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Some, probably most and perhaps all, members of the phylum Nemertea are poisonous, documented so far from marine and benthic specimens. Although the toxicity of these animals has been long known, systematic studies on the characterization of toxins, mechanisms of toxicity, and toxin evolution for this group are scarce. Here, we present the first investigation of the molecular evolution of toxins in Nemertea. Using a proteo-transcriptomic approach, we described toxins in the body and poisonous mucus of the pilidiophoran Lineus sanguineus and the hoplonemertean Nemertopsis pamelaroeae. Using these new and publicly available transcriptomes, we investigated the molecular evolution of six selected toxin gene families. In addition, we also characterized in silico the toxin genes found in the interstitial hoplonemertean, Ototyphlonemertes erneba, a meiofaunal taxa. We successfully identified over 200 toxin transcripts in each of these species. Evidence of positive selection and gene duplication was observed in all investigated toxin genes. We hypothesized that the increased rates of gene duplications observed for Pilidiophora could be involved with the expansion of toxin genes. Studies concerning the natural history of Nemertea are still needed to understand the evolution of their toxins. Nevertheless, our results show evolutionary mechanisms similar to other venomous groups.
Assuntos
Toxinas Biológicas , Peçonhas , Animais , Peçonhas/genética , Duplicação Gênica , Transcriptoma , Perfilação da Expressão Gênica , Filogenia , Evolução MolecularRESUMO
Many studies point to an association between Helicobacter pylori (H. pylori) infection and inflammatory bowel diseases (IBD). Although controversial, this association indicates that the presence of the bacterium somehow affects the course of IBD. It appears that H. pylori infection influences IBD through changes in the diversity of the gut microbiota, and hence in local chemical characteristics, and alteration in the pattern of gut immune response. The gut immune response appears to be modulated by H. pylori infection towards a less aggressive inflammatory response and the establishment of a targeted response to tissue repair. Therefore, a T helper 2 (Th2)/macrophage M2 response is stimulated, while the Th1/macrophage M1 response is suppressed. The immunomodulation appears to be associated with intrinsic factors of the bacteria, such as virulence factors - such oncogenic protein cytotoxin-associated antigen A, proteins such H. pylori neutrophil-activating protein, but also with microenvironmental changes that favor permanence of H. pylori in the stomach. These changes include the increase of gastric mucosal pH by urease activity, and suppression of the stomach immune response promoted by evasion mechanisms of the bacterium. Furthermore, there is a causal relationship between H. pylori infection and components of the innate immunity such as the NLR family pyrin domain containing 3 inflammasome that directs IBD toward a better prognosis.
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Infecções por Helicobacter , Helicobacter pylori , Doenças Inflamatórias Intestinais , Humanos , Infecções por Helicobacter/complicações , Imunidade Inata , EstômagoRESUMO
This study describes, to some extent, the VCC contribution as an early stimulation of the macrophage lineage. Regarding the onset of the innate immune response caused by infection, the ß form of IL-1 is the most important interleukin involved in the onset of the inflammatory innate response. Activated macrophages treated in vitro with VCC induced the activation of the MAPK signaling pathway in a one-hour period, with the activation of transcriptional regulators for a surviving and pro-inflammatory response, suggesting an explanation inspired and supported by the inflammasome physiology. The mechanism of IL-1ß production induced by VCC has been gracefully outlined in murine models, using bacterial knockdown mutants and purified molecules; nevertheless, the knowledge of this mechanism in the human immune system is still under study. This work shows the soluble form of 65 kDa of the Vibrio cholerae cytotoxin (also known as hemolysin), as it is secreted by the bacteria, inducing the production of IL-1ß in the human macrophage cell line THP-1. The mechanism involves triggering the early activation of the signaling pathway MAPKs pERK and p38, with the subsequent activation of (p50) NF-κB and AP-1 (cJun and cFos), determined by real-time quantitation. The evidence shown here supports that the monomeric soluble form of the VCC in the macrophage acts as a modulator of the innate immune response, which is consistent with the assembly of the NLRP3 inflammasome actively releasing IL-1ß.
Assuntos
NF-kappa B , Vibrio cholerae , Humanos , Animais , Camundongos , NF-kappa B/metabolismo , Fator de Transcrição AP-1/metabolismo , Inflamassomos/metabolismo , Vibrio cholerae/metabolismo , Ativação Transcricional , Citotoxinas/farmacologia , Transdução de Sinais , Macrófagos/metabolismo , Células THP-1 , Interleucina-1beta/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismoRESUMO
Helicobacter pylori (H. pylori) is a Gram-negative bacterium that infects about half of the world's population. H. pylori infection prevails by several mechanisms of adaptation of the bacteria and by its virulence factors including the cytotoxin associated antigen A (CagA). CagA is an oncoprotein that is the protagonist of gastric carcinogenesis associated with prolonged H. pylori infection. In this sense, small regulatory RNAs (sRNAs) are important macromolecules capable of inhibiting and activating gene expression. This function allows sRNAs to act in adjusting to unstable environmental conditions and in responding to cellular stresses in bacterial infections. Recent discoveries have shown that nickel-regulated small RNA (NikS) is a post-transcriptional regulator of virulence properties of H. pylori, including the oncoprotein CagA. Notably, high concentrations of nickel cause the reduction of NikS expression and consequently this increases the levels of CagA. In addition, NikS expression appears to be lower in clinical isolates from patients with gastric cancer when compared to patients without. With that in mind, this minireview approaches, in an accessible way, the most important and current aspects about the role of NikS in the control of virulence factors of H. pylori and the potential clinical repercussions of this modulation.
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Klebsiella oxytoca is a resident of the human gut. However, certain K. oxytoca toxigenic strains exist that secrete the nonribosomal peptide tilivalline (TV) cytotoxin. TV is a pyrrolobenzodiazepine that causes antibiotic-associated hemorrhagic colitis (AAHC). The biosynthesis of TV is driven by enzymes encoded by the aroX and NRPS operons. In this study, we determined the effect of environmental signals such as carbon sources, osmolarity, and divalent cations on the transcription of both TV biosynthetic operons. Gene expression was enhanced when bacteria were cultivated in tryptone lactose broth. Glucose, high osmolarity, and depletion of calcium and magnesium diminished gene expression, whereas glycerol increased transcription of both TV biosynthetic operons. The cAMP receptor protein (CRP) is a major transcriptional regulator in bacteria that plays a key role in metabolic regulation. To investigate the role of CRP on the cytotoxicity of K. oxytoca, we compared levels of expression of TV biosynthetic operons and synthesis of TV in wild-type strain MIT 09-7231 and a Δcrp isogenic mutant. In summary, we found that CRP directly activates the transcription of the aroX and NRPS operons and that the absence of CRP reduced cytotoxicity of K. oxytoca on HeLa cells, due to a significant reduction in TV production. This study highlights the importance of the CRP protein in the regulation of virulence genes in enteric bacteria and broadens our knowledge on the regulatory mechanisms of the TV cytotoxin.
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Shiga toxin-producing E. coli (STEC) produces Stx1 and/or Stx2, and Subtilase cytotoxin (SubAB). Since these toxins may be present simultaneously during STEC infections, the purpose of this work was to study the co-action of Stx2 and SubAB. Stx2 + SubAB was assayed in vitro on monocultures and cocultures of human glomerular endothelial cells (HGEC) with a human proximal tubular epithelial cell line (HK-2) and in vivo in mice after weaning. The effects in vitro of both toxins, co-incubated and individually, were similar, showing that Stx2 and SubAB contribute similarly to renal cell damage. However, in vivo, co-injection of toxins lethal doses reduced the survival time of mice by 24 h and mice also suffered a strong decrease in the body weight associated with a lowered food intake. Co-injected mice also exhibited more severe histological renal alterations and a worsening in renal function that was not as evident in mice treated with each toxin separately. Furthermore, co-treatment induced numerous erythrocyte morphological alterations and an increase of free hemoglobin. This work shows, for the first time, the in vivo effects of Stx2 and SubAB acting together and provides valuable information about their contribution to the damage caused in STEC infections.
Assuntos
Proteínas de Escherichia coli/toxicidade , Síndrome Hemolítico-Urêmica/etiologia , Toxina Shiga II/toxicidade , Subtilisinas/toxicidade , Animais , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Técnicas de Cocultura , Células Endoteliais/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Síndrome Hemolítico-Urêmica/patologia , Humanos , Rim/efeitos dos fármacos , Rim/patologia , Glomérulos Renais/citologia , Túbulos Renais Proximais/citologia , Masculino , Camundongos Endogâmicos BALB CRESUMO
Hemolytic uremic syndrome (HUS) is a consequence of Shiga toxin (Stx)-producing Escherichia coli (STEC) infection and is the most frequent cause of acute renal failure (ARF) in children. Subtilase cytotoxin (SubAB) has also been associated with HUS pathogenesis. We previously reported that Stx2 and SubAB cause different effects on co-cultures of human renal microvascular endothelial cells (HGEC) and human proximal tubular epithelial cells (HK-2) relative to HGEC and HK-2 monocultures. In this work we have analyzed the secretion of pro-inflammatory cytokines by co-cultures compared to monocultures exposed or not to Stx2, SubAB, and Stx2+SubAB. Under basal conditions, IL-6, IL-8 and TNF-α secretion was different between monocultures and co-cultures. After toxin treatments, high concentrations of Stx2 and SubAB decreased cytokine secretion by HGEC monocultures, but in contrast, low toxin concentrations increased their release. Toxins did not modulate the cytokine secretion by HK-2 monocultures, but increased their release in the HK-2 co-culture compartment. In addition, HK-2 monocultures were stimulated to release IL-8 after incubation with HGEC conditioned media. Finally, Stx2 and SubAB were detected in HGEC and HK-2 cells from the co-cultures. This work describes, for the first time, the inflammatory responses induced by Stx2 and SubAB, in a crosstalk model of renal endothelial and epithelial cells.
Assuntos
Citocinas/metabolismo , Células Endoteliais/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Proteínas de Escherichia coli/toxicidade , Túbulos Renais Proximais/efeitos dos fármacos , Microvasos/efeitos dos fármacos , Toxina Shiga II/toxicidade , Subtilisinas/toxicidade , Comunicação Celular/efeitos dos fármacos , Comunicação Celular/imunologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/imunologia , Células Cultivadas , Técnicas de Cocultura , Sinergismo Farmacológico , Células Endoteliais/imunologia , Células Epiteliais/imunologia , Síndrome Hemolítico-Urêmica , Humanos , Rim/irrigação sanguíneaRESUMO
BACKGROUND: Cnidarian venoms and extracts have shown a broad variety of biological activities including cytotoxic, antibacterial and antitumoral effects. Most of these studied extracts were obtained from sea anemones or jellyfish. The present study aimed to determine the toxic activity and assess the antitumor and antiparasitic potential of Palythoa caribaeorum venom by evaluating its in vitro toxicity on several models including human tumor cell lines and against the parasite Giardia intestinalis. METHODS: The presence of cytolysins and vasoconstrictor activity of P. caribaeorum venom were determined by hemolysis, PLA2 and isolated rat aortic ring assays, respectively. The cytotoxic effect was tested on HCT-15 (human colorectal adenocarcinoma), MCF-7 (human mammary adenocarcinoma), K562 (human chronic myelogenous leukemia), U251 (human glyoblastoma), PC-3 (human prostatic adenocarcinoma) and SKLU-1 (human lung adenocarcinoma). An in vivo toxicity assay was performed with crickets and the antiparasitic assay was performed against G. intestinalis at 24 h of incubation. RESULTS: P. caribaeorum venom produced hemolytic and PLA2 activity and showed specific cytotoxicity against U251 and SKLU-1 cell lines, with approximately 50% growing inhibition. The venom was toxic to insects and showed activity against G. intestinalis in a dose-dependent manner by possibly altering its membrane osmotic equilibrium. CONCLUSION: These results suggest that P. caribaeorum venom contains compounds with potential therapeutic value against microorganisms and cancer.
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Background Cnidarian venoms and extracts have shown a broad variety of biological activities including cytotoxic, antibacterial and antitumoral effects. Most of these studied extracts were obtained from sea anemones or jellyfish. The present study aimed to determine the toxic activity and assess the antitumor and antiparasitic potential of Palythoa caribaeorum venom by evaluating its in vitro toxicity on several models including human tumor cell lines and against the parasite Giardia intestinalis. Methods The presence of cytolysins and vasoconstrictor activity of P. caribaeorum venom were determined by hemolysis, PLA2 and isolated rat aortic ring assays, respectively. The cytotoxic effect was tested on HCT-15 (human colorectal adenocarcinoma), MCF-7 (human mammary adenocarcinoma), K562 (human chronic myelogenous leukemia), U251 (human glyoblastoma), PC-3 (human prostatic adenocarcinoma) and SKLU-1 (human lung adenocarcinoma). An in vivo toxicity assay was performed with crickets and the antiparasitic assay was performed against G. intestinalis at 24 h of incubation. Results P. caribaeorum venom produced hemolytic and PLA2 activity and showed specific cytotoxicity against U251 and SKLU-1 cell lines, with approximately 50% growing inhibition. The venom was toxic to insects and showed activity against G. intestinalis in a dose-dependent manner by possibly altering its membrane osmotic equilibrium. Conclusion These results suggest that P. caribaeorum venom contains compounds with potential therapeutic value against microorganisms and cancer.(AU)
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Animais , Citotoxinas/análise , Venenos de Cnidários/toxicidade , Venenos de Cnidários/uso terapêutico , Venenos de Cnidários/efeitos adversos , Antígenos de Protozoários/análise , Antígenos de Neoplasias/análise , Ensaios de Seleção de Medicamentos AntitumoraisRESUMO
Background Cnidarian venoms and extracts have shown a broad variety of biological activities including cytotoxic, antibacterial and antitumoral effects. Most of these studied extracts were obtained from sea anemones or jellyfish. The present study aimed to determine the toxic activity and assess the antitumor and antiparasitic potential of Palythoa caribaeorum venom by evaluating its in vitro toxicity on several models including human tumor cell lines and against the parasite Giardia intestinalis. Methods The presence of cytolysins and vasoconstrictor activity of P. caribaeorum venom were determined by hemolysis, PLA2 and isolated rat aortic ring assays, respectively. The cytotoxic effect was tested on HCT-15 (human colorectal adenocarcinoma), MCF-7 (human mammary adenocarcinoma), K562 (human chronic myelogenous leukemia), U251 (human glyoblastoma), PC-3 (human prostatic adenocarcinoma) and SKLU-1 (human lung adenocarcinoma). An in vivo toxicity assay was performed with crickets and the antiparasitic assay was performed against G. intestinalis at 24 h of incubation. Results P. caribaeorum venom produced hemolytic and PLA2 activity and showed specific cytotoxicity against U251 and SKLU-1 cell lines, with approximately 50% growing inhibition. The venom was toxic to insects and showed activity against G. intestinalis in a dose-dependent manner by possibly altering its membrane osmotic equilibrium. Conclusion These results suggest that P. caribaeorum venom contains compounds with potential therapeutic value against microorganisms and cancer.
Assuntos
Animais , Antígenos de Neoplasias/análise , Antígenos de Protozoários/análise , Citotoxinas/análise , Venenos de Cnidários/efeitos adversos , Venenos de Cnidários/toxicidade , Venenos de Cnidários/uso terapêutico , Ensaios de Seleção de Medicamentos AntitumoraisRESUMO
Cnidarian venoms and extracts have shown a broad variety of biological activities including cytotoxic, antibacterial and antitumoral effects. Most of these studied extracts were obtained from sea anemones or jellyfish. The present study aimed to determine the toxic activity and assess the antitumor and antiparasitic potential of Palythoa caribaeorum venom by evaluating its in vitro toxicity on several models including human tumor cell lines and against the parasite Giardia intestinalis. Methods: The presence of cytolysins and vasoconstrictor activity of P. caribaeorum venom were determined by hemolysis, PLA2 and isolated rat aortic ring assays, respectively. The cytotoxic effect was tested on HCT-15 (human colorectal adenocarcinoma), MCF-7 (human mammary adenocarcinoma), K562 (human chronic myelogenous leukemia), U251 (human glyoblastoma), PC-3 (human prostatic adenocarcinoma) and SKLU-1 (human lung adenocarcinoma). An in vivo toxicity assay was performed with crickets and the antiparasitic assay was performed against G. intestinalis at 24 h of incubation. Results: P. caribaeorum venom produced hemolytic and PLA2 activity and showed specific cytotoxicity against U251 and SKLU-1 cell lines, with approximately 50% growing inhibition. The venom was toxic to insects and showed activity against G. intestinalis in a dose-dependent manner by possibly altering its membrane osmotic equilibrium. Conclusion: These results suggest that P. caribaeorum venom contains compounds with potential therapeutic value against microorganisms and cancer.(AU)
Assuntos
Animais , Masculino , Ratos , Giardíase/terapia , Giardia lamblia/parasitologia , Venenos de Cnidários/antagonistas & inibidores , Venenos de Cnidários/toxicidade , Anticarcinógenos , Ratos Wistar , Venenos de Cnidários/uso terapêutico , HemolíticosRESUMO
Hemolytic uremic syndrome (HUS) is one of the most common causes of acute renal failure in children. The majority of cases are associated with Shiga toxin (Stx)-producing Escherichia coli (STEC). In Argentina, HUS is endemic and presents the highest incidence rate in the world. STEC strains expressing Stx type 2 (Stx2) are responsible for the most severe cases of this pathology. Subtilase cytotoxin (SubAB) is another STEC virulence factor that may contribute to HUS pathogenesis. To date, neither a licensed vaccine nor effective therapy for HUS is available for humans. Considering that Ouabain (OUA) may prevent the apoptosis process, in this study we evaluated if OUA is able to avoid the damage caused by Stx2 and SubAB on human glomerular endothelial cells (HGEC) and the human proximal tubule epithelial cell (HK-2) line. HGEC and HK-2 were pretreated with OUA and then incubated with the toxins. OUA protected the HGEC viability from Stx2 and SubAB cytotoxic effects, and also prevented the HK-2 viability from Stx2 effects. The protective action of OUA on HGEC and HK-2 was associated with a decrease in apoptosis and an increase in cell proliferation. Our data provide evidence that OUA could be considered as a therapeutic strategy to avoid the renal damage that precedes HUS.
Assuntos
Proteínas de Escherichia coli/toxicidade , Ouabaína/farmacologia , Substâncias Protetoras/farmacologia , Toxina Shiga II/toxicidade , Subtilisinas/toxicidade , Apoptose/efeitos dos fármacos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Humanos , Rim/citologia , Necrose/induzido quimicamente , Necrose/prevenção & controleRESUMO
The interleukin-17 (IL-17) family of cytokines (IL-17A to IL-17F) is involved in many inflammatory diseases. Although IL-17A is recognized as being involved in the pathophysiology of Helicobacter pylori-associated diseases, the role of other IL-17 cytokine family members remains unclear. Microarray analysis of IL-17 family cytokines was performed in H. pylori-infected and uninfected gastric biopsy specimens. IL-17C mRNA was upregulated approximately 4.5-fold in H. pylori-infected gastric biopsy specimens. This was confirmed by quantitative reverse transcriptase PCR in infected and uninfected gastric mucosa obtained from Bhutan and from the Dominican Republic. Immunohistochemical analysis showed that IL-17C expression in H. pylori-infected gastric biopsy specimens was predominantly localized to epithelial and chromogranin A-positive endocrine cells. IL-17C mRNA levels were also significantly greater among cagA-positive than cagA-negative H. pylori infections (P = 0.012). In vitro studies confirmed an increase in IL-17C mRNA and protein levels in cells infected with cagA-positive infections compared to cells infected with either cagA-negative or cag pathogenicity island (PAI) mutant. Chemical inhibition of IκB kinase (IKK), mitogen-activated protein extracellular signal-regulated kinase (MEK), and Jun N-terminal kinase (JNK) inhibited induction of IL-17C proteins in infected cells, whereas p38 inhibition had no effect on IL-17C protein secretion. In conclusion, H. pylori infection was associated with a significant increase in IL-17C expression in human gastric mucosa. The role of IL-17C in the pathogenesis of H. pylori-induced diseases remains to be determined.
Assuntos
Mucosa Gástrica/imunologia , Gastrite/imunologia , Infecções por Helicobacter/imunologia , Helicobacter pylori/imunologia , Interleucina-17/genética , Interleucina-17/imunologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteínas de Bactérias/genética , Butão , Linhagem Celular , República Dominicana , Feminino , Mucosa Gástrica/metabolismo , Mucosa Gástrica/microbiologia , Mucosa Gástrica/patologia , Gastrite/microbiologia , Gastrite/fisiopatologia , Redes Reguladoras de Genes , Ilhas Genômicas , Genótipo , Infecções por Helicobacter/epidemiologia , Infecções por Helicobacter/microbiologia , Helicobacter pylori/genética , Humanos , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Regulação para Cima , Adulto JovemRESUMO
Subtilase cytotoxin (SubAB) is a member of the AB5 cytotoxin family and is produced by certain strains of Shiga toxigenic Escherichia coli. The toxin is known to be lethal to mice, but the pathological mechanisms that contribute to Uremic Hemolytic Syndrome (HUS) are poorly understood. In this study we show that intraperitoneal injection of a sublethal dose of SubAB in rats triggers a systemic response, with ascitic fluid accumulation, heart hypertrophy and damage to the liver, colon and kidney. SubAB treated rats presented microalbuminuria 20 days post inoculation. At this time we found disruption of the glomerular filtration barrier and alteration of the protein reabsorption mechanisms of the proximal tubule. In the kidney, SubAB also triggered an epithelial to mesenchymal transition (Wuyts et al., 1996). These findings indicate that apart from direct cytotoxic effects on renal tissues, SubAB causes significant damage to the other organs, with potential consequences for HUS pathogenesis. IMPORTANCE: Uremic Hemolytic Syndrome is an endemic disease in Argentina, with over 400 hundred new cases each year. We have previously described renal effects of Shiga Toxin and its ability to alter renal protein handling. Bearing in mind that Subtilase Cytotoxin is an emerging pathogenic factor, that it is not routinely searched for in patients with HUS, and that to the date its systemic effects have not been fully clarified we decided to study both its systemic effects, and its renal effects to assess whether SubAB could be contributing to pathology seen in children.
Assuntos
Proteínas de Escherichia coli/metabolismo , Escherichia coli Shiga Toxigênica/metabolismo , Subtilisinas/metabolismo , Albuminúria/induzido quimicamente , Animais , Ascite/induzido quimicamente , Cardiomegalia/induzido quimicamente , Colo/efeitos dos fármacos , Colo/patologia , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Proteínas de Escherichia coli/toxicidade , Síndrome Hemolítico-Urêmica/etiologia , Rim/efeitos dos fármacos , Rim/metabolismo , Rim/patologia , Fígado/efeitos dos fármacos , Fígado/patologia , Masculino , Ratos Sprague-Dawley , Subtilisinas/toxicidadeRESUMO
The Lansberg's hognose pitviper, Porthidium lansbergii lansbergii, inhabits northern Colombia. A recent proteomic characterization of its venom (J. Proteomics [2015] 114, 287-299) revealed the presence of phospholipases A2 (PLA2) accounting for 16.2% of its proteins. The two most abundant PLA2s were biochemically and functionally characterized. Pllans-I is a basic, dimeric enzyme with a monomer mass of 14,136 Da, while Pllans-II is an acidic, monomeric enzyme of 13,901 Da. Both have Asp49 in their partial amino acid sequences and, accordingly, are catalytically active upon natural or synthetic substrates. Nevertheless, these two enzymes differ markedly in their bioactivities. Pllans-I induces myonecrosis, edema, and is lethal by intracerebro-ventricular injection in mice, as well as cytolytic and anticoagulant in vitro. In contrast, Pllans-II is devoid of these effects, except for the induction of a moderate edema. In spite of lacking myotoxicity, Pllans-II enhances the muscle damaging action of Pllans-I in vivo. Altogether, results further illustrate the divergent functional profiles of basic and acidic PLA2s in viperid venoms, and suggest that Pllans-I plays a myotoxic role in envenomings by P. l. lansbergii, whereas Pllans-II, apparently devoid of toxicity, enhances muscle damage caused by Pllans-I.
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Fosfolipases A2/metabolismo , Venenos de Víboras/enzimologia , Sequência de Aminoácidos , Animais , Linhagem Celular , Cromatografia Líquida de Alta Pressão , Cromatografia de Fase Reversa , Eletroforese em Gel de Poliacrilamida , Feminino , Concentração de Íons de Hidrogênio , Masculino , Camundongos , Fosfolipases A2/química , Fosfolipases A2/toxicidade , Homologia de Sequência de Aminoácidos , Venenos de Víboras/toxicidade , ViperidaeRESUMO
Culture supernatant of sepsis-associated Escherichia coli (SEPEC) isolated from patients with sepsis caused loss of intercellular junctions and elongation of human umbilical vein endothelial cells (HUVEC). The cytotoxic factor was purified from culture supernatant of SEPEC 15 (serogroup O153) by liquid chromatography process. PAGE (polyacrylamide gel electrophoresis) showed that the purified SEPEC cytotoxic factor had a molecular mass of â¼150kDa and consisted of at least two subunits. At the concentration of 1 CD50 (40µg/mL) did facilitate transcytosis through the HUVEC cells monolayer of SEPEC 15 as much as E. coli K12 within 30min without affecting cell viability. These results suggest that this cytotoxic factor, named as SPF (SEPEC's permeabilizing factor), may be an important SEPEC virulence factor that facilitates bacterial access to the bloodstream.
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Citotoxinas/metabolismo , Células Epiteliais/microbiologia , Escherichia coli , Sepse/microbiologia , Toxinas Bacterianas/toxicidade , Citotoxinas/toxicidade , Impedância Elétrica , Escherichia coli/patogenicidade , Infecções por Escherichia coli/microbiologia , Humanos , Fatores de VirulênciaRESUMO
Introduction This study compares virulence markers of Helicobacter pylori isolated from patients in 2 cities in the Brazilian Amazon. Methods The study analyzed 168 patients with chronic gastritis from Belém and 151 from Bragança, State of Pará, Brazil. Levels of bacterial DNA associated with cagA and vacA alleles were checked by PCR, and hematoxylin-eosin staining was used for histologic diagnosis. Results In Bragança 87% of patients were genotype s1m1 cagA-positive (s1m1 cagA+), compared with 76% in Belém. In samples from patients in both cities, there was an association between s1m1 cagA+ strains and gastric mucosal damage. Conclusions Both cities have a high frequency of s1m1 cagA+ strains of H. pylori. .
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Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Humanos , Pessoa de Meia-Idade , Adulto Jovem , Antígenos de Bactérias/genética , Proteínas de Bactérias/genética , Gastrite/microbiologia , Infecções por Helicobacter/microbiologia , Helicobacter pylori/patogenicidade , Brasil , Biomarcadores/análise , Doença Crônica , DNA Bacteriano/genética , Genótipo , Helicobacter pylori/genética , Helicobacter pylori/isolamento & purificação , Reação em Cadeia da Polimerase , Fatores de Virulência/genéticaRESUMO
Com o objetivo de avaliar a produção das hemolisinas alfa, beta e delta, foram analisadas 78 amostras de Staphylococcus aureusisoladas de casos de mastite bovinasubclínica. Em ágar sangue de ovino e/ouequino,100% das amostras apresentaram atividade hemolítica, com 58 amostras produzindo a toxina alfa; 73 a delta e 76 a beta, correspondendo a 74,4, 93,6 e 97,4% das amostras, respectivamente. A análise da produção combinada mostrou que em 70,5% das amostras houve co-produção de alfa+beta+delta; em 20,5% beta+delta; em 2,6% alfa+beta; e em 1,3% alfa+delta. A produção de um único tipo de toxina foi observada em apenas quatro amostras, sendo três (3,8%) produtoras datoxina beta e uma (1,3%) amostra da toxina delta. A mensuração do diâmetro dos halos hemolíticos indicou produção de uma grande quantidade de toxina. Além disso, a maioria das amostras (70,5%) demonstrou atividade hemolítica às 24 horas de incubação. Estes resultados demonstraram que vacas com mastite subclínica são um importante reservatório de Staphylococcus aureuscom habilidade de produzir hemolisinas.
Alpha, beta and delta hemolysins were evaluated in 78 samples of Staphylococcus aureusisolated from bovine subclinical mastitis. On sheep and/or horse blood agar,100% of the isolates produced hemolytic activity and 58 were positive for alfa-toxin; 73 for delta-toxin and 76 for beta-toxin, which accounted for 74.4, 93.6 and 97.4% of the isolates, respectively. The production of alpha+beta+deltatoxins was observed in 70.5% of the isolates; beta+deltain 20.5%; alpha+betain 2.6%; and alpha+deltain 1.3%. Four isolates produced one toxin type alone with three (3.8%) of them producing beta-toxin and one (1.3%) delta-toxin. The measurementof the hemolytic area showed a high amount of toxin production. Also, the majority of isolates (70.5%) produced hemolysis at 24 hours after incubation. Theseresultsdemonstrated thatcows with a subclinical mastitis are an important reservoir of hemolytic Staphylococcus aureus.
Assuntos
Animais , Bovinos , Staphylococcus aureus/isolamento & purificação , Doenças dos Bovinos , Proteínas Hemolisinas , Glândulas Mamárias Animais/anormalidades , Mastite BovinaRESUMO
AIM: To investigate age, sex, histopathology and Helicobacter pylori (H. pylori) status, as risk factors for gastroduodenal disease outcome in Brazilian dyspeptic patients. METHODS: From all 1466 consecutive dyspeptic patients submitted to upper gastroscopy at Hospital das Clinicas of Marilia, antral biopsy specimens were obtained and subjected to histopathology and H. pylori diagnosis. All patients presenting chronic gastritis (CG) and peptic ulcer (PU) disease localized in the stomach, gastric ulcer (GU) and/or duodenal ulcer (DU) were included in the study. Gastric biopsies (n = 668) positive for H. pylori by rapid urease test were investigated for vacuolating cytotoxin A (vacA) medium (m) region mosaicism by polymerase chain reaction. Logistic regression analysis was performed to verify the association of age, sex, histopathologic alterations, H. pylori diagnosis and vacA m region mosaicism with the incidence of DU, GU and CG in patients. RESULTS: Of 1466 patients submitted to endoscopy, 1060 (72.3%) presented CG [male/female = 506/554; mean age (year) ± SD = 51.2 ± 17.81], 88 (6.0%) presented DU [male/female = 54/34; mean age (year) ± SD = 51.4 ± 17.14], and 75 (5.1%) presented GU [male/female = 54/21; mean age (year) ± SD = 51.3 ± 17.12] and were included in the comparative analysis. Sex and age showed no detectable effect on CG incidence (overall χ² = 2.1, P = 0.3423). Sex [Odds ratios (OR) = 1.8631, P = 0.0058] but not age (OR = 0.9929, P = 0.2699) was associated with DU and both parameters had a highly significant effect on GU (overall χ² = 30.5, P < 0.0001). The histopathological results showed a significant contribution of ageing for both atrophy (OR = 1.0297, P < 0.0001) and intestinal metaplasia (OR = 1.0520, P < 0.0001). Presence of H. pylori was significantly associated with decreasing age (OR = 0.9827, P < 0.0001) and with the incidence of DU (OR = 3.6077, P < 0.0001). The prevalence of m1 in DU was statistically significant (OR = 2.3563, P = 0.0018) but not in CG (OR = 2.678, P = 0.0863) and GU (OR = 1.520, P= 0.2863). CONCLUSION: In our population, male gender was a risk factor for PU; ageing for GU, atrophy and metaplasia; and H. pylori of vacA m1 genotype for DU.
Assuntos
Gastrite/etiologia , Úlcera Péptica/etiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Brasil/epidemiologia , Doença Crônica , Feminino , Gastrite/diagnóstico , Gastrite/epidemiologia , Humanos , Incidência , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Úlcera Péptica/diagnóstico , Úlcera Péptica/epidemiologia , Fatores de RiscoRESUMO
A mamoneira (Ricinus communis L.) é uma oleaginosa de alto valor econômico pelo fato de apresentar um mercado bem definido para o óleo extraído de suas sementes. A torta, que é um resíduo desta extração, se destaca pelo alto teor em proteínas. Dentre as proteínas encontradas na torta destaca-se a ricina, uma citotoxina, que inviabiliza sua utilização como fonte protéica alternativa para alimentação animal. O presente trabalho tem como objetivo identificar um melhor tratamento experimental para a extração de ricina da torta de mamona, visando futuros estudos de perda de integridade da ricina, o que garantiria a inocuidade do produto. Para tanto, buscou-se identificar a solução de maior capacidade de extração de proteínas, empregando a metodologia de superfície de resposta. Um delineamento composto central rotacional foi elaborado a fim de verificar o melhor pH e concentração de NaCl para a extração. Dos cinco diferentes valores de pH (4,0; 4,6; 6,0; 7,4; 8,0) e concentração de NaCl (0,0M; 0,3M; 1,0M; 1,7M; 2,0M) utilizados, o tratamento associando fosfato de potássio 0,2M/NaCl 1,7M pH 7,4 foi escolhido como o melhor. A concentração de proteína extraída neste tratamento chegou a valores quatro vezes maiores que o encontrado no de mínima extração de proteína. Pela evidenciação do gel de eletroforese não houve extração preferencial de ricina nos tratamentos testados, entretanto etapas de purificação usando diálise e precipitação com sulfato de amônio, permitiram uma evidenciação melhor das duas cadeias polipeptídicas de ricina.
Castor bean (Ricinus communis L.) is an oilseed crop of high economic value due to the fact of presenting a clearly defined market for the oil extracted from its seeds. Castor cake, which is a residue of oil extraction, is at the moment receiving special attention because of its high protein content. However, among the proteins found in this cake it is observed the presence of ricin, a cytotoxin, which turns this seed dangerous to be used as an alternative protein source for animal feed. The present research has the objective of identifying the best experimental treatment for extraction of ricin from castor cake, with the aim of future studies of loss of ricin integrity, which would ensure the safety of the product. With this aim, it was initiated the search for a buffer of higher capacity of proteins extraction, using the response surface methodology. A central composite design was developed in order to determine the best pH and NaCl concentration for extraction. Of the five different pH values (4.0, 4.6, 6.0, 7.4, 8.0) and NaCl (0.0M, 0.3M, 1.0M, 1.7M, 2.0M) used, the treatment containing 0.2M potassium phosphate / 1.7M NaCl pH 7.4 was chosen as the best. The amount of protein extracted in this treatment reached values four times larger than the minimum found in other treatment studied. The electrophoresis analysis showed that there was no preferential extraction of ricin in the treatments; however purification steps using dialysis and precipitation with ammonium sulfate led to a better resolution of the two polypeptide chains presents in ricin.