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1.
J. venom. anim. toxins incl. trop. dis ; 27: e20200196, 2021. tab, graf, ilus
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-1346436

Resumo

Snake venoms are complex mixtures of toxic proteins or peptides encoded by various gene families that function synergistically to incapacitate prey. In the present study, in order to unravel the proteomic repertoire of Deinagkistrodon acutus venom, some trace abundance components were analyzed. Methods Shotgun proteomic approach combined with shotgun nano-LC-ESI-MS/MS were employed to characterize the medically important D. acutus venom, after collected samples were enriched with the combinatorial peptide ligand library (CPLL). Results This avenue helped us find some trace components, undetected before, in D. acutus venom. The results indicated that D. acutus venom comprised 84 distinct proteins from 10 toxin families and 12 other proteins. These results are more than twice the number of venom components obtained from previous studies, which were only 29 distinct proteins obtained through RP-HPLC for the venom of the same species. The present results indicated that in D. acutus venom, the most abundant components (66.9%) included metalloproteinases, serine proteinases, and C-type lectin proteins; the medium abundant components (13%) comprised phospholipases A2 (PLA2) and 5'-nucleotidases and nucleases; whereas least abundant components (6%) were aminopeptidases, L-amino acid oxidases (LAAO), neurotoxins and disintegrins; and the trace components. The last were undetected before the use of conventional shotgun proteomics combined with shotgun nano-LC-ESI-MS/MS, such as cysteine-rich secretory proteins Da-CRPa, phospholipases B-like 1, phospholipases B (PLB), nerve growth factors (NGF), glutaminyl-peptide cyclortransferases (QC), and vascular non-inflammatory molecules 2 (VNN2). Conclusion These findings demonstrated that the CPLL enrichment method worked well in finding the trace toxin proteins in D. acutus venom, in contrast with the previous venomic characterization of D. acutus by conventional LC-MS/MS. In conclusion, this approach combined with the CPLL enrichment was effective for allowing us to explore the hidden D. acutus venomic profile and extended the list of potential venom toxins.(AU)


Assuntos
Animais , Oxirredutases , Peptídeos , Venenos de Víboras , Proteoma , Neurotoxinas
2.
J. Venom. Anim. Toxins incl. Trop. Dis. ; 27: e20200196, 2021. tab, graf, ilus
Artigo em Inglês | VETINDEX | ID: vti-31887

Resumo

Snake venoms are complex mixtures of toxic proteins or peptides encoded by various gene families that function synergistically to incapacitate prey. In the present study, in order to unravel the proteomic repertoire of Deinagkistrodon acutus venom, some trace abundance components were analyzed. Methods Shotgun proteomic approach combined with shotgun nano-LC-ESI-MS/MS were employed to characterize the medically important D. acutus venom, after collected samples were enriched with the combinatorial peptide ligand library (CPLL). Results This avenue helped us find some trace components, undetected before, in D. acutus venom. The results indicated that D. acutus venom comprised 84 distinct proteins from 10 toxin families and 12 other proteins. These results are more than twice the number of venom components obtained from previous studies, which were only 29 distinct proteins obtained through RP-HPLC for the venom of the same species. The present results indicated that in D. acutus venom, the most abundant components (66.9%) included metalloproteinases, serine proteinases, and C-type lectin proteins; the medium abundant components (13%) comprised phospholipases A2 (PLA2) and 5'-nucleotidases and nucleases; whereas least abundant components (6%) were aminopeptidases, L-amino acid oxidases (LAAO), neurotoxins and disintegrins; and the trace components. The last were undetected before the use of conventional shotgun proteomics combined with shotgun nano-LC-ESI-MS/MS, such as cysteine-rich secretory proteins Da-CRPa, phospholipases B-like 1, phospholipases B (PLB), nerve growth factors (NGF), glutaminyl-peptide cyclortransferases (QC), and vascular non-inflammatory molecules 2 (VNN2). Conclusion These findings demonstrated that the CPLL enrichment method worked well in finding the trace toxin proteins in D. acutus venom, in contrast with the previous venomic characterization of D. acutus by conventional LC-MS/MS. In conclusion, this approach combined with the CPLL enrichment was effective for allowing us to explore the hidden D. acutus venomic profile and extended the list of potential venom toxins.(AU)


Assuntos
Animais , Oxirredutases , Peptídeos , Venenos de Víboras , Proteoma , Neurotoxinas
3.
J. venom. anim. toxins incl. trop. dis ; 27: e20200180, 2021. tab, graf
Artigo em Inglês | VETINDEX, LILACS | ID: biblio-1287094

Resumo

Snake venoms are composed of pharmacologically active proteins that are evolutionarily diverse, stable and specific to targets. Hence, venoms have been explored as a source of bioactive molecules in treating numerous diseases. Recent evidences suggest that snake venom proteins may affect the formation of new blood vessels. Excessive angiogenesis has been implicated in several pathologies including tumours, diabetic retinopathy, arthritis, inter alia. In the present study, we have examined the effects of P-I metalloproteinases isolated from Bothrops moojeni (BmMP-1) and Bothrops atrox (BaMP-1) and L-amino acid oxidases (LAAO) isolated from B. moojeni (BmLAAO) and B. atrox (BaLAAO) on biochemical and functional aspects of angiogenesis. Methods: P-I metalloproteinases and LAAO were purified from venom by molecular size exclusion and ion-exchange chromatography and subsequently confirmed using mass spectrometry. The P-I metalloproteinases were characterized by azocaseinolytic, fibrinogenolytic and gelatinase activity and LAAO activity was assessed by enzyme activity on L-amino acids. Influence of these proteins on apoptosis and cell cycle in endothelial cells was analysed by flow cytometry. The angiogenic activity was determined by in vitro 3D spheroid assay, Matrigel tube forming assay, and in vivo agarose plug transformation in mice. Results: P-I metalloproteinases exhibited azocaseinolytic activity, cleaved α and partially β chain of fibrinogen, and displayed catalytic activity on gelatin. LAAO showed differential activity on L-amino acids. Flow cytometry analysis indicated that both P-I metalloproteinases and LAAO arrested the cells in G0/G1 phase and further induced both necrosis and apoptosis in endothelial cells. In vitro, P-I metalloproteinases and LAAO exhibited significant anti-angiogenic properties in 3D spheroid and Matrigel models by reducing sprout outgrowth and tube formation. Using agarose plug transplants in mice harbouring P-I metalloproteinases and LAAO we demonstrated a marked disruption of vasculature at the periphery. Conclusion: Our research suggests that P-I metalloproteinases and LAAO exhibit anti-angiogenic properties in vitro and in vivo.(AU)


Assuntos
Animais , Oxirredutases , Bothrops/fisiologia , Inibidores da Angiogênese , Venenos de Crotalídeos , Metaloproteases
4.
J. Venom. Anim. Toxins incl. Trop. Dis. ; 27: e20200180, 2021. tab, graf
Artigo em Inglês | VETINDEX | ID: vti-31907

Resumo

Snake venoms are composed of pharmacologically active proteins that are evolutionarily diverse, stable and specific to targets. Hence, venoms have been explored as a source of bioactive molecules in treating numerous diseases. Recent evidences suggest that snake venom proteins may affect the formation of new blood vessels. Excessive angiogenesis has been implicated in several pathologies including tumours, diabetic retinopathy, arthritis, inter alia. In the present study, we have examined the effects of P-I metalloproteinases isolated from Bothrops moojeni (BmMP-1) and Bothrops atrox (BaMP-1) and L-amino acid oxidases (LAAO) isolated from B. moojeni (BmLAAO) and B. atrox (BaLAAO) on biochemical and functional aspects of angiogenesis. Methods: P-I metalloproteinases and LAAO were purified from venom by molecular size exclusion and ion-exchange chromatography and subsequently confirmed using mass spectrometry. The P-I metalloproteinases were characterized by azocaseinolytic, fibrinogenolytic and gelatinase activity and LAAO activity was assessed by enzyme activity on L-amino acids. Influence of these proteins on apoptosis and cell cycle in endothelial cells was analysed by flow cytometry. The angiogenic activity was determined by in vitro 3D spheroid assay, Matrigel tube forming assay, and in vivo agarose plug transformation in mice. Results: P-I metalloproteinases exhibited azocaseinolytic activity, cleaved α and partially β chain of fibrinogen, and displayed catalytic activity on gelatin. LAAO showed differential activity on L-amino acids. Flow cytometry analysis indicated that both P-I metalloproteinases and LAAO arrested the cells in G0/G1 phase and further induced both necrosis and apoptosis in endothelial cells. In vitro, P-I metalloproteinases and LAAO exhibited significant anti-angiogenic properties in 3D spheroid and Matrigel models by reducing sprout outgrowth and tube formation. Using agarose plug transplants in mice harbouring P-I metalloproteinases and LAAO we demonstrated a marked disruption of vasculature at the periphery. Conclusion: Our research suggests that P-I metalloproteinases and LAAO exhibit anti-angiogenic properties in vitro and in vivo.(AU)


Assuntos
Animais , Oxirredutases , Bothrops/fisiologia , Inibidores da Angiogênese , Venenos de Crotalídeos , Metaloproteases
5.
J. venom. anim. toxins incl. trop. dis ; 26: e20190103, 2020. tab, graf
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-1101265

Resumo

The Brazil's lancehead, Bothrops brazili, is a poorly studied pit viper distributed in lowlands of the equatorial rainforests of southern Colombia, northeastern Peru, eastern Ecuador, southern and southeastern Venezuela, Guyana, Suriname, French Guiana, Brazil, and northern Bolivia. Few studies have been reported on toxins isolated from venom of Ecuadorian and Brazilian B. brazili. The aim of the present study was to elucidate the qualitative and quantitative protein composition of B. brazili venom from Pará (Brazil), and to carry out a comparative antivenomics assessment of the immunoreactivity of the Brazilian antibothropic pentavalent antivenom [soro antibotrópico (SAB) in Portuguese] against the venoms of B. brazili and reference species, B. jararaca. Methods: We have applied a quantitative snake venomics approach, including reverse-phase and two-dimensional electrophoretic decomplexation of the venom toxin arsenal, LC-ESI-MS mass profiling and peptide-centric MS/MS proteomic analysis, to unveil the overall protein composition of B. brazili venom from Pará (Brazil). Using third-generation antivenomics, the specific and paraspecific immunoreactivity of the Brazilian SAB against homologous (B. jararaca) and heterologous (B. brazili) venoms was investigated. Results: The venom proteome of the Brazil's lancehead (Pará) is predominantly composed of two major and three minor acidic (19%) and two major and five minor basic (14%) phospholipase A2 molecules; 7-11 snake venom metalloproteinases of classes PI (21%) and PIII (6%); 10-12 serine proteinases (14%), and 1-2 L-amino acid oxidases (6%). Other toxins, including two cysteine-rich secretory proteins, one C-type lectin-like molecule, one nerve growth factor, one 5'-nucleotidase, one phosphodiesterase, one phospholipase B, and one glutaminyl cyclase molecule, represent together less than 2.7% of the venom proteome. Third generation antivenomics profile of the Brazilian pentabothropic antivenom showed paraspecific immunoreactivity against all the toxin classes of B. brazili venom, with maximal binding capacity of 132.2 mg venom/g antivenom. This figure indicates that 19% of antivenom's F(ab')2 antibodies bind B. brazili venom toxins. Conclusion: The proteomics outcome contribute to a deeper insight into the spectrum of toxins present in the venom of the Brazil's lancehead, and rationalize the pathophysiology underlying this snake bite envenomings. The comparative qualitative and quantitative immunorecognition profile of the Brazilian pentabothropic antivenom toward the venom toxins of B. brazili and B. jararaca (the reference venom for assessing the bothropic antivenom's potency in Brazil), provides clues about the proper use of the Brazilian antibothropic polyvalent antivenom in the treatment of bites by the Brazil's lancehead.(AU)


Assuntos
Animais , Oxirredutases , Mordeduras de Serpentes , Venenos de Serpentes , Mordeduras e Picadas , Antivenenos , Bothrops , Proteoma
6.
Sci. agric ; 76(6): 463-472, Nov.-Dec. 2019. ilus, tab, graf
Artigo em Inglês | VETINDEX | ID: biblio-1497819

Resumo

Gossypol easily pairs with lysine side chains and enzymes involved in the cellular growth process. The effect of gossypol (a compound present in cotton co-products) in ruminant metabolism and meat quality is not yet clear. This study was undertaken in order to evaluate the effects of cotton co-products in lamb muscle proteome. Twenty-four Santa Inês ram lambs, 5-months old (20.6 ± 1.9 kg), were randomly assigned to four treatments: control (without cottonseed), whole cottonseed, cottonseed meal and high oil cottonseed meal. At 95 days into the experiment, lambs were slaughtered and samples from Longissimus dorsi were collected. Proteins were extracted and analyzed by 2-D electrophoresis. Spots showing a significant effect from the treatment (the “treatment effect”) and present in more than 90 % of the samples were identified using mass spectrometry. Cotton co-products decreased the abundance of aldehyde and malate dehydrogenases, creatine kinase M-type and Adenosine triphosphate (ATP) synthase. They also increased four proteins related to muscle contraction. Thus, feeding cotton co-products to lambs changed the abundance of important muscle proteins. A cotton co-product diet induced a negative impact on the energy supply of muscle cells and, consequently, the abundance of ATP dependent proteins (contractile apparatus) increased, probably in order to offset and maintain muscle function. These proteomic changes can promote our understanding of alterations in the sensorial properties of meat due to cotton co-product diets in further investigations.


Assuntos
Animais , Carne Vermelha/análise , Oxirredutases , Proteoma , Óleo de Sementes de Algodão/administração & dosagem , Óleo de Sementes de Algodão/análise , Ovinos
7.
Bol. ind. anim. (Impr.) ; 76: 1-12, 2019. tab
Artigo em Português | VETINDEX | ID: biblio-1466976

Resumo

A atividade enzimática do solo reflete a atividade dos microrganismos que decompõem a matéria orgânica e liberam nutrientes às plantas. O objetivo foi avaliar a atividade enzimática no solo em pastagens de cultivares de Urochloa manejados sob as ofertas de forragem de 4, 7, 10 e 13 kg de matéria seca (MS)/100 kg de peso corporal (PC). O método de pastejo adotado foi mob-stocking, com quatro ciclos de pastejo (entre dezembro de 2008 e fevereiro de 2009). Também foi realizada uma amostragem em junho de 2009 para melhor caracterização, pois a mineralização da matéria orgânica no solo ocorre em longo prazo. Assim, as datas de coleta de solo para análise foram descritas como dias de avaliações e não como pastejos: 1 (de 17 a 20 de dezembro de 2008), 21 (de 07 a 10 de janeiro de 2009), 42 (de 28 a 31 de janeiro de 2009), 63 (de 18 a 21 de fevereiro de 2009), 183 (de 18 a 20 de junho de 2009). Foi avaliada a atividade enzimática no solo da desidrogenase, protease, arilsulfatase e celulase. O delineamento experimental foi inteiramente casualizado, com três repetições por oferta. Foi realizada a análise de variância e teste de Tukey ao nível de 5% de probabilidade, para comparação de médias, utilizando-se o procedimento o programa estatístico R. As análises foram realizadas de maneira individual para cada cultivar. De modo geral, nas pastagens das três cultivares estudadas, a atividade da desidrogenase foi maior (P<0,05) aos 63 dias e da protease no último dia de avaliação. O decorrer do período experimental implicou em mudanças na atividade da arilsulfatase e da celulase em relação às ofertas de forragem, mas sem um padrão de comportamento definido. As ofertas de forragem mostraram efeito sobre a atividade das enzimas ao longo do período de avaliação, porém sem padrão de comportamento definido, sendo observadas oscilações cíclicas da atividade dessas enzimas. As ofertas de forragem estudadas afetaram a atividade enzimática da desidrogenase, protease, arilsulfatase e...


Soil enzymatic activity reflects the activity microorganisms that decompose the organic matter and release nutrients into the soil that can be used by plants. The objective of this study was to evaluate soil enzymatic activity in Urochloa cultivar pastures managed under forage allowances of 4, 7, 10 and 13 kg dry matter/100 kg body weight. The grazing method adopted was mob-stocking, with four grazing cycles (between December 2008 and February 2009). Soil sampling was performed in June 2009 for better characterization since the mineralization of soil organic matter is a long-term process. Thus, the dates of soil sampling for analysis are reported as evaluation days and not as grazing: 1 (December 17 to 20, 2008), 21 (January 7 to 10, 2009), 42 (January 28 to 31, 2009), 63 (February 18 to 21, 2009), 183 (June 18 to 20, 2009). The soil enzymatic activity of dehydrogenase, protease, arylsulfatase, and cellulase was evaluated. The experimental design was completely randomized, with three replications per forage allowance. Analysis of variance and Tukey’s test at the 5% probability level were applied to compare means using the R statistical program. The analyses were performed individually for each cultivar. Overall, in pastures of the three cultivars studied, dehydrogenase activity was higher (P<0.05) on day 63 and protease on the last day of evaluation. The experimental period implied changes in the activity of arylsulfatase and cellulase according to forage allowance, but without a defined pattern. Forage allowances exerted an effect on the activity of the enzymes throughout the evaluation period, but there was no defined pattern. Cyclic oscillations in the activity of these enzymes were observed. The forage allowances studied affected the soil enzymatic activity of dehydrogenase, protease, arylsulfatase and cellulase in Urochloa pastures of the Marandu, Xaraes and Mulato varieties, but no pattern was found in...


Assuntos
Ativação Enzimática , Brachiaria , Enzimas/análise , Pastagens , Química do Solo/análise , Arilsulfatases , Celulase , Oxirredutases , Peptídeo Hidrolases
8.
Sci. agric. ; 76(6): 463-472, Nov.-Dec. 2019. ilus, tab, graf
Artigo em Inglês | VETINDEX | ID: vti-24529

Resumo

Gossypol easily pairs with lysine side chains and enzymes involved in the cellular growth process. The effect of gossypol (a compound present in cotton co-products) in ruminant metabolism and meat quality is not yet clear. This study was undertaken in order to evaluate the effects of cotton co-products in lamb muscle proteome. Twenty-four Santa Inês ram lambs, 5-months old (20.6 ± 1.9 kg), were randomly assigned to four treatments: control (without cottonseed), whole cottonseed, cottonseed meal and high oil cottonseed meal. At 95 days into the experiment, lambs were slaughtered and samples from Longissimus dorsi were collected. Proteins were extracted and analyzed by 2-D electrophoresis. Spots showing a significant effect from the treatment (the “treatment effect”) and present in more than 90 % of the samples were identified using mass spectrometry. Cotton co-products decreased the abundance of aldehyde and malate dehydrogenases, creatine kinase M-type and Adenosine triphosphate (ATP) synthase. They also increased four proteins related to muscle contraction. Thus, feeding cotton co-products to lambs changed the abundance of important muscle proteins. A cotton co-product diet induced a negative impact on the energy supply of muscle cells and, consequently, the abundance of ATP dependent proteins (contractile apparatus) increased, probably in order to offset and maintain muscle function. These proteomic changes can promote our understanding of alterations in the sensorial properties of meat due to cotton co-product diets in further investigations.(AU)


Assuntos
Animais , Óleo de Sementes de Algodão/administração & dosagem , Óleo de Sementes de Algodão/análise , Carne Vermelha/análise , Proteoma , Oxirredutases , Ovinos
9.
B. Indústr. Anim. ; 76: 1-12, 2019. tab
Artigo em Português | VETINDEX | ID: vti-24483

Resumo

A atividade enzimática do solo reflete a atividade dos microrganismos que decompõem a matéria orgânica e liberam nutrientes às plantas. O objetivo foi avaliar a atividade enzimática no solo em pastagens de cultivares de Urochloa manejados sob as ofertas de forragem de 4, 7, 10 e 13 kg de matéria seca (MS)/100 kg de peso corporal (PC). O método de pastejo adotado foi mob-stocking, com quatro ciclos de pastejo (entre dezembro de 2008 e fevereiro de 2009). Também foi realizada uma amostragem em junho de 2009 para melhor caracterização, pois a mineralização da matéria orgânica no solo ocorre em longo prazo. Assim, as datas de coleta de solo para análise foram descritas como dias de avaliações e não como pastejos: 1 (de 17 a 20 de dezembro de 2008), 21 (de 07 a 10 de janeiro de 2009), 42 (de 28 a 31 de janeiro de 2009), 63 (de 18 a 21 de fevereiro de 2009), 183 (de 18 a 20 de junho de 2009). Foi avaliada a atividade enzimática no solo da desidrogenase, protease, arilsulfatase e celulase. O delineamento experimental foi inteiramente casualizado, com três repetições por oferta. Foi realizada a análise de variância e teste de Tukey ao nível de 5% de probabilidade, para comparação de médias, utilizando-se o procedimento o programa estatístico R. As análises foram realizadas de maneira individual para cada cultivar. De modo geral, nas pastagens das três cultivares estudadas, a atividade da desidrogenase foi maior (P<0,05) aos 63 dias e da protease no último dia de avaliação. O decorrer do período experimental implicou em mudanças na atividade da arilsulfatase e da celulase em relação às ofertas de forragem, mas sem um padrão de comportamento definido. As ofertas de forragem mostraram efeito sobre a atividade das enzimas ao longo do período de avaliação, porém sem padrão de comportamento definido, sendo observadas oscilações cíclicas da atividade dessas enzimas. As ofertas de forragem estudadas afetaram a atividade enzimática da desidrogenase, protease, arilsulfatase e...(AU)


Soil enzymatic activity reflects the activity microorganisms that decompose the organic matter and release nutrients into the soil that can be used by plants. The objective of this study was to evaluate soil enzymatic activity in Urochloa cultivar pastures managed under forage allowances of 4, 7, 10 and 13 kg dry matter/100 kg body weight. The grazing method adopted was mob-stocking, with four grazing cycles (between December 2008 and February 2009). Soil sampling was performed in June 2009 for better characterization since the mineralization of soil organic matter is a long-term process. Thus, the dates of soil sampling for analysis are reported as evaluation days and not as grazing: 1 (December 17 to 20, 2008), 21 (January 7 to 10, 2009), 42 (January 28 to 31, 2009), 63 (February 18 to 21, 2009), 183 (June 18 to 20, 2009). The soil enzymatic activity of dehydrogenase, protease, arylsulfatase, and cellulase was evaluated. The experimental design was completely randomized, with three replications per forage allowance. Analysis of variance and Tukeys test at the 5% probability level were applied to compare means using the R statistical program. The analyses were performed individually for each cultivar. Overall, in pastures of the three cultivars studied, dehydrogenase activity was higher (P<0.05) on day 63 and protease on the last day of evaluation. The experimental period implied changes in the activity of arylsulfatase and cellulase according to forage allowance, but without a defined pattern. Forage allowances exerted an effect on the activity of the enzymes throughout the evaluation period, but there was no defined pattern. Cyclic oscillations in the activity of these enzymes were observed. The forage allowances studied affected the soil enzymatic activity of dehydrogenase, protease, arylsulfatase and cellulase in Urochloa pastures of the Marandu, Xaraes and Mulato varieties, but no pattern was found in...(AU)


Assuntos
Brachiaria , Pastagens , Química do Solo/análise , Enzimas/análise , Ativação Enzimática , Arilsulfatases , Celulase , Oxirredutases , Peptídeo Hidrolases
10.
Acta cir. bras. ; 34(11): e201901104, 2019. graf
Artigo em Inglês | VETINDEX | ID: vti-24134

Resumo

Purpose: Myocardial ischemia/reperfusion (Ml/R) injury is a leading cause of damage in cardiac tissues, with high rates of mortality and disability. Biochanin A (BCA) is a main constituent of Trifolium pratense L. This study was intended to explore the effect of BCA on Ml/R injury and explore the potential mechanism. Methods: In vivo MI/R injury was established by transient coronary ligation in Sprague-Dawley rats. Triphenyltetrazolium chloride staining (TTC) was used to measure myocardial infarct size. ELISA assay was employed to evaluate the levels of myocardial enzyme and inflammatory cytokines. Western blot assay was conducted to detect related protein levels in myocardial tissues. Results: BCA significantly ameliorated myocardial infarction area, reduced the release of myocardial enzyme levels including aspartate transaminase (AST), creatine kinase (CK-MB) and lactic dehydrogenase (LDH). It also decreased the production of inflammatory cytokines (IL-1, IL-18, IL-6 and TNF-) in serum of Ml/R rats. Further mechanism studies demonstrated that BCA inhibited inflammatory reaction through blocking TLR4/NF-kB/NLRP3 signaling pathway. Conclusion: The present study is the first evidence demonstrating that BCA attenuated Ml/R injury through suppressing TLR4/NF-kB/NLRP3 signaling pathway-mediated anti-inflammation pathway.(AU)


Assuntos
Animais , Ratos , Ratos/lesões , Isquemia Miocárdica/veterinária , Oxirredutases/análise
11.
Pesqui. vet. bras ; 38(5): 883-888, May 2018. graf
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-955401

Resumo

Theileria parva is the causative agent of East Coast Fever (ECF), a tick borne disease, which results in major economic losses in cattle. Major problems in dealing with this illness are the high cost of drugs, development of resistance, and absence of effective vaccines. Thus, exploiting new targets for cost effective and higher therapeutic value drugs are imperative. Glycolysis is the main pathway for generation of ATP in T. parva, given its development inside erythrocytes. Thus, the enzymes of this pathway may prove potential targets for designing new-generation anti-theilerials. Lactate dehydrogenase of T. parva (TpLDH) has the highest activity of all glycolytic enzymes and thus we selected this enzyme as the potential therapeutic target. Our study is the first to report the isolation, removal of introns through directed mutagenesis, and cloning of TpLDH and showing that amino acid insertions or deletions most notably corresponded to a 5-amino acid sequence (Asn-91A, Glu-91B, Glu-91C, Trp-91D, Asn-91E) between Ser-91 ve Arg-92 of the enzyme. This region is also present in other apicomplexan such as Babesia bovis, a pathogen of cattle and Plasmodium falciparum, a human pathogen. Providing as the attachment site for the enzyme inhibitors and not being present in LDH of respective hosts, we propose this site as an attractive drug target. The work here is expected to lead new studies on detailed structural and kinetic aspects of apicomplexan LDHs and development of new inhibitors.(AU)


Assuntos
Engenharia Genética/veterinária , Theileria parva/genética , Oxirredutases/análise
12.
Pesqui. vet. bras ; 38(5): 883-888, May 2018. ilus
Artigo em Inglês | VETINDEX | ID: vti-20690

Resumo

Theileria parva is the causative agent of East Coast Fever (ECF), a tick borne disease, which results in major economic losses in cattle. Major problems in dealing with this illness are the high cost of drugs, development of resistance, and absence of effective vaccines. Thus, exploiting new targets for cost effective and higher therapeutic value drugs are imperative. Glycolysis is the main pathway for generation of ATP in T. parva, given its development inside erythrocytes. Thus, the enzymes of this pathway may prove potential targets for designing new-generation anti-theilerials. Lactate dehydrogenase of T. parva (TpLDH) has the highest activity of all glycolytic enzymes and thus we selected this enzyme as the potential therapeutic target. Our study is the first to report the isolation, removal of introns through directed mutagenesis, and cloning of TpLDH and showing that amino acid insertions or deletions most notably corresponded to a 5-amino acid sequence (Asn-91A, Glu-91B, Glu-91C, Trp-91D, Asn-91E) between Ser-91 ve Arg-92 of the enzyme. This region is also present in other apicomplexan such as Babesia bovis, a pathogen of cattle and Plasmodium falciparum, a human pathogen. Providing as the attachment site for the enzyme inhibitors and not being present in LDH of respective hosts, we propose this site as an attractive drug target. The work here is expected to lead new studies on detailed structural and kinetic aspects of apicomplexan LDHs and development of new inhibitors.(AU)


Assuntos
Engenharia Genética/veterinária , Theileria parva/genética , Oxirredutases/análise
13.
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-984692

Resumo

L-amino acid oxidases isolated from snake venoms (SV-LAAOs) are enzymes that have great therapeutic potential and are currently being investigated as tools for developing new strategies to treat various diseases, including cancer and bacterial infections. The main objective of this study was to make a brief evaluation of the enzymatic stability of two Bothrops LAAOs, one isolated from Bothrops jararacussu (BjussuLAAO-II) and the other from Bothrops moojeni (BmooLAAO-I) venoms. Methods and results: The enzymatic activity and stability of both LAAOs were evaluated by microplate colorimetric assays, for which BjussuLAAO-II and BmooLAAO-I were incubated with different L-amino acid substrates, in the presence of different ions, and at different pH ranges and temperatures. BjussuLAAO-II and BmooLAAO-I demonstrated higher affinity for hydrophobic amino acids, such as Phe and Leu. The two enzymes showed high enzymatic activity in a wide temperature range, from 25 to 75 °C, and presented optimum pH around 7.0. Additionally, Zn2+, Al3+, Cu2+ and Ni2+ ions negatively modulated the enzymatic activity of both LAAOs. As to stability, BjussuLAAO-II and BmooLAAO-I showed high enzymatic activity for 42 days stored at 4°C in neutral pH solution. Moreover, the glycan portions of both LAAOs were analyzed by capillary electrophoresis, which revealed that BjussuLAAO-II presented two main glycan portions with relative masses of 7.78 and 8.13 CGU, while BmooLAAO-I showed three portions of 7.58, 7.94 and 8.37 CGU. Conclusions: Our results showed that, when stored properly, BjussuLAAO-II and BmooLAAO-I present enzymatic stability over a long time period, which is very important to allow the use of these enzymes in pharmacological studies of great impact in the medical field.(AU)


Assuntos
Animais , Oxirredutases , Polissacarídeos , Venenos de Serpentes , Infecções Bacterianas , Bothrops , Aminoácidos
14.
Neotrop. ichthyol ; 15(1): e160102, 2017. tab, graf
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-841887

Resumo

Two Coryphaena hippurus morphotypes (dourado and palombeta) are found along the Brazilian coast and are considered by Rio de Janeiro’s fisherman and fishmongers as two different species. Furthermore, these morphotypes are commercialized under different values and suffer different fishing pressure. Therefore, a definition of their taxonomic status is an important economic and biological matter. In order to investigate this problem, allozyme electrophoresis method was undertaken for seventeen loci on 117 individuals of C. hippurus sampled at Cabo Frio/RJ (Brazil). The data indicate homogeneity between the morphotypes gene pools. Nevertheless, differences were found for genetic variation among dourado and palombeta, especially due to alcohol dehydrogenase locus. Natural selection hypothesis is discussed in explaining these findings.(AU)


Dois morfotipos de Coryphaena hippurus (dourado e palombeta) encontrados ao longo da costa brasileira são considerados espécies diferentes por pescadores e mercadores das regiões de desembarque do estado do Rio de Janeiro. Além disso, esses morfotipos são comercializados por valores diferentes e sofrem diferentes pressões de pesca. Desta forma, a definição do status taxonômico desses morfotipos é importante, tanto em termos econômicos quanto biológicos. A fim de investigar esse problema foi utilizado o método de eletroforese de aloenzimas com a amostragem de dezessete loci para 117 indivíduos dos dois morfotipos de C. hippurus obtidos em desembarques pesqueiros na região de Cabo Frio/RJ (Brasil). Os dados indicaram uma homogeneidade entre os conjuntos gênicos dos morfotipos. A despeito disso, diferenças entre os conjuntos gênicos de dourado e palombeta foram encontradas, devido, especialmente, ao locus álcool desidrogenase. A hipótese de seleção natural é discutida como possível explicação para esses resultados.(AU)


Assuntos
Animais , Classificação , Perciformes/anatomia & histologia , Perciformes/classificação , Bioquímica , Pesqueiros , Oxirredutases
15.
Neotrop. ichthyol ; 15(1): [e160102], Abril 6, 2017. tab, graf
Artigo em Inglês | VETINDEX | ID: vti-16441

Resumo

Two Coryphaena hippurus morphotypes (dourado and palombeta) are found along the Brazilian coast and are considered by Rio de Janeiros fisherman and fishmongers as two different species. Furthermore, these morphotypes are commercialized under different values and suffer different fishing pressure. Therefore, a definition of their taxonomic status is an important economic and biological matter. In order to investigate this problem, allozyme electrophoresis method was undertaken for seventeen loci on 117 individuals of C. hippurus sampled at Cabo Frio/RJ (Brazil). The data indicate homogeneity between the morphotypes gene pools. Nevertheless, differences were found for genetic variation among dourado and palombeta, especially due to alcohol dehydrogenase locus. Natural selection hypothesis is discussed in explaining these findings.(AU)


Dois morfotipos de Coryphaena hippurus (dourado e palombeta) encontrados ao longo da costa brasileira são considerados espécies diferentes por pescadores e mercadores das regiões de desembarque do estado do Rio de Janeiro. Além disso, esses morfotipos são comercializados por valores diferentes e sofrem diferentes pressões de pesca. Desta forma, a definição do status taxonômico desses morfotipos é importante, tanto em termos econômicos quanto biológicos. A fim de investigar esse problema foi utilizado o método de eletroforese de aloenzimas com a amostragem de dezessete loci para 117 indivíduos dos dois morfotipos de C. hippurus obtidos em desembarques pesqueiros na região de Cabo Frio/RJ (Brasil). Os dados indicaram uma homogeneidade entre os conjuntos gênicos dos morfotipos. A despeito disso, diferenças entre os conjuntos gênicos de dourado e palombeta foram encontradas, devido, especialmente, ao locus álcool desidrogenase. A hipótese de seleção natural é discutida como possível explicação para esses resultados.(AU)


Assuntos
Animais , Perciformes/anatomia & histologia , Perciformes/classificação , Classificação , Pesqueiros , Bioquímica , Oxirredutases
16.
J. venom. anim. toxins incl. trop. dis ; 21: 26, 31/03/2015. tab, ilus, graf
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-954737

Resumo

Background Crotalus durissus terrificus venom (CdtV) is one of the most studied snake venoms in Brazil. Despite presenting several well known proteins, its L-amino acid oxidase (LAAO) has not been studied previously. This study aimed to isolate, characterize and evaluate the enzyme stability of bordonein-L, an LAAO from CdtV.Methods The enzyme was isolated through cation exchange, gel filtration and affinity chromatography, followed by a reversed-phase fast protein liquid chromatography to confirm its purity. Subsequently, its N-terminal amino acid sequence was determined by Edman degradation. The enzyme activity and stability were evaluated by a microplate colorimetric assay and the molecular mass was estimated by SDS-PAGE using periodic acid-Schiff staining and determined by mass spectrometry.Results The first 39 N-terminal amino acid residues exhibited high identity with other snake venom L-amino acid oxidases. Bordonein-L is a homodimer glycoprotein of approximately 101 kDa evaluated by gel filtration. Its monomer presents around 53 kDa estimated by SDS-PAGE and 58,702 Da determined by MALDI-TOF mass spectrometry. The enzyme exhibited maximum activity at pH 7.0 and lost about 50 % of its activity after five days of storage at 4 °C. Bordonein-L's activity was higher than the control when stored in 2.8 % mannitol or 8.5 % sucrose.Conclusions This research is pioneering in its isolation, characterization and enzyme stability evaluation of an LAAO from CdtV, denominated bordonein-L. These results are important because they increase the knowledge about stabilization of LAAOs, aiming to increase their shelf life. Since the maintenance of enzymatic activity after long periods of storage is essential to enable their biotechnological use as well as their functional studies.(AU)


Assuntos
Animais , Oxirredutases , Venenos de Serpentes , Estabilidade Enzimática , L-Aminoácido Oxidase , Aminoácidos
17.
Artigo em Inglês | VETINDEX | ID: vti-10953

Resumo

The L-amino acid oxidases (LAAOs) constitute a major component of snake venoms and have been widely studied due to their widespread presence and various effects, such as apoptosis induction, cytotoxicity, induction and/or inhibition of platelet aggregation, hemorrhage, hemolysis, edema, as well as antimicrobial, antiparasitic and anti-HIV activities. The isolated and characterized snake venom LAAOs have become important research targets due to their potential biotechnological applications in pursuit for new drugs of interest in the scientific and medical fields. The current study discusses the antitumor effects of snake venom LAAOs described in the literature to date, highlighting the mechanisms of apoptosis induction proposed for this class of proteins.(AU)


Assuntos
Animais , Oxirredutases/análise , L-Aminoácido Oxidase/análise , Venenos/administração & dosagem , Serpentes/classificação
18.
J. venom. anim. toxins incl. trop. dis ; 20: 1-7, 04/02/2014. tab, ilus
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-1484558

Resumo

The L-amino acid oxidases (LAAOs) constitute a major component of snake venoms and have been widely studied due to their widespread presence and various effects, such as apoptosis induction, cytotoxicity, induction and/or inhibition of platelet aggregation, hemorrhage, hemolysis, edema, as well as antimicrobial, antiparasitic and anti-HIV activities. The isolated and characterized snake venom LAAOs have become important research targets due to their potential biotechnological applications in pursuit for new drugs of interest in the scientific and medical fields. The current study discusses the antitumor effects of snake venom LAAOs described in the literature to date, highlighting the mechanisms of apoptosis induction proposed for this class of proteins.


Assuntos
Animais , L-Aminoácido Oxidase/análise , Oxirredutases/análise , Venenos/administração & dosagem , Serpentes/classificação
19.
J. venom. anim. toxins incl. trop. dis ; 16(3): 449-455, 2010. tab
Artigo em Inglês | LILACS, VETINDEX | ID: lil-557173

Resumo

The present study aimed to investigate the effects of Mesobuthus tamulus gangeticus Pocock (Buthidae) venom on albino mice (NIH strain). Whole venom was obtained by electrical stimulation and its toxicity was determined in albino mice by subcutaneous envenomation. The venom LD50 was 2.5 mg kg-1 of mouse body weight. Toxic effects on different biochemical and enzymatic parameters in blood serum and other tissues of albino mice were determined after experimental envenomation with sublethal doses of M. tamulus gangeticus venom. Increased levels of glucose, uric acid and cholesterol, as well as decreased serum total proteins, were observed at 2 and 4 hours after the envenomation. In the liver and muscles, glycogen content dropped after venom injection. Moreover, M. tamulus gangeticus venom elevated the enzymatic activity of acid phosphatase (ACP), lactic dehydrogenase (LDH) and alanine aminotransferase (ALT) in the serum of albino mice. In conclusion, M. tamulus gangeticus can be considered a lethal scorpion species.(AU)


Assuntos
Animais , Camundongos , Oxirredutases , Venenos de Escorpião/intoxicação , Escorpiões , Estimulação Elétrica , Padrões de Referência , Dose Letal Mediana
20.
J. venom. anim. toxins incl. trop. dis ; 15(2): 226-235, 2009. tab
Artigo em Inglês | LILACS, VETINDEX | ID: lil-517283

Resumo

In the present study, Heterometrus fastigiousus venom (HFV) was employed as antigen to produce species-specific scorpion antivenom (SAV) in albino mice (NIH) strain. To determine SAV efficacy, it was pre-incubated with 10 LD50 of HFV and then injected subcutaneously into mice. Subsequently, mortality was observed after 24 hours. Minimum effective dose (MED) was 12.5 LD50 of HFV/mL of SAV. SAV effectiveness to reverse HFV-induced biochemical alterations in mice was analyzed by challenge method. Simultaneously, mice received subcutaneously 40 percent of 24-hour-LD50 of HFV and intravenously SAV. After four hours, changes in serum glucose, free amino acids, uric acids, pyruvic acid, cholesterol, total protein, alkaline phosphatase, acid phosphatase, lactic dehydrogenase and glutamate-pyruvate transaminase enzyme level were determined. Treatment with species-specific SAV resulted in the reversal of HFV-induced biochemical alterations.(AU)


Assuntos
Animais , Camundongos , Oxirredutases , Escorpiões , Antivenenos , Soro , Transaminases , Dose Letal Mediana , Ácido Pirúvico
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