IFN-γ stimulates Paneth cell secretion through necroptosis mTORC1 dependent.

Immune mediators affect multiple biological functions of intestinal epithelial cells (IECs) and, like Paneth and Paneth-like cells, play an important role in intestinal epithelial homeostasis. IFN-γ a prototypical proinflammatory cytokine disrupts intestinal epithelial homeostasis. However, the mechanism underlying the process remains unknown. In this study, using in vivo and in vitro models we demonstrate that IFN-γ is spontaneously secreted in the small intestine. Furthermore, we observed that this cytokine stimulates mitochondrial activity, ROS production, and Paneth and Paneth-like cell secretion. Paneth and Paneth-like secretion downstream of IFN-γ, as identified here, is mTORC1 and necroptosis-dependent. Thus, our findings revealed that the pleiotropic function of IFN-γ also includes the regulation of Paneth cell function in the homeostatic gut.

Effect of IL-17A on the immune response to pulmonary tuberculosis induced by high- and low-virulence strains of Mycobacterium bovis.

Tuberculosis (TB) is an infectious, chronic, and progressive disease occurring globally. Human TB is caused mainly by Mycobacterium tuberculosis (M. tuberculosis), while the main causative agent of bovine TB is Mycobacterium bovis (M. bovis). The latter is one of the most important cattle pathogens and is considered the main cause of zoonotic TB worldwide. The mechanisms responsible for tissue damage (necrosis) during post-primary TB remain elusive. Recently, IL-17A was reported to be important for protection against M. tuberculosis infection, but it is also related to the production of an intense inflammatory response associated with necrosis. We used two M. bovis isolates with different levels of virulence and high IL-17A production to study this important cytokine's contrasting functions in a BALB/c mouse model of pulmonary TB. In the first part of the study, the gene expression kinetics and cellular sources of IL-17A were determined by real time PCR and immunohistochemistry respectively. Non-infected lungs showed low production of IL-17A, particularly by the bronchial epithelium, while lungs infected with the low-virulence 534 strain showed high IL-17A expression on Day 3 post-infection, followed by a decrease in expression in the early stage of the infection and another increase during late infection, on Day 60, when very low bacillary burdens were found. In contrast, infection with the highly virulent strain 04-303 induced a peak of IL-17A expression on Day 14 of infection, 1 week before extensive pulmonary necrosis was seen, being lymphocytes and macrophages the most important sources. In the second part of the study, the contribution of IL-17A to immune protection and pulmonary necrosis was evaluated by suppressing IL-17A via the administration of specific blocking antibodies. Infection with M. bovis strain 534 and treatment with IL-17A neutralizing antibodies did not affect mouse survival but produced a significant increase in bacillary load and a non-significant decrease in inflammatory infiltrate and granuloma area. In contrast, mice infected with the highly virulent 04-303 strain and treated with IL-17A blocking antibodies showed a significant decrease in survival, an increase in bacillary loads on Day 24 post-infection, and significantly more and earlier necrosis. Our results suggest that high expression of IL-17A is more related to protection than necrosis in a mouse model of pulmonary TB induced by M. bovis strains.

Necrosis plays a role in the concentration of mycobacterial antigens in granulomas from Mycobacterium bovis naturally infected cattle.

The dynamics that develop between cells and molecules in the host against infection by Mycobacterium bovis, leads to the formation of granulomas mainly present in the lungs and regional lymph nodes in cattle. Cell death is one of the main features in granuloma organization, however, it has not been characterized in granulomatous lesions caused by M. bovis. In this study we aimed to identify the profiles of cell death in the granuloma stages and its relationship with the accumulation of bacteria. We identified necrosis, activated caspase-3, LC3B/p62 using immunohistochemistry and digital pathology analysis on 484 granulomatous lesions in mediastinal lymph nodes from 23 naturally infected cattle. Conclusions: greater amounts of mycobacterial antigens were identified in granulomas from calves compared with adult cattle. The highest percentage of necrosis and quantity of mycobacterial antigens were identified in granuloma stages (III/IV) from adults. The LC3B/p62 profile was heterogeneous in granulomas between adults and calves. Our data suggest that necrosis is associated with a higher amount of mycobacterial antigens in the late stages of granuloma and the development of autophagy appears to play an heterogeneous effector response against infection in adults and calves. These results represent one of the first approaches in the identification of cell death in the four stages of granulomas in bovine tuberculosis.

Hypoxia inducible transcription factor (HIF-1) regulates CXCL13 expression: Clinical implications in pediatric non-Hodgkin's lymphoma

Background: Lymphomas are B and/or T cell clonal neoplasms in various states of differentiation, characteristically compromising lymph nodes. They are constituted by B and T lymphocytes that reach the node by chemokine-mediated recruitment including CXCL13. Hypoxia-inducible transcription factor (HIF-1α) plays a role in cellular adaptation to oxygen concentration changes. It also regulates expression of chemokines such as CXCL12, CCL20, and CCL5 as well as some of their receptors such as CCR7 and CXCR4. Methods: We performed in silico analysis of the CXCL13 promoter, pharmacologic modulation of HIF-1α activity and, using reporter plasmids, site-directed mutation and DNA-protein interaction analysis we analyzed the relation between HIF-1α activity and CXCL13 expression. Moreover, we did tissue microarray and immunohistochemistry to see the expression of HIF-1α and CXCL13. Results: This study detected three possible HIF-1α binding sites suggesting that this chemokine may be regulated by the CXCL13 transcription factor. We showed that CXCL13 expression is directly dependent, whereby an increase in HIF-1α activity increases CXCL13 expression and decreased HIF-1α activity in turn decreases CXCL13 expression. We proved that HIF-1α transcriptionally regulates the expression of CXCL13 in a direct manner. We established that HIF-1α and CXCL13 are greatly overexpressed in the most aggressive pediatric lymphomas. Conclusions: For the first time, this study showed that HIF-1α directly regulates transcriptional CXCL13 and that both proteins are overexpressed in the most aggressive forms of pediatric lymphoma. This suggests that they may play a significant role in the pathogenesis of pediatric non-Hodgkin's lymphoma.

Evaluación de la expresión del factor de transcripción Yin-Yang-1 y su asociación con TGF-β en un modelo murino de inflamación alérgica pulmonar con diferentes grados de severidad / Evaluation of the expression of the transcription factor Yin-Yang-1: association with TGF-β in a lung allergic inflammation in a mouse model with different severity grades

Introducción. El asma alérgica es una de las enfermedades más prevalecientes en la edad pediátrica. Los mecanismos implicados en este padecimiento no han sido esclarecidos totalmente. Se sabe que el factor de crecimiento transformante-beta (TGF-β) juega un papel muy importante en la fisiopatología de esta enfermedad y que la activación del factor de trascripción Yin-Yang-1 (YY1) induce un aumento en la expresión de esta citocina. El factor YY1 también regula la expresión de otras citocinas involucradas en el asma tales como la IL-4 y la IL-10. El objetivo de este trabajo fue evaluarla asociación entre YY1 y TGF-β en un modelo murino de inflamación alérgica pulmonar. Métodos. Se trabajó con un modelo murino de inflamación alérgica pulmonar con diferentes grados de severidad empleando ovalbúmina como alérgeno. Posteriormente se obtuvo el tejido pulmonar, que fue incluido en parafina, se construyó un microarreglo del tejido en un equipo semiautomático y, mediante inmunohistoquímica, se evaluó la expresión de YY1 y de TGF-β La densidad de la expresión se midió de manera cuantitativa por métodos computarizados. Resultados. Se observó inflamación alérgica pulmonar diferencial acorde con el grado de severidad del modelo; se observó el mismo patrón con la producción de moco. La expresión de ambas proteínas se correlacionó de manera directa con el grado de severidad de la inflamación alérgica pulmonar. Conclusiones. Los resultados obtenidos corroboran el papel que juegan ambas proteínas en la fisiopatología de la inflamación alérgica pulmonar.

Background. Allergic asthma is one of the most prevalent childhood diseases. This disease is characterized by airway inflammation and remodelling. The mechanisms implicated in the pathogenesis of this disease remain unclear. Several studies have shown that TGF-β plays an important role in the pathogenesis of asthma. In addition, the polymorphism of the TGF-β promoter region results in the overexpression of TGF-β via regulation of the transcription factor Yin-Yang-1 (YY1). It is has recently been demonstrated that YY1 may be involved in the pathogenesis of asthma by the regulation of IL-4 and IL-10. The aim of this study was to evaluate the association between the YY1 and TGF-β expression levels in a murine model of lung allergic inflammation. Methods. In this study we used a lung allergic inflammatory murine model with different severity degrees. Tissue microarray technology and immunohistochemistry were used to evaluate YY1 and TGF-p expression. The density expression was measured by quantitative methods using specific software. Results. Expression of both proteins correlated with the degrees of severity of lung allergic inflammation. A similar result was observed with mucus production. Conclusions. These results corroborate the role of YY1 and TGF-p in the pathogenesis of this disease.