaques. This correlated with a 30-fold elevated incidence of elastic lamina degradation, with matrix metalloproteinase-9 accumulation, underneath plaques and manifestation of ectasia, exclusively in Tsp1−/−ApoE−/− mice. At 9 months, the necrotic core was 1.4-fold larger and 4-fold higher numbers of undigested disintegrated apoptotic cells were found in Tsp1−/−ApoE−/− plaques. Phagocytosis of platelets by cultured Tsp1−/− macrophages revealed the instrumental role of TSP1 in phagocytosis, corroborating the defective intraplaque phagocytosis of apoptotic cells. Hence, the altered smooth muscle cell phenotype in Tsp1−/−ApoE−/− mice has limited quantitative impact on atherosclerosis, but defective TSP1-mediated phagocytosis enhanced plaque necrotic core formation, accelerating inflammation and macrophage-induced elastin degradation by metalloproteinases, speeding up plaque maturation and vessel wall degeneration.

The evolving role of thrombospondin-1 in hemostasis and vascular biology

Abstract.Thrombospondin-1 (TSP1) is a multi-domain, multi-functional glycoprotein synthesized by many cells. Matricellular TSP1 modulates cell adhesion and proliferation. TSP1 is involved in angiogenesis, inflammation, wound healing and cancer. As a major platelet protein, for a long time it was postulated to control hemostasis via platelet aggregate stabilization. However, these in vitro findings have been questioned in the absence of corroborating clinical data and of obvious hemostatic defects in TSP1 gene-deficient mice.Yet, the past few years have provided indices to implicate TSP1 in hemostasis. In clinical studies, a correlation exists between a welldefined TSP1 polymorphism and a significant risk of myocardial infarction.At the same time, recent in vivo animal model data imply TSP1 in the multimer size control of von Willebrand factor, in smooth muscle cell regulation and in vascular perfusion. These findings shed new light on the role of TSP1 in hemostasis and prothrombotic vascular pathologies. (Part of a Multi-author Review)

Membrane-anchored uPAR regulates the proliferation, marrow pool size, engraftment, and mobilization of mouse hematopoietic stem/progenitor cells

The mechanisms of BM hematopoietic stem/progenitor cell (HSPC) adhesion, engraftment, and mobilization remain incompletely identified. Here, using WT and transgenic mice, we have shown that membrane-anchored plasminogen activator, urokinase receptor (MuPAR) marks a subset of HSPCs and promotes the preservation of the size of this pool of cells in the BM. Loss or inhibition of MuPAR increased HSPC proliferation and impaired their homing, engraftment, and adhesion to the BM microenvironment. During mobilization, MuPAR was inactivated by plasmin via proteolytic cleavage. Cell-autonomous loss of the gene encoding MuPAR also impaired long-term engraftment and multilineage repopulation in primary and secondary recipient mice. These findings identify MuPAR and plasmin as regulators of the proliferation, marrow pool size, homing, engraftment, and mobilization of HSPCs and possibly also of HSCs.

The Fragile X Protein binds mRNAs involved in cancer progression and modulates metastasis formation

The role of the fragile X mental retardation protein (FMRP) is well established in brain, where its absence leads to the fragile X syndrome (FXS). FMRP is almost ubiquitously expressed, suggesting that, in addition to its effects in brain, it may have fundamental roles in other organs. There is evidence that FMRP expression can be linked to cancer. FMR1 mRNA, encoding FMRP, is overexpressed in hepatocellular carcinoma cells. A decreased risk of cancer has been reported in patients with FXS while a patient‐case with FXS showed an unusual decrease of tumour brain invasiveness. However, a role for FMRP in regulating cancer biology, if any, remains unknown. We show here that FMRP and FMR1 mRNA levels correlate with prognostic indicators of aggressive breast cancer, lung metastases probability and triple negative breast cancer (TNBC). We establish that FMRP overexpression in murine breast primary tumours enhances lung metastasis while its reduction has the opposite effect regulating cell spreading and invasion. FMRP binds mRNAs involved in epithelial mesenchymal transition (EMT) and invasion including E‐cadherin and Vimentin mRNAs, hallmarks of EMT and cancer progression.

Thrombospondins: from structure to therapeutics

Abstract.Thrombospondin-1 (TSP1) is a multi-domain, multi-functional glycoprotein synthesized by many cells. Matricellular TSP1 modulates cell adhesion and proliferation. TSP1 is involved in angiogenesis, inflammation, wound healing and cancer. As a major platelet protein, for a long time it was postulated to control hemostasis via platelet aggregate stabilization. However, these in vitro findings have been questioned in the absence of corroborating clinical data and of obvious hemostatic defects in TSP1 gene-deficient mice.Yet, the past few years have provided indices to implicate TSP1 in hemostasis. In clinical studies, a correlation exists between a welldefined TSP1 polymorphism and a significant risk of myocardial infarction.At the same time, recent in vivo animal model data imply TSP1 in the multimer size control of von Willebrand factor, in smooth muscle cell regulation and in vascular perfusion. These findings shed new light on the role of TSP1 in hemostasis and prothrombotic vascular pathologies. (Part of a Multi-author Review)

Thrombospondin-1 Activates Medial Smooth Muscle Cells and Triggers Neointima Formation Upon Mouse Carotid Artery Ligation

Objective— Thrombospondin-1 (TSP1) is described as a positive regulator of vascular smooth muscle growth in cell culture. However, insight into the in vivo effects of TSP1 on smooth muscle cell (SMC) function is lacking. Methods and Results— We analyzed wild-type (WT) and TSP1-deficient (Tsp1−/−) mice in a carotid artery ligation model, in which neointimal lesions form without overt mechanical damage to the endothelium. On ligation, the expression of TSP1 increased strongly in the matrix of neointima and adventitia. In the early phase after ligation (day 3 to 7), activation, proliferation, and migration of medial SMCs were delayed and impaired in Tsp1−/− mice, in parallel with defective upregulation of metalloproteinase (MMP)-2 activity. As a result, Tsp1−/− arteries developed smaller neointimal lesions, a thicker media but comparably attenuated patency as in WT arteries, 28 days after ligation. Furthermore, medial and neointimal SMCs in Tsp1−/− mice produced more collagen, more osteopontin, and displayed weaker smooth muscle actin staining than WT SMCs, indicative of a modified SMC phenotype in Tsp1−/− mice. Conclusions— Arterial SMC activation in the absence of TSP1 is delayed and dysregulated, reducing neointima formation, on mild vascular injury.

Fibrinolysis-independent role of plasmin and its activators in the haematopoietic recovery after myeloablation.

Proteinases have been implicated in the mobilization of haematopoietic progenitor cells (HPCs) from the bone marrow (BM). Here, we report the involvement of the plasminogen (Plg) system in the haematopoietic recovery following chemotherapy. By using gene‐deficient mice, we found that plasmin and its activators tPA and uPA play a role in the haematopoietic recovery upon delivery of the cytotoxic agent 5‐fluoro‐uracil (5‐FU). The impaired haematopoietic recovery of Plg‐deficient (Plg−/−) mice after 5‐FU was not rescued by depletion of fibrinogen, indicating that it was not due to defective fibrinolysis. Instead, loss of Plg impaired breakdown of fibronectin, VCAM‐1 and laminin‐BM matrix proteins involved in adhesion of HPCs to their BM microenvironment and in transendothelial migration of HPCs. These findings provide novel insights in how plasmin regulates haematopoietic recovery upon cytotoxic myeloablation.

Thrombospondins: from structure to therapeutics: The evolving role of thrombospondin-1 in hemostasis and vascular biology

Abstract.Thrombospondin-1 (TSP1) is a multi-domain, multi-functional glycoprotein synthesized by many cells. Matricellular TSP1 modulates cell adhesion and proliferation. TSP1 is involved in angiogenesis, inflammation, wound healing and cancer. As a major platelet protein, for a long time it was postulated to control hemostasis via platelet aggregate stabilization. However, these in vitro findings have been questioned in the absence of corroborating clinical data and of obvious hemostatic defects in TSP1 gene-deficient mice.Yet, the past few years have provided indices to implicate TSP1 in hemostasis. In clinical studies, a correlation exists between a welldefined TSP1 polymorphism and a significant risk of myocardial infarction.At the same time, recent in vivo animal model data imply TSP1 in the multimer size control of von Willebrand factor, in smooth muscle cell regulation and in vascular perfusion. These findings shed new light on the role of TSP1 in hemostasis and prothrombotic vascular pathologies. (Part of a Multi-author Review)

The Fragile X Protein binds mRNAs involved in cancer progression and modulates metastasis formation [EMBO Mol Med, 5, (2013) 1523-1536, DOI: 10.1002/emmm.201302847]

The Fragile X Protein binds mRNAs involved in cancer progression and modulates metastasis formation Rossella Lucá1,2y, Michele Averna1,2y, Francesca Zalfa3,4y, Manuela Vecchi5,6y, Fabrizio Bianchi, Giorgio La Fata, Franca Del Nonno, Roberta Nardacci, Marco Bianchi, Paolo Nuciforo, Sebastian Munck, Paola Parrella, Rute Moura, Emanuela Signori, Robert Alston, Anna Kuchnio, Maria Giulia Farace, Vito Michele Fazio, Mauro Piacentini, Bart De Strooper, Tilmann Achsel, Giovanni Neri, Patrick Neven, D. Gareth Evans, Peter Carmeliet, Massimiliano Mazzone, Claudia Bagni*