Circular RNA protein tyrosine kinase 2 aggravates pyroptosis and inflammation in septic lung tissue by promoting microRNA-766/eukaryotic initiation factor 5A axis-mediated ATP efflux

Purpose: Sepsis is characterized by an acute inflammatory response to infection, often with multiple organ failures, especially severe lung injury. This study was implemented to probe circular RNA (circRNA) protein tyrosine kinase 2 (circPTK2)-associated regulatory mechanisms in septic acute lung injury (ALI). Methods: A cecal ligation and puncture-based mouse model and an lipopolysaccharides (LPS)-based alveolar type II cell (RLE-6TN) model were generated to mimic sepsis. In the two models, inflammation- and pyroptosisrelated genes were measured. Results: The degree of lung injury in mice was analyzed by hematoxylin and eosin (H&E) staining and the apoptosis was by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling staining. In addition, pyroptosis and toxicity were detected in cells. Finally, the binding relationship between circPTK2, miR-766, and eukaryotic initiation factor 5A (eIF5A) was detected. Data indicated that circPTK2 and eIF5A were up-regulated and miR-766 was down-regulated in LPS-treated RLE-6TN cells and lung tissue of septic mice. Lung injury in septic mice was ameliorated after inhibition of circPTK2. Conclusion: It was confirmed in the cell model that knockdown of circPTK2 effectively ameliorated LPS-induced ATP efflux, pyroptosis, and inflammation. Mechanistically, circPTK2 mediated eIF5A expression by competitively adsorbing miR-766. Taken together, circPTK2/ miR-766/eIF5A axis ameliorates septic ALI, developing a novel therapeutic target for the disease.

Biochemical and immunohistochemical examination of the effects of ephedrine in rat ovary tissue

Purpose: It was aimed to investigate the biochemical and immunohistochemical effects of ephedrine (EPH) in bilateral ovariectomized rats. Methods: Twenty-four Sprague Dawley female rats were divided into three groups: control group: The abdomen was opened and closed without any treatment; ischemia-reperfusion (IR) group: 2 h of ischemia followed by 2 h of reperfusion were allowed to cause IR injury; IR+EPH group: oral EPH solution (5 mg/kg) was administered for 28 days. Results: Biochemical parameters were statistically significant in group comparisons. Increased interleukin-6 (IL-6) expression, degenerative preantral and antral follicle cells and inflammatory cells around blood vessels were seen in IR group. Negative IL-6 expression was observed in seminal epithelial cells, preantral and antral follicle cells in IR+EPH group. While caspase-3 activity increased in granulosa cells and stromal cells in IR group, caspase-3 expression was negative in preantral and antral follicle cells in the germinal epithelium and cortex in IR+EPH group. Conclusion: The effect of apoptosis, which occurs with the signaling that starts in the cell nucleus, caused the cessation of the stimulating effect at the nuclear level after EPH administration, and a decrease in the antioxidative effect in IR damage and inflammation in the apoptotic process.

Columbianadin attenuates doxorubicin-induced cardiac injury, oxidative stress, and apoptosis via Sirt1/FOXO1 signaling pathway

Purpose: Oxidative stress and apoptosis contribute to the pathological basis of doxorubicin (DOX)-induced cardiotoxicity. Columbianadin (CBN) is one of the main bioactive constituents isolated from the root of Angelica pubescens. Herein, we intended to explore the potential role and molecular basis of CBN in DOX-induced cardiotoxicity. Methods: C57BL/6 mice were subjected to DOX (15 mg/kg/day, i.p.) to generate DOX-induced cardiotoxicity. CBN (10 mg/kg/day, i.p.) was administered for four week following DOX injection. Results: DOX administered markedly dampened cardiac function, increased cardiac injury, excessive reactive oxygen species (ROS) production, and cardiomyocyte loss. These alterations induced by DOX significantly alleviated by CBN treatment. Mechanistically, our results demonstrated that the CBN exerts cardioprotection role against DOX by up-regulating silent information regulator 1 (Sirt1) and decreasing acetylation of forkhead box O1 (FOXO1). Moreover, Sirt1 inhibition with Ex-527 significantly blunt the beneficial effect of CBN on DOX-induced cardiotoxicity, including cardiac dysfunction, ROS, and apoptosis. Conclusion: Collectively, CBN attenuated oxidative stress and cardiomyocyte apoptosis in DOX-induced cardiotoxicity through maintaining Sirt1/FOXO1 signaling pathway. Our results demonstrated that CBN might be used to treat DOX-related cardiotoxicity.

Effects of neutrophil extracellular traps during human respiratory syncytial virus infection in vitro / Efeitos das redes extracelulares dos neutrófilos durante a infecção in vitro pelo vírus sincicial respiratório

Abstract The human respiratory syncytial virus (hRSV) is the most common cause of severe lower respiratory tract diseases in young children worldwide, leading to a high number of hospitalizations and significant expenditures for health systems. Neutrophils are massively recruited to the lung tissue of patients with acute respiratory diseases. At the infection site, they release neutrophil extracellular traps (NETs) that can capture and/or inactivate different types of microorganisms, including viruses. Evidence has shown that the accumulation of NETs results in direct cytotoxic effects on endothelial and epithelial cells. Neutrophils stimulated by the hRSV-F protein generate NETs that are able to capture hRSV particles, thus reducing their transmission. However, the massive production of NETs obstructs the airways and increases disease severity. Therefore, further knowledge about the effects of NETs during hRSV infections is essential for the development of new specific and effective treatments. This study evaluated the effects of NETs on the previous or posterior contact with hRSV-infected Hep-2 cells. Hep-2 cells were infected with different hRSV multiplicity of infection (MOI 0.5 or 1.0), either before or after incubation with NETs (0.5-16 μg/mL). Infected and untreated cells showed decreased cellular viability and intense staining with trypan blue, which was accompanied by the formation of many large syncytia. Previous contact between NETs and cells did not result in a protective effect. Cells in monolayers showed a reduced number and area of syncytia, but cell death was similar in infected and non-treated cells. The addition of NETs to infected tissues maintained a similar virus-induced cell death rate and an increased syncytial area, indicating cytotoxic and deleterious damages. Our results corroborate previously reported findings that NETs contribute to the immunopathology developed by patients infected with hRSV.

Resumo O vírus sincicial respiratório humano (hRSV) é a causa mais comum de doenças graves do trato respiratório inferior em crianças pequenas em todo o mundo, resultando em grande número de hospitalizações e gastos significativos para os sistemas de saúde. Neutrófilos são recrutados em massa para o tecido pulmonar de pacientes com doenças respiratórias agudas. No local da infecção, eles liberam armadilhas extracelulares de neutrófilos (NETs) que podem capturar e/ou inativar diferentes tipos de microrganismos, incluindo vírus. Evidências demonstraram que o acúmulo de NETs resulta em efeitos citotóxicos diretos nas células endoteliais e epiteliais. Os neutrófilos estimulados pela proteína F do vírus sincicial respiratório (hRSV-F) geram NETs que são capazes de capturar partículas virais, reduzindo assim sua transmissão. No entanto, a produção maciça de NETs obstrui as vias aéreas e aumenta a gravidade da doença. Assim, um maior conhecimento sobre os efeitos das NETs durante as infecções por hRSV é essencial para o desenvolvimento de novos tratamentos específicos e eficazes. Este estudo avaliou os efeitos das NETs no contato prévio ou posterior à infecção de células Hep-2 com hRSV. As células Hep-2 foram infectadas com diferentes quantidades de hRSV (multiplicidade de infecção ou MOI 0,5 ou 1,0), antes ou após a incubação com NETs (0,5-16 μg/mL). Células infectadas e não tratadas mostraram redução da viabilidade celular e intensa coloração com azul de tripano, que foi acompanhada pela formação de sincícios numerosos e grandes. O contato prévio entre as NETs e as células não resultou em efeito protetor. As células em monocamadas mostraram um número e área de sincícios reduzidos, mas a morte celular foi semelhante àquela apresentada por células infectadas e não tratadas. A adição de NETs aos tecidos infectados manteve taxa de morte celular e formação de sincícios semelhantes àqueles induzidos pelo vírus em células não tratadas, indicando danos citotóxicos e deletérios. Nossos resultados corroboram achados relatados anteriormente de que as NETs contribuem para a imunopatologia desenvolvida por pacientes infectados com hRSV.

Cytological grading of canine mast cell tumors: correlation with histologic grading and survival time / Graduação citológica de mastocitomas caninos: correlação com graduação histológica e sobrevida

ABSTRACT: Mast cell tumors are one of the most common neoplasia in dogs and cytopathology and/or histopathology examinations are used for diagnosis. Histologic grading is considered the gold standard test to predict the prognosis of this neoplasia. However, studies have been conducted using the cytological grading system to provide similar information in a faster, less invasive, and more accessible way. This study aimed to investigate cytological graduation and correlate it with histological grading and the survival time of dogs diagnosed with cutaneous mast cell tumors at the Veterinary Hospital of "Universidade Federal de Uberlândia" over five years. For that, cytological and histological slides from 72 animals were reviewed. The statistical methods used were the kappa test for agreement between grading systems, the Kaplan-Meier for survival time, Cox regression for comparison of cytological and histological grades and survival time. The cytological grading when compared to the two-tier histologic grading, high and low grades, had a moderate agreement (kappa 0.566). When the correlation between survival time and the cytological grade was evaluated, there was a higher death rate in the group with high-grade mast cell tumors compared to low grade, pointing to a correlation between survival time and cytological grade (p=0.009). In conclusion, the cytological grade is useful to treatment planning and providing prognostic information that precedes tumor removal, showing a good correlation with the two-tier histologic grading and with the survival time of the animals.

RESUMO: O mastocitoma é uma das neoplasias cutâneas mais comum nos cães e os exames citopatológicos e/ou histopatológicos são utilizados para diagnóstico. A graduação histológica é considerada padrão ouro para prever o prognóstico dessa neoplasia. Contudo, estudos têm sido realizados visando utilizar graduação citológica para fornecer informações semelhantes de maneira rápida, menos invasiva e mais acessível. Esse trabalho objetivou realizar graduação citológica e correlacionar com as graduações histológicas e com a sobrevida de cães diagnosticados com mastocitoma cutâneo no Hospital Veterinário da Universidade Federal de Uberlândia durante 5 anos. Para isso, lâminas de citologia e histologia de mastocitomas de 72 animais foram revisadas. Os métodos estatísticos utilizados foram teste kappa para concordância entre os sistemas de graduação, método Kaplan-Meier para tempo de sobrevida dos animais, e análise pela regressão de Cox para comparação do grau citológico e grau histológico e o tempo de sobrevida global. A graduação citológica quando comparada com a histológica de dois níveis, alto grau e baixo grau, obteve uma concordância moderada (kappa 0,566). Na avaliação da correlação entre sobrevida e grau citológico, houve maior taxa de óbito no grupo de cães com mastocitoma de alto grau, comparado aos de baixo grau, sendo observada correlação entre a sobrevida e o grau citológico (p=0,009). Esse estudo concluiu que o grau citológico é útil para o planejamento do tratamento e para fornecer informações prognósticas que antecedem a exérese do tumor, tendo boa correlação com a graduação histológica de dois níveis e com a sobrevida dos animais.

In vitro cytotoxicity of curcuminoids against head and neck cancer HNO97 cell line / Citotoxicidade in vitro de curcuminoides contra a linhagem de células HNO97 de câncer de cabeça e pescoço

Oral squamous cell carcinoma (OSCC) is a malignant tumour of Head and Neck Cancer (HNC). The recent therapeutic approaches used to treat cancer have adverse side effects. The natural agents exhibiting anticancer activities are generally considered to have a robust therapeutic potential. Curcuminoids, one of the major active compounds of the turmeric herb, are used as a therapeutic agent for several diseases including cancer. In this study, the cytotoxicity of curcuminoids was investigated against OSCC cell line HNO97. Our data showed that curcuminoids significantly inhibits the proliferation of HNO97 in a time and dose-dependent manner (IC50=35 μM). Cell cycle analysis demonstrated that curcuminoids increased the percentage of G2/M phase cell populations in the treated groups. Treating HNO97 cells with curcuminoids led to cell shrinking and increased detached cells, which are the typical appearance of apoptotic cells. Moreover, flow cytometry analysis revealed that curcuminoids significantly induced apoptosis in a time-dependent manner. Furthermore, as a response to curcuminoids treatment, comet tails were formed in cell nuclei due to the induction of DNA damage. Curcuminoids treatment reduced the colony formation capacity of HNO97 cells and induced morphological changes. Overall, these findings demonstrate that curcuminoids can in vitro inhibit HNC proliferation and metastasis and induce apoptosis.

O carcinoma de células escamosas oral (OSCC) é um tumor maligno do câncer de cabeça e pescoço (HNC). As recentes abordagens terapêuticas usadas para tratar o câncer têm efeitos colaterais adversos. Os agentes naturais que exibem atividades anticâncer são geralmente considerados como tendo um potencial terapêutico robusto. Curcuminoides, um dos principais compostos ativos da erva cúrcuma, são usados como agente terapêutico para várias doenças, incluindo câncer. Neste estudo, a citotoxicidade dos curcuminoides foi investigada contra a linha de células OSCC HNO97. Nossos dados mostraram que os curcuminoides inibem significativamente a proliferação de HNO97 de forma dependente do tempo e da dose (IC50 = 35 μM). A análise do ciclo celular demonstrou que os curcuminoides aumentaram a porcentagem de populações de células da fase G2 / M nos grupos tratados. O tratamento das células HNO97 com curcuminoides levou ao encolhimento celular e ao aumento das células destacadas, que são a aparência típica das células apoptóticas. Além disso, a análise de citometria de fluxo revelou que os curcuminoides induziram significativamente a apoptose de uma maneira dependente do tempo. Além disso, em resposta ao tratamento com curcuminoides, caudas de cometa foram formadas nos núcleos das células devido à indução de danos ao DNA. O tratamento com curcuminoides reduziu a capacidade de formação de colônias das células HNO97 e induziu alterações morfológicas. No geral, esses achados demonstram que os curcuminoides podem inibir in vitro a proliferação e metástase de HNC e induzir apoptose.

In vitro cytotoxicity of curcuminoids against head and neck cancer HNO97 cell line / Citotoxicidade in vitro de curcuminoides contra a linhagem de células HNO97 de câncer de cabeça e pescoço

Oral squamous cell carcinoma (OSCC) is a malignant tumour of Head and Neck Cancer (HNC). The recent therapeutic approaches used to treat cancer have adverse side effects. The natural agents exhibiting anticancer activities are generally considered to have a robust therapeutic potential. Curcuminoids, one of the major active compounds of the turmeric herb, are used as a therapeutic agent for several diseases including cancer. In this study, the cytotoxicity of curcuminoids was investigated against OSCC cell line HNO97. Our data showed that curcuminoids significantly inhibits the proliferation of HNO97 in a time and dose-dependent manner (IC50=35 μM). Cell cycle analysis demonstrated that curcuminoids increased the percentage of G2/M phase cell populations in the treated groups. Treating HNO97 cells with curcuminoids led to cell shrinking and increased detached cells, which are the typical appearance of apoptotic cells. Moreover, flow cytometry analysis revealed that curcuminoids significantly induced apoptosis in a time-dependent manner. Furthermore, as a response to curcuminoids treatment, comet tails were formed in cell nuclei due to the induction of DNA damage. Curcuminoids treatment reduced the colony formation capacity of HNO97 cells and induced morphological changes. Overall, these findings demonstrate that curcuminoids can in vitro inhibit HNC proliferation and metastasis and induce apoptosis.(AU)

O carcinoma de células escamosas oral (OSCC) é um tumor maligno do câncer de cabeça e pescoço (HNC). As recentes abordagens terapêuticas usadas para tratar o câncer têm efeitos colaterais adversos. Os agentes naturais que exibem atividades anticâncer são geralmente considerados como tendo um potencial terapêutico robusto. Curcuminoides, um dos principais compostos ativos da erva cúrcuma, são usados como agente terapêutico para várias doenças, incluindo câncer. Neste estudo, a citotoxicidade dos curcuminoides foi investigada contra a linha de células OSCC HNO97. Nossos dados mostraram que os curcuminoides inibem significativamente a proliferação de HNO97 de forma dependente do tempo e da dose (IC50 = 35 μM). A análise do ciclo celular demonstrou que os curcuminoides aumentaram a porcentagem de populações de células da fase G2 / M nos grupos tratados. O tratamento das células HNO97 com curcuminoides levou ao encolhimento celular e ao aumento das células destacadas, que são a aparência típica das células apoptóticas. Além disso, a análise de citometria de fluxo revelou que os curcuminoides induziram significativamente a apoptose de uma maneira dependente do tempo. Além disso, em resposta ao tratamento com curcuminoides, caudas de cometa foram formadas nos núcleos das células devido à indução de danos ao DNA. O tratamento com curcuminoides reduziu a capacidade de formação de colônias das células HNO97 e induziu alterações morfológicas. No geral, esses achados demonstram que os curcuminoides podem inibir in vitro a proliferação e metástase de HNC e induzir apoptose.(AU)

Effects of neutrophil extracellular traps during human respiratory syncytial virus infection in vitro

Abstract The human respiratory syncytial virus (hRSV) is the most common cause of severe lower respiratory tract diseases in young children worldwide, leading to a high number of hospitalizations and significant expenditures for health systems. Neutrophils are massively recruited to the lung tissue of patients with acute respiratory diseases. At the infection site, they release neutrophil extracellular traps (NETs) that can capture and/or inactivate different types of microorganisms, including viruses. Evidence has shown that the accumulation of NETs results in direct cytotoxic effects on endothelial and epithelial cells. Neutrophils stimulated by the hRSV-F protein generate NETs that are able to capture hRSV particles, thus reducing their transmission. However, the massive production of NETs obstructs the airways and increases disease severity. Therefore, further knowledge about the effects of NETs during hRSV infections is essential for the development of new specific and effective treatments. This study evaluated the effects of NETs on the previous or posterior contact with hRSV-infected Hep-2 cells. Hep-2 cells were infected with different hRSV multiplicity of infection (MOI 0.5 or 1.0), either before or after incubation with NETs (0.516 g/mL). Infected and untreated cells showed decreased cellular viability and intense staining with trypan blue, which was accompanied by the formation of many large syncytia. Previous contact between NETs and cells did not result in a protective effect. Cells in monolayers showed a reduced number and area of syncytia, but cell death was similar in infected and non-treated cells. The addition of NETs to infected tissues maintained a similar virus-induced cell death rate and an increased syncytial area, indicating cytotoxic and deleterious damages. Our results corroborate previously reported findings that NETs contribute to the immunopathology developed by patients infected with hRSV.

Resumo O vírus sincicial respiratório humano (hRSV) é a causa mais comum de doenças graves do trato respiratório inferior em crianças pequenas em todo o mundo, resultando em grande número de hospitalizações e gastos significativos para os sistemas de saúde. Neutrófilos são recrutados em massa para o tecido pulmonar de pacientes com doenças respiratórias agudas. No local da infecção, eles liberam armadilhas extracelulares de neutrófilos (NETs) que podem capturar e/ou inativar diferentes tipos de microrganismos, incluindo vírus. Evidências demonstraram que o acúmulo de NETs resulta em efeitos citotóxicos diretos nas células endoteliais e epiteliais. Os neutrófilos estimulados pela proteína F do vírus sincicial respiratório (hRSV-F) geram NETs que são capazes de capturar partículas virais, reduzindo assim sua transmissão. No entanto, a produção maciça de NETs obstrui as vias aéreas e aumenta a gravidade da doença. Assim, um maior conhecimento sobre os efeitos das NETs durante as infecções por hRSV é essencial para o desenvolvimento de novos tratamentos específicos e eficazes. Este estudo avaliou os efeitos das NETs no contato prévio ou posterior à infecção de células Hep-2 com hRSV. As células Hep-2 foram infectadas com diferentes quantidades de hRSV (multiplicidade de infecção ou MOI 0,5 ou 1,0), antes ou após a incubação com NETs (0,516 g/mL). Células infectadas e não tratadas mostraram redução da viabilidade celular e intensa coloração com azul de tripano, que foi acompanhada pela formação de sincícios numerosos e grandes. O contato prévio entre as NETs e as células não resultou em efeito protetor. As células em monocamadas mostraram um número e área de sincícios reduzidos, mas a morte celular foi semelhante àquela apresentada por células infectadas e não tratadas. A adição de NETs aos tecidos infectados manteve taxa de morte celular e formação de sincícios semelhantes àqueles induzidos pelo vírus em células não tratadas, indicando danos citotóxicos e deletérios. Nossos resultados corroboram achados relatados anteriormente de que as NETs contribuem para a imunopatologia desenvolvida por pacientes infectados com hRSV.

Effects of neutrophil extracellular traps during human respiratory syncytial virus infection in vitro / Efeitos das redes extracelulares dos neutrófilos durante a infecção in vitro pelo vírus sincicial respiratório

The human respiratory syncytial virus (hRSV) is the most common cause of severe lower respiratory tract diseases in young children worldwide, leading to a high number of hospitalizations and significant expenditures for health systems. Neutrophils are massively recruited to the lung tissue of patients with acute respiratory diseases. At the infection site, they release neutrophil extracellular traps (NETs) that can capture and/or inactivate different types of microorganisms, including viruses. Evidence has shown that the accumulation of NETs results in direct cytotoxic effects on endothelial and epithelial cells. Neutrophils stimulated by the hRSV-F protein generate NETs that are able to capture hRSV particles, thus reducing their transmission. However, the massive production of NETs obstructs the airways and increases disease severity. Therefore, further knowledge about the effects of NETs during hRSV infections is essential for the development of new specific and effective treatments. This study evaluated the effects of NETs on the previous or posterior contact with hRSV-infected Hep-2 cells. Hep-2 cells were infected with different hRSV multiplicity of infection (MOI 0.5 or 1.0), either before or after incubation with NETs (0.5–16 μg/mL). Infected and untreated cells showed decreased cellular viability and intense staining with trypan blue, which was accompanied by the formation of many large syncytia. Previous contact between NETs and cells did not result in a protective effect. Cells in monolayers showed a reduced number and area of syncytia, but cell death was similar in infected and non-treated cells. The addition of NETs to infected tissues maintained a similar virus-induced cell death rate and an increased syncytial area, indicating cytotoxic and deleterious damages. Our results corroborate previously reported findings that NETs contribute to the immunopathology developed by patients infected with hRSV.

O vírus sincicial respiratório humano (hRSV) é a causa mais comum de doenças graves do trato respiratório inferior em crianças pequenas em todo o mundo, resultando em grande número de hospitalizações e gastos significativos para os sistemas de saúde. Neutrófilos são recrutados em massa para o tecido pulmonar de pacientes com doenças respiratórias agudas. No local da infecção, eles liberam armadilhas extracelulares de neutrófilos (NETs) que podem capturar e/ou inativar diferentes tipos de microrganismos, incluindo vírus. Evidências demonstraram que o acúmulo de NETs resulta em efeitos citotóxicos diretos nas células endoteliais e epiteliais. Os neutrófilos estimulados pela proteína F do vírus sincicial respiratório (hRSV-F) geram NETs que são capazes de capturar partículas virais, reduzindo assim sua transmissão. No entanto, a produção maciça de NETs obstrui as vias aéreas e aumenta a gravidade da doença. Assim, um maior conhecimento sobre os efeitos das NETs durante as infecções por hRSV é essencial para o desenvolvimento de novos tratamentos específicos e eficazes. Este estudo avaliou os efeitos das NETs no contato prévio ou posterior à infecção de células Hep-2 com hRSV. As células Hep-2 foram infectadas com diferentes quantidades de hRSV (multiplicidade de infecção ou MOI 0,5 ou 1,0), antes ou após a incubação com NETs (0,5–16 μg/mL). Células infectadas e não tratadas mostraram redução da viabilidade celular e intensa coloração com azul de tripano, que foi acompanhada pela formação de sincícios numerosos e grandes. O contato prévio entre as NETs e as células não resultou em efeito protetor. As células em monocamadas mostraram um número e área de sincícios reduzidos, mas a morte celular foi semelhante àquela apresentada por células infectadas e não tratadas. A adição de NETs aos tecidos infectados manteve taxa de morte celular e formação de sincícios [...].

Effects of neutrophil extracellular traps during human respiratory syncytial virus infection in vitro / Efeitos das redes extracelulares dos neutrófilos durante a infecção in vitro pelo vírus sincicial respiratório

The human respiratory syncytial virus (hRSV) is the most common cause of severe lower respiratory tract diseases in young children worldwide, leading to a high number of hospitalizations and significant expenditures for health systems. Neutrophils are massively recruited to the lung tissue of patients with acute respiratory diseases. At the infection site, they release neutrophil extracellular traps (NETs) that can capture and/or inactivate different types of microorganisms, including viruses. Evidence has shown that the accumulation of NETs results in direct cytotoxic effects on endothelial and epithelial cells. Neutrophils stimulated by the hRSV-F protein generate NETs that are able to capture hRSV particles, thus reducing their transmission. However, the massive production of NETs obstructs the airways and increases disease severity. Therefore, further knowledge about the effects of NETs during hRSV infections is essential for the development of new specific and effective treatments. This study evaluated the effects of NETs on the previous or posterior contact with hRSV-infected Hep-2 cells. Hep-2 cells were infected with different hRSV multiplicity of infection (MOI 0.5 or 1.0), either before or after incubation with NETs (0.5–16 μg/mL). Infected and untreated cells showed decreased cellular viability and intense staining with trypan blue, which was accompanied by the formation of many large syncytia. Previous contact between NETs and cells did not result in a protective effect. Cells in monolayers showed a reduced number and area of syncytia, but cell death was similar in infected and non-treated cells. The addition of NETs to infected tissues maintained a similar virus-induced cell death rate and an increased syncytial area, indicating cytotoxic and deleterious damages. Our results corroborate previously reported findings that NETs contribute to the immunopathology developed by patients infected with hRSV.(AU)

O vírus sincicial respiratório humano (hRSV) é a causa mais comum de doenças graves do trato respiratório inferior em crianças pequenas em todo o mundo, resultando em grande número de hospitalizações e gastos significativos para os sistemas de saúde. Neutrófilos são recrutados em massa para o tecido pulmonar de pacientes com doenças respiratórias agudas. No local da infecção, eles liberam armadilhas extracelulares de neutrófilos (NETs) que podem capturar e/ou inativar diferentes tipos de microrganismos, incluindo vírus. Evidências demonstraram que o acúmulo de NETs resulta em efeitos citotóxicos diretos nas células endoteliais e epiteliais. Os neutrófilos estimulados pela proteína F do vírus sincicial respiratório (hRSV-F) geram NETs que são capazes de capturar partículas virais, reduzindo assim sua transmissão. No entanto, a produção maciça de NETs obstrui as vias aéreas e aumenta a gravidade da doença. Assim, um maior conhecimento sobre os efeitos das NETs durante as infecções por hRSV é essencial para o desenvolvimento de novos tratamentos específicos e eficazes. Este estudo avaliou os efeitos das NETs no contato prévio ou posterior à infecção de células Hep-2 com hRSV. As células Hep-2 foram infectadas com diferentes quantidades de hRSV (multiplicidade de infecção ou MOI 0,5 ou 1,0), antes ou após a incubação com NETs (0,5–16 μg/mL). Células infectadas e não tratadas mostraram redução da viabilidade celular e intensa coloração com azul de tripano, que foi acompanhada pela formação de sincícios numerosos e grandes. O contato prévio entre as NETs e as células não resultou em efeito protetor. As células em monocamadas mostraram um número e área de sincícios reduzidos, mas a morte celular foi semelhante àquela apresentada por células infectadas e não tratadas. A adição de NETs aos tecidos infectados manteve taxa de morte celular e formação de sincícios [...].(AU)

Criopreservação e cultivo de tecido testicular em mamíferos - estado da arte, desafios e perspectivas / Cryopreservation and culture of testicular tissue in mammals - state of the art, challenges and perspectives

O tecido testicular contém células germinativas, que possuem potencial para se desenvolver em espermatozoides viáveis por meio do cultivo in vitro ou xenotransplante, sendo uma alternativa interessante a ser utilizada na formação de biobancos. Esta revisão compila as atualizações, desafios e perspectivas relacionadas às técnicas de criopreservação e cultivo de tecido testicular como estratégia para a conservação de espécies mamíferas. O tecido testicular pode ser obtido tanto de indivíduos adultos como pré púberes, seja após orquiectomia ou até mesmo após a sua morte. O tecido fragmentado pode ser criopreservado por congelação lenta ou rápida e por métodos de vitrificação. Os crioprotetores são indispensáveis durante a criopreservação e podem variar o tipo e concentração de acordo com a espécie. Com os avanços da criopreservação deste material, espermatozoides podem ser obtidos por transplante de fragmentos testiculares ou células germinativas isoladas em camundongos imunodeficientes. No entanto, a obtenção de espermatozoides no cultivo in vitro ainda é um desafio.(AU)

The testicular tissue contains germ cells, which have the potential to develop into viable spermatozoa through in vitro culture or xenotransplantation, being an interesting alternative to be used in the formation of biobanks. This review compiles updates, challenges and perspectives related to cryopreservation techniques and testicular tissue culture as a strategy for the conservation of mammalian species. Testicular tissue can be obtained from both adult and pre-pubertal individuals, either after orchiectomy or even after their death. Fragmented tissue can be cryopreserved by slow or fast freezing and by vitrification methods. Cryoprotectants are indispensable during cryopreservation and may vary in type and concentration according to the species. With advances in cryopreservation of this material, spermatozoa can be obtained by transplanting testicular fragments or isolated germ cells into immunodeficient mice. However, obtaining spermatozoa in in vitro culture is still a challenge.(AU)

Protective effect of Mucuna pruriens (L) DC. var. pruriens seed extract on apoptotic germ cells in ethanolic male rats / Efeito protetor de Mucuna pruriens (L) DC. var. extrato de semente de pruriens, em células germinativas apoptóticas em ratos machos etanólicos

Thai Mucuna pruriens (L.) DC. var pruriens (T-MP) seed containing levodopa (L-DOPA) and antioxidant capacity has been shown to improve sexual behavior and male reproductive parameters in rats treated with ethanol (Eth). However, its protective effect on testicular apoptotic germ cells has never been reported. This study aimed to investigate the potential effects of T-MP seed extract on expressions of caspase, proliferating cell nuclear antigen (PCNA), and dopamine D2 receptor (D2R) proteins in Eth rats. Thirty-six male Wistar rats were divided into four groups (9 animals/group), including control, Eth, T-MP150+Eth, and T-MP300+Eth, respectively. Control rats received distilled water, and Eth rats received Eth (3g/kg BW; 40%v/v). The T-MP groups were treated with T-MP seed extract at a dose of 150 or 300 mg/kg before Eth administration for 56 consecutive days. The results showed that the seminiferous tubule diameter and epithelial height were significantly increased in both T-MP treated groups compared to the Eth group. Additionally, the caspase-9 and -3, and PCNA expressions were decreased, but D2R expression was markedly increased in T-MP groups. It was concluded that T-MP seed extract could protect testicular apoptosis induced by Eth via changes in caspase, PCNA, and D2R protein expressions.

Thai Mucuna pruriens (L.) DC. var pruriens (T-MP) contendo levodopa (L-DOPA) e capacidade antioxidante demonstrou melhorar o comportamento sexual e os parâmetros reprodutivos masculinos em ratos tratados com etanol (Eth). No entanto, seu efeito protetor sobre células germinativas apoptóticas testiculares nunca foi relatado. Este estudo teve como objetivo investigar os efeitos potenciais do extrato de semente de T-MP na expressão de proteínas de caspase, antígeno nuclear de proliferação celular (PCNA) e receptor de dopamina D2 (D2R) em ratos Eth. Trinta e seis ratos Wistar machos foram divididos em quatro grupos (9 animais/grupo), incluindo controle, Eth, T-MP150+Eth e T-MP300+Eth, respectivamente. Ratos controle receberam água destilada e ratos Eth receberam Eth (3g/kg PC; 40%v/v). Os grupos T-MP foram tratados com extrato de semente de T-MP na dose de 150 ou 300 mg/kg antes da administração de Eth por 56 dias consecutivos. Os resultados mostraram que o diâmetro dos túbulos seminíferos e a altura epitelial foram significativamente aumentados em ambos os grupos tratados com T-MP em comparação com o grupo Eth. Além disso, as expressões de caspase-9 e -3 e de PCNA diminuíram, mas a expressão de D2R aumentou acentuadamente nos grupos T-MP. Concluiu-se que o extrato de semente de T-MP pode proteger a apoptose testicular induzida por Eth através de alterações na expressão de proteínas caspase, PCNA e D2R.

Histopathological examination of the protective effect of intense exercise in apoptotic germ cell damage due to diabetes

Purpose: The aim of this study was to determine the protective and antioxidative effects of intensive exercise on streptozotocin (STZ)-induced testicular damage, apoptotic spermatognial cells death, and oxidative stress. Methods: 36 male Sprague Dawley rats were divided into three groups: control, diabetes, and diabetes+intensive exercise (IE) groups. Testicular tissues were examined histopathologically and antioxidant enzymes, including catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), and malondialdehyde (MDA) activity, as well as serum testosterone level, were measured. Results: Seminiferous tubules and germ cells were found to be better in the testis tissue of the intense exercise group than in the diabetes group. Diabetes suppressed antioxidant enzymes CAT, SOD, GPx and testosterone levels were significantly decreased, and increased MDA level in the diabetic group compared to diabetes+IE group (p < 0.001). Following four weeks of treatment, intensive exercise improved the antioxidant defense, significantly decreased MDA activity, and increased testosterone levels in testicular tissue in the diabetic group compared to diabetes+IE group (p < 0.01). Conclusion: STZ-induced diabetes causes damage to the testis tissue. In order to prevent these damages, exercise practice has become very popular nowadays. In present study, our intensive exercise protocol, histological, and biochemical analysis of the effect of diabetes on the testicular tissues is shown.

Urethral obstruction secondary to Staphylococcus felis chronic urethritis and hepatoid gland adenoma in a captive ocelot (Leopardus pardalis)

A male adult ocelot (Leopardus pardalis) kept under human care developed anuria, which progressed to death. Grossly, the urinary bladder was markedly dilated and filled with red discolored urine containing blood clots. In addition, the animal had a hepatoid cell adenoma adjacent to the urethra, which likely caused partial urethral occlusion. Microscopically, there was a predominantly neutrophilic, fibrinous and hemorrhagic urethritis, cystitis, and pyelonephritis with intralesional gram-positive cocci. Microbiologic culture followed by MALDI-TOF MS analysis resulted in the identification of isolates from the urine and urethra as Staphylococcus felis.(AU)

Lipopolysaccharide effects on activation and apoptosis of primordial ovarian follicles in heifers / Efeitos de lipopolissacarídeos na ativação e apoptose de folículos ovarianos primordiais em novilhas

The objective of the present study was to evaluate the effect of lipopolysaccharide (LPS) administration on activation and apoptosis of primordial follicles. There was no difference in the total number of follicles as well as in the different types of follicles. Furthermore, the LPS challenge didn't modulate the expression of genes related with ovarian reserve (HAM), oocyte survival (Survivin), activation rate (Pten, KIT, KITL1, KITL2, AKT1, SIRT1), and follicular abnormalities. Therefore, the LPS exposure with 24h interval had no effect on activation rate and primordial follicles abnormalities, and also had no effect on expression of anti-apoptotic genes and genes related with ovarian reserve, oocyte survival, activation rate, and primordial follicles abnormalities.

O objetivo do presente estudo foi avaliar o efeito da administração de lipopolissacarídeo (LPS) na ativação e a apoptose de folículos primordiais. Dez novilhas saudáveis (Bos taurus taurus), com idade média de 14 meses, alojadas em sistema de confinamento e alimentadas com TMR, foram utilizadas neste experimento. Os animais foram distribuídos aleatoriamente em dois grupos: grupo LPS (LPS; n = 5), que recebeu duas injeções intravenosas de 0,5µg/kg de peso corporal de lipopolissacarídeo (Sigma Aldrich®) diluído em 2mL de solução salina (0,9% de NaCl), com intervalo de 24h; e grupo controle (CTR; n = 5), que recebeu duas injeções intravenosas de 2mL de solução salina (0,9% de NaCl), com intervalo de 24h. A primeira injeção de LPS foi realizada no d 1, e no d 5 os animais foram abatidos, os ovários foram pesados e as amostras dos ovários foram coletadas para avaliação histológica e molecular. Não houve diferença no número total de folículos, bem como nos diferentes tipos de folículos. Além disso, o desafio com LPS não modulou a expressão de genes relacionados à reserva ovariana (HAM), à sobrevivência oocitária (Survivin), à taxa de ativação (Pten, KIT, KITL1, KITL2, AKT1, SIRT1) e às anormalidades foliculares. Portanto, a exposição ao LPS com intervalo de 24h não teve efeito sobre a taxa de ativação e as anormalidades dos folículos primordiais, bem como não teve efeito sobre a expressão de genes antiapoptóticos e de genes relacionados com a reserva ovariana, a sobrevivência oocitária, a taxa de ativação e as anormalidades dos folículos primordiais.

Protective effect of Guanxin Danshen formula on myocardial ischemiareperfusion injury in rats

Purpose: Myocardial ischemia/reperfusion injury (MIRI) leads to myocardial tissue necrosis, which will increase the size of myocardial infarction. The study examined the protective effect and mechanism of the Guanxin Danshen formula (GXDSF) on MIRI in rats. Methods: MIRI model was performed in rats; rat H9C2 cardiomyocytes were hypoxia-reoxygenated to establish a cell injury model. Results: The GXDSF significantly reduced myocardial ischemia area, reduced myocardial structural injury, decreased the levels of interleukin (IL-1ß, IL-6) in serum, decreased the activity of myocardial enzymes, increased the activity of superoxide dismutase (SOD), and reduced glutathione in rats with MIRI. The GXDSF can reduce the expression of nucleotide- binding oligomerization domain, leucine-rich repeat and pyrin domain containing nod-like receptor family protein 3 (NLRP3), IL-1ß, caspase-1, and gasdermin D (GSDMD) in myocardial tissue cells. Salvianolic acid B and notoginsenoside R1 protected H9C2 cardiomyocytes from hypoxia and reoxygenation injury and reduced the levels of tumor necrosis factor α (TNF-α) and IL-6 in the cell supernatant, decreasing the NLRP3, IL-18, IL-1ß, caspase-1, and GSDMD expression in H9C2 cardiomyocytes. GXDSF can reduce the myocardial infarction area and alleviate the damage to myocardial structure in rats with MIRI, which may be related to the regulation of the NLRP3. Conclusion: GXDSF reduces MIRI in rat myocardial infarction injury, improves structural damage in myocardial ischemia injury, and reduces myocardial tissue inflammation and oxidative stress by lowering inflammatory factors and controlling focal cell death signaling pathways.

Enhancement of Annexin V in response to combination of epigallocatechin gallate and quercetin as a potent arrest the cell cycle of colorectal cancer / Aumento da anexina V em resposta à combinação de galato de epigalocatequina e quercetina como uma potente parada do ciclo celular do câncer colorretal

Colorectal cancer (CRC) is one of the most common cancers leading to comorbidities and mortalities globally. The rational of current study was to evaluate the combined epigallocatechin gallate and quercetin as a potent antitumor agent as commentary agent for therapeutic protocol. The present study investigated the effect of epigallocatechin Gallate (EGCG) (150mg) and quercetin (200mg) at different proportions on proliferation and induction of apoptosis in human colon cancer cells (HCT-116). Cell growth, colonogenic, Annexin V in addition cell cycle were detected in response to phytomolecules. Data obtained showed that, the colony formation was inhibited significantly in CRC starting from the lowest concentration tested of 10 µg/mL resulting in no colonies as visualized by a phase-contrast microscope. Data showed a significant elevation in the annexin V at 100 µg/mL EGCG(25.85%) and 150 µg/mL quercetin (48.35%). Moreover, cell cycle analysis showed that this combination caused cell cycle arrest at the G1 phase at concentration of 100 µg/mL (72.7%) and 150 µg/mL (75.25%). The combined effect of epigallocatechin Gallate and quercetin exert antiproliferative activity against CRC, it is promising in alternative conventional chemotherapeutic agent.

O câncer colorretal (CCR) é um dos cânceres mais comuns, levando a comorbidades e mortalidade em todo o mundo. O racional do presente estudo foi avaliar a combinação de galato de epigalocatequina e quercetina como um agente antitumoral potente como agente de comentário para protocolo terapêutico. O presente estudo investigou o efeito de galato de epigalocatequina (EGCG) (150 mg) e quercetina (200 mg) em diferentes proporções na proliferação e indução de apoptose em células de câncer de cólon humano (HCT-116). O crescimento celular, colonogênico, anexina V, além do ciclo celular foram detectados em resposta a fitomoléculas. Os dados obtidos mostraram que a formação de colônias foi inibida significativamente no CRC a partir da concentração mais baixa testada de 10 µg/mL, resultando em nenhuma colônia conforme visualizado por um microscópio de contraste de fase. Os dados mostraram uma elevação significativa na anexina V a 100 µg/mL de EGCG (25,85%) e 150 µg/mL de quercetina (48,35%). Além disso, a análise do ciclo celular mostrou que essa combinação causou parada do ciclo celular na fase G1 na concentração de 100 µg/mL (72,7%) e 150 µg/mL (75,25%). O efeito combinado da epigalocatequina galato e quercetina exerce atividade antiproliferativa contra o CCR, é promissor como agente quimioterápico alternativo convencional.

Enhancement of Annexin V in response to combination of epigallocatechin gallate and quercetin as a potent arrest the cell cycle of colorectal cancer / Aumento da anexina V em resposta à combinação de galato de epigalocatequina e quercetina como uma potente parada do ciclo celular do câncer colorretal

Colorectal cancer (CRC) is one of the most common cancers leading to comorbidities and mortalities globally. The rational of current study was to evaluate the combined epigallocatechin gallate and quercetin as a potent antitumor agent as commentary agent for therapeutic protocol. The present study investigated the effect of epigallocatechin Gallate (EGCG) (150mg) and quercetin (200mg) at different proportions on proliferation and induction of apoptosis in human colon cancer cells (HCT-116). Cell growth, colonogenic, Annexin V in addition cell cycle were detected in response to phytomolecules. Data obtained showed that, the colony formation was inhibited significantly in CRC starting from the lowest concentration tested of 10 µg/mL resulting in no colonies as visualized by a phase-contrast microscope. Data showed a significant elevation in the annexin V at 100 µg/mL EGCG(25.85%) and 150 µg/mL quercetin (48.35%). Moreover, cell cycle analysis showed that this combination caused cell cycle arrest at the G1 phase at concentration of 100 µg/mL (72.7%) and 150 µg/mL (75.25%). The combined effect of epigallocatechin Gallate and quercetin exert antiproliferative activity against CRC, it is promising in alternative conventional chemotherapeutic agent.(AU)

O câncer colorretal (CCR) é um dos cânceres mais comuns, levando a comorbidades e mortalidade em todo o mundo. O racional do presente estudo foi avaliar a combinação de galato de epigalocatequina e quercetina como um agente antitumoral potente como agente de comentário para protocolo terapêutico. O presente estudo investigou o efeito de galato de epigalocatequina (EGCG) (150 mg) e quercetina (200 mg) em diferentes proporções na proliferação e indução de apoptose em células de câncer de cólon humano (HCT-116). O crescimento celular, colonogênico, anexina V, além do ciclo celular foram detectados em resposta a fitomoléculas. Os dados obtidos mostraram que a formação de colônias foi inibida significativamente no CRC a partir da concentração mais baixa testada de 10 µg/mL, resultando em nenhuma colônia conforme visualizado por um microscópio de contraste de fase. Os dados mostraram uma elevação significativa na anexina V a 100 µg/mL de EGCG (25,85%) e 150 µg/mL de quercetina (48,35%). Além disso, a análise do ciclo celular mostrou que essa combinação causou parada do ciclo celular na fase G1 na concentração de 100 µg/mL (72,7%) e 150 µg/mL (75,25%). O efeito combinado da epigalocatequina galato e quercetina exerce atividade antiproliferativa contra o CCR, é promissor como agente quimioterápico alternativo convencional.(AU)

Neuropathology of natural canid alphaherpesvirus 1 infection in dogs

The neuropathology of canid alphaherpesvirus 1 (CaHV-1) infection in dogs has been reported, but the importance of the cerebellar lesions in the routine diagnosis remains relatively understated in the veterinary literature. Here we characterize the neuropathology of natural CaHV-1 infection in 25 dogs. The disease affected predominantly male dogs (22 cases). Clinical signs in 15 cases varied and sudden death was reported in 10 cases. All dogs had the typical areas of necrosis and hemorrhage in multiple organs with eosinophilic intranuclear viral inclusions in epithelial and/or inflammatory cells in multiple organs in 21 cases. Cerebral swelling and reddening were reported in 3 cases. Neurohistologic changes consisted of mild lymphoplasmacytic meningoencephalitis with occasional vasculitis and widespread glial nodules. Rare intranuclear viral inclusions (4 cases) were observed in endothelial cells or inflammatory cells in glial nodules. Cerebellum was examined in 11 cases and had extensive segmental necrosis of Purkinje cell layer (8 cases) and granule neurons in the internal (9 cases) or external (2 cases) granular cell layer. Intranuclear viral inclusions were observed in 8 cases and occurred in necrotic granule neurons in the inner granular cell layer (7 cases), outer granular cell layer (2 cases), Purkinje neurons (1 case), or endothelial cells (1 case). Diagnostic confirmation in 23 cases was based on routine histology and fluorescent antibody testing for CaHV-1 on fresh or frozen tissue samples. A routine diagnosis was based solely on the visualization of intranuclear viral inclusions in 2 cases.(AU)

Investigation of the histopathological level of Ki-67, caspase-3 expressions of the effects of hesperidin on wound healing in the rat esophagus

Purpose: The effects of hesperidin application on the wound caused by esophageal burns were investigated in this study. Methods: Wistar albino rats were divided into three groups: Control group: only 1 mL of 0.09% NaCl was administered i.p. for 28 days; Burn group: An alkaline esophageal burn model was created with 0.2 mL of 25% NaOH orally by gavage­1 mL of 0.09% NaCl was administered i.p. for 28 days; Burn+Hesperidin group: 1 mL of 50 mL/kg of hesperidin was given i.p. for 28 days to rats after burn injury. Blood samples were collected for biochemical analysis. Esophagus samples were processed for histochemical staining and immunohistochemistry. Results: Malondialdehyde (MDA) and myeloperoxidase (MPO) levels were significantly increased in Burn group. Glutathione (GSH) content and histological scores of epithelialization, collagen formation, neovascularization was decreased. After hesperidin treatment, these values were significantly improved in the Burn+Hesperidin group. In the Burn group, epithelial cells and muscular layers were degenerated. Hesperidin treatment restored these pathologies in Burn+Hesperidin group. Ki-67 and caspase-3 expressions were mainly negative in control group; however, the expression was increased in the Burn group. In the Burn+Hesperidin group, Ki-67 and caspase-3 immune activities were reduced. Conclusion: Hesperidin dosage and application methods can be developed as an alternative treatment for burn healing and treatment.