Increased Expression Profile and Functionality of TLR6 in Peripheral Blood Mononuclear Cells and Hepatocytes of Morbidly Obese Patients with Non-Alcoholic Fatty Liver Disease.

Current evidence suggests that gut dysbiosis drives obesity and non-alcoholic fatty liver disease (NAFLD) pathogenesis. Toll-like receptor 2 (TLR2) and TLR6 specifically recognize components of Gram-positive bacteria. Despite the potential implications of TLR2 in NAFLD pathogenesis, the role of TLR6 has not been addressed. Our aim is to study a potential role of TLR6 in obesity-related NAFLD. Forty morbidly obese patients undergoing bariatric surgery were prospectively studied. Cell surface expression of TLR2 and TLR6 was assessed on peripheral blood mononuclear cells (PBMCs) by flow cytometry. Freshly isolated monocytes were cultured with specific TLR2/TLR6 agonists and intracellular production of cytokines was determined by flow-cytometry. In liver biopsies, the expression of TLR2 and TLR6 was analyzed by immunohistochemistry and cytokine gene expression using RT-qPCR. TLR6 expression in PBMCs from non-alcoholic steatohepatitis (NASH) patients was significantly higher when compared to those from simple steatosis. The production of pro-inflammatory cytokines in response to TLR2/TLR6 stimulation was also significantly higher in patients with lobular inflammation. Hepatocyte expression of TLR6 but not that of TLR2 was increased in NAFLD patients compared to normal liver histology. Deregulated expression and activity of peripheral TLR6 in morbidly obese patients can mirror the liver inflammatory events that are well known drivers of obesity-related NASH pathogenesis. Moreover, TLR6 is also significantly overexpressed in the hepatocytes of NAFLD patients compared to their normal counterparts. Thus, deregulated TLR6 expression may potentiate TLR2-mediated liver inflammation in NAFLD pathogenesis, and also serve as a potential peripheral biomarker of obesity-related NASH.

Absence of core autophagy gene expression in an ex vivo central nervous system model infected with Listeria monocytogenes

Estudios recientes han evidenciado que la autofagia puede actuar como un mecanismo inmune protector frente a la infección con Listeria monocytogenes. L. monocytogenes es una bacteria grampositiva, intracelular facultativa, que causa enfermedades invasivas en humanos y animales, especialmente en el sistema nervioso central (SNC). La listeriosis humana en el SNC puede manifestarse de diferentes maneras, incluyendo meningitis y abscesos cerebrales. La línea principal de defensa frente a las infecciones bacterianas es proporcionada por la microglía, fagocitos residentes del parénquima del SNC. Las células de microglía son conocidas, también, por eliminar las células dañadas o muertas tras un daño cerebral, y por lo tanto desempeñan un papel clave en las enfermedades infecciosas y neurodegenerativas. Se sabe poco sobre el papel de la autofagia en las interacciones entre el hospedador y el patógeno, debido a que la mayoría de los estudios in vitro han usado macrófagos o células epiteliales. En el presente trabajo hemos utilizado matrices de PCR en tiempo real para analizar la expresión de genes de autofagia en un modelo organotípico de cerebro de rata infectado con L. monocytogenes. Hemos observado que, en general, la expresión de genes centrales de la autofagia no está modulada por la infección, a pesar de la presencia de una intensa actividad fagocítica de la microglía en la superficie del tejido cerebral, observada mediante microscopia electrónica de barrido. Concluimos que, en nuestro modelo, la autofagia podría desempeñar un papel clave en la homeostasis del tejido dañado en lugar de tener un papel inmune relevante(AU)

Recent studies have suggested that autophagy can act as a protective immune mechanism against Listeria monocytogenes infection. L. monocytogenes is a Gram-positive, facultative intracellular bacterium that causes invasive diseases in humans and animals, particularly in the central nervous system (CNS). Human listeriosis of the CNS can manifest in many ways, including meningitis and brain abscesses. The initial line of defence against bacterial colonisation is provided by microglia, resident phagocytes of the CNS parenchyma. Microglial cells are also well known for clearing dead and dying neural cells after injury, and therefore play a key role in infectious diseases and neurodegeneration. Little is known about the role of the autophagy pathway in host–pathogen interactions in the brain as most in vitro studies have used macrophages or epithelial cells to study this interaction. In the present work, a quantitative real time-PCR array analysis was performed to assess autophagy-related gene expression in a brain rat ex vivo organotypic nervous system model during L. monocytogenes infection. We found that, in brief, core autophagy gene expression is not modulated by the infection, despite the presence of intense microglial phagocytic activity on the brain tissue surface that can be seen by scanning electron microscopy. We conclude that, in our model, autophagy could play a role in homeostasis in the damaged brain tissue instead of an immune-relevant pathway (AU)

Relevancia de TLR4 en la infección por virus de la hepatitis C / Relevance of TLR4 in HCV infection

Objetivo: Se evaluó la frecuencia de los polimorfismos Asp-299Gly y Thr-399Ile en el gen deTLR4 en sujetos infectados por VHC y coinfectados por VIH y VHC y se estudió la expresión y función de TLR4 en función de esos polimorfismos. Material y métodos: El estudio incluyó 53 pacientes con infección por VHC, de los que 28 eran coinfectados además por VIH, más 30 sujetos controles sanos. Los polimorfismos Asp-299Glyy Thr-399Ile se determinaron mediante PCR-RFLP, las poblaciones celulares y la expresión deTLR4 se estudiaron mediante citometría de flujo, mientras que las citocinas proinflamatoriasse midieron en suero mediante CBA. Resultados: Se encontró un descenso significativo de células T CD4+ en los pacientes coinfectados por VHC y VIH. No se demostró una mayor susceptibilidad a sufrir la infección para ninguno de los 2 polimorfismos analizados. TLR4 estaba disminuido en células B, mientras que aumentaba en células T y monocitos. Se comprobó un aumento significativo de las citocinas proinflamatorias que fue el doble en coinfectados que en los portadores de la infección aislada por VHC. Estos cambios no mostraron relación con el polimorfismo estudiado. Discusión: La expresión de TLR4 en células T y monocitos se encuentra aumentada en la infección por VIH y se acompaña de un aumento en suero de citocinas proinflamatorias. Estos hallazgos no guardan relación con los polimorfismos Asp-299Gly y Thr-399Ile de TLR4 (AU)

Objective: The presence of the Asp-299Gly and Thr-399Ile polymorphisms in the toll-like receptor 4 (TLR4) gene was studied in subjects with HCV infection and HCV+HIV coinfection. The expression and function of TLR4 as regards these polymorphisms is assessed. Material and methods: The study included 53 patients infected with HCV, among whom 27had coinfection HCV+HIV, and 30 healthy subjects. The polymorphisms were studied by PCR-RFLP. The number of lymphocyte subsets, as well as TLR4 expression, was determined by flow cytometry, and the concentration of cytokines was measured in serum by (cytometricbead assay) CBA. Results: CD4+T cells were significantly decreased in patients coinfected with HCV+HIV. There was no association between the presence of any of the two studied polymorphisms and the susceptibility to suffer from infection. TLR4 was less expressed in B cells, whereas it was increased in T cells and, in particular, monocytes. A significant increase in the levels of circulating pro-inflammatory cytokines was found, being two-fold increased incoinfected subjects as compared with patients with isolated HCV infection. None of the findings in cell subsets, TLR4 expression and cytokines was associated with the studied TLR4polymorphism.Discussion: The expression of TLR4 in T cells and monocytes is increased in HCV infection and is accompanied by increased serum levels of proinflammatory cytokines. These findings do not have any relationship with the Asp-299Gly and Thr-399Ile polymorphism (AU)