Sterile inflammation drives multiple programmed cell death pathways in the gut.

Intestinal epithelial cells have a rapid turnover, being rapidly renewed by newly differentiated enterocytes, balanced by massive and constant removal of damaged cells by programmed cell death (PCD). The main forms of PCD are apoptosis, pyroptosis, and necroptosis, with apoptosis being a noninflammatory process, whereas the others drive innate immune responses. Although apoptosis is thought to be the principal means of cell death in the healthy intestine, which mechanisms are responsible for PCD during inflammation are not fully understood. To address this question, we used an in vivo model of enteropathy in wild-type mice induced by a single intragastric administration of the p31-43 gliadin peptide, which is known to elicit transient MyD88, NLRP3, and caspase-1-dependent mucosal damage and inflammation in the small intestine. Here, we found increased numbers of TUNEL+ cells in the mucosa as early as 2 h after p31-43 administration. Western blot and immunofluorescence analysis showed the presence of caspase-3-mediated apoptosis in the epithelium and lamina propria. In addition, the presence of mature forms of caspase-1, IL-1ß, and gasdermin D showed activation of pyroptosis and inhibition of caspase-1 led to decreased enterocyte death in p31-43-treated mice. There was also up-regulation of RIPK3 in crypt epithelium, suggesting that necroptosis was also occurring. Taken together, these results indicate that the inflammatory response induced by p31-43 can drive multiple PCD pathways in the small intestine.

Innate lymphoid cells. New players in tissue homeostasis and inflammatory responses.

In recent times, our understanding of the role of the immune system in different physiopathological situations has increased markedly. A new set of cells, generically known as innate lymphoid cells (ILC), has been discovered in the lymphoid compartment. Five ILC subsets can be recognized according to phenotypic and functional similarities with different subpopulations of T lymphocytes. Unlike T and B lymphocytes, ILC do not express antigen receptors nor undergo selection and clonal expansion upon activation. Instead, they respond rapidly to cytokines and danger signals in infected or inflamed tissues, producing cytokines that direct the immune response toward a type suitable for controlling the initial insult. In addition, ILC establish a crosstalk with other cells of the microenvironment that contributes to the maintenance and restoration of tissue homeostasis. Although many evidences on ILC were obtained from animal models, solid data confirm their existence in humans and their role in various inflammatory disorders. In this article, we address new knowledge on ILC, particularly on their role in the homeostasis of the immune system and in various inflammatory pathologies, in order to present new actors regulating immunity and immunopathology and affecting human health.

En tiempos recientes, nuestra comprensión del rol del sistema inmune en diferentes situaciones fisiopatológicas ha aumentado notablemente. En el compartimiento linfoide se ha descubierto un conjunto de células denominadas células linfoides innatas o innate lymphoid cells (ILC). Las ILC incluyen cinco grupos, clasificados según su similitud fenotípica y funcional con diferentes subpoblaciones de linfocitos T. A diferencia de los linfocitos T y B, las ILC no expresan receptores de antígeno ni sufren selección y expansión clonal cuando se activan. En cambio, responden rápidamente frente a citoquinas y señales de peligro en tejidos infectados o inflamados produciendo citoquinas que dirigen la respuesta inmune hacia un tipo adecuado para controlar la noxa original. Además, las ILC establecen un diálogo cruzado con otras células del microambiente que contribuye al mantenimiento y la restauración de la homeostasis tisular. Si bien muchas evidencias acerca de las ILC fueron obtenidas en modelos animales, existen datos sólidos que confirman su existencia en seres humanos y su papel en diversos trastornos inflamatorios. En este artículo, abordamos los nuevos conocimientos acerca de las ILC, y su rol en la homeostasis del sistema inmune y en diversas patologías inflamatorias, con el fin de presentar nuevos actores que regulan la inmunidad y la inmunopatología, lo que repercute en la salud humana.

Modulation of Intestinal Immune and Barrier Functions by Vitamin A: Implications for Current Understanding of Malnutrition and Enteric Infections in Children.

The micronutrient vitamin A refers to a group of compounds with pleiotropic effects on human health. These molecules can modulate biological functions, including development, vision, and regulation of the intestinal barrier. The consequences of vitamin A deficiency and supplementation in children from developing countries have been explored for several years. These children live in an environment that is highly contaminated by enteropathogens, which can, in turn, influence vitamin A status. Vitamin A has been described to modulate gene expression, differentiation and function of diverse immune cells; however, the underlying mechanisms are not fully elucidated. This review aims to summarize the most updated advances on elucidating the vitamin A effects targeting intestinal immune and barrier functions, which may help in further understanding the burdens of malnutrition and enteric infections in children. Specifically, by covering both clinical and in vivo/in vitro data, we describe the effects of vitamin A related to gut immune tolerance/homeostasis, intestinal barrier integrity, and responses to enteropathogens in the context of the environmental enteric dysfunction. Some of the gaps in the literature that require further research are also highlighted.

Immunobiotics for the Bovine Host: Their Interaction with Intestinal Epithelial Cells and Their Effect on Antiviral Immunity.

The scientific community has reported several cases of microbes that exhibit elevated rates of antibiotic resistance in different regions of the planet. Due to this emergence of antimicrobial resistant microorganisms, the use of antibiotics as promoters of livestock animals' growth is being banned in most countries around the world. One of the challenges of agricultural immunology therefore is to find alternatives by modulating the immune system of animals in drug-independent safe food production systems. In this regard, in an effort to supplant antibiotics from bovine feeds, several alternatives were proposed including the use of immunomodulatory probiotics (immunobiotics). The purpose of this review is to provide an update of the status of the modulation of intestinal antiviral innate immunity of the bovine host by immunobiotics, and the beneficial impact of immunobiotics on viral infections, focused on intestinal epithelial cells (IECs). The results of our group, which demonstrate the capacity of immunobiotic strains to beneficially modulate Toll-like receptor 3-triggered immune responses in bovine IECs and improve the resistance to viral infections, are highlighted. This review provides comprehensive information on the innate immune response of bovine IECs against virus, which can be further investigated for the development of strategies aimed to improve defenses in the bovine host.

Long polar fimbriae participates in the induction of neutrophils transepithelial migration across intestinal cells infected with enterohemorrhagic E. coli O157:H7.

Enterohemorrhagic Escherichia coli (EHEC) strains are causative agents of diarrhea and hemorrhagic colitis, both diseases associated with intestinal inflammation and cell damage. Several studies have correlated EHEC virulence factors to high levels of intestinal pro-inflammatory cytokines and we have previously described that the Long polar fimbriae (Lpf) is involved in the secretion of interleukin-8 (IL-8) and up-regulation of genes belonging to the NF-κB pathway using non-polarized epithelial intestinal T84 cells. In the current study, we evaluated the two EHEC O157 Lpf fimbriae (Lpf1 and Lpf2) for their ability to induce intestinal secretion of IL-8 and the activation of IL8, CCL20, and ICAM1 genes on polarized T84 cells. We also determined the participation of Lpf1 and Lpf2 in transepithelial migration of polymorphonuclear neutrophils (PMNs). Polarized T84 cells infected with EHEC revealed that both, Lpf1 and Lpf2, were required for the secretion of IL-8 and the induction of IL8, CCL20, and ICAM1 genes. Both fimbriae also played a role in the migration of PMNs trough the intestinal cells monolayer. Overall, the present work further demonstrated that the fimbriae Lpf1 and Lpf2 are important bacterial virulence factors that might be involved in the inflammatory responses associated with EHEC infections.

Transcriptional changes in the expression of chemokines related to natural killer and T-regulatory cells in patients with deep infiltrative endometriosis.

OBJECTIVE: To evaluate the expression of chemokines that regulate natural killer (NK) and T-regulatory (T-reg) cell activity in eutopic and ectopic endometrial tissue samples from endometriosis patients. DESIGN: Case-control study (Canadian Task Force classification II-2). SETTING: Tertiary referral hospital. PATIENT(S): Sixty-four consecutive patients with and without endometriosis. INTERVENTION(S): After videolaparoscopy, patients were divided into three groups: bowel endometriosis (n = 22), retrocervical endometriosis (n = 10), and endometriosis-free women (n = 32). MAIN OUTCOME MEASURE(S): Gene expression of the chemokines that regulate NK (CXCL9, CXCL10, CXCL11, CXCL12, XCL1, and CX3CL1) and T-reg cell activity (CCL17 and CCL21) evaluated by real-time polymerase chain reaction. RESULT(S): Of the chemokines associated with NK cells, CX3CL1 and CXCL12 expression was statistically significantly greater in the foci of endometriosis compared with the eutopic endometrium in patients and controls. From the chemokines associated with T-reg cells, CCL17 expression was statistically significantly greater in the eutopic endometrium of the patients with rectosigmoid endometriosis compared with the foci of endometriosis or eutopic endometrium of the patients with retrocervical endometriosis or the disease-free women. CONCLUSION(S): Both T-reg and NK cells mediate inflammatory response and may play a fundamental role in endometriosis by causing an impaired clearing of endometrial cells. Establishing how CCL17, CXCL12, and CX3CL1 modulate this response is essential to understanding inflammatory responses in endometriosis.

The long polar fimbriae of STEC O157:H7 induce expression of pro-inflammatory markers by intestinal epithelial cells.

Infection with Shiga toxin-producing Escherichia coli (STEC) O157:H7 is characterized by acute inflammation of the colonic mucosa. STEC O157:H7 contains two non-identical loci encoding long polar fimbriae (Lpf), which play a role in the STEC colonization of the intestinal epithelial cells. However, no information is available regarding the involvement of Lpf in the STEC-induced host inflammatory response. Hence, in this study we assess the role of Lpf as an inducer of inflammation on intestinal epithelial cells. Secretion of pro-inflammatory cytokines in response to STEC wild type and lpf isogenic mutants was evaluated on intestinal T84 cells. Of the 27 cytokines assayed, IL-6, IL-8, IL-15, FGF, GM-CSF and IP-10 were significantly reduced, when compared to the wild-type strain, in the lpfA1 lpfA2 double mutant. Further, the host intracellular signaling pathways activated in response to Lpf were determined by using an array containing genes representative of 18 different signal transduction pathways. The analysis indicated that the NF-κB pathway is activated in response to Lpf-expressing STEC. Therefore, our study supports the role of Lpf as a STEC factor mediating intestinal inflammation.

Associations between atopic markers in asthma and intestinal helminth infections in Cuban schoolchildren.

Total serum IgE (tIgE), allergen-specific IgE (sIgE), and skin prick test (SPT) are commonly used markers for atopy and atopic disease. The association between these measures and their relationship to clinical symptoms differs in affluent and non-affluent countries. We investigated the role of intestinal helminth infections in observed variations in atopic markers and asthma, and possible diagnostic and epidemiological consequences. A cross-sectional study was conducted in Cuban schoolchildren (n = 1285; 4-14 yrs). Atopy was determined by SPT, sIgE, and tIgE; asthma by International Study of Asthma and Allergies in Childhood questionnaire; and intestinal helminth infections by stool examination. Percentages of tIgE, sIgE, and SPT positives were 88.9%, 25.5%, and 16.5%, respectively. Asthma was found in 20.8%, and helminth infections in 20.9% of the children. All three atopic markers were significantly associated with each other and with asthma. Median tIgE levels were higher in helminth-infected than in uninfected children, irrespective of their status of atopy/asthma. Discordant results between SPT and sIgE were observed in 22.6% of the children. Among SPT positives, 41% were sIgE negative. The proportion of SPT negatives among sIgE positives was 74% in helminth-infected and 58.4% in uninfected children (p < 0.05). Helminth infections affected tIgE levels, reconfirming the limited value of tIgE for diagnosis of atopy and asthma in tropical areas. Higher frequencies of sIgE than positive SPTs were observed, especially in helminth-infected children. This corresponds with current hypothesis on the role of helminths in atopy. However, the observed proportion of sIgE negatives among children with positive SPT suggests that other mechanisms may also be involved.

Enteric pathology and Salmonella-induced cell death in healthy and SIV-infected rhesus macaques.

The goal of this study was to morphologically characterize a ligated ileal loop model of Salmonella enterica serotype Typhimurium infection in rhesus macaques (Macaca mulatta) and to verify the occurrence of Salmonella-induced cell death in vivo. Eight adult healthy male rhesus macaques were used for ligated ileal loop surgery. Four macaques had been intravenously inoculated with simian immunodeficiency virus (SIV) mac251. Ileal ligated loops were inoculated with wild-type (WT) S. Typhimurium strain IR715 (ATCC14028 nal (r)), an isogenic noninvasive mutant strain (ATCC14028 nal (r) ΔsipAΔsopABDE2), or sterile Luria Bertani broth. Loops were surgically removed at 2, 5, and 8 hours post-inoculation (hpi). Intestinal samples were processed for histopathology, immunohistochemistry for detecting Salmonella, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL), and transmission electron microscopy. Combined histopathology scores were similar between SIV-infected and control macaques. As expected, the invasion-deficient mutant was less pathogenic than WT S. Typhimurium. Neutrophil infiltrate in the intestinal mucosa correlated with bacterial loads (r = 0.7148; P < .0001) and fluid accumulation (r = 0.6019; P < .0001) in the lumen of the intestinal loops. Immunolabeled WT S. Typhimurium was observed in the epithelium and lamina propria at the tip of the villi at 2 hpi, progressing toward deeper lamina propria at 5-8 hpi. Most TUNEL-positive cells localized to the lamina propria, and some had morphological features of macrophages. Ultrastructurally, bacteria were observed intracellularly in the lamina propria as well as within apoptotic bodies. This study provides morphological evidence of Salmonella-induced cell death in vivo in a relevant nonhuman primate model.

Immunomodulatory activity of Lactobacillus rhamnosus strains isolated from goat milk: impact on intestinal and respiratory infections.

The immune stimulation induced by Lactobacillus rhamnosus CRL1505 (Lr05) and L. rhamnosus CRL1506 (Lr06) on the resistance to infection with an intestinal pathogen (Salmonella typhimurium) and a respiratory pathogen (Streptococcus pneumoniae) was studied in swiss-albine mice experimental models. The cytokine profiles that induced the innate and specific immune response in both infectious processes were investigated. Both strains were able to improve resistance against the intestinal pathogen. Only Lr05 was able to induce a significant decrease in the number of S. pneumoniae in the lung, prevent its dissemination into the blood and induce a significant increase in Th1 (INF-gamma) and Th2 (IL-6, IL-4 and IL-10) cytokine levels in the bronchoalveolar lavages (BAL). The changes in the cytokines profiles in BAL were associated with an increase in the number and activity of phagocytic cells and with the increase in specific antibodies in serum and BAL, which would explain the increased resistance to the challenge. The administration of Lr06 did not induce significant effects at the respiratory mucosal level. The results described in the present paper showed that certain LAB strains can share certain functional properties, although some of them can perform a functional role better than others, so that it is important to perform careful studies on specific strains, according to their therapeutic use.

Diferença de patogenicidade entre Escherichia coli enteroinvasora e Shigella flexneri em modelo experimental de infecção intestinal / Diference in pathogenicity between enteroinvasive Escherichia coli and Shigella flexneri in an experimental model of intestinal infection

Neste trabalho, esclarecemos tópicos da patogenicidade de EIEC que sustentam a sua menor virulência quando comparada à S. flexneri, e mostramos a importância das células dendríticas (CD) nesse processo. Estudou-se o comportamento de EIEC e S. flexneri quando em contato com células Caco-2, avaliando-se uma cinética de expressão dos genes envolvidos na invasão e disseminação bacteriana. Em geral, todos os genes foram menos expressos em EIEC, fato corroborado pelo fenótipo de disseminação bacteriana, onde EIEC foi menos eficiente do que Shigella. Também foi avaliada a modulação da resposta inflamatória de células dendríticas intestinais murinas pela produção de citocinas, expressão de moléculas co-estimulatórias e apresentação de antígenos, após desafio das células com as bactérias. Os resultados sugerem que EIEC induz a uma resposta protetora ao hospedeiro, enquanto que Shigella estaria "driblando" o sistema imune, além de provavelmente super-estimular o sistema imune adaptativo, fato que poderia levar a um agravamento da doença. As ações integradas das células Caco-2, células dendríticas e estímulos bacterianos foram estudadas em cocultura celular. Observou-se que EIEC e suas proteínas secretadas induzem a migração das CDs ao compartimento apical da co-cultura; nada foi observado quando o desafio se deu com Shigella...

Intestinal involvement is not sufficient to explain hypertransaminasemia in celiac disease?

Chronic unexplained hypertransaminasemia is an isolated clinical manifestation of celiac disease (CD) and lacks of a clear physiopathological explanation. Since CD and tropical sprue (TS) have similar intestinal functional and histological pattern of injury and that an increased inflammatory response has been reported to occur in patients with irritable bowel syndrome (IBS), liver involvement might be expected to occur either in TS or IBS. However, according to author's prior observations, the frequency of hypertransaminasemia is significantly higher in CD than in TS and IBS-diarrhea predominant patients (IBS-D). Thus, based on current knowledge, intestinal mucosal damage, increased intestinal permeability and/or an active intestinal inflammatory response do not completely explain liver damage in CD. We hypothesize that other factors, unique to CD not present in TS or IBS-D, like gluten toxicity and the presence of tissular transglutaminase (tTG) an auto-antigen with pro-inflammatory and remodeling properties, act in addition to intestinal mucosal injury and account to hypertransaminasemia in CD. Further research focusing on the mechanisms of gluten and tTG hepatic toxicity, and/or the characterization of the expression, secretion and enteral-hepatic transport of certain pro-inflammatory cytokines is needed, to understand the possible links between intestinal and liver disorders seen in CD.

Cell tropism of Salmonella enterica.

Salmonella serotypes are able to actively cross the intestinal epithelium, mainly but not exclusively through M cells in the follicle-associated epithelium of Peyer's patches. Once reaching the basal side of the epithelium, Salmonella serotypes are internalized by macrophages, dendritic cells, and neutrophils but are not found in fibroblasts or other mesenchymal cells. The outcome of the interaction between Salmonella serotypes and dendritic cells or neutrophils is detrimental to the pathogen. In some host species Salmonella serotypes find a safe haven from humoral defenses and neutrophils within macrophages, and replication within this niche appears to be a prerequisite for the development of a systemic infection. In other host species, macrophages can control bacterial growth and the infection remains localized to the intestine and mesenteric lymph nodes. This review summarizes our knowledge on the cellular tropism of Salmonella serotypes and the bacterial and host factors relevant for these interactions.

Modulation of the immune system and the response against pathogens with bovine colostrum concentrates.

The growth, development and health conditions for children living under deprived conditions in developing countries are so adverse that immediate public health measures to reduce morbidity and improve nutrition are urgently needed. Preventing and shortening the course of diarrhoeal episodes, eliminating protozoal colonization, and balancing intestinal microflora would all contribute to these goals. The consumption by humans of part of the colostrum produced when a dairy animal gives birth is an established tradition in many traditional societies. Recent advances in food technology in industrial dairying allow for continuous availability of stabilized bovine colostrum concentrate, both natural and hyperimmunized against specific human pathogens. This is safe for the calves of the producers themselves, for laboratory animals, and generally for humans, with the caveat of the milk-allergic. Moreover, substantial amounts of orally ingested bovine colostrum concentrate survive their passage through the stomach to remain intact and active in the lower reaches of the bowel. Studies in animals, human volunteers and naturally infected humans have demonstrated a therapeutic efficacy of oral bovine colostrum with certain infections. Similarly, attempts to prevent gastrointestinal infections in animals, exposed volunteers and at-risk populations have met with limited success with specific pathogens. It is time to begin to assess the feasibility and potential effectiveness and efficiency of employing seasonal or chronic bovine colostrum feeding in populations of deprived infantile populations to reduce the rates of recurrent gastroenteritis and decrease immunostimulation to improve vitality and nutritional status in early life.

Cow's milk protein-sensitive enteropathy at school age.

OBJECTIVES: Cow's milk protein-sensitive enteropathy (CMSE) may persist in children to school age. We sought to define the morphologic and immunohistochemical features of persistent CMSE. STUDY DESIGN: We studied 15 children with a definite diagnosis of CMSE on the basis of a blind challenge, 12 children with suspected cases of CMSE, 11 children with celiac disease, and 12 control children. RESULTS: Typical findings in children with CMSE were endoscopically visible lymphonodular hyperplasia of the duodenal bulb and lymphoid follicles without villous atrophy in biopsy samples. The patients with definite CMSE showed significantly increased densities of intraepithelial T cells skewed clearly to gammadelta(+) cells, compared with the control patients but fewer than in the patients with celiac disease. The study children showed no aberrant upregulation of HLA-DR expression in the duodenal mucosa, and the prevalence of HLA DQ2 antigen among them was equal to that in the control children. CONCLUSIONS: Our observations corroborated the claim that CMSE at school age is an identifiable clinical entity. Immunohistochemical findings suggest the abrogation of antigen tolerance locally on the gastrointestinal mucosa. A careful clinical assessment that includes a long elimination-challenge test supported by typical endoscopic and histologic findings form the basis for diagnosis.