Neuronal regulation of diverse physiological functions requires complex molecular interactions in innervated tissues to maintain proper organ function. Here we show that loss of the neuronal cell surface adhesion/recognition molecule Contactin-1 (Cntn1) directly impairs intestinal function causing wasting that subsequently results in global immune defects. Loss of Cntn1 results in hematologic alterations and changes in blood metabolites associated with malnourishment. We found thymus and spleen of Cntn1-deficient animals atrophied with severe reductions in lymphocyte populations. Elevated thymic Gilz expression indicated ongoing glucocorticoid signaling in Cntn1-deficient animals, consistent with the malnourishment phenotype. Intestinal Contactin-1 was localized to neurons in the villi and the submucosal/myenteric plexus that innervates smooth muscle. Loss of Cntn1 was associated with reduced intestinal Bdnf and Adrb2, indicating reduced neuromuscular crosstalk. Additionally, loss of Cntn1 resulted in reduced recruitment of CD3+ T cells to villi within the small intestine. Together, these data illustrate the critical role of Contactin-1 function within the gut, and how this is required for normal systemic immune functions.

Although immunity induced by typhoid fever is moderated and short-lived, typhoid vaccination with the attenuated Ty21a oral vaccine generates long-lasting protection rates reaching up to 92%. Thus, there are important differences on how wild-type Salmonella and typhoid vaccine strains stimulate host immunity. We hypothesize that vaccine strains with different mutations might affect gut inflammation and intestinal permeability by different mechanisms. To test this hypothesis, we used an in vitro organotypic model of the human intestinal mucosa composed of human intestinal epithelial cells, lymphocytes/monocytes, endothelial cells, and fibroblasts. We also used six Salmonella enterica serovar Typhi (S. Typhi) strains: the licensed Ty21a oral vaccine, four typhoid vaccine candidates (i.e., CVD 908, CVD 909, CVD 910, and CVD 915) and the wild-type Ty2 strain. We found that genetically engineered S. Typhi vaccine strains elicit differential host changes not only in the intestinal permeability and secretion of inflammatory cytokines, but also in the phenotype and activation pathways of innate cells. These changes were distinct from those elicited by the parent wild-type S. Typhi and depended on the genetic manipulation. In sum, these results emphasize the importance of carefully selecting specific manipulations of the Salmonella genome in the development of typhoid vaccines.

One of the leading causes of infectious diarrhea in newborn calves is the apicomplexan protozoan Cryptosporidium parvum (C. parvum). However, little is known about its immunopathogenesis. Using next generation sequencing, this study investigated the immune transcriptional response to C. parvum infection in neonatal calves. Neonatal male Holstein-Friesian calves were either orally infected (N = 5) or not (CTRL group, N = 5) with C. parvum oocysts (gp60 subtype IIaA15G2R1) at day 1 of life and slaughtered on day 7 after infection. Total RNA was extracted from the jejunal mucosa for short read. Differentially expressed genes (DEGs) between infected and CTRL groups were assessed using DESeq2 at a false discovery rate < 0.05. Infection did not affect plasma immunohematological parameters, including neutrophil, lymphocyte, monocyte, leucocyte, thrombocyte, and erythrocyte counts as well as hematocrit and hemoglobin concentration on day 7 post infection. The immune-related DEGs were selected according to the UniProt immune system process database and were used for gene ontology (GO) and pathway enrichment analysis using Cytoscape (v3.9.1). Based on GO analysis, DEGs annotated to mucosal immunity, recognizing and presenting antigens, chemotaxis of neutrophils, eosinophils, natural killer cells, B and T cells mediated by signaling pathways including toll like receptors, interleukins, tumor necrosis factor, T cell receptor, and NF-KB were upregulated, while markers of macrophages chemotaxis and cytosolic pattern recognition were downregulated. This study provides a holistic snapshot of immune-related pathways induced by C. parvum in calves, including novel and detailed feedback and feedforward regulatory mechanisms establishing the crosstalk between innate and adaptive immune response in neonate calves, which could be utilized further to develop new therapeutic strategies.

The intestinal mucosa is enriched for unconventional T-cells, including mucosal associated invariant T-cells (MAIT), invariant natural killer T-cells (iNKT) and γδ T-cells. These cells are activated by bacterial metabolites, lipid antigens and cytokines, and are important for intestinal barrier integrity. The loss of gut homeostasis observed in HIV infection is central to disease pathogenesis, and studies have highlighted impairment of particular unconventional T-cell subsets within a specific gut compartment. However, although the small and large intestine are distinct niches, the overall impact of HIV on unconventional T-cells across the gut mucosal has not been well-studied. We hypothesized that compartment specific differences in the unconventional T-cell repertoire would exist between the small and large intestine, due to increasing bacterial loads and microbial diversity; and that the impact of HIV infection might differ depending on the compartment examined. We used mass cytometry, flow cytometry and unbiased T-cell receptor profiling to quantify unconventional T-cells in blood and tissue from the small (duodenum) and large (colon) intestine in HIV infected and uninfected participants undergoing examination for a range of intestinal conditions. Overall, we find distinct compartmentalisation of T-cells between blood, duodenum and colon, with iNKT cells significantly enriched in the duodenum and δ-1 expressing γδ T-cells in the colon. In addition, we observe greater clonal expansion of conventional TCRs in the duodenum, suggestive of stronger adaptive immunity in this compartment. Conversely, we find evidence of an expanded unconventional TCR repertoire in the colon, which contained far more overlapping "donor unrestricted" sequences than the duodenum. Twelve of these TCRs were highly "MAIT-like" and 3 were unique to the colon, suggesting an enrichment of donor unrestricted T-cells (DURTs) in this compartment. Unexpectedly, however, no significant impact of HIV infection on any of the unconventional T-cell subsets measured was observed in either mucosal site in terms of frequency or TCR repertoire. Further studies are required to investigate the importance of these unconventional T-cell subsets to intestinal homeostasis within the different gut compartments and determine if they are functionally impaired during HIV infection.

Background and Aims: Despite the negative results of blocking IL-17 in Crohn's disease (CD) patients, selective modulation of Th17-dependent responses warrants further study. Inhibition of retinoic acid-related orphan receptor gamma (RORγt), the master regulator of the Th17 signature, is currently being explored in inflammatory diseases. Our aim was to determine the effect of a novel oral RORγt antagonist (BI119) in human CD and on an experimental model of intestinal inflammation. Methods: 51 CD patients and 11 healthy subjects were included. The effects of BI119 were tested on microbial-stimulated peripheral blood mononuclear cells (PBMCs), intestinal crypts and biopsies from CD patients. The ability of BI119 to prevent colitis in vivo was assessed in the CD4+CD45RBhigh T cell transfer model. Results: In bacterial antigen-stimulated PBMCs from CD patients, BI119 inhibits Th17-related genes and proteins, while upregulating Treg and preserving Th1 and Th2 signatures. Intestinal crypts cultured with supernatants from BI119-treated commensal-specific CD4+ T cells showed decreased expression of CXCL1, CXCL8 and CCL20. BI119 significantly reduced IL17 and IL26 transcription in colonic and ileal CD biopsies and did not affect IL22. BI119 has a more profound effect in ileal CD with additional significant downregulation of IL23R, CSF2, CXCL1, CXCL8, and S100A8, and upregulation of DEFA5. BI119 significantly prevented development of clinical, macroscopic and molecular markers of colitis in the T-cell transfer model. Conclusions: BI119 modulated CD-relevant Th17 signatures, including downregulation of IL23R while preserving mucosa-associated IL-22 responses, and abrogated experimental colitis. Our results provide support to the use of RORγt antagonists as a novel therapy to CD treatment.

Aryl hydrocarbon receptor (AhR), a transcription factor activated by a large number of natural and synthetic agents, modulates the activity of immune cells in the gut and represents an important link between the environment and immune-mediated pathologies. In this study, we investigated the role of AhR in celiac disease (CD), a gluten-driven enteropathy. AhR expression was evaluated in intestinal biopsies taken from patients with CD and controls by real-time polymerase chain reaction (PCR), immunohistochemistry and flow cytometry. AhR was also analyzed in ex vivo organ cultures of duodenal biopsies taken from inactive CD patients incubated in presence or absence of peptic-tryptic digest of gliadin. IFN-γ, TNF-α, granzyme B, and perforin expression was evaluated in anti-CD3/CD28-activated intestinal lamina propria mononuclear cells (LPMC) and intestinal intra-epithelial cells (IEL) of active CD patients cultured in the presence or absence of the AhR agonist 6-formylindolo(3, 2-b)carbazole (Ficz). Finally, the protective role of AhR was evaluated in a mouse model of poly I:C-driven small intestine damage. AhR RNA transcripts were reduced in active CD samples as compared to inactive CD and normal controls. Flow cytometry confirmed such results and showed a reduction of AhR in both IEL and LPMC of active CD patients. The addition of a peptic-tryptic digest of gliadin to ex vivo organ cultures of duodenal biopsies taken from inactive CD patients reduced AhR expression. Treatment of CD IEL and LPMC with Ficz reduced the levels of inflammatory cytokines, granzyme B and perforin. Mice injected with Ficz were protected against poly I:C-induced intestinal lesions. Our findings suggest that defective AhR-driven signals could contribute to amplify pathogenic responses in the gut of CD patients.

Background: Chlorogenic acid (CGA), a natural bioactive polyphenol, exerts anti-inflammatory, antioxidant, and antibacterial effects that support the maintenance of intestinal health. However, the influence of CGA on gut microbiota and their metabolites, as well as its potential effects and mechanism of action in inflammatory bowel disease, remain to be elucidated. Methods: First, an oral gavage was used to administer CGA to indomethacin-treated mice. Then, fecal microbiota transplantation was performed to explore the role of intestinal microbiota in indomethacin-induced inflammation. Results: CGA treatment protected against body weight loss, damage to intestinal morphology and integrity, inflammation, and alteration of microbiota composition in indomethacin-treated mice. Interestingly, CGA failed to inhibit inflammation or protect intestine integrity in mice treated with antibiotics. Notably, mice who had been colonized with intestinal microbiota from CGA-treated or CGA-and-indomethacin-treated mice, through the fecal microbiota transplantation program, were protected from indomethacin-induced inflammation, growth of Bacteroides, and the accumulation of Bacteroides-derived LPS, in congruence with those who had been treated with CGA. Conclusion: The results suggest that CGA may protect intestine integrity and alleviate inflammatory responses, primarily by inhibiting the growth of Bacteroides and the accumulation of Bacteroides-derived LPS, in indomethacin-induced colitis. This newly identified mechanism broadens our knowledge of how CGA exerts protective effects on intestinal inflammation and provides strategies for the prevention of gastrointestinal mucosal damage in patients treated with indomethacin.

Gut acute graft-versus-host disease (aGVHD) is a serious complication after allogeneic hematopoietic stem cell transplantation (allo-HSCT) and is associated with high mortality. Mucosa-associated invariant T (MAIT) cells are a group of innate-like T cells enriched in the intestine that can be activated by riboflavin metabolites from various microorganisms. However, little is known about the function or mechanism of action of MAIT cells in the occurrence of gut aGVHD in humans. In our study, multiparameter flow cytometry (FCM) was used to evaluate the number of MAIT cells and functional cytokines. 16S V34 region amplicon sequencing analysis was used to analyze the intestinal flora of transplant patients. In vitro stimulation and coculture assays were used to study the activation and function of MAIT cells. The number and distribution of MAIT cells in intestinal tissues were analyzed by immunofluorescence technology. Our study showed that the number and frequency of MAIT cells in infused grafts in gut aGVHD patients were lower than those in no-gut aGVHD patients. Recipients with a high number of MAITs in infused grafts had a higher abundance of intestinal flora in the early posttransplantation period (+14 days). At the onset of gut aGVHD, the number of MAIT cells decreased in peripheral blood, and the activation marker CD69, chemokine receptors CXCR3 and CXCR4, and transcription factors Rorγt and T-bet tended to increase. Furthermore, when gut aGVHD occurred, the proportion of MAIT17 was higher than that of MAIT1. The abundance of intestinal flora with non-riboflavin metabolic pathways tended to increase in gut aGVHD patients. MAIT cells secreted more granzyme B, tumor necrosis factor (TNF)-α, and interferon (IFN)-γ under the interleukin (IL)-12/IL-18 stimulation [non-T-cell receptor (TCR) signal] and secreted most of the IL-17 under the cluster of differentiation (CD)3/CD28 stimulation (TCR signal). MAIT cells inhibited the proliferation of CD4+ T cells in vitro. In conclusion, the lower number of MAIT cells in infused grafts was related to the higher incidence of gut aGVHD, and the number of MAIT cells in grafts may affect the composition of the intestinal flora of recipients early after transplantation. The flora of the riboflavin metabolism pathway activated MAIT cells and promoted the expression of intestinal protective factors to affect the occurrence of gut aGVHD in humans.

Innate lymphoid cells (ILCs) are tissue-resident lymphocytes that lack antigen-specific receptors and exhibit innate effector functions such as cytokine production that play an important role in immediate responses to pathogens especially at mucosal sites. Mouse and human ILC subsets have been extensively characterized in various tissues and in blood. In this study, we present the first characterization of ILCs and ILC subsets in rat gut and secondary lymphoid organs using flow cytometry and single cell RNA sequencing. Our results show that phenotype and function of rat ILC subsets are conserved as compared to human and mouse ILCs. However, and in contrast to human and mouse, our study unexpectedly revealed that ILC2 and not ILC3 was the dominant ILC subset in the rat intestinal lamina propria. ILC2 predominance in the gut was independent of rat strain, sex or housing facility. In contrast, ILC3 was the predominant ILC subset in mesenteric lymph nodes and Peyer patches. In conclusion, our study demonstrates that in spite of highly conserved phenotype and function between mice, rat and humans, the distribution of ILC subsets in the intestinal mucosa is dependent on the species likely in response to both genetic and environmental factors.

Small heterodimer partner interacting leucine zipper protein (SMILE) is an orphan nuclear receptor and a member of the bZIP family of proteins. We investigated the mechanism by which SMILE suppressed the development of inflammatory bowel disease (IBD) using a DSS-induced colitis mouse model and peripheral blood mononuclear cells (PBMCs) from patients with ulcerative colitis (UC). Metformin, an antidiabetic drug and an inducer of AMPK, upregulated the level of SMILE in human intestinal epithelial cells and the number of SMILE-expressing cells in colon tissues from DSS-induced colitis mice compared to control mice. Overexpression of SMILE using a DNA vector reduced the severity of DSS-induced colitis and colitis-associated intestinal fibrosis compared to mock vector. Furthermore, SMILE transgenic mice showed ameliorated DSS-induced colitis compared with wild-type mice. The mRNA levels of SMILE and Foxp3 were downregulated and SMILE expression was positively correlated with Foxp3 in PBMCs from patients with UC and an inflamed mucosa. Metformin increased the levels of SMILE, AMPK, and Foxp3 but decreased the number of interleukin (IL)-17-producing T cells among PBMCs from patients with UC. These data suggest that SMILE exerts a therapeutic effect on IBD by modulating IL-17 production.

The mechanism underlying the pathology of neuromyelitis optica spectrum disorders (NMOSD) remains unclear even though antibodies to the water channel protein aquaporin-4 (AQP4) on astrocytes play important roles. Our previous study showed that dysbiosis occurred in the fecal microbiota of NMOSD patients. In this study, we further investigated whether the intestinal barrier and mucosal flora balance are also interrupted in NMOSD patients.

The etiology and pathogenesis of inflammatory bowel disease (IBD), including ulcerative colitis (UC) and Crohn's disease (CD), are generally believed to be related to immune dysfunction and intestinal microbiota disorder. However, the exact mechanism is not yet fully understood. The pathological changes associated with dextran sodium sulfate (DSS)-induced colitis are similar to those in human UC. As a subgroup of the innate immune system, group 3 innate lymphoid cells (ILC3s) are widely distributed in the lamina propria of the intestinal mucosa, and their function can be regulated by a variety of molecules. Musashi2 (MSI2) is a type of evolutionarily conserved RNA-binding protein that maintains the function of various tissue stem cells and is essential for postintestinal epithelial regeneration. The effect of MSI2 deficiency in ILC3s on IBD has not been reported. Thus, mice with conditional MSI2 knockout in ILC3s were used to construct a DSS-induced colitis model and explore its effects on the pathogenesis of IBD and the species, quantity and function of the intestinal microbiota.

Members of the Regulator of G-protein signaling (Rgs) family regulate the extent and timing of G protein signaling by increasing the GTPase activity of Gα protein subunits. The Rgs family member Rgs1 is one of the most up-regulated genes in tissue-resident memory (TRM) T cells when compared to their circulating T cell counterparts. Functionally, Rgs1 preferentially deactivates Gαq, and Gαi protein subunits and can therefore also attenuate chemokine receptor-mediated immune cell trafficking. The impact of Rgs1 expression on tissue-resident T cell generation, their maintenance, and the immunosurveillance of barrier tissues, however, is only incompletely understood. Here we report that Rgs1 expression is readily induced in naïve OT-I T cells in vivo following intestinal infection with Listeria monocytogenes-OVA. In bone marrow chimeras, Rgs1 -/- and Rgs1 +/+ T cells were generally present in comparable frequencies in distinct T cell subsets of the intestinal mucosa, mesenteric lymph nodes, and spleen. After intestinal infection with Listeria monocytogenes-OVA, however, OT-I Rgs1 +/+ T cells outnumbered the co-transferred OT-I Rgs1- /- T cells in the small intestinal mucosa already early after infection. The underrepresentation of the OT-I Rgs1 -/- T cells persisted to become even more pronounced during the memory phase (d30 post-infection). Remarkably, upon intestinal reinfection, mice with intestinal OT-I Rgs1 +/+ TRM cells were able to prevent the systemic dissemination of the pathogen more efficiently than those with OT-I Rgs1 -/- TRM cells. While the underlying mechanisms are not fully elucidated yet, these data thus identify Rgs1 as a critical regulator for the generation and maintenance of tissue-resident CD8+ T cells as a prerequisite for efficient local immunosurveillance in barrier tissues in case of reinfections with potential pathogens.

Defensins have attracted considerable research interest worldwide because of their potential to serve as a substitute for antibiotics. In this study, we characterized a novel porcine β-defensin (pBD129) and explored its role in alleviating bacterial endotoxin-induced inflammation and intestinal epithelium atrophy. The pBD129 gene was cloned and expressed in Escherichia coli. A recombinant pBD129 protein was also purified. To explore its role in alleviating the endotoxin-induced inflammation, mice, with or without lipopolysaccharide (LPS) challenge were treated by pBD129 at different doses. The recombinant pBD129 showed significant antimicrobial activities against the E. coli and Streptococcus with a minimal inhibitory concentration (MICs) of 32 μg/mL. Hemolytic assays showed that the pBD129 had no detrimental impact on cell viabilities. Interestingly, we found that pBD129 attenuated LPS-induced inflammatory responses by decreasing serum concentrations of inflammatory cytokines, such as the IL-1β, IL-6, and TNF-α (P < 0.05). Moreover, pBD129 elevated the intestinal villus height (P < 0.05) and enhanced the expression and localization of the major tight junction-associated protein ZO-1 in LPS-challenged mice. Additionally, pDB129 at a high dose significantly decreased serum diamine oxidase (DAO) concentration (P < 0.05) and reduced intestinal epithelium cell apoptosis (P < 0.05) in LPS-challenged mice. Importantly, pBD129 elevated the expression level of Bcl-2-associated death promoter (Bcl-2), but down-regulated the expression levels of apoptosis-related genes such as the B-cell lymphoma-2-associated X protein (Bax), BH3-interacting domain death agonist (Bid), cysteinyl aspartate-specific proteinase-3 (Caspase-3), and caspase-9 in the intestinal mucosa (P < 0.05). These results suggested a novel function of the mammalian defensins, and the anti-bacterial and anti-inflammatory properties of pBD129 may allow it a potential substitute for conventionally used antibiotics or drugs.

The human intestine contains numerous mononuclear phagocytes (MNP), including subsets of conventional dendritic cells (cDC), macrophages (Mf) and monocytes, each playing their own unique role within the intestinal immune system and homeostasis. The ability to isolate and interrogate MNPs from fresh human tissue is crucial if we are to understand the role of these cells in homeostasis, disease settings and immunotherapies. However, liberating these cells from tissue is problematic as many of the key surface identification markers they express are susceptible to enzymatic cleavage and they are highly susceptible to cell death. In addition, the extraction process triggers immunological activation/maturation which alters their functional phenotype. Identifying the evolving, complex and highly heterogenous repertoire of MNPs by flow cytometry therefore requires careful selection of digestive enzyme blends that liberate viable cells and preserve recognition epitopes involving careful selection of antibody clones to enable analysis and sorting for functional assays. Here we describe a method for the anatomical separation of mucosa and submucosa as well as isolating lymphoid follicles from human jejunum, ileum and colon. We also describe in detail the optimised enzyme digestion methods needed to acquire functionally immature and biologically functional intestinal MNPs. A comprehensive list of screened antibody clones is also presented which allows for the development of high parameter flow cytometry panels to discriminate all currently identified human tissue MNP subsets including pDCs, cDC1, cDC2 (langerin+ and langerin-), newly described DC3, monocytes, Mf1, Mf2, Mf3 and Mf4. We also present a novel method to account for autofluorescent signal from tissue macrophages. Finally, we demonstrate that these methods can successfully be used to sort functional, immature intestinal DCs that can be used for functional assays such as cytokine production assays.

Intestinal immunity exists as a complex relationship among immune cells, epithelial cells, and microbiota. CCR6 and its ligand-CCL20 are highly expressed in intestinal mucosal tissues, such as Peyer's patches (PPs) and isolated lymphoid follicles (ILFs). In this study, we investigated the role of the CCR6-CCL20 axis in intestinal immunity under homeostatic conditions. CCR6 deficiency intrinsically affects germinal center reactions in PPs, leading to impairments in IgA class switching, IgA affinity, and IgA memory B cell production and positioning in PPs, suggesting an important role for CCR6 in T-cell-dependent IgA generation. CCR6 deficiency impairs the maturation of ILFs. In these follicles, group 3 innate lymphoid cells are important components and a major source of IL-22, which stimulates intestinal epithelial cells (IECs) to produce antimicrobial peptides (AMPs). We found that CCR6 deficiency reduces IL-22 production, likely due to diminished numbers of group 3 innate lymphoid cells within small-sized ILFs. The reduced IL-22 levels subsequently decrease the production of AMPs, suggesting a critical role for CCR6 in innate intestinal immunity. Finally, we found that CCR6 deficiency impairs the production of IgA and AMPs, leading to increased levels of Alcaligenes in PPs, and segmented filamentous bacteria in IECs. Thus, the CCR6-CCL20 axis plays a crucial role in maintaining intestinal symbiosis by limiting the overgrowth of mucosa-associated commensal bacteria.

The intestinal microbiota is a complex ecosystem implicated in host health and disease. Inflammatory bowel disease (IBD) is a multifactorial chronic disorder of the gastrointestinal mucosa. Even though the exact mechanisms are still unknown, the intestinal microbiota is crucial in IBD development. We previously showed that murine norovirus (MNV) induces colitis in the Il10-deficient (Il10-/-) mouse model of IBD in a microbiota-dependent manner. Thus, in this study we analyzed whether distinct minimal bacterial consortia influence the outcome of MNV-triggered colitis in Il10-/- mice. Gnotobiotic Il10-/- mice associated with Oligo-Mouse-Microbiota 12 (OMM12) or Altered Schaedler Flora (ASF) developed little to no inflammatory lesions in the colon and cecum. MNV infection exacerbated colitis severity only in ASF-colonized mice, but not in those associated with OMM12. Four weeks after MNV infection, inflammatory lesions in ASF-colonized Il10-/- mice were characterized by epithelial hyperplasia, infiltration of inflammatory cells, and increased barrier permeability. Co-colonization of ASF-colonized Il10-/- mice with segmented filamentous bacteria (SFB) abolished MNV-induced colitis, whereas histopathological scores in SFB-OMM12-co-colonized mice stayed unchanged. Moreover, SFB only colonized mice associated with ASF. The SFB-mediated protective effects in ASF-colonized mice involved enhanced activation of intestinal barrier defense mechanisms and mucosal immune responses in the chronic and acute phase of MNV infection. SFB colonization strengthened intestinal barrier function by increasing expression of tight junction proteins, antimicrobial peptides and mucus. Furthermore, SFB colonization enhanced the expression of pro-inflammatory cytokines such as Tnfα, Il1β, and Il12a, as well as the expression of the regulatory cytokine Tgfβ. Altogether, our results showed that MNV-triggered colitis depends on the microbial context.

Increasing evidence supports the ability of eugenol to maintain intestinal barrier integrity and anti-inflammatory in vitro and in vivo; however, whether eugenol alleviates virus-mediated intestinal barrier damage and inflammation remains a mystery. Transmissible gastroenteritis virus (TGEV), a coronavirus, is one of the main causative agents of diarrhea in piglets and significantly impacts the global swine industry. Here, we found that eugenol could alleviate TGEV-induced intestinal functional impairment and inflammatory responses in piglets. Our results indicated that eugenol improved feed efficiency in TGEV-infected piglets. Eugenol not only increased serum immunoglobulin concentration (IgG) but also significantly decreased serum inflammatory cytokine concentration (TNF-α) in TGEV-infected piglets. In addition, eugenol also significantly decreased the expression of NF-κB mRNA and the phosphorylation level of NF-κB P65 protein in the jejunum mucosa of TGEV-infected piglets. Eugenol increased villus height and the ratio of villus height to crypt depth in the jejunum and ileum, and decreased serum D-lactic acid levels. Importantly, eugenol increased tight junction protein (ZO-1) and mRNA expression levels of nutrient transporter-related genes (GluT-2 and CaT-1) in the jejunum mucosa of TGEV-infected piglets. Meanwhile, compared with TGEV-infected IPEC-J2 cells, treatment with eugenol reduced the cell cytopathic effect, attenuated the inflammatory response. Interestingly, eugenol did not increase the expression of ZO-1 and Occludin in IPEC-J2 cells. However, western blot and immunofluorescence results showed that eugenol restored TGEV-induced down-regulation of ZO-1 and Occludin, while BAY11-7082 (The NF-κB specific inhibitor) enhanced the regulatory ability of eugenol. Our findings demonstrated that eugenol attenuated TGEV-induced intestinal injury by increasing the expression of ZO-1 and Occludin, which may be related to the inhibition of NF-κB signaling pathway. Eugenol may offer some therapeutic opportunities for coronavirus-related diseases.

Typhoid fever, caused by S. Typhi, is responsible for approximately 200,000 deaths per year worldwide. Little information is available regarding epithelium-bacterial interactions in S. Typhi infection. We have evaluated in vitro the effects of wild-type S. Typhi, the licensed Ty21a typhoid vaccine and the leading strains CVD 908-htrA and CVD 909 vaccine candidates on intestinal barrier function and immune response. Caco2 monolayers infected with wild-type S. Typhi exhibited alterations in the organization of tight junctions, increased paracellular permeability, and a rapid decrease in Trans-Epithelial Electrical Resistance as early as 4 h post-exposure. S. Typhi triggered the secretion of interleukin (IL)-8 and IL-6. Caco2 cells infected with the attenuated strains exhibited a milder pro-inflammatory response with minimal disruption of the barrier integrity. We conclude that wild-type S. Typhi causes marked transient alterations of the intestinal mucosa that are more pronounced than those observed with Ty21a or new generation attenuated typhoid vaccine candidates.

Patients with Crohn disease (CD) and ulcerative colitis (UC) suffer from chronic relapsing intestinal inflammation. While many studies focused on adaptive immunity, less is known about the role of innate immune cells in these diseases. Innate lymphoid cells (ILCs) are recently identified cells with a high cytokine-producing capacity at mucosal barriers. The aim was to study the impact of biological treatment on ILC in CD and UC. Patients initiating anti-tumor necrosis factor (TNF), ustekinumab, or vedolizumab treatment were prospectively followed up and peripheral and intestinal ILCs were determined. In the inflamed gut tissue of patients with inflammatory bowel disease, we found an increase of ILC1 and in immature NKp44- ILC3, whereas there was a decrease of mature NKp44+ ILC3 when compared to healthy controls (HCs). Similar but less pronounced changes in ILC1 were observed in blood, whereas circulating NKp44- ILC3 were decreased. Fifteen percent of CD patients had NKp44+ ILC3 in blood and these cells were not detected in blood of HCs or UC patients. Therapy with three different biologicals (ustekinumab targeting the IL-12/23 cytokines, anti-TNF and vedolizumab) partly restored intestinal ILC subset equilibrium with a decrease of ILC1 (except for ustekinumab) and an increase of NKp44+ ILC3. Anti-TNF also mobilized more NKp44+ ILC3 in circulation. As ILC1 are proinflammatory cells and as NKp44+ ILC3 contribute to homeostasis of intestinal mucosa, the observed effects of biologicals on ILCs might contribute to their clinical efficacy.