IFN-gamma-dependent transcription of MHC class II IA is impaired in macrophages from aged mice

To determine the effect of aging on IFN-gamma-induced MHC class II antigen expression, we produced bone marrow¿derived macrophages in vitro. In these conditions, we analyzed the effect of aging on the genomic expression of macrophages without the influence of other cell types that may be affected by aging. Although macrophages from young and aged mice showed an identical degree of differentiation, after incubation with IFN-gamma, the expression at the cell surface of the IA complex and the levels of IAbeta protein and mRNA were lower in aged macrophages. Moreover, the transcription of the IAbeta gene was impaired in aged macrophages. The amount of transcription factors that bound to the W and X, but not to the Y, boxes of the IAbeta promoter gene was lower in aged macrophages. Similar levels of CIITA mRNA were found after IFN-gamma treatment of both young and aged macrophages. This shows that neither the initial cascade that starts after the interaction of IFN-gamma with the receptor nor the second signals involved in the expression of CIITA are impaired in aged macrophages. These data indicate that aging is associated with low levels of MHC class II gene induction by IFN-gamma because of impaired transcription.

Neutralizing antibodies in Multiple Sclerosis a model-based analysis of Interferon beta signaling pathway in macrophages

Multiple Sclerosis is the most common non-traumatic cause of neurologicaldisability in young people. There is no cure yet, and until recently, few long-termtherapies existed. Interferon beta (IFNβ) was the first treatment, and remains the mostcommonly prescribed. One of the most significant problems of IFNβ therapy is theproduction of drug specific antibodies. Up to 45% of patients develop neutralizingantibodies (NAbs) to IFNβ products. The neutralizing antibody binds to the biologicalagent preventing its interaction with its receptor, inhibiting the biological action of theprotein, which abrogates the clinical efficacy of IFNβ treatment. Interferon-betamediates its response by binding to its high affinity cell surface receptor and initiatingthe JAK/STAT signalling cascade. In this project we have analyzed the IFNβ signalingpathway in macrophages when neutralizing antibodies are present. The response tothis pathway after IFNβ stimulation shows a transient oscillatory rhythm of STAT1phosphorylation, which varies as NAbs concentration increases. To improve ourunderstanding of that behavior, we extended an existing mathematical model based onnonlinear ordinary differential equations of JAK/STAT pathway by including IFN-NAbassociation and IFN-activation receptor. Combining our theoretical model withexperimental data we could study the role of neutralizing antibodies on the molecularresponse and determine its lifetime after cytokine stimulation.

IFN-gamma-dependent transcription of MHC class II IA is impaired in macrophages from aged mice

To determine the effect of aging on IFN-gamma-induced MHC class II antigen expression, we produced bone marrow¿derived macrophages in vitro. In these conditions, we analyzed the effect of aging on the genomic expression of macrophages without the influence of other cell types that may be affected by aging. Although macrophages from young and aged mice showed an identical degree of differentiation, after incubation with IFN-gamma, the expression at the cell surface of the IA complex and the levels of IAbeta protein and mRNA were lower in aged macrophages. Moreover, the transcription of the IAbeta gene was impaired in aged macrophages. The amount of transcription factors that bound to the W and X, but not to the Y, boxes of the IAbeta promoter gene was lower in aged macrophages. Similar levels of CIITA mRNA were found after IFN-gamma treatment of both young and aged macrophages. This shows that neither the initial cascade that starts after the interaction of IFN-gamma with the receptor nor the second signals involved in the expression of CIITA are impaired in aged macrophages. These data indicate that aging is associated with low levels of MHC class II gene induction by IFN-gamma because of impaired transcription.

Enhanced fatty acid oxidation in adipocytes and macrophages reduces lipid-induced triglyceride accumulation and inflammation

Lipid overload in obesity and type 2 diabetes is associated with adipocyte dysfunction, inflammation, macrophage infiltration, and decreased fatty acid oxidation (FAO). Here, we report that the expression of carnitine palmitoyltransferase 1A (CPT1A), the rate-limiting enzyme in mitochondrial FAO, is higher in human adipose tissue macrophages than in adipocytes and that it is differentially expressed in visceral vs. subcutaneous adipose tissue in both an obese and a type 2 diabetes cohort. These observations led us to further investigate the potential role of CPT1A in adipocytes and macrophages. We expressed CPT1AM, a permanently active mutant form of CPT1A, in 3T3-L1 CARΔ1 adipocytes and RAW 264.7 macrophages through adenoviral infection. Enhanced FAO in palmitate-incubated adipocytes and macrophages reduced triglyceride content and inflammation, improved insulin sensitivity in adipocytes, and reduced endoplasmic reticulum stress and ROS damage in macrophages. We conclude that increasing FAO in adipocytes and macrophages improves palmitate-induced derangements. This indicates that enhancing FAO in metabolically relevant cells such as adipocytes and macrophages may be a promising strategy for the treatment of chronic inflammatory pathologies such as obesity and type 2 diabetes.

Enhanced fatty acid oxidation in adipocytes and macrophages reduces lipid-induced triglyceride accumulation and inflammation

Lipid overload in obesity and type 2 diabetes is associated with adipocyte dysfunction, inflammation, macrophage infiltration, and decreased fatty acid oxidation (FAO). Here, we report that the expression of carnitine palmitoyltransferase 1A (CPT1A), the rate-limiting enzyme in mitochondrial FAO, is higher in human adipose tissue macrophages than in adipocytes and that it is differentially expressed in visceral vs. subcutaneous adipose tissue in both an obese and a type 2 diabetes cohort. These observations led us to further investigate the potential role of CPT1A in adipocytes and macrophages. We expressed CPT1AM, a permanently active mutant form of CPT1A, in 3T3-L1 CARΔ1 adipocytes and RAW 264.7 macrophages through adenoviral infection. Enhanced FAO in palmitate-incubated adipocytes and macrophages reduced triglyceride content and inflammation, improved insulin sensitivity in adipocytes, and reduced endoplasmic reticulum stress and ROS damage in macrophages. We conclude that increasing FAO in adipocytes and macrophages improves palmitate-induced derangements. This indicates that enhancing FAO in metabolically relevant cells such as adipocytes and macrophages may be a promising strategy for the treatment of chronic inflammatory pathologies such as obesity and type 2 diabetes.

Papel de la Interleucina 6 (IL-6) en la regulación de la expresión de Osteopontina (OPN) y CD44 tras axotomía del nervio facial

Investigación producida a partir de una estancia en la University of Sidney, Australia, entre octubre del 2008 y enero del 2009. Se ha desarrollado el proyecto titulado "Papel de la interleucina 6 (IL6) en la regulación de la expresión de Osteopontina (OPN) y de CD44 tras axotomía del nervio facial". Tras efectuar una transección del nervio facial, se indujo una reactividad glial en el núcleo facial (NF) localizado en el tronco cerebral, utilizando ratones transgénicos que sobrexpresan IL6 bajo promotor GFAP (tg GFAP-IL6), es decir selectivamente en astrocitos. Se han utilizado técnicas histoquímicas e inmunohistoquímicas, así como también se ha completado el estudio utilizando análisis de RPA, western blotting y citometría de flujo para la identificación de poblaciones celulares. Los resultados obtenidos indican que la OPN se expresa constitutivamente en las neuronas del NF. Tras axotomía del nervio facial, la expresión de OPN y CD44 incrementa en los ratones WT, mientras que en los tg GFAP-IL6 disminuye significativamente, sugiriendo que la IL6 podría estar involucrada en la modulación de la expresión de ambas moléculas. Sin embargo, no se ha visto diferencias en otros receptores de OPN como la integrina Alpha-5. La ctometría de flujo corroboró algunos de los resultados histológicos sobre la reactividad microglial y permitió concluir que la proporción de microglía activada (CD11b+/CD45+mid) y macrófagos (CD11b+/CD45+high) que expresan CD44 incrementa en in los tg GFAP-IL6 versus WT donde la mayor parte de microglia activada mostraba un perfil CD11b+/CD45+low.