Navegando por Palavras-chave "Plasminogen"
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- ItemSomente MetadadadosCorneal angiogenesis modulation by cysteine cathepsins: in vitro and in vivo studies(Elsevier B.V., 2015-05-01) Coppini, Larissa Pereira [UNIFESP]; Visniauskas, Bruna [UNIFESP]; Costa, Elaine F.; Filho, Milton N. [UNIFESP]; Rodrigues, Eduardo B. [UNIFESP]; Chagas, Jair R. [UNIFESP]; Farah, Michel E. [UNIFESP]; Barros, Nilana M. T. [UNIFESP]; Carmona, Adriana K. [UNIFESP]; Universidade Federal de São Paulo (UNIFESP); Univ Fed MaranhaoCorneal avascularization is essential for normal vision. Several antiangiogenic factors were identified in cornea such as endostatin and angiostatin. Cathepsin V, which is highly expressed in the cornea, can hydrolyze human plasminogen to release angiostatin fragments. Herein, we describe a detailed investigation of the expression profile of cathepsins B, L, S and V in the human cornea and the role of cysteine peptidases in modulating angiogenesis both in vitro and in vivo. We used various methodological tools for this purpose, including real-time PCR, SDS-PAGE, western blotting, catalytic activity assays, cellular assays and induction of corneal neovascularity in rabbit eyes. Human corneal enzymatic activity assays revealed the presence of cysteine proteases that were capable of processing endogenous corneal plasminogen to produce angiostatin-like fragments. Comparative real-time analysis of cathepsin B, L, S and V expression revealed that cathepsin V was the most highly expressed, followed by cathepsins L, B and S. However, cathepsin V depletion revealed that this enzyme is not the major cysteine protease responsible for plasminogen degradation under non-pathological conditions. Furthermore, western blotting analysis indicated that only cathepsins B and S were present in their enzymatically active forms. in vivo analysis of angiogenesis demonstrated that treatment with the cysteine peptidase inhibitor E64 caused a reduction in neovascularization. Taken together, our results show that human corneal cysteine proteases are critically involved in angiogenesis. (C) 2015 Elsevier B.V. All rights reserved.
- ItemAcesso aberto (Open Access)Estudo da internalização de macromoléculas em eritrócitos infectados pelo Plasmodium falciparum(Universidade Federal de São Paulo (UNIFESP), 2019-08-29) Maluf, Sarah El Chamy [UNIFESP]; Carmona, Adriana Karaoglanovic [UNIFESP]; Gazarini, Marcos Leoni [UNIFESP]; http://lattes.cnpq.br/2620389100131312; http://lattes.cnpq.br/2122863342403909; http://lattes.cnpq.br/3990204365027336; Universidade Federal de São Paulo (UNIFESP)The intraerythrocytic development of the malaria parasite Plasmodium falciparum is dependent on the uptake of essential nutrients from the host cell cytoplasm and blood plasma. It is widely recognized that it imports low molecular mass nutrients, such as polyols, amino acids, lipids, nucleosides, organic anions and cations from the plasma. However, although some studies show that P. falciparum is capable of importing macromolecules from the host, the route for internalization continues to be subject of debate between different groups, mainly due to the necessity of these macromolecules to pass through three membranes, namely the erythrocyte, parasitophorous vacuole and the parasite membrane itself. To better understand this process, this study provides new evidence to help elucidate the mechanism used by the parasite to internalize macromolecules with a focus on protein uptake. Using different experimentais approaches, we demonstrated that P. falciparum imports human plasminogen and the exogenous protein crotamine, at different stages of its intraeritrocitic development. In the infected erythrocyte, plasminogen is located in the parasite's cytosol, and is also present in the Maurer’s Cleft and tubular structures in the erythrocyte cytosol, which are described as being associated with the export of proteins. In addition, some interactions may be involved with the internalization of plasminogen, such as with Rabs proteins, kinesin, myosin, PfMC-2TM, Pf113 and heat shock proteins. There was a significant decrease in the uptake of plasminogen by the parasites after cytochalasin D treatment, suggesting a participation of the actin-myosin motor system in protein trafficking. Crotamine was selectively internalized by infected erythrocytes, and presented a colocalization with the parasite nucleus and part of the protein remains associated with the erythrocyte plasma membrane. The uptake was 2-deoxy-D-glucose sensitive, indicating that it is a mechanism dependent on energy via glycolysis. The results obtained here offer new elements involved in macromolecular uptake pathways in infected erythrocytes, which may constitute a potential target for the development of new drugs, in addition to identifying the permeability characteristics of the infected cell.
- ItemAcesso aberto (Open Access)Glycosaminoglycans affect the interaction of human plasma kallikrein with plasminogen, factor XII and inhibitors(Associação Brasileira de Divulgação Científica, 2003-08-01) Gozzo, Andrezza Justino [UNIFESP]; Nunes, V.a. [UNIFESP]; Nader, Helena Bonciani [UNIFESP]; Dietrich, Carl Peter [UNIFESP]; Carmona, Adriana Karaoglanovic [UNIFESP]; Sampaio, Misako Uemura [UNIFESP]; Sampaio, C.a.m. [UNIFESP]; Araujo, Mariana da Silva [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)Human plasma kallikrein, a serine proteinase, plays a key role in intrinsic blood clotting, in the kallikrein-kinin system, and in fibrinolysis. The proteolytic enzymes involved in these processes are usually controlled by specific inhibitors and may be influenced by several factors including glycosaminoglycans, as recently demonstrated by our group. The aim of the present study was to investigate the effect of glycosaminoglycans (30 to 250 µg/ml) on kallikrein activity on plasminogen and factor XII and on the inhibition of kallikrein by the plasma proteins C1-inhibitor and antithrombin. Almost all available glycosaminoglycans (heparin, heparan sulfate, bovine and tuna dermatan sulfate, chondroitin 4- and 6-sulfates) reduced (1.2 to 3.0 times) the catalytic efficiency of kallikrein (in a nanomolar range) on the hydrolysis of plasminogen (0.3 to 1.8 µM) and increased (1.9 to 7.7 times) the enzyme efficiency in factor XII (0.1 to 10 µM) activation. On the other hand, heparin, heparan sulfate, and bovine and tuna dermatan sulfate improved (1.2 to 3.4 times) kallikrein inhibition by antithrombin (1.4 µM), while chondroitin 4- and 6-sulfates reduced it (1.3 times). Heparin and heparan sulfate increased (1.4 times) the enzyme inhibition by the C1-inhibitor (150 nM).
- ItemAcesso aberto (Open Access)Human tissue kallikreins 3 and 5 can act as plasminogen activator releasing active plasmin(Elsevier B.V., 2013-04-12) Souza, Lucas R. de; Melo, Pollyana Maria Saud [UNIFESP]; Paschoalin, Thaysa [UNIFESP]; Carmona, Adriana Karaoglanovic [UNIFESP]; Kondo, Marcia Yuri [UNIFESP]; Hirata, Izaura Yoshico [UNIFESP]; Blaber, Michael; Tersariol, Ivarne Luis dos Santos [UNIFESP]; Takatsuka, Joyce; Juliano, Maria Aparecida [UNIFESP]; Juliano, Luiz [UNIFESP]; Gomes, Roseli A.; Puzer, Luciano; Universidade Federal do ABC (UFABC); Universidade Federal de São Paulo (UNIFESP); Florida State Univ; Univ Mogi das Cruzes; Univ Fed Triangulo MineiroHuman tissue kallikreins (KLKs) are a group of serine proteases found in many tissues and biological fluids and are differentially expressed in several specific pathologies. Here, we present evidences of the ability of these enzymes to activate plasminogen. Kallikreins 3 and 5 were able to induce plasmin activity after hydrolyzing plasminogen, and we also verified that plasminogen activation was potentiated in the presence of glycosaminoglycans compared with plasminogen activation by tPA. This finding can shed new light on the plasminogen/plasmin system and its involvement in tumor metastasis, in which kallikreins appear to be upregulated. (C) 2013 Elsevier Inc. All rights reserved.
- ItemSomente MetadadadosPlasmodium falciparum proteases hydrolyze plasminogen, generating angiostatin-like fragments(Elsevier B.V., 2014-01-01) Melo, Pollyana Maria Saud [UNIFESP]; Bagnaresi, Piero [UNIFESP]; Paschoalin, Thaysa [UNIFESP]; Hirata, Izaura Yoshico [UNIFESP]; Gazarini, Marcos Leoni [UNIFESP]; Carmona, Adriana Karaoglanovic [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)Malaria is a disease caused by Plasmodium parasites and remains one of the most prevalent and persistent maladies, affecting hundreds of millions of people. in the present work, we evaluated the capability of Plasmodium falciparum proteases to hydrolyze the multifunctional protein plasminogen, which is implicated in angiogenesis and coagulation processes by the generation of angiostatin and plasmin, respectively. Using fluorescence microscopy, we visualized the internalization of FITC-labeled plasminogen in erythrocytes infected by P. falciparum and showed that the parasites are able to hydrolyze the protein. the cleavage of plasminogen by the P. falciparum proteases was also observed by SDS-PAGE, followed by immunoblotting with anti-angiostatin antibody. N-terminal sequencing of the main generated fragments indicated that they are comprised in the five plasminogen kringle domains, suggesting as being angiostatin-like peptides. This assumption was reinforced by the demonstration that the products of plasminogen processing mimic angiostatin functions, including the capability to inhibit angiogenesis and to stimulate calcium response in endothelial cells in vitro. However, no plasmin activity was detected after plasminogen hydrolysis by P. falciparum. Nonetheless, exogenous tissue plasminogen activator (tPA) activated plasmin in infected erythrocytes, suggesting that the uptake of plasminogen by P. falciparum may be modulated by the vertebrate host. Taken together, the data presented here provide evidence for the processing of host plasminogen by malaria parasites to generate active fragments that may modulate host physiology events during malaria infection. (C) 2014 Elsevier B.V. All rights reserved.