RESUMO
In December 2021, the United States Food and Drug Administration (FDA) issued the final guidance for industry titled Pathology Peer Review in Nonclinical Toxicology Studies: Questions and Answers. The stated purpose of the FDA guidance is to provide information to sponsors, applicants, and nonclinical laboratory personnel regarding the management and conduct of histopathology peer review as part of nonclinical toxicology studies conducted in compliance with good laboratory practice (GLP) regulations. On behalf of and in collaboration with global societies of toxicologic pathology and the Society of Quality Assurance, the Scientific and Regulatory Policy Committee (SRPC) of the Society of Toxicologic Pathology (STP) initiated a review of this FDA guidance. The STP has previously published multiple papers related to the scientific conduct of a pathology peer review of nonclinical toxicology studies and appropriate documentation practices. The objectives of this review are to provide an in-depth analysis and summary interpretation of the FDA recommendations and share considerations for the conduct of pathology peer review in nonclinical toxicology studies that claim compliance to GLP regulations. In general, this working group is in agreement with the recommendations from the FDA guidance that has added clear expectations for pathology peer review preparation, conduct, and documentation.
Assuntos
Patologia , Revisão por Pares , Toxicologia , United States Food and Drug Administration , Estados Unidos , Toxicologia/normas , Toxicologia/legislação & jurisprudência , Toxicologia/métodos , Revisão por Pares/normas , Patologia/normas , Guias como Assunto , Animais , Testes de Toxicidade/normas , Testes de Toxicidade/métodosRESUMO
Samples of biologic specimens and their derivatives (eg, wet tissues, paraffin-embedded tissue blocks, histology slides, frozen tissues, whole blood, serum/plasma, and urine) are routinely collected during the course of nonclinical toxicity studies. Good Laboratory Practice regulations and/or guidance specify minimum requirements for specimen retention duration, with the caveat that retention of biologic specimens need not extend beyond the duration of sample stability. However, limited availability of published data regarding stability for various purposes following storage of each specimen type has resulted in confusion, uncertainty, and inconsistency as to the appropriate duration for storage of these specimens. To address these issues, a working group of the Society of Toxicologic Pathology Scientific and Regulatory Policy Committee was formed to review published information, regulations, and guidance pertinent to this topic and to summarize the current practices and rationales for retention duration through a survey-based approach. Information regarding experiences reaccessing biologic specimens and performing sample stability investigations was also collected. Based on this combined information, the working group developed several points to consider that may be referenced when developing or revising sample retention practices. [Box: see text].
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Políticas , Projetos de PesquisaRESUMO
Anatomic pathology and clinical pathology end points are standard components of almost every nonclinical general toxicity study conducted during the risk assessment of novel pharmaceuticals and chemicals. On occasion, an ultrastructural pathology evaluation using transmission electron microscopy (TEM) may be included in nonclinical toxicity studies. Transmission electron microscopy is most commonly used when a light microscopic finding may require further characterization that could inform on the pathogenesis and/or mechanism of action. Regulatory guidance do not address the use of TEM in general study designs nor whether these assessments should be performed in laboratories conducted in compliance with Good Laboratory Practices. The Scientific and Regulatory Policy Committee of the Society of Toxicologic Pathology (STP) formed a Working Group to assess the current practices on the use of TEM in nonclinical toxicity studies. The Working Group constructed a survey sent to members of societies of toxicologic pathology in the United States, Europe, Britain, and Japan, and responses were collected through the STP for evaluation by the Working Group. The survey results and regulatory context are discussed, as are "points to consider" from the collective experience of the Working Group. This survey indicates that TEM remains an essential diagnostic option for complementing toxicologic pathology evaluations. *This Points to Consider article is a product of a Society of Toxicologic Pathology (STP) Working Group commissioned by the Scientific and Regulatory Policy Committee (SRPC) of the STP. It has been reviewed and approved by the SRPC and Executive Committee of the STP but it does not represent a formal Best Practice recommendation of the Society; rather, it is intended to provide key "points to consider" in designing nonclinical studies or interpreting data from toxicity and safety studies intended to support regulatory submissions. The points expressed in this document are those of the authors and do not reflect views or policies of the employing institutions. Readers of Toxicologic Pathology are encouraged to send their thoughts on these articles or ideas for new topics to the Editor.
Assuntos
Microscopia Eletrônica de Transmissão , Patologia Clínica/métodos , Toxicologia/métodos , Comitês Consultivos , Animais , Avaliação Pré-Clínica de Medicamentos/métodos , Avaliação Pré-Clínica de Medicamentos/normas , Guias como Assunto , Humanos , Microscopia Eletrônica de Transmissão/métodos , Microscopia Eletrônica de Transmissão/normas , Patologia Clínica/legislação & jurisprudência , Patologia Clínica/normas , Sociedades Científicas , Testes de Toxicidade/métodos , Testes de Toxicidade/normas , Toxicologia/legislação & jurisprudência , Toxicologia/normas , Estados Unidos , United States Food and Drug AdministrationRESUMO
Short interfering RNAs (siRNAs) and antisense oligonucleotides (ASOs) are the most clinically advanced oligonucleotide-based platforms. A number of N-acetylgalactosamine (GalNAc)-conjugated siRNAs (GalNAc-siRNAs), also referred to as RNA interference (RNAi) therapeutics, are currently in various stages of development, though none is yet approved. While the safety of ASOs has been the subject of extensive review, the nonclinical safety profiles of GalNAc-siRNAs have not been reported. With the exception of sequence differences that confer target RNA specificity, GalNAc-siRNAs are largely chemically uniform, containing limited number of phosphorothioate linkages, and 2'-O-methyl and 2'-deoxy-2'-fluoro ribose modifications. Here, we present the outcomes of short-term (3-5 week) rat and monkey weekly repeat-dose toxicology studies of six Enhanced Stabilization Chemistry GalNAc-siRNAs currently in clinical development. In nonclinical studies at supratherapeutic doses, these molecules share similar safety signals, with histologic findings in the organ of pharmacodynamic effect (liver), the organ of elimination (kidney), and the reticuloendothelial system (lymph nodes). The majority of these changes are nonadverse, partially to completely reversible, correlate well with pharmacokinetic parameters and tissue distribution, and often reflect drug accumulation. Furthermore, all GalNAc-siRNAs tested to date have been negative in genotoxicity and safety pharmacology studies.
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Acetilgalactosamina/toxicidade , Aberrações Cromossômicas/induzido quimicamente , Fígado/efeitos dos fármacos , RNA Interferente Pequeno/toxicidade , Acetilgalactosamina/química , Acetilgalactosamina/farmacologia , Animais , Células CHO , Cricetulus , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Fígado/patologia , Linfócitos/efeitos dos fármacos , Linfócitos/patologia , Macaca fascicularis , Testes de Mutagenicidade , RNA Interferente Pequeno/química , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Ratos Sprague-Dawley , Especificidade da Espécie , Testes de Toxicidade SubagudaRESUMO
Doxorubicin (DOX) is a potent and effective broad-spectrum anthracycline antitumor agent, but its clinical usefulness is restricted by cardiotoxicity. This study compared pharmacokinetic, functional, structural and biochemical effects of single dose DOX bolus or 3-h continuous iv infusion (3-h iv) in the HanWistar rat to characterize possible treatment-related differences in drug safety over a 72 h observation period. Both DOX dosing paradigms significantly altered blood pressure, core body temperature and QA interval (indirect measure of cardiac contractility); however, there was no recovery observed in the bolus iv treatment group. Following the 3-h iv treatment, blood pressures and QA interval normalized by 36 h then rose above baseline levels over 72 h. Both treatments induced biphasic changes in heart rate with initial increases followed by sustained decreases. Cardiac injury biomarkers in plasma were elevated only in the bolus iv treatment group. Tissue cardiac injury biomarkers, cardiac mitochondrial complexes I, III and V and cardiac mitochondrial sphingolipids were decreased only in the bolus iv treatment group. Results indicate that each DOX dosing paradigm deregulates sinus rhythm.However, slowing the rate of infusion allows for functional compensation of blood pressure and may decrease the likelihood of cardiac myocyte necrosis via a mechanism associated with reduced mitochondrial damage.
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Doxorrubicina/administração & dosagem , Coração/efeitos dos fármacos , Rim/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/patologia , Esfingolipídeos/metabolismo , Administração Intravenosa/métodos , Animais , Biomarcadores/metabolismo , Pressão Sanguínea/efeitos dos fármacos , Esquema de Medicação , Coração/fisiopatologia , Rim/metabolismo , Rim/patologia , Ratos , Ratos WistarRESUMO
Colistin and polymyxin B are effective treatment options for Gram-negative resistant bacteria but are used as last-line therapy due to their dose-limiting nephrotoxicity. A critical factor in developing safer polymyxin analogues is understanding accumulation of the drugs and their metabolites, which is currently limited due to the lack of effective techniques for analysis of these challenging molecules. Mass spectrometry imaging (MSI) allows direct detection of targets (drugs, metabolites, and endogenous compounds) from tissue sections. The presented study exemplifies the utility of MSI by measuring the distribution of polymyxin B1, colistin, and polymyxin B nonapeptide (PMBN) within dosed rat kidney tissue sections. The label-free MSI analysis revealed that the nephrotoxic compounds (polymyxin B1 and colistin) preferentially accumulated in the renal cortical region. The less nephrotoxic analogue, polymyxin B nonapeptide, was more uniformly distributed throughout the kidney. In addition, metabolites of the dosed compounds were detected by MSI. Kidney homogenates were analyzed using LC/MS/MS to determine total drug exposure and for metabolite identification. To our knowledge, this is the first time such techniques have been utilized to measure the distribution of polymyxin drugs and their metabolites. By simultaneously detecting the distribution of drug and drug metabolites, MSI offers a powerful alternative to tissue homogenization analysis and label or antibody-based imaging.
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Rim/efeitos dos fármacos , Polimixinas/toxicidade , Espectrometria de Massas por Ionização por Electrospray/métodos , Animais , Cromatografia Líquida , Masculino , Polimixinas/farmacocinética , Ratos , Ratos WistarRESUMO
Recombinant interleukin-2 (rIL-2) administration in oncology indications is hampered by vascular toxicity, which presents as a vascular leak syndrome. We used this aspect of the toxicity of rIL-2 to evaluate candidate biomarkers of drug-induced vascular injury (DIVI) in rats given 0.36 mg/kg rIL-2 daily. Groups of rats were given either 2 or 5 doses of rIL-2 or 5 doses of rIL-2 followed by a 7-day recovery. The histomorphologic lexicon and grading scheme developed by the Vascular Injury Working Group of the Predictive Safety Testing Consortium of the Critical Path Institute were utilized to enable semiquantitative integration with circulating biomarker levels. The administration of rIL-2 was associated with time-dependent endothelial cell hyperplasia and hypertrophy and perivascular inflammation that correlated with increases in circulating angiopoietin-2, lipocalin-2, monocyte chemotactic protein-1, tissue inhibitor of metalloproteinase-1, vascular endothelial growth factor A, E-selectin, and chemokine (C-X-C motif) ligand-1, and the microRNAs miR-21, miR-132, and miR-155. The dose groups were differentially identified by panels comprising novel candidate biomarkers and traditional hematologic parameters. These results identify biomarkers of the early stages of DIVI prior to the onset of vascular smooth muscle necrosis.
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Interleucina-2/toxicidade , Lesões do Sistema Vascular/sangue , Lesões do Sistema Vascular/induzido quimicamente , Animais , Biomarcadores/sangue , Imuno-Histoquímica , Hibridização In Situ , Masculino , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase em Tempo Real , Proteínas Recombinantes/toxicidadeRESUMO
OBJECTIVE: Bruton's tyrosine kinase (BTK) plays a critical role in B cell development and function. We recently described a selective BTK inhibitor, RN486, that blocks B cell receptor (BCR) and Fcγ receptor signaling and is efficacious in animal models of arthritis. The aim of this study was to examine the potential efficacy of BTK in systemic lupus erythematosus (SLE), using an NZB × NZW mouse model of spontaneous SLE. METHODS: Mice received RN486 or its vehicle (administered in chow) at a final concentration of 30 mg/kg for 8 weeks, starting at 32 weeks of age. RESULTS: The administration of RN486 completely stopped disease progression, as determined by histologic and functional analyses of glomerular nephritis. The efficacy was associated with striking inhibition of B cell activation, as demonstrated by a significant reduction in CD69 expression in response to BCR crosslinking. RN486 markedly reduced the secretion of IgG anti-double-stranded DNA (anti-dsDNA) secretion, as determined by enzyme-linked immunosorbent and enzyme-linked immunospot assays. Flow cytometric analysis demonstrated depletion of CD138(high) B220(low) plasma cells in the spleen. RN486 inhibited secretion of IgG anti-dsDNA but not IgM anti-dsDNA, suggesting that pharmacologic blockade of BTK resembles the reported transgenic expression of low levels of endogenous BTK in B cells. In addition, RN486 may also impact the effector function of autoantibodies, as evidenced by a significant reduction in immune complex-mediated activation of human monocytes in vitro and down-regulation of the expression of macrophage-related and interferon-inducible genes in both the kidneys and spleens of treated mice. CONCLUSION: Collectively, our data suggest that BTK inhibitors may simultaneously target autoantibody-producing and effector cells in SLE, thus constituting a promising therapeutic alternative for this disease.
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Linfócitos B/patologia , Glomerulonefrite/tratamento farmacológico , Glomérulos Renais/patologia , Lúpus Eritematoso Sistêmico/tratamento farmacológico , Proteínas Tirosina Quinases/antagonistas & inibidores , Tirosina Quinase da Agamaglobulinemia , Animais , Complexo Antígeno-Anticorpo/metabolismo , Antígenos CD/metabolismo , Antígenos de Diferenciação de Linfócitos T/metabolismo , Linfócitos B/efeitos dos fármacos , Linfócitos B/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Regulação para Baixo , Glomerulonefrite/metabolismo , Glomerulonefrite/patologia , Glomérulos Renais/efeitos dos fármacos , Glomérulos Renais/metabolismo , Lectinas Tipo C/metabolismo , Lúpus Eritematoso Sistêmico/metabolismo , Lúpus Eritematoso Sistêmico/patologia , Ativação Linfocitária/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos NZB , Receptores de IgG/metabolismoRESUMO
Ficolins are collagenous lectins that bind N-acetylated glycans and participate in innate immune responses, including phagocytosis and complement activation. Related collagenous lectins such as mannan binding lectin (MBL) and surfactant proteins A and D possess antiviral activity, but this activity has not been demonstrated for ficolins. In these studies, we used purified porcine plasma ficolin alpha and recombinant ficolin alpha to assess their ability to bind and neutralize porcine reproductive and respiratory virus (PRRSV) in various assays. Recombinant ficolin alpha was designed with a C-terminal 6-histidine tag using a pcDNA3.1 expression vector system in CHO K1 cells. Plasma-purified and recombinant ficolin alpha reduced cytopathic effect of PRRSV-infected Marc-145 cells in neutralization assays and inhibited replication of infectious viral particles in a GlcNAc-dependent manner. In vitro replication determined by plaque assay was inhibited in the presence of plasma-purified ficolin alpha and recombinant ficolin. Immunoreactive plasma ficolin alpha and recombinant ficolin alpha also bound PRRSV-coated wells in a GlcNAc-dependent manner. These studies indicate that porcine ficolin can bind and neutralize a common arterivirus that is a major pathogen of swine.
Assuntos
Lectinas/metabolismo , Lectinas/farmacologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/efeitos dos fármacos , Animais , Linhagem Celular , Efeito Citopatogênico Viral/efeitos dos fármacos , Ponto Isoelétrico , Lectinas/sangue , Lectinas/isolamento & purificação , Vírus da Síndrome Respiratória e Reprodutiva Suína/fisiologia , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacologia , Suínos , Ensaio de Placa Viral , FicolinasRESUMO
Small interfering RNAs (siRNAs) conjugated to a trivalent N-acetylgalactosamine (GalNAc) ligand are being evaluated in investigational clinical studies for a variety of indications. The typical development candidate selection process includes evaluation of the most active compounds for toxicity in rats at pharmacologically exaggerated doses. The subset of GalNAc-siRNAs that show rat hepatotoxicity is not advanced to clinical development. Potential mechanisms of hepatotoxicity can be associated with the intracellular accumulation of oligonucleotides and their metabolites, RNA interference (RNAi)-mediated hybridization-based off-target effects, and/or perturbation of endogenous RNAi pathways. Here we show that rodent hepatotoxicity observed at supratherapeutic exposures can be largely attributed to RNAi-mediated off-target effects, but not chemical modifications or the perturbation of RNAi pathways. Furthermore, these off-target effects can be mitigated by modulating seed-pairing using a thermally destabilizing chemical modification, which significantly improves the safety profile of a GalNAc-siRNA in rat and may minimize the occurrence of hepatotoxic siRNAs across species.
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Acetilgalactosamina/química , Fígado/efeitos dos fármacos , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/toxicidade , Acetilgalactosamina/toxicidade , Animais , Fígado/metabolismo , Masculino , Interferência de RNA , RNA Interferente Pequeno/química , RNA Interferente Pequeno/metabolismo , Ratos , Ratos Sprague-DawleyRESUMO
Previous studies showed that low expression of mannan-binding lectin C (MBL-C) in pigs was not due to single-nucleotide polymorphisms (SNPs) in the coding region of pig MBL2. In these studies, we compared the 5' flanking regions of porcine MBL1 (1907 bp) and MBL2 (1880 bp) in normal and diseased pigs with low or high hepatic expression of MBL2. Hepatic expression of MBL-C was very low in all pigs submitted for postmortem diagnosis. In various European pig breeds, a G(-1081)A substitution was linked to very low hepatic MBL-C expression, and was more frequent in diseased pigs. A C(-251)T substitution with less influence on MBL-C expression was more common in various breeds but was not associated with disease. MBL2 polymorphisms were associated with some disease groups and with the presence of some etiologic agents. These findings indicate that some promoter polymorphisms impair MBL-C expression in pigs and may increase their susceptibility to disease.
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Fígado/metabolismo , Lectina de Ligação a Manose/biossíntese , Lectina de Ligação a Manose/genética , Polimorfismo de Nucleotídeo Único , Doenças dos Suínos/genética , Suínos/genética , Animais , Sequência de Bases , Predisposição Genética para Doença , Lectina de Ligação a Manose/imunologia , Dados de Sequência Molecular , Regiões Promotoras Genéticas , Análise de Sequência de DNA , Suínos/imunologia , Suínos/metabolismo , Doenças dos Suínos/imunologiaRESUMO
A 10-year-old, spayed female, obese golden retriever, presented for immune-mediated thrombocytopenia, was successfully managed with the administration of vincristine and prednisone. However, 6 mo after discontinuing corticosteroid therapy because of suspected iatrogenic hyperglucocorticoidism, the patient was presented with multiple, firm, bilaterally symmetric, dermal masses composed histologically of differentiated cortical bone.
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Neoplasias Ósseas/veterinária , Doenças do Cão/diagnóstico , Osteoma/veterinária , Corticosteroides/efeitos adversos , Corticosteroides/uso terapêutico , Animais , Neoplasias Ósseas/diagnóstico , Neoplasias Ósseas/patologia , Doenças do Cão/patologia , Cães , Feminino , Neoplasias Primárias Múltiplas/diagnóstico , Neoplasias Primárias Múltiplas/patologia , Neoplasias Primárias Múltiplas/veterinária , Osteoma/diagnóstico , Osteoma/patologia , Púrpura Trombocitopênica Idiopática/tratamento farmacológico , Púrpura Trombocitopênica Idiopática/veterináriaRESUMO
Collagenous lectins such as mannan-binding lectins (MBLs), ficolins (FCNs), surfactant proteins A and D (SP-A, SP-D), conglutinin (CG), and related ruminant lectins are multimeric proteins with carbohydrate-binding domains aligned in a manner that facilitates binding to microbial surface polysaccharides. MBLs and FCNs are structurally related to C1q, but activate the lectin complement pathway via interaction with MBL-associated serine proteases (MASPs). MBLs, FCNs, and other collagenous lectins also bind to some host macromolecules and contribute to their removal. While there is evidence that some lectins and the lectin complement pathway are conserved in vertebrates, many differences in collagenous lectins have been observed among humans, rodents, and other vertebrates. For example, humans have only one MBL but three FCNs, whereas most other species express two FCNs and two MBLs. Bovidae express CG and other SP-D-related collectins that are not found in monogastric species. Some dysfunctions of human MBL are due to single nucleotide polymorphisms (SNPs) that affect its expression or structure and thereby increase susceptibility to some infections. Collagenous lectins have well-established roles in innate immunity to various microorganisms, so it is possible that some lectin genotypes or induced phenotypes influence resistance to some infectious or inflammatory diseases in animals.
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Colágeno/genética , Colágeno/imunologia , Lectinas/genética , Lectinas/imunologia , Animais , Colágeno/química , Genótipo , Humanos , Imunidade Inata/genética , Lectinas/química , Estrutura Molecular , Fenótipo , Especificidade da Espécie , Vertebrados/genética , Vertebrados/imunologiaRESUMO
BACKGROUND: The reovirus M1 genome segment encodes the mu2 protein, a structurally minor component of the viral core, which has been identified as a transcriptase cofactor, nucleoside and RNA triphosphatase, and microtubule-binding protein. The mu2 protein is the most poorly understood of the reovirus structural proteins. Genome segment sequences have been reported for 9 of the 10 genome segments for the 3 prototypic reoviruses type 1 Lang (T1L), type 2 Jones (T2J), and type 3 Dearing (T3D), but the M1 genome segment sequences for only T1L and T3D have been previously reported. For this study, we determined the M1 nucleotide and deduced mu2 amino acid sequences for T2J, nine other reovirus field isolates, and various T3D plaque-isolated clones from different laboratories. RESULTS: Determination of the T2J M1 sequence completes the analysis of all ten genome segments of that prototype. The T2J M1 sequence contained a 1 base pair deletion in the 3' non-translated region, compared to the T1L and T3D M1 sequences. The T2J M1 gene showed approximately 80% nucleotide homology, and the encoded mu 2 protein showed approximately 71% amino acid identity, with the T1L and T3D M1 and mu2 sequences, respectively, making the T2J M1 gene and mu2 proteins amongst the most divergent of all reovirus genes and proteins. Comparisons of these newly determined M1 and mu2 sequences with newly determined M1 and mu2 sequences from nine additional field isolates and a variety of laboratory T3D clones identified conserved features and/or regions that provide clues about mu2 structure and function. CONCLUSIONS: The findings suggest a model for the domain organization of mu2 and provide further evidence for a role of mu2 in viral RNA synthesis. The new sequences were also used to explore the basis for M1/mu2-determined differences in the morphology of viral factories in infected cells. The findings confirm the key role of Ser/Pro208 as a prevalent determinant of differences in factory morphology among reovirus isolates and trace the divergence of this residue and its associated phenotype among the different laboratory-specific clones of type 3 Dearing.
Assuntos
Genoma Viral/genética , Reoviridae/genética , Proteínas Virais/genética , Animais , Sequência de Bases , Linhagem Celular , Camundongos , Dados de Sequência Molecular , Filogenia , Reoviridae/isolamento & purificaçãoRESUMO
Traumatic detachment of the mitral valve from the annulus fibrosis occurred in a dog following blunt chest trauma. Euthanasia was elected approximately 7 months posttrauma due to refractory, chronic left heart failure. This is the first reported case of traumatic mitral valve rupture in a dog.
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Doenças do Cão/etiologia , Insuficiência Cardíaca/veterinária , Valva Mitral/lesões , Ferimentos não Penetrantes/veterinária , Doença Aguda , Animais , Cães , Evolução Fatal , Fibrose , Insuficiência Cardíaca/etiologia , MasculinoRESUMO
Streptococcus iniae, the etiological agent of streptococcosis in fish, is an important pathogen of cultured and wild fish worldwide. During the last decade outbreaks of streptococcosis have occurred in a wide range of cultured and wild fish in the Americas and Caribbean islands. To gain a better understanding of the epizootiology of S. iniae in the western hemisphere, over 30 S. iniae isolates recovered from different fish species and geographic locations were characterized phenotypically and genetically. Species identities were determined biochemically and confirmed by amplification and sequencing of the 16S rRNA gene. Repetitive-element palindromic PCR fingerprinting as well as biochemical and antimicrobial susceptibility profiles suggest that a single strain of S. iniae was responsible for two different disease outbreaks among reef fishes in the Caribbean, one in 1999 and another in 2008. Interestingly, a majority of the isolates recovered from cultured fish in the Americas were genetically distinct from the Caribbean isolates and exhibited a trend toward higher minimal inhibitory concentration with respect to several antibiotics as well as greater genetic variability. The biological significance of this genetic variability is unclear, but it could have implications for future vaccine development and treatment.
Assuntos
Doenças dos Peixes/microbiologia , Infecções Estreptocócicas/veterinária , Streptococcus intermedius/classificação , Streptococcus intermedius/genética , América/epidemiologia , Animais , Doenças dos Peixes/epidemiologia , Peixes , Filogenia , Infecções Estreptocócicas/epidemiologia , Infecções Estreptocócicas/microbiologia , Índias Ocidentais/epidemiologiaRESUMO
Despite six decades of clinical experience with the polymyxin class of antibiotics, their dose-limiting nephrotoxicity remains difficult to predict due to a paucity of sensitive biomarkers. Here, we evaluate the performance of standard of care and next-generation biomarkers of renal injury in the detection and monitoring of polymyxin-induced acute kidney injury in male Han Wistar rats using colistin (polymyxin E) and a polymyxin B (PMB) derivative with reduced nephrotoxicity, PMB nonapeptide (PMBN). This study provides the first histopathological and biomarker analysis of PMBN, an important test of the hypothesis that fatty acid modifications and charge reductions in polymyxins can reduce their nephrotoxicity. The results indicate that alterations in a panel of urinary kidney injury biomarkers can be used to monitor histopathological injury, with Kim-1 and α-GST emerging as the most sensitive biomarkers outperforming clinical standards of care, serum or plasma creatinine and blood urea nitrogen. To enable the prediction of polymyxin-induced nephrotoxicity, an in vitro cytotoxicity assay was employed using human proximal tubule epithelial cells (HK-2). Cytotoxicity data in these HK-2 cells correlated with the renal toxicity detected via safety biomarker data and histopathological evaluation, suggesting that in vitro and in vivo methods can be incorporated within a screening cascade to prioritize polymyxin class analogs with more favorable renal toxicity profiles.
Assuntos
Antibacterianos/toxicidade , Colistina/toxicidade , Nefropatias/urina , Polimixina B/análogos & derivados , Animais , Antibacterianos/administração & dosagem , Antibacterianos/farmacocinética , Biomarcadores/urina , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Colistina/administração & dosagem , Colistina/farmacocinética , Interpretação Estatística de Dados , Relação Dose-Resposta a Droga , Diagnóstico Precoce , Nefropatias/induzido quimicamente , Nefropatias/metabolismo , Nefropatias/patologia , Masculino , Polimixina B/administração & dosagem , Polimixina B/farmacocinética , Polimixina B/toxicidade , Prognóstico , Ratos , Ratos WistarRESUMO
Klebsiella pneumoniae is a zoonotic, Gram-negative member of the family Enterobacteriaceae and is the causative agent of nosocomial septicemic, pneumonic, and urinary tract infections. Recently, pathogenic strains of K. pneumoniae sharing a hypermucoviscosity (HMV) phenotype have been attributed to multisystemic abscessation in both human and nonhuman primates. Although K. pneumoniae is a well-recognized zoonotic agent, there is a lack of general information including adequate diagnostic methods or treatments for nonhuman primates. In an effort to increase the body of knowledge of this enigmatic pathogen, K. pneumoniae isolates from African green monkeys (Chlorocebus aethiops sabaeus) on the island of St. Kitts, West Indies were genotypically and phenotypically characterized. Genetic fingerprints generated by PCR-mediated genomic fingerprinting, phenotypic characterization, and antimicrobial susceptibility all identified a high degree of similarity between the HMV and non-HMV K. pneumoniae isolates. The results obtained from this work will help establish a baseline for the development of efficacious diagnostic methods and treatment strategies for both human and nonhuman primates.