RESUMEN
Immunotoxicology/immunosafety science is rapidly evolving, with novel modalities and immuno-oncology among the primary drivers of new tools and technologies. The Immunosafety Working Group of IQ/DruSafe sought to better understand some of the key challenges in immunosafety evaluation, gaps in the science, and current limitations in methods and data interpretation. A survey was developed to provide a baseline understanding of the needs and challenges faced in immunosafety assessments, the tools currently being applied across the industry, and the impact of feedback received from regulatory agencies. This survey also focused on current practices and challenges in conducting the T-cell-dependent antibody response (TDAR) and the cytokine release assay (CRA). Respondents indicated that ICH S8 guidance was insufficient for the current needs of the industry portfolio of immunomodulators and novel modalities and should be updated. Other challenges/gaps identified included translation of nonclinical immunosafety assessments to the clinic, and lack of relevant nonclinical species and models in some cases. Key areas of emerging science that will add future value to immunotoxicity assessments include development of additional in vitro and microphysiological system models, as well as application of humanized mouse models. Efforts are ongoing in individual companies and consortia to address some of these gaps and emerging science.
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Factores Inmunológicos , Humanos , Animales , Encuestas y Cuestionarios , Factores Inmunológicos/efectos adversos , Factores Inmunológicos/toxicidad , Citocinas/inmunología , Medición de Riesgo , Evaluación Preclínica de Medicamentos/métodos , Pruebas de Toxicidad/métodosRESUMEN
With the growth of monoclonal antibodies and other proteins as major modalities in the pharmaceutical industry, there has been an increase in pharmacology and toxicity testing of biotherapeutics in animals. Animals frequently mount an immune response to human therapeutic proteins. This can result in asymptomatic anti-drug antibody formation, immune complexes that affect drug disposition and/or organ function such as kidney, cytokine release responses, fatal hypersensitivity, or a range of reactions in between. In addition, an increasing number of oncology therapeutics are being developed that enhance or directly stimulate immune responses by a variety of mechanisms, which could increase the risk of autoreactivity and an autoimmune-like syndrome in animals and humans. When evaluating the risk of biotherapeutics prior to entering the clinic, the nonclinical safety data may include any of these responses and it is critical to understand whether they represent a safety liability for humans. The DruSafe Leadership group of the IQ Consortium conducted a survey of industry to understand sponsors' experiences with these immune reactions in nonclinical studies related to both immunogenicity and pharmacologically-mediated immune perturbations. The survey covered what pathways were affected, how the immune responses were presented, how the company and health authorities interpreted the data and whether the immune responses were observed in the clinic. Additionally, the survey gathered information on association of these findings with anti-drug antibodies as well as sponsor's use of immunogenicity predictive tools. The data suggests that the ability of a biotherapeutic to activate the immune system, intended or not, plays a significant role on characteristics of the response and whether theys are translatable.
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Productos Biológicos/toxicidad , Sistema Inmunológico/efectos de los fármacos , Animales , Anticuerpos/inmunología , Productos Biológicos/inmunología , Evaluación Preclínica de Medicamentos , Industria Farmacéutica , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos , Macaca fascicularis , Ratones , Ratas , Encuestas y Cuestionarios , Pruebas de ToxicidadRESUMEN
Hydraulic fracturing ("fracking") is a process used to enhance retrieval of gas from subterranean natural gas-laden rock by fracturing it under pressure. Sand used to stabilize fissures and facilitate gas flow creates a potential occupational hazard from respirable fracking sand dust (FSD). As studies of the immunotoxicity of FSD are lacking, the effects of whole-body inhalation (6 h/d for 4 d) of a FSD, i.e., FSD 8, was investigated at 1, 7, and 27 d post-exposure in rats. Exposure to 10 mg/m3 FSD 8 resulted in decreased lung-associated lymph node (LLN) cellularity, total B-cells, CD4+ T-cells, CD8+ T-cells and total natural killer (NK) cells at 7-d post exposure. The frequency of CD4+ T-cells decreased while the frequency of B-cells increased (7 and 27 d) in the LLN. In contrast, increases in LLN cellularity and increases in total CD4+ and CD8+ T-cells were observed in rats following 30 mg/m3 FSD 8 at 1 d post-exposure. Increases in the frequency and number of CD4+ T-cells and NK cells were observed in bronchial alveolar lavage fluid at 7-d post-exposure (10 mg/m3) along with an increase in total CD4+ T-cells, CD11b + cells, and NK cells at 1-day post-exposure (30 mg/m3). Increases in the numbers of B-cells and CD8+ T-cells were observed in the spleen at 1-day post 30 mg/m3 FSD 8 exposure. In addition, NK cell activity was suppressed at 1 d (30 mg/m3) and 27 d post-exposure (10 mg/m3). No change in the IgM response to sheep red blood cells was observed. The findings indicate that FSD 8 caused alterations in cellularity, phenotypic subsets, and impairment of immune function.
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Polvo , Fracking Hidráulico , Arena , Administración por Inhalación , Animales , Líquido del Lavado Bronquioalveolar/inmunología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Línea Celular , Eritrocitos , Inmunoglobulina M/inmunología , Células Asesinas Naturales/inmunología , Ganglios Linfáticos/inmunología , Masculino , Ratones , Ratas Sprague-Dawley , Ovinos , Bazo/inmunologíaRESUMEN
CD4 T cells can differentiate into multiple effector subsets, including ThCTL that mediate MHC class II-restricted cytotoxicity. Although CD4 T cell-mediated cytotoxicity has been reported in multiple viral infections, their characteristics and the factors regulating their generation are unclear, in part due to a lack of a signature marker. We show in this article that, in mice, NKG2C/E identifies the ThCTL that develop in the lung during influenza A virus infection. ThCTL express the NKG2X/CD94 complex, in particular the NKG2C/E isoforms. NKG2C/E+ ThCTL are part of the lung CD4 effector population, and they mediate influenza A virus-specific cytotoxic activity. The phenotype of NKG2C/E+ ThCTL indicates they are highly activated effectors expressing high levels of binding to P-selectin, T-bet, and Blimp-1, and that more of them secrete IFN-γ and readily degranulate than non-ThCTL. ThCTL also express more cytotoxicity-associated genes including perforin and granzymes, and fewer genes associated with recirculation and memory. They are found only at the site of infection and not in other peripheral sites. These data suggest ThCTL are marked by the expression of NKG2C/E and represent a unique CD4 effector population specialized for cytotoxicity.
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Linfocitos T CD4-Positivos/inmunología , Citotoxicidad Inmunológica , Virus de la Influenza A , Subfamília C de Receptores Similares a Lectina de Células NK/análisis , Infecciones por Orthomyxoviridae/inmunología , Animales , Biomarcadores/análisis , Linfocitos T CD4-Positivos/química , Linfocitos T CD4-Positivos/clasificación , Interferón gamma/biosíntesis , Pulmón/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Factor 1 de Unión al Dominio 1 de Regulación Positiva , Factores de Transcripción/análisisRESUMEN
Activation of the aryl hydrocarbon receptor (AhR) by immunosuppressive ligands promotes the development of regulatory T (Treg) cells. Although AhR-induced Foxp3+ Treg cells have been well studied, much less is known about the development and fate of AhR-induced Type 1 Treg (AhR-Tr1) cells. In the current study, we identified the unique transcriptional and functional changes in murine CD4+ T cells that accompany the differentiation of AhR-Tr1 cells during the CD4+ T-cell-dependent phase of an allospecific cytotoxic T lymphocyte (allo-CTL) response. AhR activation increased the expression of genes involved in T-cell activation, immune regulation and chemotaxis, as well as a global downregulation of genes involved in cell cycling. Increased IL-2 production was responsible for the early AhR-Tr1 activation phenotype previously characterized as CD25+ CTLA4+ GITR+ on day 2. The AhR-Tr1 phenotype was further defined by the coexpression of the immunoregulatory receptors Lag3 and Tim3 and non-overlapping expression of CCR4 and CCR9. Consistent with the increased expression of CCR9, real-time imaging showed enhanced migration of AhR-Tr1 cells to the lamina propria of the small intestine and colon. The discovery of mucosal imprinting of AhR-Tr1 cells provides an additional mechanism by which therapeutic AhR ligands can control immunopathology.
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Diferenciación Celular/inmunología , Interleucina-2/biosíntesis , Receptores de Hidrocarburo de Aril/inmunología , Linfocitos T Reguladores/inmunología , Aloinjertos , Animales , Linfocitos T CD4-Positivos/inmunología , Movimiento Celular/inmunología , Mucosa Intestinal/inmunología , Activación de Linfocitos/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones TransgénicosRESUMEN
BACKGROUND: Aspergillus fumigatus-induced allergic airway disease has been shown to involve conidial germination in vivo, but the immunological mechanisms remain uncharacterized. OBJECTIVE: A subchronic murine exposure model was used to examine the immunological mediators that are regulated in response to either culturable or nonculturable A fumigatus conidia. METHODS: Female B6C3F1/N mice were repeatedly dosed via inhalation with 1 × 105 viable or heat-inactivated conidia (HIC), twice per week for 13 weeks (26 exposures). Control mice inhaled high-efficiency particulate arrestor-filtered air. The influence of A fumigatus conidial germination on the pulmonary immunopathological outcomes was evaluated by flow cytometry analysis of cellular infiltration in the airways, assessment of lung messenger RNA expression, quantitative proteomics, and histopathology of whole lung tissue. RESULTS: Repeated inhalation of viable conidia, but not HIC, resulted in allergic inflammation marked by vascular remodeling, extensive eosinophilia, and accumulation of alternatively activated macrophages (AAMs) in the murine airways. More specifically, mice that inhaled viable conidia resulted in a mixed TH1 and TH2 (IL-13) cytokine response. Recruitment of eosinophils corresponded with increased Ccl11 transcripts. Furthermore, genes associated with M2 or alternatively activated macrophage polarization (eg, Arg1, Chil3, and Retnla) were significantly up-regulated in viable A fumigatus-exposed mice. In mice inhaling HIC, CD4+ T cells expressing IFN-γ (TH1) dominated the lymphocytic infiltration. Quantitative proteomics of the lung revealed metabolic reprogramming accompanied by mitochondrial dysfunction and endoplasmic reticulum stress stimulated by oxidative stress from repetitive microbial insult. CONCLUSION: Our studies demonstrate that A fumigatus conidial viability in vivo is critical to the immunopathological presentation of chronic fungal allergic disease.
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Alérgenos/inmunología , Antígenos Fúngicos/inmunología , Aspergilosis/inmunología , Aspergillus fumigatus/fisiología , Hipersensibilidad/inmunología , Esporas Fúngicas/inmunología , Células Th2/inmunología , Administración por Inhalación , Animales , Supervivencia Celular , Células Cultivadas , Modelos Animales de Enfermedad , Eosinofilia , Femenino , Humanos , Interleucina-13/metabolismo , Activación de Macrófagos , RatonesRESUMEN
Over the last decade, the known spectrum of CD4(+) T-cell effector subsets has become much broader, and it has become clear that there are multiple dimensions by which subsets with a particular cytokine commitment can be further defined, including their stage of differentiation, their location, and, most importantly, their ability to carry out discrete functions. Here, we focus on our studies that highlight the synergy among discrete subsets, especially those defined by helper and cytotoxic function, in mediating viral protection, and on distinctions between CD4(+) T-cell effectors located in spleen, draining lymph node, and in tissue sites of infection. What emerges is a surprising multiplicity of CD4(+) T-cell functions that indicate a large arsenal of mechanisms by which CD4(+) T cells act to combat viruses.
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Linfocitos T CD4-Positivos/inmunología , Memoria Inmunológica , Infecciones del Sistema Respiratorio/inmunología , Infecciones del Sistema Respiratorio/virología , Virosis/inmunología , Virus/inmunología , Animales , Citocinas/inmunología , Citocinas/metabolismo , Regulación de la Expresión Génica , Humanos , Virus de la Influenza A/inmunología , Gripe Humana/inmunología , Gripe Humana/prevención & control , Infecciones por Orthomyxoviridae/inmunología , Infecciones por Orthomyxoviridae/prevención & control , Infecciones del Sistema Respiratorio/genética , Infecciones del Sistema Respiratorio/metabolismo , Factores de Transcripción/metabolismo , Virosis/genética , Virosis/metabolismoRESUMEN
N-Butylbenzene sulfonamide (NBBS) is a commonly used plasticizer found in numerous products. Due to its extensive use, lack of adequate toxicological data, and suspicion of toxicity based on the presence of structural alerts, it was nominated to the National Toxicology Program for comprehensive toxicological testing. The purpose of this study was to evaluate the potential for hypersensitivity and immune suppression following dermal exposure to NBBS using a murine model. NBBS tested negative in a combined irritancy/local lymph node assay (LLNA), classifying it as nonirritating and nonsensitizing. To estimate the immunosuppressive potential of NBBS, assays that assessed immunotoxicity were performed, including the immumnoglobulin (Ig) M response to T-cell-dependent antigen sheep red blood cells (SRBC), using the plaque-forming cell (PFC) assay and immune cell phenotyping. After a 28-d treatment with NBBS, mice exposed to the lowest concentration (25% NBBS) showed a significant increase in IgM-producing B cells in the spleen. No marked changes were identified in immune cell markers in the lymph node. In contrast to body weight, a significant elevation in kidney and liver weight was observed following dermal exposure to all concentrations of NBBS. These results demonstrate that dermal exposure to NBBS, other than liver and kidney toxicity, did not apparently induce immunotoxicity in a murine model.
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Plastificantes/toxicidad , Sulfonamidas/toxicidad , Administración Cutánea , Animales , Femenino , Inmunoglobulina G/inmunología , Inmunosupresores/farmacología , Ganglios Linfáticos/inmunología , Ratones , Ratones Endogámicos BALB C , Ovinos , Bazo/efectos de los fármacos , Bazo/inmunología , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Pruebas de ToxicidadRESUMEN
Spot welding is used in the automotive and aircraft industries, where high-speed, repetitive welding is needed to join thin sections of metal. Epoxy adhesives are applied as sealers to the metal seams. Pulmonary function abnormalities and airway irritation have been reported in spot welders, but no animal toxicology studies exist. Therefore, the goal of this study was to investigate vascular, immune and lung toxicity measures after exposure to these metal fumes in an animal model. Male Sprague-Dawley rats were exposed by inhalation to 25 mg/m³ to either mild-steel spot welding aerosols with sparking (high metal, HM) or without sparking (low metal, LM) for 4 h/d for 3, 8 and 13 d. Shams were exposed to filtered air. Bronchoalveolar lavage (BAL), lung gene expression and ex vivo BAL cell challenge were performed to assess lung toxicity. Lung resistance (R(L)) was evaluated before and after challenge with inhaled methacholine (MCh). Functional assessment of the vascular endothelium in isolated rat tail arteries and leukocyte differentiation in the spleen and lymph nodes via flow cytometry was also done. Immediately after exposure, baseline R(L) was significantly elevated in the LM spot welding aerosols, but returned to control level by 24 h postexposure. Airway reactivity to MCh was unaffected. Lung inflammation and cytotoxicity were mild and transient. Lung epithelial permeability was significantly increased after 3 and 8 d, but not after 13 d of exposure to the HM aerosol. HM aerosols also caused vascular endothelial dysfunction and increased CD4+, CD8+ and B cells in the spleen. Only LM aerosols caused increased IL-6 and MCP-1 levels compared with sham after ex vivo LPS stimulation in BAL macrophages. Acute inhalation of mild-steel spot welding fumes at occupationally relevant concentrations may act as an irritant as evidenced by the increased R(L) and result in endothelial dysfunction, but otherwise had minor effects on the lung.
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Contaminantes Ocupacionales del Aire/toxicidad , Endotelio Vascular/efectos de los fármacos , Exposición por Inhalación/efectos adversos , Pulmón/efectos de los fármacos , Mucosa Respiratoria/efectos de los fármacos , Vasculitis/inducido químicamente , Soldadura , Adhesivos/química , Aerosoles , Animales , Células Cultivadas , Endotelio Vascular/inmunología , Endotelio Vascular/fisiopatología , Incendios , Hematopoyesis Extramedular/efectos de los fármacos , Inmunidad Mucosa/efectos de los fármacos , Leucocitos/efectos de los fármacos , Leucocitos/inmunología , Leucocitos/patología , Pulmón/irrigación sanguínea , Pulmón/inmunología , Pulmón/patología , Macrófagos Alveolares/efectos de los fármacos , Macrófagos Alveolares/inmunología , Macrófagos Alveolares/patología , Masculino , Ratas Sprague-Dawley , Mucosa Respiratoria/inmunología , Mucosa Respiratoria/patología , Organismos Libres de Patógenos Específicos , Bazo/efectos de los fármacos , Bazo/inmunología , Bazo/patología , Acero/química , Pruebas de Toxicidad Aguda , Vasculitis/inmunología , Vasculitis/patología , Vasculitis/fisiopatología , Soldadura/métodosRESUMEN
Immune responses are heavily involved in the regulation and pathogenesis of human diseases, including infectious diseases, inflammatory and autoimmune conditions, cancer, neurological disorders, and cardiometabolic syndromes. The immune system is considered a double-edged sword serving as a powerful host defense mechanism against infection and cancerous cells and causing detrimental tissue damage when the immune response is exaggerated or uncontrollable. One of the challenges in studying the efficacy and toxicity of drugs that target or modulate the immune system is the lack of suitable preclinical human models that are predictive of human response. Recent advancements in human microphysiological systems (MPS) have provided a promising in vitro platform to evaluate the response of immune organs ex vivo, to investigate the interaction of immune cells with non-lymphoid tissue cells, and to reduce the reliance on animals in preclinical studies. The development, regulation, trafficking, and responses of immune cells have been extensively studied in preclinical animal models and clinically, providing a wealth of knowledge by which to evaluate new in vitro models. Therefore, the application of immunocompetent MPS in drug discovery and development should first verify that the immune response in an MPS model recapitulates the complexity of the human immune physiology. This manuscript reviews biological functions of immune organ systems and tissue-resident immune cells and discusses contexts-of-use for commonly used immunocompetent and immune organ MPS models. Current perspective and recommendations are provided to guide the continued development of immune organ and immunocompetent MPS models and their application in drug discovery and development.
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CD4 T cells that acquire cytotoxic phenotype and function have been repeatedly identified in humans, mice, and other species in response to many diverse pathogens. Since CD4 cytotoxic T cells are able to recognize antigenic determinants unique from those recognized by the parallel CD8 cytotoxic T cells, they can potentially contribute additional immune surveillance and direct effector function by lysing infected or malignant cells. Here, we briefly review much of what is known about the generation of cytotoxic CD4 T cells and describe our current understanding of their role in antiviral immunity. Furthering our understanding of the many roles of CD4 T cells during an anti-viral response is important for developing effective vaccine strategies that promote long-lasting protective immunity.
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Linfocitos T CD4-Positivos/inmunología , Linfocitos T Citotóxicos/inmunología , Animales , Linfocitos T CD8-positivos/inmunología , Granzimas/química , Antígenos de Histocompatibilidad Clase II/química , Humanos , Sistema Inmunológico , Proteína 1 de la Membrana Asociada a los Lisosomas/química , Ratones , Modelos Biológicos , FenotipoRESUMEN
Triclosan is an antimicrobial chemical used in healthcare settings that can be absorbed through the skin. Exposure to triclosan has been positively associated with food and aeroallergy and asthma exacerbation in humans and, although not directly sensitizing, has been demonstrated to augment the allergic response in a mouse model of asthma. The skin barrier and microbiome are thought to play important roles in regulating inflammation and allergy and disruptions may contribute to development of allergic disease. To investigate potential connections of the skin barrier and microbiome with immune responses to triclosan, SKH1 mice were exposed dermally to triclosan (0.5-2%) or vehicle for up to 7 consecutive days. Exposure to 2% triclosan for 5-7 days on the skin was shown to increase transepidermal water loss levels. Seven days of dermal exposure to triclosan decreased filaggrin 2 and keratin 10 expression, but increased filaggrin and keratin 14 protein along with the danger signal S100a8 and interleukin-4. Dermal exposure to triclosan for 7 days also altered the alpha and beta diversity of the skin and gut microbiome. Specifically, dermal triclosan exposure increased the relative abundance of the Firmicutes family, Lachnospiraceae on the skin but decreased the abundance of Firmicutes family, Ruminococcaceae in the gut. Collectively, these results demonstrate that repeated dermal exposure to the antimicrobial chemical triclosan alters the skin barrier integrity and microbiome in mice, suggesting that these changes may contribute to the increase in allergic immune responses following dermal exposure to triclosan.
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Antiinfecciosos , Microbiota , Triclosán , Animales , Inmunidad , Ratones , Piel , Triclosán/toxicidadRESUMEN
Companion animals are susceptible to SARS-CoV-2 infection and sporadic cases of pet infections have occurred in the United Kingdom. Here we present the first large-scale serological survey of SARS-CoV-2 neutralising antibodies in dogs and cats in the UK. Results are reported for 688 sera (454 canine, 234 feline) collected by a large veterinary diagnostic laboratory for routine haematology during three time periods; pre-COVID-19 (January 2020), during the first wave of UK human infections (April-May 2020) and during the second wave of UK human infections (September 2020-February 2021). Both pre-COVID-19 sera and those from the first wave tested negative. However, in sera collected during the second wave, 1.4% (n=4) of dogs and 2.2% (n=2) cats tested positive for neutralising antibodies. The low numbers of animals testing positive suggests pet animals are unlikely to be a major reservoir for human infection in the UK. However, continued surveillance of in-contact susceptible animals should be performed as part of ongoing population health surveillance initiatives.
RESUMEN
Companion animals are susceptible to SARS-CoV-2 infection and sporadic cases of pet infections have occurred in the United Kingdom. Here we present the first large-scale serological survey of SARS-CoV-2 neutralising antibodies in dogs and cats in the UK. Results are reported for 688 sera (454 canine, 234 feline) collected by a large veterinary diagnostic laboratory for routine haematology during three time periods; pre-COVID-19 (January 2020), during the first wave of UK human infections (April-May 2020) and during the second wave of UK human infections (September 2020-February 2021). Both pre-COVID-19 sera and those from the first wave tested negative. However, in sera collected during the second wave, 1.4% (n â= â4) of dogs and 2.2% (n â= â2) of cats tested positive for neutralising antibodies. The low numbers of animals testing positive suggests pet animals are unlikely to be a major reservoir for human infection in the UK. However, continued surveillance of in-contact susceptible animals should be performed as part of ongoing population health surveillance initiatives.
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Although the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) are mediated through binding and activation of the aryl hydrocarbon receptor (AhR), the subsequent biochemical and molecular changes that confer immune suppression are not well understood. Mice exposed to TCDD during an acute B6-into-B6D2F1 graft-vs-host response do not develop disease, and recently this has been shown to correlate with the generation of CD4(+) T cells that express CD25 and demonstrate in vitro suppressive function. The purpose of this study was to further characterize these CD4(+) cells (TCDD-CD4(+) cells) by comparing and contrasting them with both natural regulatory CD4(+) T cells (T-regs) and vehicle-treated cells. Cellular anergy, suppressive functions, and cytokine production were examined. We found that TCDD-CD4(+) cells actively proliferate in response to various stimuli but suppress IL-2 production and the proliferation of effector T cells. Like natural T-regs, TCDD-CD4(+) cells do not produce IL-2 and their suppressive function is contact dependent but abrogated by costimulation through glucocorticoid-induced TNFR (GITR). TCDD-CD4(+) cells also secrete significant amounts of IL-10 in response to both polyclonal and alloantigen stimuli. Several genes were significantly up-regulated in TCDD-CD4(+) cells including TGF-beta3, Blimp-1, and granzyme B, as well as genes associated with the IL12-Rb2 signaling pathway. TCDD-CD4(+) cells demonstrated an increased responsiveness to IL-12 as indicated by the phosphorylation levels of STAT4. Only 2% of TCDD-CD4(+) cells express Foxp3, suggesting that the AhR does not rely on Foxp3 for suppressive activity. The generation of CD4(+) cells with regulatory function mediated through activation of the AhR by TCDD may represent a novel pathway for the induction of T-regs.
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Regulación de la Expresión Génica/efectos de los fármacos , Inmunosupresores/farmacología , Activación de Linfocitos/efectos de los fármacos , Dibenzodioxinas Policloradas/farmacología , Linfocitos T Reguladores/efectos de los fármacos , Linfocitos T Reguladores/inmunología , Enfermedad Aguda , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/inmunología , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Factores de Transcripción Forkhead/biosíntesis , Regulación de la Expresión Génica/inmunología , Reacción Injerto-Huésped/efectos de los fármacos , Reacción Injerto-Huésped/inmunología , Inmunosupresores/administración & dosificación , Activación de Linfocitos/inmunología , Ratones , Ratones Endogámicos C57BL , Dibenzodioxinas Policloradas/administración & dosificación , Receptores de Hidrocarburo de Aril/metabolismo , Linfocitos T Reguladores/citología , Linfocitos T Reguladores/metabolismoRESUMEN
Healthcare workers concurrently may be at a higher risk of developing respiratory infections and allergic disease, such as asthma, than the general public. Increased incidence of allergic diseases is thought to be caused, in part, due to occupational exposure to chemicals that induce or augment Th2 immune responses. However, whether exposure to these chemical antimicrobials can influence immune responses to respiratory pathogens is unknown. Here, we use a BALB/c murine model to test if the Th2-promoting antimicrobial chemical triclosan influences immune responses to influenza A virus. Mice were dermally exposed to 2% triclosan for 7 days prior to infection with a sub-lethal dose of mouse adapted PR8 A(H1N1) virus (50 pfu); triclosan exposure continued until 10 days post infection (dpi). Infected mice exposed to triclosan did not show an increase in morbidity or mortality, and viral titers were unchanged. Assessment of T cell responses at 10 dpi showed a decrease in the number of total and activated (CD44hi) CD4+ and CD8+ T cells at the site of infection (BAL and lung) in triclosan exposed mice compared to controls. Influenza-specific CD4+ and CD8+ T cells were assessed using MHCI and MHCII tetramers, with reduced populations, although not reaching statistical significance at these sites following triclosan exposure. Reductions in the Th1 transcription factor T-bet were seen in both activated and tetramer+ CD4+ and CD8+ T cells in the lungs of triclosan exposed infected mice, indicating reduced Th1 polarization and providing a potential mechanism for numerical reduction in T cells. Overall, these results indicate that the immune environment induced by triclosan exposure has the potential to influence the developing immune response to a respiratory viral infection and may have implications for healthcare workers who may be at an increased risk for developing infectious diseases.
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Inmunidad Adaptativa/efectos de los fármacos , Personal de Salud , Gripe Humana/inmunología , Exposición Profesional/efectos adversos , Células TH1/efectos de los fármacos , Triclosán/efectos adversos , Administración Tópica , Animales , Modelos Animales de Enfermedad , Femenino , Humanos , Subtipo H1N1 del Virus de la Influenza A/inmunología , Gripe Humana/prevención & control , Gripe Humana/virología , Ratones , Células TH1/inmunología , Triclosán/administración & dosificaciónRESUMEN
The anti-microbial compound triclosan is incorporated into numerous consumer products and is detectable in the urine of 75% of the general United States population. Recent epidemiological studies report positive associations with urinary triclosan levels and allergic disease. Although not sensitizing, earlier studies previously found that repeated topical application of triclosan augments the allergic response to ovalbumin (OVA) though a thymic stromal lymphopoietin (TSLP) pathway in mice. In the present study, early immunological effects following triclosan exposure were further evaluated following topical application in a murine model. These investigations revealed abundant expression of S100A8/A9, which reportedly acts as an endogenous ligand for Toll-like Receptor 4 (TLR4), in skin tissues and in infiltrating leukocytes during topical application of 0.75-3.0% triclosan. Expression of Tlr4 along with Tlr1, Tlr2 and Tlr6 increased in skin tissues over time with triclosan exposure; high levels of TLR4 were expressed on skin-infiltrating leukocytes. In vivo antibody blockade of the TLR4/MD-2 receptor complex impaired local inflammatory responses after four days, as evidenced by decreased Il6, Tnfα, S100a8, S100a9, Tlr1, Tlr2, Tlr4 and Tlr6 expression in the skin and decreased lymph node cellularity and production of IL-4 and IL-13 by lymph node T-cells. After nine days of triclosan exposure with TLR4/MD-2 blockade, impaired T-helper cell type 2 (TH2) cytokine responses were sustained, but other early effects on skin and lymph node cellularity were lost; this suggested alternative ligands/receptors compensated for the loss of TLR4 signaling. Taken together, these data suggest the S100A8/A9-TLR4 pathway plays an early role in augmenting immunomodulatory responses with triclosan exposure and support a role for the innate immune system in chemical adjuvancy.
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Antiinfecciosos/administración & dosificación , Calgranulina A/metabolismo , Calgranulina B/metabolismo , Hipersensibilidad/inmunología , Piel/inmunología , Receptor Toll-Like 4/metabolismo , Triclosán/administración & dosificación , Administración Tópica , Alérgenos/efectos adversos , Animales , Antiinfecciosos/efectos adversos , Anticuerpos Bloqueadores/administración & dosificación , Células Cultivadas , Citocinas/metabolismo , Modelos Animales de Enfermedad , Femenino , Humanos , Inmunidad Innata , Inmunomodulación , Ratones , Ratones Endogámicos BALB C , Transducción de Señal , Células Th2/inmunología , Triclosán/efectos adversosRESUMEN
Antisense oligomers have been shown to be effective tools for inhibiting gene expression in a highly specific manner. This technology has proven to be invaluable for determining gene function in conventional molecular and cellular studies. However, the promise of an antisense-based drug technology, suggested by antiviral efficacy shown nearly 25 years ago, is just now coming of age. Since then, numerous antisense approaches have been shown to be effective in animal models against numerous viruses and some tumors. Not surprisingly, antisense agents targeting these diseases are taking the lead in human clinical trials and FDA approval. Although comparatively smaller in scope, approaches for modulating immune responses to treat Crohn's disease, diabetes, multiple sclerosis and transplant rejection appear to be the next burgeoning phase of development in antisense therapy.
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Enfermedades Autoinmunes/tratamiento farmacológico , Enfermedades Autoinmunes/inmunología , Supervivencia de Injerto/efectos de los fármacos , Supervivencia de Injerto/inmunología , Oligonucleótidos Antisentido/farmacología , Oligonucleótidos Antisentido/uso terapéutico , Enfermedades Autoinmunes/genética , Supervivencia de Injerto/genética , Humanos , Oligonucleótidos Antisentido/genética , Linfocitos T Reguladores/efectos de los fármacos , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismoRESUMEN
Toluene diisocyanate (TDI) is a potent low molecular weight chemical sensitizer and a leading cause of chemical-induced occupational asthma. The regulatory potential of microRNAs (miRNAs) has been recognized in a variety of disease states, including allergic disease; however, the roles of miRNAs in chemical sensitization are largely unknown. In a previous work, increased expression of multiple miRNAs during TDI sensitization was observed and several putative mRNA targets identified for these miRNAs were directly related to regulatory T-cell (Treg) differentiation and function including Foxp3 and Runx3. In this work, we show that miR-210 expression is increased in the mouse draining lymph node (dLN) and Treg subsets following dermal TDI sensitization. Alterations in dLN mRNA and protein expression of Treg related genes/putative miR-210 targets (foxp3, runx3, ctla4, and cd25) were observed at multiple time points following TDI exposure and in ex vivo systems. A Treg suppression assay, including a miR-210 mimic, was utilized to investigate the suppressive ability of Tregs. Cells derived from TDI sensitized mice treated with miR-210 mimic had less expression of miR-210 compared to the acetone control suggesting other factors, such as additional miRNAs, might be involved in the regulation of the functional capabilities of these cells. These novel findings indicate that miR-210 may have an inhibitory role in Treg function during TDI sensitization. Because the functional roles of miRNAs have not been previously elucidated in a model of chemical sensitization, these data contribute to the understanding of the potential immunologic mechanisms of chemical induced allergic disease.
RESUMEN
Triclosan is an antimicrobial chemical commonly used occupationally and by the general public. Using select immune function assays, the purpose of these studies was to evaluate the immunotoxicity of triclosan following dermal exposure using a murine model. Triclosan was not identified to be a sensitizer in the murine local lymph node assay (LLNA) when tested at concentrations ranging from 0.75-3.0%. Following a 28-day exposure, triclosan produced a significant increase in liver weight at concentrations of ≥ 1.5%. Exposure to the high dose (3.0%) also produced a significant increase in spleen weights and number of platelets. The absolute number of B-cells, T-cells, dendritic cells and NK cells were significantly increased in the skin draining lymph node, but not the spleen. An increase in the frequency of dendritic cells was also observed in the lymph node following exposure to 3.0% triclosan. The IgM antibody response to sheep red blood cells (SRBC) was significantly increased at 0.75% - but not at the higher concentrations - in the spleen and serum. These results demonstrate that dermal exposure to triclosan induces stimulation of the immune system in a murine model and raise concerns about potential human exposure.