RESUMO
Breast cancer is characterized by a hypoxic microenvironment inside the tumor mass, contributing to cell metastatic behavior. Hypoxia induces the expression of hypoxia-inducible factor (HIF-1α), a transcription factor for genes involved in angiogenesis and metastatic behavior, including the vascular endothelial growth factor (VEGF), matrix metalloproteinases (MMPs), and integrins. Integrin receptors play a key role in cell adhesion and migration, being considered targets for metastasis prevention. We investigated the migratory behavior of hypoxia-cultured triple-negative breast cancer cells (TNBC) and endothelial cells (HUVEC) upon αvß3 integrin blocking with DisBa-01, an RGD disintegrin with high affinity to this integrin. Boyden chamber, HUVEC transmigration, and wound healing assays in the presence of DisBa-01 were performed in hypoxic conditions. DisBa-01 produced similar effects in the two oxygen conditions in the Boyden chamber and transmigration assays. In the wound healing assay, hypoxia abolished DisBa-01's inhibitory effect on cell motility and decreased the MMP-9 activity of conditioned media. These results indicate that αvß3 integrin function in cell motility depends on the assay and oxygen levels, and higher inhibitor concentrations may be necessary to achieve the same inhibitory effect as in normoxia. These versatile responses add more complexity to the role of the αvß3 integrin during tumor progression.
Assuntos
Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Células Endoteliais/metabolismo , Integrina alfaVbeta3/antagonistas & inibidores , Integrina alfaVbeta3/metabolismo , Hipóxia Tumoral , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Venenos de Crotalídeos/farmacologia , Meios de Cultivo Condicionados/farmacologia , Desintegrinas/farmacologia , Células Endoteliais/patologia , Feminino , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Metaloproteinases da Matriz/metabolismo , Neovascularização Fisiológica/efeitos dos fármacos , Oxigênio , Subunidades Proteicas/metabolismo , Hipóxia Tumoral/efeitos dos fármacosRESUMO
Disintegrins comprise a family of small proteins that bind to and alter the physiological function of integrins, especially integrins that mediate platelet aggregation in blood. Here, we report a lysine-glycine-aspartic acid (KGD) disintegrin-like motif present in a 15-amino acid residue peptide identified in a cDNA library of the amphibian Hypsiboas punctatus skin. The original peptide sequence was used as a template from which five new analogs were designed, chemically synthesized by solid phase, and tested for disintegrin activity and tridimensional structural studies using NMR spectroscopy. The original amphibian peptide had no effect on integrin-mediated responses. Nevertheless, derived peptide analogs inhibited integrin-mediated platelet function, including platelet spreading on fibrinogen.
Assuntos
Desintegrinas , Peptídeos , Anfíbios/genética , Anfíbios/metabolismo , Animais , DNA Complementar/genética , Desintegrinas/química , Desintegrinas/genética , Desintegrinas/farmacologia , Peptídeos/química , Peptídeos/genética , Peptídeos/farmacologia , Agregação Plaquetária/fisiologiaRESUMO
Jararhagin-C (Jar-C) is a disintegrin-like protein, isolated from the venom of B. jararaca, with affinity for α2ß1 integrin and the ability to incite processes such as angiogenesis and collagen deposition in vivo. Thus, we raised the hypothesis that this protein could be used as a therapeutic strategy for stimulating the healing of excisional wounds in mice. Four wounds were made on the back of Swiss mice, treated with daily intradermal injections of PBS (control group) or Jar-C (200 ng). Ten animals from each experimental group were euthanized and the tissue from the wounds and skin around them were collected for further biochemical, histological and molecular analysis. Wounds treated with Jar-C showed a faster closure rate, accompanied by a reduction in neutrophil infiltrate (MPO), pro-inflammatory cytokine levels (TNF, CXCL1 and CCL2) and an accumulation of macrophages in the analyzed tissues. It was also observed a greater expression of genes associated with the phenotype of alternatively activated macrophages (M2). Concomitantly, the administration of Jar-C holds an angiogenic potential, increasing the density of blood vessels and the synthesis of pro-angiogenic cytokines (VEGF and FGF). We also observed an increase in collagen deposition, accompanied by higher levels of the pro-fibrogenic cytokine TGF-ß1. Our data suggests Jar-C stimulates wound healing through stimulation of M2-like macrophage, angiogenesis and collagen deposition. Jar-C may be explored as a therapeutic strategy for wound healing, including the treatment of chronic wounds, where processes such as inflammation, angiogenesis and the deposition / remodeling of the matrix constituents are unregulated.
Assuntos
Colágeno , Venenos de Crotalídeos , Desintegrinas , Neovascularização Fisiológica , Cicatrização , Animais , Humanos , Camundongos , Sobrevivência Celular/efeitos dos fármacos , Colágeno/metabolismo , Venenos de Crotalídeos/química , Citocinas/genética , Citocinas/metabolismo , Desintegrinas/química , Desintegrinas/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana , Leucócitos/efeitos dos fármacos , Leucócitos/fisiologia , Macrófagos , Neovascularização Fisiológica/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Cicatrização/efeitos dos fármacos , Veneno de Bothrops jararacaRESUMO
Disintegrins are low molecular weight cysteine-rich proteins (4-14 kDa) that are isolated mainly from viperid snake venom. Due to their potential as lead compounds for binding and blocking integrin receptors, snake venom disintegrins have become one of the most studied venom protein families. The aim of this study was to obtain disintegrins from C. totonacus venom and evaluate their capability to bind and block integrin receptors. The C. totonacus disintegrin fraction (totonacin) represents two disintegrin isoforms obtained from C. totonacus venom. These disintegrins showed extracellular-matrix (ECM) protein adhesion and migration inhibitory effects on MDA-MB-231 and HMEC-1 cells. Totonacin (3 µM) inhibited MDA-MB-231 cell adhesion to the ECM proteins, fibronectin, vitronectin, and laminin by 31.2, 44.0, and 32.1, respectively. Adhesion inhibition to fibronectin, vitronectin, and laminin observed on HMEC-1 cells was 42.8, 60.8, and 51%, respectively. In addition, totonacin (3 µM) significantly inhibited MDA-MB-231 and HMEC-1 cell migration (41.4 and 48.3%, respectively). Totonacin showed more potent cell adhesion inhibitory activity toward vitronectin in both cell lines. These results suggest a major affinity of totonacin toward αVß3, α8ß1, αVß5, αVß1, and αIIbß3 integrins. In addition, the inhibitory effect observed on MDA-MB-231 and HMEC-1 cell migration reinforces the evidence of an interaction between these disintegrins and αVß3 integrin, which plays a key role in migration and angiogenesis.
Assuntos
Venenos de Crotalídeos/química , Desintegrinas/farmacologia , Proteínas de Répteis/farmacologia , Animais , Adesão Celular/efeitos dos fármacos , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Crotalus , Desintegrinas/isolamento & purificação , Humanos , Proteínas de Répteis/isolamento & purificação , Cicatrização/efeitos dos fármacosRESUMO
BACKGROUND: Disintegrins from snake venoms bind with high specificity cell surface integrins, which are important pharmacological targets associated with cancer development and progression. OBJECTIVE: In this study, we isolated a disintegrin from the Porthidium lansbergii lansbergii venom and evaluated its antitumoral effects on breast cancer cells. METHODS: The isolation of the disintegrin was performed on RP-HPLC and the inhibition of platelet aggregation was evaluated on human platelet-rich plasma. The inhibition of cell adhesion was also evaluated in vitro on cultures of cell lines by the MTT method as well as the inhibition of breast cancer cell migration by the wound healing assay. The binding of the disintegrin to integrin subunits was verified by flow cytometry and confocal microscopy. Finally, inhibition of angiogenesis was assessed in vitro on HUVEC cells and the concentration of VEGF was measured in the cellular supernatants. RESULTS: The disintegrin, named Lansbermin-I, is a low molecular weight protein (< 10 kDa) that includes an RGD on its sequence identified previously. Lansbermin-I showed potent inhibition of ADP and collagen-induced platelet aggregation on human plasma and also displayed inhibitory effects on the adhesion and migration of breast cancer MCF7 and MDA-MB 231cell lines, without affecting nontumorigenic breast MCF-10A and lung BEAS cells. Additionally, Lansbermin-I prevented MCF7 cells to adhere to fibronectin and collagen, and also inhibited in vitro angiogenesis on human endothelial HUVEC cells. CONCLUSION: Our results display the first report on the antitumor and anti-metastatic effects of an RGDdisintegrin isolated from a Porthidium snake venom by possibly interfering with α2 and/or ß1-containing integrins. Thus, Lansbermin-I could be an attractive model to elucidate the role of disintegrins against breast cancer development.
Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Venenos de Crotalídeos/farmacologia , Desintegrinas/farmacologia , Animais , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Adesão Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Células Cultivadas , Venenos de Crotalídeos/química , Venenos de Crotalídeos/isolamento & purificação , Desintegrinas/química , Desintegrinas/isolamento & purificação , Relação Dose-Resposta a Droga , Feminino , Humanos , Integrinas/análise , Integrinas/metabolismo , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Relação Estrutura-Atividade , Viperidae , Cicatrização/efeitos dos fármacosRESUMO
Alternagin-C (ALT-C), a disintegrin-like protein obtained from the venom of Bothrops alternatus, is able to modulate cellular behaviors such as adhesion, migration and proliferation, as well as the production of various growth factors via α2ß1 integrin, important processes during inflammation, angiogenesis and fibrogenesis, which although appear as distinct events, act concomitantly in several chronic inflammatory diseases. Our objective was to investigate the effects of ALT-C on components of the sponge-induced inflammatory response in balb/c mice. The polyester-polyurethane sponges were implanted in mice's subcutaneous layer of the dorsal region and daily injected with saline (control group) or ALT-C (10, 100 or 1000â¯ng). Nine days after implantation the implants were removed and processed. ALT-C inhibited the inflammatory response, observed through mast cell reduction, NAG-activity and also by the inhibition of TNF-α, CXCL-1 and CCL2/JE/MCP-1 cytokines. ALT-C was also able to reduce hemoglobin content, number of vessels and the concentrations of VEGF and FGF cytokines. Finally, at its highest dose (1000â¯ng), ALT-C increased all evaluated markers associated with fibrogenesis (collagen production and TGF-ß1 levels). All these factors reveal that ALT-C is a strong candidate to be exploited in the development of anti-inflammatory and anti-angiogenic therapies in chronic inflammatory processes.
Assuntos
Bothrops/metabolismo , Colágeno/metabolismo , Venenos de Crotalídeos/farmacologia , Desintegrinas/farmacologia , Inflamação/tratamento farmacológico , Neovascularização Patológica/tratamento farmacológico , Animais , Hemoglobinas/metabolismo , Inflamação/metabolismo , Masculino , Mastócitos/efeitos dos fármacos , Mastócitos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Neovascularização Patológica/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
BACKGROUND: Integrins mediate cell adhesion, migration, and survival by connecting the intracellular machinery with the surrounding extracellular matrix. Previous studies demonstrated the interaction between αvß3 integrin and VEGF type 2 receptor (VEGFR2) in VEGF-induced angiogenesis. DisBa-01, a recombinant His-tag fusion, RGD-disintegrin from Bothrops alternatus snake venom, binds to αvß3 integrin with nanomolar affinity blocking cell adhesion to the extracellular matrix. Here we present in vitro evidence of a direct interference of DisBa-01 with αvß3/VEGFR2 cross-talk and its downstream pathways. METHODS: Human umbilical vein (HUVECs) were cultured in plates coated with fibronectin (FN) or vitronectin (VN) and tested for migration, invasion and proliferation assays in the presence of VEGF, DisBa-01 (1000 nM) or VEGF and DisBa-01 simultaneously. Phosphorylation of αvß3/VEGFR2 receptors and the activation of intracellular signaling pathways were analyzed by western blotting. Morphological alterations were observed and quantified by fluorescence confocal microscopy. RESULTS: DisBa-01 treatment of endothelial cells inhibited critical steps of VEGF-mediated angiogenesis such as migration, invasion and tubulogenesis. The blockage of αvß3/VEGFR2 cross-talk by this disintegrin decreases protein expression and phosphorylation of VEGFR2 and ß3 integrin subunit, regulates FAK/SrC/Paxillin downstream signals, and inhibits ERK1/2 and PI3K pathways. These events result in actin re-organization and inhibition of HUVEC migration and adhesion. Labelled-DisBa-01 colocalizes with αvß3 integrin and VEGFR2 in treated cells. CONCLUSIONS: Disintegrin inhibition of αvß3 integrin blocks VEGFR2 signalling, even in the presence of VEGF, which impairs the angiogenic mechanism. These results improve our understanding concerning the mechanisms of pharmacological inhibition of angiogenesis.
Assuntos
Movimento Celular/efeitos dos fármacos , Venenos de Crotalídeos/farmacologia , Desintegrinas/farmacologia , Células Endoteliais da Veia Umbilical Humana , Integrina alfaVbeta3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Adesão Celular , Células Cultivadas , Quinase 1 de Adesão Focal/metabolismo , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Paxilina/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Quinases da Família src/metabolismoRESUMO
Disintegrins are non-enzymatic proteins that interfere on cell-cell interactions and signal transduction, contributing to the toxicity of snake venoms and play an essential role in envenomations. Most of their pharmacological and toxic effects are the result of the interaction of these molecules with cell surface ligands, which has been widely described and studied. These proteins may act on platelets, leading to hemorrhage, and may also induce apoptosis and cytotoxicity, which highlights a high pharmacological potential for the development of thrombolytic and antitumor agents. Additionally, these molecules interfere with the functions of integrins by altering various cellular processes such as migration, adhesion and proliferation. This review gathers information on functional characteristics of disintegrins isolated from snake venoms, emphasizing a comprehensive view of the possibility of direct use of these molecules in the development of new drugs, or even indirectly as structural models.
Assuntos
Desintegrinas/farmacologia , Integrinas/metabolismo , Venenos de Serpentes/metabolismo , Animais , Plaquetas/citologia , Plaquetas/efeitos dos fármacos , Adesão Celular/efeitos dos fármacos , Comunicação Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Humanos , Transdução de Sinais/efeitos dos fármacosRESUMO
Disintegrins are a family of small cysteine-rich peptides, found in a wide variety of snake venoms of different phylogenetic origin. These peptides selectively bind to integrins, which are heterodimeric adhesion receptors that play a fundamental role in the regulation of many physiological and pathological processes, such as hemostasis and tumor metastasis. Most disintegrins interact with integrins through the RGD (Arg-Gly-Asp) sequence loop, resulting in an active site that modulates the integrin activity. Some variations in the tripeptide sequence and the variability in its neighborhood result in a different specificity or affinity toward integrin receptors from platelets, tumor cells or neutrophils. Recombinant forms of these proteins are obtained mainly through Escherichia coli, which is the most common host used for heterologous expression. Advances in the study of the structure-activity relationship and importance of some regions of the molecule, especially the hairpin loop and the C-terminus, rely on approaches such as site-directed mutagenesis and the design and expression of chimeric peptides. This review provides highlights of the biological relevance and contribution of recombinant disintegrins to the understanding of their binding specificity, biological activities and therapeutic potential. The biological and pharmacological relevance on the newest discoveries about this family of integrin-binding proteins are discussed.
Assuntos
Desintegrinas , Animais , Pesquisa Biomédica , Desintegrinas/química , Desintegrinas/farmacologia , Desintegrinas/fisiologia , Humanos , Proteínas Recombinantes/farmacologia , Venenos de Serpentes/químicaRESUMO
Metastasis involves the migration of cancer cells from a primary tumor to invade and establish secondary tumors in distant organs, and it is the main cause for cancer-related deaths. Currently, the conventional cytostatic drugs target the proliferation of malignant cells, being ineffective in metastatic disease. This highlights the need to find new anti-metastatic drugs. Toxins isolated from snake venoms are a natural source of potentially useful molecular scaffolds to obtain agents with anti-migratory and anti-invasive effects in cancer cells. While there is greater evidence concerning the mechanisms of cell death induction of several snake toxin classes on cancer cells; only a reduced number of toxin classes have been reported on (i.e., disintegrins/disintegrin-like proteins, C-type lectin-like proteins, C-type lectins, serinproteases, cardiotoxins, snake venom cystatins) as inhibitors of adhesion, migration, and invasion of cancer cells. Here, we discuss the anti-metastatic mechanisms of snake toxins, distinguishing three targets, which involve (1) inhibition of extracellular matrix components-dependent adhesion and migration, (2) inhibition of epithelial-mesenchymal transition, and (3) inhibition of migration by alterations in the actin/cytoskeleton network.
Assuntos
Desintegrinas/farmacologia , Metástase Neoplásica/patologia , Metástase Neoplásica/prevenção & controle , Venenos de Serpentes/farmacologia , Animais , Adesão Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Desintegrinas/isolamento & purificação , Transição Epitelial-Mesenquimal/efeitos dos fármacos , HumanosRESUMO
Alternagin-C (ALT-C) is a disintegrin-like protein isolated from Rhinocerophis alternatus snake venom, which induces endothelial cell proliferation and angiogenesis. The aim of this study was to evaluate the systemic effects of a single dose of alternagin-C (0.5 mg·kg-1, via intra-arterial) on oxidative stress biomarkers, histological alterations, vascular endothelial growth factor (VEGF) production, and the degree of vascularization in the liver of the freshwater fish traíra, Hoplias malabaricus, seven days after the initiation of therapy. ALT-C treatment increased VEGF levels and hepatic angiogenesis. ALT-C also enhanced hepatic antioxidant enzymes activities such as superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase, decreasing the basal oxidative damage to lipids and proteins in the fish liver. These results indicate that ALT-C improved hepatic tissue and may play a crucial role in tissue regeneration mechanisms.
Assuntos
Caraciformes/metabolismo , Desintegrinas/farmacologia , Fígado/efeitos dos fármacos , Neovascularização Fisiológica/efeitos dos fármacos , Animais , Vasos Sanguíneos/efeitos dos fármacos , Bothrops , Catalase/metabolismo , Caraciformes/fisiologia , Venenos de Crotalídeos , Glutationa/metabolismo , Glutationa Peroxidase/metabolismo , Glutationa Redutase/metabolismo , Glutationa Transferase/metabolismo , Fígado/irrigação sanguínea , Fígado/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Superóxido Dismutase/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
PURPOSE: To investigate the changes induced by DisBa-01 on repair of wound healing after induced incisional hernia (IH) in rats. METHODS: Thirty two male albino rats were submitted to IH and divided into four experimental groups: G1, placebo control; G2, DisBa-01-treated; G3, anti-αvß3 antibodies-treated and G4, anti-α2 antibodies-treated. Histological, biochemical and extracellular matrix remodeling analysis of abdominal wall were evaluated. RESULTS: After 14 days, 100% of the G2 did not present hernia, and the hernia ring was closed by a thin membrane. In contrast, all groups maintained incisional hernia. DisBa-01 also increased the number macrophages and fibroblasts and induced the formation of new vessels. Additionally, MMP-2 was strongly activated only in G2 (p<0.05). Anti- αvß3-integrin antibodies produced similar results than DisBa-01 but not anti-α2 integrin blocking antibodies. CONCLUSION: DisBa-01 has an important role in the control of wound healing and the blocking of this integrin may be an interesting therapeutically strategy in incisional hernia.
Assuntos
Desintegrinas/farmacologia , Hérnia Ventral/patologia , Integrina alfaVbeta3/antagonistas & inibidores , Inibidores da Agregação Plaquetária/farmacologia , Cicatrização/efeitos dos fármacos , Parede Abdominal/patologia , Animais , Colágeno/análise , Colágeno/efeitos dos fármacos , Modelos Animais de Doenças , Fibroblastos/efeitos dos fármacos , Hérnia Ventral/tratamento farmacológico , Hérnia Ventral/cirurgia , Masculino , Metaloproteinase 2 da Matriz/análise , Metaloproteinase 2 da Matriz/fisiologia , Distribuição Aleatória , Ratos Wistar , Reprodutibilidade dos Testes , Fatores de Tempo , Resultado do TratamentoRESUMO
PURPOSE: To investigate the changes induced by DisBa-01 on repair of wound healing after induced incisional hernia (IH) in rats. METHODS: Thirty two male albino rats were submitted to IH and divided into four experimental groups: G1, placebo control; G2, DisBa-01-treated; G3, anti-αvβ3 antibodies-treated and G4, anti-α2 antibodies-treated. Histological, biochemical and extracellular matrix remodeling analysis of abdominal wall were evaluated. RESULTS: After 14 days, 100% of the G2 did not present hernia, and the hernia ring was closed by a thin membrane. In contrast, all groups maintained incisional hernia. DisBa-01 also increased the number macrophages and fibroblasts and induced the formation of new vessels. Additionally, MMP-2 was strongly activated only in G2 (p<0.05). Anti- αvβ3-integrin antibodies produced similar results than DisBa-01 but not anti-α2 integrin blocking antibodies. CONCLUSION: DisBa-01 has an important role in the control of wound healing and the blocking of this integrin may be an interesting therapeutically strategy in incisional hernia. .
Assuntos
Animais , Masculino , Desintegrinas/farmacologia , Hérnia Ventral/patologia , /antagonistas & inibidores , Inibidores da Agregação Plaquetária/farmacologia , Cicatrização/efeitos dos fármacos , Parede Abdominal/patologia , Colágeno/análise , Colágeno/efeitos dos fármacos , Modelos Animais de Doenças , Fibroblastos/efeitos dos fármacos , Hérnia Ventral/tratamento farmacológico , Hérnia Ventral/cirurgia , /análise , /fisiologia , Distribuição Aleatória , Ratos Wistar , Reprodutibilidade dos Testes , Fatores de Tempo , Resultado do TratamentoRESUMO
Integrins are involved in a number of physio-pathological processes including wound healing, chronic inflammation and neoplasias. Blocking its activity is potentially of therapeutic value in these conditions. We investigated whether DisBa-01, a recombinant His-tag RGD-disintegrin from Bothrops alternatus snake venom, could modulate key events (inflammatory cell recruitment/activation, neovascularization and extracellular matrix deposition) of the proliferative fibrovascular tissue induced by polyether polyurethane sponge implants in mice. The hemoglobin content (µg/mg wet tissue), blood flow measurements (laser Doppler perfusion imaging) and number of vessels in the implants, used as indices of vascularization, showed that the disintegrin dose-dependently reduced angiogenesis in the implants relative to the Saline-treated group. DisBa-01 inhibited neutrophil and macrophage content as determined by the myeloperoxidase (MPO) and N-acetyl-ß-D-glucosaminidase (NAG) activities, respectively. Similarly, down regulation of the fibrogenic component studied (collagen deposition) was observed in DisBa-01-treated implants. VEGF, bFGF, TNF-α, CXCL1 and CCL2 levels were also decreased by the disintegrin. The inhibitory effect of this αvß3-blocking disintegrin on the angiogenic, inflammatory, and fibrogenic components of the fibrovascular tissue induced by the synthetic matrix extends the range of DisBa-01 actions and may indicate its therapeutic potential in controlling angiogenesis in fibroproliferative diseases.
Assuntos
Bothrops/metabolismo , Venenos de Crotalídeos/análise , Desintegrinas/farmacologia , Matriz Extracelular/efeitos dos fármacos , Inflamação/prevenção & controle , Neovascularização Fisiológica/efeitos dos fármacos , Proteínas Recombinantes/farmacologia , Acetilglucosaminidase/metabolismo , Animais , Venenos de Crotalídeos/farmacologia , Desintegrinas/análise , Avaliação Pré-Clínica de Medicamentos , Fluxometria por Laser-Doppler , Macrófagos/efeitos dos fármacos , Camundongos , Peroxidase/metabolismo , Poliuretanos , Fluxo Sanguíneo Regional/efeitos dos fármacosRESUMO
Four disintegrins were isolated from the venoms of the Central American viperid snakes Atropoides mexicanus (atropoimin), Bothrops asper (bothrasperin), Cerrophidion sasai (sasaimin), and Crotalus simus (simusmin). Purifications were performed by reverse-phase HPLC. The four disintegrins have biochemical characteristics, i.e. molecular mass and location of Cys, which allow their classification within the group of medium-size disintegrins. All of them present the canonical RGD sequence, which determines their interaction with integrins in cell membranes. The disintegrins inhibited ADP and collagen-induced human platelet aggregation, with similar IC50s in the nM range. In addition, disintegrins inhibited the adhesion of an endothelial cell line and a melanoma cell line to the extracellular matrix proteins type I collagen, laminin, fibronectin, and vitronectin, albeit showing variable ability to exert this activity. This study expands the inventory of this family of viperid venom proteins, and reports, for the first time, disintegrins from the venoms of species of the genera Atropoides and Cerrophidion.
Assuntos
Desintegrinas/química , Desintegrinas/isolamento & purificação , Desintegrinas/farmacologia , Inibidores da Agregação Plaquetária/farmacologia , Venenos de Víboras/química , Sequência de Aminoácidos , Animais , Adesão Celular/efeitos dos fármacos , Linhagem Celular/efeitos dos fármacos , Costa Rica , Venenos de Crotalídeos/química , Proteínas da Matriz Extracelular/metabolismo , Fibronectinas/metabolismo , Humanos , Concentração Inibidora 50 , Laminina/metabolismo , Camundongos , Dados de Sequência Molecular , Peso Molecular , Oligopeptídeos/química , Agregação Plaquetária/efeitos dos fármacos , Inibidores da Agregação Plaquetária/químicaRESUMO
PURPOSE: Incisional hernia (IH) is characterized by defective wound healing process. Disba-01, a αvb3 integrin blocker has shown to control the rate of wound repair and therefore it could be a target for new wound healing therapies.The objective of the study was to determine the changes induced by Disba-01 on repair of wound healing after induced IH in rats. METHODS: Thirty two male albino rats were submitted to IH and divided into 4 experimental groups: G1, placebo control; G2, DisBa-01-treated; G3, anti-αvß3 antibodies-treated and G4, anti-α2 antibodies-treated. Histological. biochemical and extracellular matrix remodeling analysis of abdominal wall were evaluated. RESULTS: After 14 days, 100% of the G2 did not present hernia, and the hernia ring was closed by a thin membrane. In contrast, all groups maintained incisional hernia. DisBa-01 also increased the number macrophages and fibroblasts and induced the formation of new vessels. Additionally, MMP-2 was strongly activated only in G2 (P<0.05). Anti- αvß3-integrin antibodies produced similar results than Disba-01 but not anti-α2 integrin blocking antibodies. CONCLUSION: These results strongly indicate that Disba-01 has an important role in the control of wound healing and the blocking of this integrin may be an interesting therapeutical strategy in IH.
Assuntos
Parede Abdominal , Desintegrinas/farmacologia , Hérnia Ventral/tratamento farmacológico , Integrina alfaVbeta3/antagonistas & inibidores , Metaloproteinase 2 da Matriz/farmacologia , Cicatrização/efeitos dos fármacos , Animais , Colágeno/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Hérnia Ventral/metabolismo , Integrina alfaVbeta3/química , Integrina alfaVbeta3/metabolismo , Macrófagos/efeitos dos fármacos , Masculino , Metaloproteinase 2 da Matriz/metabolismo , Complicações Pós-Operatórias/prevenção & controle , Distribuição Aleatória , Ratos Wistar , Cicatrização/fisiologiaRESUMO
Integrin αvß3 is most likely the foremost modulator of angiogenesis among all known integrins. Recombinant disintegrin DisBa-01, originally obtained from snake venom glands, binds to αvß3, thereby significantly inhibiting adhesion and generating in vivo anti-metastatic ability. However, its function in mediator production is not clear. Here, we observed that the mediators VEGF-A, IL-8, and TGF-ß are not produced by human umbilical vein endothelial cells (HUVEC cell line) or monocyte/macrophage cells (SC cell line) when cells adhered to vitronectin. However, when exposed to DisBa-01, HUVECs produced higher levels of TGF-ß, and SC cells produced higher levels of VEGF-A. Nonetheless, HUVECs also showed an enhancement of apoptosis after losing adherence when exposed to disintegrin, which is a characteristic of anoikis. We propose that disintegrin DisBa-01 could be used to modulate integrin αvß3 functions.
Assuntos
Inibidores da Angiogênese/farmacologia , Citocinas/biossíntese , Desintegrinas/farmacologia , Integrina alfaVbeta3/metabolismo , Apoptose/efeitos dos fármacos , Adesão Celular , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Ensaios de Seleção de Medicamentos Antitumorais , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/fisiologia , Humanos , Proteínas Recombinantes/farmacologia , Vitronectina/metabolismoRESUMO
PURPOSE: Incisional hernia (IH) is characterized by defective wound healing process. Disba-01, a αvb3 integrin blocker has shown to control the rate of wound repair and therefore it could be a target for new wound healing therapies.The objective of the study was to determine the changes induced by Disba-01 on repair of wound healing after induced IH in rats. METHODS: Thirty two male albino rats were submitted to IH and divided into 4 experimental groups: G1, placebo control; G2, DisBa-01-treated; G3, anti-αvβ3 antibodies-treated and G4, anti-α2 antibodies-treated. Histological. biochemical and extracellular matrix remodeling analysis of abdominal wall were evaluated. RESULTS: After 14 days, 100% of the G2 did not present hernia, and the hernia ring was closed by a thin membrane. In contrast, all groups maintained incisional hernia. DisBa-01 also increased the number macrophages and fibroblasts and induced the formation of new vessels. Additionally, MMP-2 was strongly activated only in G2 (P<0.05). Anti- αvβ3-integrin antibodies produced similar results than Disba-01 but not anti-α2 integrin blocking antibodies. CONCLUSION: These results strongly indicate that Disba-01 has an important role in the control of wound healing and the blocking of this integrin may be an interesting therapeutical strategy in IH. .
Assuntos
Animais , Masculino , Parede Abdominal , Desintegrinas/farmacologia , Hérnia Ventral/tratamento farmacológico , /antagonistas & inibidores , /farmacologia , Cicatrização/efeitos dos fármacos , Colágeno/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Hérnia Ventral/metabolismo , /química , /metabolismo , Macrófagos/efeitos dos fármacos , /metabolismo , Complicações Pós-Operatórias/prevenção & controle , Distribuição Aleatória , Ratos Wistar , Cicatrização/fisiologiaRESUMO
Cancer can develop to the extent tumor cells grow, divide and grow into other body tissues. Integrins are a family of cell-surface heterodimeric receptors that play an important role in the development of tumor angiogenesis, growth and metastasis, thus being recognized as an attractive therapeutic target. Snake venom contains low-molecular weight peptides known as "disintegrins" that bind to integrins with high affinity, and prevent their action in cancer. In the next article, we go over the results of investigations, both in vitro and in vivo, which have shown promising results, thus revealing that the use of disintegrins could be a promising alternative for the treatment of different neoplasias.
Assuntos
Desintegrinas/farmacologia , Desintegrinas/uso terapêutico , Integrinas/antagonistas & inibidores , Neoplasias/tratamento farmacológico , Venenos de Serpentes , Desintegrinas/análise , Humanos , Integrinas/fisiologia , Neoplasias/etiologia , Neovascularização Patológica , Venenos de Serpentes/químicaRESUMO
El desarrollo del cáncer es posible en la medida que las células tumorales proliferen, se dispersen e invadan otros tejidos del cuerpo. Las integrinas son una familia de receptores heterodiméricos de superficie celular que cumplen un papel crucial en el desarrollo de la angiogénesis, crecimiento y metástasis de un tumor señalándolas como un atractivo blanco terapéutico. Los venenos de serpientes contienen péptidos de bajo peso molecular conocidos como desintegrinas, las que se unen con una alta afinidad a las integrinas e inhiben su accionar en un proceso cancerígeno. En el siguiente articulo revisamos los resultados de investigaciones, tanto in vitro como in vivo, que han mostrado resultados promisorios, por lo cual el uso de las desintegrinas podrían constituir una alternativa promisoria para el tratamiento de diversas neoplasias.
Cancer can develop to the extent tumor cells grow, divide and grow into other body tissues. Integrins are a family of cell-surface heterodimeric receptors that play an important role in the development of tumor angiogenesis, growth and metastasis, thus being recognized as an attractive therapeutic target. Snake venom contains low-molecular weight peptides known as disintegrins that bind to integrins with high affinity, and prevent their action in cancer. In the next article, we go over the results of investigations, both in vitro and in vivo, which have shown promising results, thus revealing that the use of disintegrins could be a promising alternative for the treatment of different neoplasias.