Cisplatin Toxicity Causes Neutrophil-Mediated Inflammation in Zebrafish Larvae.

Cisplatin is an antineoplastic agent used to treat various tumors. In mammals, it can cause nephrotoxicity, tissue damage, and inflammation. The release of inflammatory mediators leads to the recruitment and infiltration of immune cells, particularly neutrophils, at the site of inflammation. Cisplatin is often used as an inducer of acute kidney injury (AKI) in experimental models, including zebrafish (Danio rerio), due to its accumulation in kidney cells. Current protocols in larval zebrafish focus on studying its effect as an AKI inducer but ignore other systematic outcomes. In this study, cisplatin was added directly to the embryonic medium to assess its toxicity and impact on systemic inflammation using locomotor activity analysis, qPCR, microscopy, and flow cytometry. Our data showed that larvae exposed to cisplatin at 7 days post-fertilization (dpf) displayed dose-dependent mortality and morphological changes, leading to a decrease in locomotion speed at 9 dpf. The expression of pro-inflammatory cytokines such as interleukin (il)-12, il6, and il8 increased after 48 h of cisplatin exposure. Furthermore, while a decrease in the number of neutrophils was observed in the glomerular region of the pronephros, there was an increase in neutrophils throughout the entire animal after 48 h of cisplatin exposure. We demonstrate that cisplatin can have systemic effects in zebrafish larvae, including morphological and locomotory defects, increased inflammatory cytokines, and migration of neutrophils from the hematopoietic niche to other parts of the body. Therefore, this protocol can be used to induce systemic inflammation in zebrafish larvae for studying new therapies or mechanisms of action involving neutrophils.

HLA-DM constrains epitope selection in the human CD4 T cell response to vaccinia virus by favoring the presentation of peptides with longer HLA-DM-mediated half-lives.

HLA-DM (DM) is a nonclassical MHC class II (MHC II) protein that acts as a peptide editor to mediate the exchange of peptides loaded onto MHC II during Ag presentation. Although the ability of DM to promote peptide exchange in vitro and in vivo is well established, the role of DM in epitope selection is still unclear, especially in human response to infectious disease. In this study, we addressed this question in the context of the human CD4 T cell response to vaccinia virus. We measured the IC(50), intrinsic dissociation t(1/2), and DM-mediated dissociation t(1/2) for a large set of peptides derived from the major core protein A10L and other known vaccinia epitopes bound to HLA-DR1 and compared these properties to the presence and magnitude of peptide-specific CD4(+) T cell responses. We found that MHC II-peptide complex kinetic stability in the presence of DM distinguishes T cell epitopes from nonrecognized peptides in A10L peptides and also in a set of predicted tight binders from the entire vaccinia genome. Taken together, these analyses demonstrate that DM-mediated dissociation t(1/2) is a strong and independent factor governing peptide immunogenicity by favoring the presentation of peptides with greater kinetic stability in the presence of DM.

Human CD4+ T cell response to human herpesvirus 6.

Following primary infection, human herpesvirus 6 (HHV-6) establishes a persistent infection for life. HHV-6 reactivation has been associated with transplant rejection, delayed engraftment, encephalitis, muscular dystrophy, and drug-induced hypersensitivity syndrome. The poor understanding of the targets and outcome of the cellular immune response to HHV-6 makes it difficult to outline the role of HHV-6 in human disease. To fill in this gap, we characterized CD4 T cell responses to HHV-6 using peripheral blood mononuclear cell (PBMC) and T cell lines generated from healthy donors. CD4(+) T cells responding to HHV-6 in peripheral blood were observed at frequencies below 0.1% of total T cells but could be expanded easily in vitro. Analysis of cytokines in supernatants of PBMC and T cell cultures challenged with HHV-6 preparations indicated that gamma interferon (IFN-γ) and interleukin-10 (IL-10) were appropriate markers of the HHV-6 cellular response. Eleven CD4(+) T cell epitopes, all but one derived from abundant virion components, were identified. The response was highly cross-reactive between HHV-6A and HHV-6B variants. Seven of the CD4(+) T cell epitopes do not share significant homologies with other known human pathogens, including the closely related human viruses human herpesvirus 7 (HHV-7) and human cytomegalovirus (HCMV). Major histocompatibility complex (MHC) tetramers generated with these epitopes were able to detect HHV-6-specific T cell populations. These findings provide a window into the immune response to HHV-6 and provide a basis for tracking HHV-6 cellular immune responses.

Differences in systemic and skin migrating-specific CD4 T cells in papular urticaria by flea bite.

BACKGROUND: Papular urticaria by flea bite is a chronic allergic condition in which clinical improvement may occur at the age of 7 years, thus representing a natural model of acquired immunologic tolerance in humans. The aim of this study was to characterize regulatory cells and specific responses to flea antigens of CD4(+) T lymphocytes expressing cutaneous migration markers in patients with papular urticaria caused by flea bite and with different disease evolution times. METHODS: Cell populations were characterized by flow cytometry in samples from patients and healthy controls. Specific cell stimulation was performed with a complete flea body extract. The Mann-Whitney U test was used for comparisons. RESULTS: Total dendritic cells were lower in patients than in healthy controls. No quantitative differences were found in CD4 regulatory T cells. CD4(+) T cells from patients produced more IL-4, lL-10, IL-17, and IFN-γ. Patients who experienced the onset of symptoms within the first 5 years of age showed a greater percentage of local (cutaneous lymphocyte antigen +) IL-4- and IL-17-producing cells, while patients who experienced the onset of symptoms after the age of 5 years had a higher percentage of systemic (cutaneous lymphocyte antigen -) IL-10-producing cells. CONCLUSION: Analysis of the cellular immune response against whole flea antigen in patients with papular urticaria by flea bites suggests a possible participation of inflammatory cytokines in the skin reaction (Th17) and a systemic control mechanism (IL-10). This pattern of cytokine production in patients could be a consequence of an impaired dendritic cell population.

Telecommuting and COVID-19: how has the pandemic changed workers' perception on physical and mental health?

Due to the current pandemic situation, work from home, or telecommuting, has been implemented as part of public health measures to prevent the spread of SARS-CoV-2. Although this measure was introduced rapidly, it is likely to remain in effect for some time to prevent further outbreaks of COVID-19. Despite being few, various studies have addressed the relationship between telecommuting and workers' health in the context of the current pandemic. Some aspects observed include fatigue, dietary changes, reduced levels of physical activity, and pain. Other conditions observed are associated with "techno-stress," namely work overload, invasion of privacy, pace of information technology changes, decreased job autonomy, emotional exhaustion, and being constantly in electronic contact with work. Generally speaking, the COVID-19 pandemic has created a new environment for considering work and family life within the discussion on telecommuting. Likewise, a contextualized understanding of factors related to physical and mental well-being is essential to ensure positive impacts on workers. It is important to develop studies and discussions within organizations that allow knowing, analyzing and reformulating strategies and policies aimed at aspects such as changes in workers' physical and mental well-being in the pandemic context and the way how occupational environments at home affect these components.

Debido a la situación actual de la pandemia, el trabajo en casa, o home office, se ha implementado como parte de las medidas de salud pública para prevenir la propagación del SARS-CoV-2. Aunque esta medida se introdujo rápidamente, es probable que permanezca en vigor durante algún tiempo para evitar nuevos brotes de la COVID-19. A pesar de ser pocos, diversos estudios han abordado la relación entre el home office y la salud de los trabajadores en el contexto de la pandemia actual. Algunos aspectos observados incluyen fatiga, alteraciones en la dieta, reducción de la actividad física y dolor. Otras condiciones observadas se encuentran asociadas con el tecnoestrés, a saber: sobrecarga de trabajo, invasión de la privacidad, ritmo del cambio de las tecnologías de la información, menor autonomía laboral, agotamiento emocional y contacto electrónico constante con el trabajo. En términos generales, la pandemia de COVID-19 ha creado un nuevo entorno para considerar la vida laboral y familiar dentro de la discusión del home office. Así mismo, realizar una lectura en contexto de los factores que se relacionan con el bienestar físico y mental es fundamental para garantizar impactos positivos en los trabajadores. Es importante realizar estudios y discusiones al interior de las organizaciones que permitan conocer, analizar y reformular estrategias y políticas orientadas a aspectos como las alteraciones en el bienestar físico y mental de los trabajadores en el contexto de la pandemia y la forma en que los entornos ocupacionales en el hogar afectan estos componentes.

Cardiovascular Risk Factors in Colombian Penitentiary Staff: An Interdisciplinary View of a High-Risk Occupation.

INTRODUCTION: Cardiovascular risk factors have been measured under different conditions, there is some missing information related to specific occupations, such as penitentiary staff which due to their characteristics could have an increased cardiovascular risk. Objective: To determine cardiovascular risk factors on military staff from penitentiary institutions in Santander-Colombia. Methods: Cross-sectional study conducted with 182 workers. Anthropometric parameters, blood pressure, serum lipid profile, and glucose levels were measured. Univariate and bivariate analyses were carried out to establish differences between individuals. Results: Anthropometric and biochemical measures showed that 71.3% participants were overweight or obese, 29.4% presented high blood pressure with increased levels of total cholesterol (27.5%), triglycerides (40.7%), glucose (9.3%), and 84.1% presented low levels of HDL cholesterol. Bivariate analysis found a negative correlation between BMI and HDL cholesterol (p < .05) and a positive correlation between BMI with triglycerides (p < .01), systolic and diastolic blood pressure (DBP) (p < .01). Conclusion: The studied military population presented increased levels of cardiovascular risk in comparison with a similar group in age, gender of nonmilitary individuals. However, it is important to carry out comparative studies between military staff in order to determine the prevalence and other risk predicting factors present in this specific population.

Melanopsin (Opn4) is an oncogene in cutaneous melanoma.

The search for new therapeutical targets for cutaneous melanoma and other cancers is an ongoing task. We expanded this knowledge by evaluating whether opsins, light- and thermo-sensing proteins, could display tumor-modulatory effects on melanoma cancer. Using different experimental approaches, we show that melanoma cell proliferation is slower in the absence of Opn4, compared to Opn4WT due to an impaired cell cycle progression and reduced melanocyte inducing transcription factor (Mitf) expression. In vivo tumor progression of Opn4KO cells is remarkably reduced due to slower proliferation, and higher immune system response in Opn4KO tumors. Using pharmacological assays, we demonstrate that guanylyl cyclase activity is impaired in Opn4KO cells. Evaluation of Tumor Cancer Genome Atlas (TCGA) database confirms our experimental data as reduced MITF and OPN4 expression in human melanoma correlates with slower cell cycle progression and presence of immune cells in the tumor microenvironment (TME). Proteomic analyses of tumor bulk show that the reduced growth of Opn4KO tumors is associated with reduced Mitf signaling, higher translation of G2/M proteins, and impaired guanylyl cyclase activity. Conversely, in Opn4WT tumors increased small GTPase and an immune-suppressive TME are found. Such evidence points to OPN4 as an oncogene in melanoma, which could be pharmacologically targeted.

An Immunometabolic Shift Modulates Cytotoxic Lymphocyte Activation During Melanoma Progression in TRPA1 Channel Null Mice.

Melanoma skin cancer is extremely aggressive with increasing incidence and mortality. Among the emerging therapeutic targets in the treatment of cancer, the family of transient receptor potential channels (TRPs) has been reported as a possible pharmacological target. Specifically, the ankyrin subfamily, representing TRPA1 channels, can act as a pro-inflammatory hub. These channels have already been implicated in the control of intracellular metabolism in several cell models, but little is known about their role in immune cells, and how it could affect tumor progression in a process known as immune surveillance. Here, we investigated the participation of the TRPA1 channel in the immune response against melanoma tumor progression in a mouse model. Using Trpa1 +/+ and Trpa1 -/- animals, we evaluated tumor progression using murine B16-F10 cells and assessed isolated CD8+ T cells for respiratory and cytotoxic functions. Tumor growth was significantly reduced in Trpa1 -/- animals. We observed an increase in the frequency of circulating lymphocytes. Using a dataset of CD8+ T cells isolated from metastatic melanoma patients, we found that TRPA1 reduction correlates with several immunological pathways. Naïve CD8+ T cells from Trpa1 +/+ and Trpa1 -/- animals showed different mitochondrial respiration and glycolysis profiles. However, under CD3/CD28 costimulatory conditions, the absence of TRPA1 led to an even more extensive metabolic shift, probably linked to a greater in vitro killling ability of Trpa1 -/- CD8+ T cells. Therefore, these data demonstrate an unprecedented role of TRPA1 channel in the metabolism control of the immune system cells during carcinogenesis.

Cholesterol-Ester Transfer Protein Alters M1 and M2 Macrophage Polarization and Worsens Experimental Elastase-Induced Pulmonary Emphysema.

Cholesterol-ester transfer protein (CETP) plays a role in atherosclerosis, the inflammatory response to endotoxemia and in experimental and human sepsis. Functional alterations in lipoprotein (LP) metabolism and immune cell populations, including macrophages, occur during sepsis and may be related to comorbidities such as chronic obstructive pulmonary disease (COPD). Macrophages are significantly associated with pulmonary emphysema, and depending on the microenvironment, might exhibit an M1 or M2 phenotype. Macrophages derived from the peritoneum and bone marrow reveal CETP that contributes to its plasma concentration. Here, we evaluated the role of CETP in macrophage polarization and elastase-induced pulmonary emphysema (ELA) in human CETP-expressing transgenic (huCETP) (line 5203, C57BL6/J background) male mice and compared it to their wild type littermates. We showed that bone marrow-derived macrophages from huCETP mice reduce polarization toward the M1 phenotype, but with increased IL-10. Compared to WT, huCETP mice exposed to elastase showed worsened lung function with an increased mean linear intercept (Lm), reflecting airspace enlargement resulting from parenchymal destruction with increased expression of arginase-1 and IL-10, which are M2 markers. The cytokine profile revealed increased IL-6 in plasma and TNF, and IL-10 in bronchoalveolar lavage (BAL), corroborating with the lung immunohistochemistry in the huCETP-ELA group compared to WT-ELA. Elastase treatment in the huCETP group increased VLDL-C and reduced HDL-C. Elastase-induced pulmonary emphysema in huCETP mice promotes lung M2-like phenotype with a deleterious effect in experimental COPD, corroborating the in vitro result in which CETP promoted M2 macrophage polarization. Our results suggest that CETP is associated with inflammatory response and influences the role of macrophages in COPD.

Immunometabolism: A target for the comprehension of immune response toward transplantation.

Organ transplantation is a life-saving procedure, however predicting graft survival is still challenging. Understanding immune-cell pathobiology is critical to the development of effective therapies to prevent rejection. Over the recent years it has become progressively evident that the complex nature of immune cell behavioral dynamics is strongly dependent on cellular metabolism, which in turn, relies on competition for nutrients, oxygen and metabolites with other immune cells and microbiota. Furthermore, the influence of the inflammatory state can lead to substantial changes in conditions within the tissue micro-environment. Considering the context of immunity, alterations in metabolic pathways (glycolysis, the tricarboxylic acid cycle, the pentose phosphate pathway, the fatty acid oxidation and synthesis, and the amino acid metabolic pathways) will influence the production of different sets of cytokines and affect transplantation outcome. It is now known that naïve, resting and effector cells acquire different metabolic profiles and studies have shown that specifically targeting some of these metabolic routes can prevent differentiation of effector T cells in favor of Tregs. Ultimately, to develop effective therapies that will prevent graft loss and understanding how cell metabolism impacts the fate and function of immune cells is now a critical point of discussion. The distinct metabolic features and requirements observed in effector and suppressive cell subsets offer promising opportunities for selective regulation of the immune responses in transplantation and will be discussed in this review.

Aprendizaje basado en proyectos como una estrategia para la enseñanza en ciencias de la salud / Project-based learning as a teaching strategy in health sciences

Introducción: En los procesos de enseñanza-aprendizaje, los profesores deben reflexionar continuamente sobre las estrategias que se aplican directamente en el aula. Se ha estudiado el efecto de metodologías como el aprendizaje basado en problemas, el aprendizaje basado en proyectos, las simulaciones apoyadas en la tecnología, entre otras, en comparación con las metodologías tradicionales. Objetivo: Describir las fortalezas del aprendizaje basado en proyectos como estrategia educativa en el contexto de las ciencias de la salud. Desarrollo: El proceso de enseñanza-aprendizaje de las ciencias de la salud requiere de estrategias dinámicas que permitan articular los conocimientos adquiridos, con una visión práctica de las disciplinas en un contexto basado en la solución de problemas y el pensamiento crítico. Se destaca el aprendizaje basado en proyectos como una metodología que exige una participación activa, investigativa y reflexiva, para la solución de un problema; así, el conocimiento es consecuencia de la discusión, la argumentación y la toma de decisiones, que implican la construcción de dicha solución. Conclusiones: El aprendizaje basado en proyectos puede introducir mejoras y cambios con respecto a las estrategias tradicionales, al fomentar la motivación, el trabajo colaborativo, la adquisición y el afianzamiento de conocimientos en los estudiantes. Asimismo, es una metodología basada en la autoformación del estudiante que fortalece su capacidad de expresión oral y escrita, y la planificación del tiempo, por lo que se facilita la conformación y participación en equipos interdisciplinarios, competencias esenciales para el desempeño ideal de los profesionales de la salud(AU)

Introduction: In teaching-learning processes, professors must continually reflect on the strategies applied directly in the classroom. The effect of methodologies such as problem-based learning, project-based learning, technology-supported simulations, among others, has been studied in comparison with traditional methodologies. Objective: To describe the strengths of project-based learning as an educational strategy in the context of health sciences. Development: The teaching-learning process of health sciences requires dynamic strategies that allow the articulation of the acquired knowledge, with a practical vision of disciplines in a context based on problem solving and critical thinking. Project-based learning is highlighted as a methodology that requires active, investigative and reflective participation to solve a problem; thus, knowledge is the consequence of discussion, argumentation and decision-making, which imply the construction of such a solution. Conclusions: Project-based learning can introduce improvements and changes with respect to traditional strategies, by promoting motivation, collaborative work, as well as knowledge acquisition and consolidation in students. Likewise, it is a methodology based on student self-training and strengthening their capacity for oral and written expression, as well as time planning, thus facilitating the formation and participation in interdisciplinary teams, essential competences for an ideal performance of health professionals(AU)

A Simple Proteomics-Based Approach to Identification of Immunodominant Antigens from a Complex Pathogen: Application to the CD4 T Cell Response against Human Herpesvirus 6B.

Most of humanity is chronically infected with human herpesvirus 6 (HHV-6), with viral replication controlled at least in part by a poorly characterized CD4 T cell response. Identification of viral epitopes recognized by CD4 T cells is complicated by the large size of the herpesvirus genome and a low frequency of circulating T cells responding to the virus. Here, we present an alternative to classical epitope mapping approaches used to identify major targets of the T cell response to a complex pathogen like HHV-6B. In the approach presented here, extracellular virus preparations or virus-infected cells are fractionated by SDS-PAGE, and eluted fractions are used as source of antigens to study cytokine responses in direct ex vivo T cell activation studies. Fractions inducing significant cytokine responses are analyzed by mass spectrometry to identify viral proteins, and a subset of peptides from these proteins corresponding to predicted HLA-DR binders is tested for IFN-γ production in seropositive donors with diverse HLA haplotypes. Ten HHV-6B viral proteins were identified as immunodominant antigens. The epitope-specific response to HHV-6B virus was complex and variable between individuals. We identified 107 peptides, each recognized by at least one donor, with each donor having a distinctive footprint. Fourteen peptides showed responses in the majority of donors. Responses to these epitopes were validated using in vitro expanded cells and naturally expressed viral proteins. Predicted peptide binding affinities for the eight HLA-DRB1 alleles investigated here correlated only modestly with the observed CD4 T cell responses. Overall, the response to the virus was dominated by peptides from the major capsid protein U57 and major antigenic protein U11, but responses to other proteins including glycoprotein H (U48) and tegument proteins U54 and U14 also were observed. These results provide a means to follow and potentially modulate the CD4 T-cell immune response to HHV-6B.

Differences in IgE mediated basophil degranulation induced by proteic fractions from whole flea body extract in patients with papular urticaria by flea bite and healthy controls.

BACKGROUND: Papular urticaria by flea bite (PUFB) is a chronic inflammatory disease in children. The aim of this study was to assess the functional activity of IgE to protein fractions from flea body extract, through basophil degranulation in PUFB patients and controls. METHODS: Basophil degranulation, measured by overexpression of CD63 surface molecules, was evaluated by flow cytometry in samples from patients and controls. Cell stimulation was performed with three fractions with different molecular weight from flea body extract using a Basotest® modified protocol. Mann-Whitney U-test was used for comparisons. RESULTS: Specific IgE from PUFB patients and healthy controls induced basophil degranulation to flea body extract with no significant differences between them (16.2 ± 3.1% vs 13.6 ± 2.8% p = 0.77). However, when flea extract was analyzed in fractions with proteins ranging different molecular weights, significant differences were observed on the response from patients compared with controls to <50 kD (14.9 ± 5.1% vs 9.7 ± 2.1% p = 0.0058) and 50-100 kD proteic fractions (8.3 ± 3.2% vs 2.8 ± 1.6% p = 0.0021). CONCLUSION: In this study, was established that the differential response by IgE, in PUFB, depends from the molecular weight of the antigens contained in the flea extract. These antigens may be related to 30-35 kD proteins previously described as major allergens.

Proteína C reactiva ultrasensible (PCR-us)como marcador de riesgo de enfermedad cardiovascular / C-reactive protein (hs-CRP) as a marker of cardiovascular disease risk

La importancia de los factores de riesgo clásicos ha sido demostrada para el desarrollo de la enfermedad cardiovascular. Aun así, se han identificado nuevos factores de riesgo, llamados factores de riesgo emergentes, que optimizan el manejo y la detección del riesgo cardiovascular y en consecuencia benefician la población con más programas eficientes de prevención. Para mejorar la predicción del riesgo cardiovascular, la comunidad científica ha centrado su interés en la determinación de la proteína C reactiva ultrasensible (PCR-us), como un marcador de inflamación, puesto que diferentes estudios epidemiológicos prospectivos han demostrado su utilidad en la predicción de la incidencia de algunas enfermedades cardiovasculares, tales como infarto agudo de miocardio, accidente cerebrovascular y enfermedad vascular periférica. Adicionalmente, varias investigaciones de los últimos años han mostrado la participación de diferentes células y moléculas de la respuesta inmune, como mediadores de la lesión vascular relacionada con la ateroesclerosis. La proteína C reactiva es una proteína de fase aguda que ha sido clásicamente considerada como un marcador de inflamación. Bajo condiciones normales, su síntesis hepática es menor a 1 mg/L, la cual se ve incrementada en una persona que desarrolla un proceso inflamatorio o infeccioso. Esta elevación puede ser hasta de 100 veces el valor normal durante las primeras 24 a 48 horas y se mantiene así por varios días. Se ha demostrado que la PCR-us brinda información en cada uno de los niveles de riesgo cardiovascular de acuerdo a la escala de Framingham; niveles de PCR-us menores a 1 mg/L, entre 1 y 3 mg/L, y mayores a 3 mg/L, corresponden al riesgo cardiovascular bajo, medio y alto, respectivamente. El valor predictor de la PCR-us se incrementa cuando es evaluado en asociación con el perfil lipídico y se realiza una correlación apropiada.