Intestinal Homing Research Articles

Stem Cell Therapy Using Bone Marrow-Derived Muse Cells Repairs Radiation-Induced Intestinal Injury Through Their Intestine-Homing via Sphingosine Monophosphate-Sphingosine Monophosphate Receptor 2 Interaction

PurposeThere is still no effective treatment for the gastrointestinal side effects of radiation therapy. Multilineage-differentiating stress enduring (Muse) cells are tissue stem cells that have the ability to spontaneously homing in on injured tissues and repairing them. Several clinical trials have shown that stem cell therapy using human bone marrow-derived Muse (hBM-Muse) cells is effective in treating various diseases, but it is not known whether they are effective in treating radiation-induced intestinal injury. In this study, we investigated whether hBM-Muse cells homing to the radiation-damaged intestine and promote its repair. Methods and MaterialshBM-Muse cells were injected into the tail vein of mice 2 hours after high-dose total body irradiation. Then, homing analysis, crypt assay, BrdU assay, TUNEL assay, immunostaining, and survival time measurements were performed. In addition, we analyzed the expression of sphingosine monophosphate (S1P), a Muse cell inducing factor, in the mouse small intestine after irradiation. Finally, we investigated whether administration of JTE-013, an S1P receptor 2 (S1PR2)-specific antagonist, inhibits hBM-Muse cells homing to the injured intestine. ResultsS1P expression increased in mouse intestine after irradiation, with hBM-Muse cells homing in on the injured intestine. Injection of hBM-Muse cells after radiation exposure significantly increased the number of crypts, proliferating cells in the crypts, and small intestinal component cells such as intestinal stem cells, inhibited radiation-induced apoptosis, and prolonged mouse survival. Treatment with JTE-013 significantly inhibited intestinal homing and therapeutic effects of hBM-Muse cells. These findings indicate that hBM-Muse cells homed in on the injured intestine through the S1P-S1PR2 interaction to exert therapeutic effects on the radiation-induced intestinal injury. ConclusionThis study indicates that hBM-Muse cells are effective in treating radiation-induced intestinal injury, suggesting that hBM-Muse cell-based stem cell therapy has the potential to overcome the gastrointestinal side effects that limit the indications for radiation therapy.

Exploring the therapeutic potential of porphyran extracted from Porphyra haitanensis in the attenuation of DSS-induced intestinal inflammation

Ulcerative colitis is a chronic, spontaneous inflammatory bowel disease that primarily affects the colon. This study aimed to explore how Porphyra haitanensis porphyran (PHP) modulates the immune response and the associated mechanisms that alleviate dextran sulphate sodium-induced colitis in mice. Histological assessments via H&E staining and AB-PAS staining revealed that PHP intervention partially restored the number of goblet cells and improved intestinal mucosal function. Immunohistochemical and Western blot analyses of claudin-1, occludin, and MUC-2 demonstrated that PHP could repair the intestinal barrier and reduce colon damage by upregulating the expression of these proteins. PHP intervention was associated with a decrease in pro-inflammatory cytokine expression and an increase in anti-inflammatory cytokine expression. Moreover, the expression of proteins involved in intestinal immune homing, such as CCR-9, CCL-25, MAdCAM-1, and α4β7, was significantly suppressed in response to PHP treatment. Conversely, PHP upregulates the expression of CD40 and TGF-β1, both of these can promote healing and reduce inflammation in the gut lining. This study demonstrates that PHP can ameliorate ulcerative colitis by enhancing the intestinal barrier and modulating immune responses. These findings offer valuable insights into the potential utility of P. haitanensis as a promising natural product for managing ulcerative colitis.

Polysaccharide from Atractylodes macrocephala Koidz Binding with Zinc Oxide Nanoparticles as a Novel Mucosal Immune Adjuvant for H9N2 Inactivated Vaccine.

H9N2 avian influenza poses a significant public health risk, necessitating effective vaccines for mass immunization. Oral inactivated vaccines offer advantages like the ease of administration, but their efficacy often requires enhancement through mucosal adjuvants. In a previous study, we established a novel complex of polysaccharide from Atractylodes macrocephala Koidz binding with zinc oxide nanoparticles (AMP-ZnONPs) and preliminarily demonstrated its immune-enhancing function. This work aimed to evaluate the efficacy of AMP-ZnONPs as adjuvants in an oral H9N2-inactivated vaccine and the vaccine's impact on intestinal mucosal immunity. In this study, mice were orally vaccinated on days 0 and 14 after adapting to the environment. AMP-ZnONPs significantly improved HI titers, the levels of specific IgG, IgG1 and IgG2a in serum and sIgA in intestinal lavage fluid; increased the number of B-1 and B-2 cells and dendritic cell populations; and enhanced the mRNA expression of intestinal homing factors and immune-related cytokines. Interestingly, AMP-ZnONPs were more likely to affect B-1 cells than B-2 cells. AMP-ZnONPs showed mucosal immune enhancement that was comparable to positive control (cholera toxin, CT), but not to the side effect of weight loss caused by CT. Compared to the whole-inactivated H9N2 virus (WIV) group, the WIV + AMP-ZnONP and WIV + CT groups exhibited opposite shifts in gut microbial abundance. AMP-ZnONPs serve as an effective and safe mucosal adjuvant for oral WIV, improving cellular, humoral and mucosal immunity and microbiota in the gastrointestinal tract, avoiding the related undesired effects of CT.

P108 Response to vedolizumab in ulcerative colitis is associated with reduced neutrophil extracellular traps formation and IL-1β production: insights from NEUTROVED study

Abstract Background Vedolizumab (VDZ) is a humanized mAb against integrin α4β7. VDZ inhibits gut inflammation by impeding intestinal T-cell homing, however, recent data suggest that effects of α4β7 blocking extend beyond T-cells to alterations in innate immunity. Neutrophils are key components of innate immune response and inflammation in ulcerative colitis (UC). Studies have provided evidence linking neutrophils with UC pathogenesis, through autophagy induction and production of IL-1β-enriched neutrophil extracellular traps (NETs). NEUTROVED aimed to investigate the effects of VDZ treatment on neutrophils that may be associated with clinical and endoscopic response. Methods Patients with moderate-to-severe UC who received VDZ in standard medical care were evaluated in two timepoints, 24 hours before treatment initiation (T1) and at week 14 of treatment (T2). Clinical/endoscopic scores and common laboratory indexes were evaluated. In parallel, RNA-sequencing in peripheral neutrophils was performed, and markers of NETs in plasma and intestinal biopsies were assessed. Results In total 5 patients completed the study until today (Table 1). In T2, 4 patients demonstrated complete remission, and one patient had mild disease. No adverse events were recorded. RNA-sequencing analysis of paired samples revealed 248 significantly downregulated and 109 upregulated genes in peripheral neutrophils after VDZ treatment. Bioinformatics analysis using the GeneCodis4 web-based tool revealed that downregulated DEGs clustered mainly in immune-related pathways, including neutrophil degranulation, Toll-like receptor cascades, signaling by interleukins, NETs formation, and HIF-1 signaling pathway, while upregulated DEGs were involved in RNA metabolism-related processes, and the ribosome biogenesis pathway. A comparative analysis between these differentially expressed genes (DEGs) and our previous RNA-sequencing data of peripheral neutrophils isolated from active UC patients (n=24) highlighted a set of 168 DEGs as restored targets after VDZ therapy. Of note, several unique genes implicated in neutrophil activation, including IL-1 signaling components, were found significantly downregulated after VDZ therapy. In line with the abovementioned transcriptome alterations, markers of NETs and IL1β-bearing NETs in plasma and colonic tissue were significantly reduced, recapitalizing clinical and endoscopic response to VDZ. Conclusion Response to VDZ treatment in UC is related to anti-inflammatory actions on neutrophils characterized by reduced NETosis and IL-1β production through NETs. Further mechanistic studies investigating the possible effects of VDZ on innate immunity and neutrophil-mediated responses are warranted.

Administration of intestinal mesenchymal stromal cells reduces colitis-associated cancer in C57BL/6J mice modulating the immune response and gut dysbiosis

BackgroundPatients with inflammatory bowel disease (IBD) have a higher risk of developing colitis-associated colorectal cancer (CAC) with poor prognosis. IBD etiology remains undefined but involves environmental factors, genetic predisposition, microbiota imbalance (dysbiosis) and mucosal immune defects. Mesenchymal stromal cell (MSC) injections have shown good efficacy in reducing intestinal inflammation in animal and human studies. However, their effect on tumor growth in CAC and their capacity to restore gut dysbiosis are not clear. MethodsThe outcome of systemic administrations of in vitro expanded human intestinal MSCs (iMSCs) on tumor growth in vivo was evaluated using the AOM/DSS model of CAC in C57BL/6J mice. Innate and adaptive immune responses in blood, mesenteric lymph nodes (MLNs) and colonic tissue were analyzed by flow cytometry. Intestinal microbiota composition was evaluated by 16S rRNA amplicon sequencing. ResultsiMSCs significantly inhibited colitis and intestinal tumor development, reducing IL-6 and COX-2 expression, and IL-6/STAT3 and PI3K/Akt signaling. iMSCs decreased colonic immune cell infiltration, and partly restored intestinal monocyte homing and differentiation. iMSC administration increased the numbers of Tregs and IFN-γ+CD8+ T cells in the MLNs while decreasing the IL-4+Th2 response. It also ameliorated intestinal dysbiosis in CAC mice, increasing diversity and Bacillota/Bacteroidota ratio, as well as Akkermansia abundance, while reducing Alistipes and Turicibacter, genera associated with inflammation. ConclusionAdministration of iMSCs protects against CAC, ameliorating colitis and partially reverting intestinal dysbiosis, supporting the use of MSCs for the treatment of IBD.

IRF4 is required for migration of CD4+ T cells to the intestine but not for Th2 and Th17 cell maintenance.

The transcription factor Interferon Regulatory Factor 4 (IRF4) is central in control of T cell activation and differentiation. Deficiency of IRF4 results in severe immune deficiency and affects maturation and function of most if not all T cell subsets. Here we use mouse infection models for Citrobacter rodentium and Strongyloides ratti to analyze the function of IRF4 in T helper (Th) 17 and Th2 cell responses, respectively. IRF4 deficient mice were impaired in the control of both pathogens, failed to mount Th17 and Th2 cell responses and showed impaired recruitment of T helper cells to the intestine, the infection site of both pathogens. Compromised intestinal migration was associated with reduced expression of the intestinal homing receptors α4β7 integrin, CCR9 and GPR15. Identification of IRF4 binding sites in the gene loci of these receptors suggests a direct control of their expression by IRF4. Competitive T cell transfer assays further demonstrated that loss of one functional Irf4 allele already affected intestinal accumulation and Th2 and Th17 cell generation, indicating that lower IRF4 levels are of disadvantage for Th2 and Th17 cell differentiation as well as their migration to the intestine. Conversion of peripheral CD4+ T cells from an Irf4 wildtype to an Irf4 heterozygous or from an Irf4 heterozygous to a homozygous mutant genotype after C. rodentium or S. ratti infection did not reduce their capacity to produce Th17 or Th2 cytokines and only partially affected their persistence in the intestine, revealing that IRF4 is not essential for maintenance of the Th2 and Th17 phenotype and for survival of these T helper cells in the intestine. In conclusion, we demonstrate that the expression levels of IRF4 determine Th2 and Th17 cell differentiation and their intestinal accumulation but that IRF4 expression is not crucial for Th2 and Th17 cell survival.

Vascular traffic control of neutrophil recruitment to the liver by microbiota-endothelium crosstalk

During bloodstream infections, neutrophils home to the liver as part of an intravascular immune response to eradicate blood-borne pathogens, but the mechanisms regulating this crucial response are unknown. Using invivo imaging of neutrophil trafficking in germ-free and gnotobiotic mice, we demonstrate that the intestinal microbiota guides neutrophil homing to the liver in response to infection mediated by the microbial metabolite D-lactate. Commensal-derived D-lactate augments neutrophil adhesion in the liver independent of granulopoiesis in bone marrow or neutrophil maturation and activation in blood. Instead, gut-to-liver D-lactate signaling primes liver endothelial cells to upregulate adhesion molecule expression in response to infection and promote neutrophil adherence. Targeted correction of microbiota D-lactate production in a model of antibiotic-induced dysbiosis restores neutrophil homing to the liver and reduces bacteremia in a model of Staphylococcus aureus infection. These findings reveal long-distance traffic control of neutrophil recruitment to the liver by microbiota-endothelium crosstalk.

Expression of Homing Receptors in IgM+IgD+CD27+ B Cells and Their Frequencies in Appendectomized and/or Tonsillectomized Individuals

ABSTRACT Background In humans, blood circulating IgM+IgD+CD27+ B cells are considered analogous to those described in the marginal zone of the spleen and are involved in important immunological processes. The homing receptors they express, and the organs involved in their development (for example, intestinal organs in rabbits) are only partially known. We recently reported that this population is heterogeneous and composed of at least two subsets: one expressing high levels of IgM – IgMhi B cells – and another low levels – IgMlo B cells. Objectives To evaluate the expression of homing receptors on IgD+CD27+ IgMhi and IgMlo B cells and quantify their frequencies in blood of control and appendectomized and/or tonsillectomized volunteers. Materials and Methods Using spectral flow cytometry, the simultaneous expression of 12 previously reported markers that differentiate IgMhi B cells and IgMlo B cells and of α4β7, CCR9, CD22 and CCR10 were evaluated in blood circulating B cells of control and appendectomized and/or tonsillectomized volunteers. Results The existence of phenotypically defined IgMlo and IgMhi B cell subsets was confirmed. They differentially expressed intestinal homing receptors, and the expression of α4β7 and CCR9 seems to determine new IgM subpopulations. IgMlo and IgMhi B cells were detected at lower frequencies in the appendectomized and/or tonsillectomized volunteers relative to controls. Conclusions Human blood circulating IgD+CD27+ IgMlo and IgMhi B cell subsets differentially express homing receptors, and it is necessary to investigate if mucosal organs are important in their development.

CD103+ Cells and Chemokine Receptor Expression in Breast Cancer.

Mucosal environments harbour lymphocytes, which express several adhesion molecules, including intestinal homing receptors and integrin αE/β7 (CD103). CD103 binds E-cadherin, an integrin receptor expressed in intestinal endothelial cells. Its expression not only enables homing or retention of T lymphocytes at these sites but is also associated with increased T lymphocyte activation. However, it is not yet clear how CD103 expression is related to the clinical staging of breast cancer, which is determined by factors such as the size of the tumor (T), the involvement of nearby lymph nodes (N), and presence of metastasis (M). We examined the prognostic significance of CD103 by FACS in 53 breast cancer patients and 46 healthy controls enrolled, and investigated its expression, which contributes to lymphocyte recruitment in tumor tissue. Patients with breast cancer showed increased frequencies of CD103+, CD4+CD103+, and CD8+CD103+ cells compared to controls. CD103 was expressed at a high level on the surfaces of tumor-infiltrating lymphocytes in patients with breast cancer. Its expression in peripheral blood was not correlated with clinical TNM stage. To determine the localisation of CD103+ cells in breast tissue, tissue sections of breast tumors were stained for CD103. In tissue sections of breast tumors stained for CD103, its expression in T lymphocytes was higher compared to normal breast tissue. In addition, CD103+ cells expressed higher levels of receptors for inflammatory chemokines, compared to CD103- cells. CD103+ cells in peripheral blood and tumor tissue might be an important source of tumor-infiltrating lymphocyte trafficking, homing, and retention in cancer patients.

The microbiome restrains melanoma bone growth by promoting intestinal NK and Th1 cell homing to bone.

Bone metastases are frequent complications of malignant melanoma leading to reduced quality of life and significant morbidity. Regulation of immune cells by the gut microbiome influences cancer progression, but the role of the microbiome in tumor growth in bone is unknown. Using intracardiac or intratibial injections of B16-F10 melanoma cells into mice, we showed that gut microbiome depletion by broad-spectrum antibiotics accelerated intraosseous tumor growth and osteolysis. Microbiome depletion blunted melanoma-induced expansion of intestinal NK cells and Th1 cells and their migration from the gut to tumor-bearing bones. Demonstrating the functional relevance of immune cell trafficking from the gut to the bone marrow (BM) in bone metastasis, blockade of S1P-mediated intestinal egress of NK and Th1 cells, or inhibition of their CXCR3/CXCL9-mediated influx into the BM, prevented the expansion of BM NK and Th1 cells and accelerated tumor growth and osteolysis. Using a mouse model, this study revealed mechanisms of microbiota-mediated gut-bone crosstalk that are relevant to the immunological restraint of melanoma metastasis and tumor growth in bone. Microbiome modifications induced by antibiotics might have negative clinical consequences in patients with melanoma.

Antibody secreting cells are critically dependent on integrin \u03b14\u03b27/MAdCAM-1 for intestinal recruitment and control of the microbiota during chronic colitis

T and B cells employ integrin α4β7 to migrate to intestine under homeostatic conditions. Whether those cells differentially rely on α4β7 for homing during inflammatory conditions has not been fully examined. This may have implications for our understanding of the mode of action of anti-integrin therapies in inflammatory bowel disease (IBD). Here, we examined the role of α4β7 integrin during chronic colitis using IL-10−/− mice, β7-deficient IL-10−/−, IgA-deficient IL-10−/− mice, and antibody blockade of MAdCAM-1. We found that α4β7 was predominantly expressed by B cells. β7 deficiency and MAdCAM-1 blockade specifically depleted antibody secreting cells (ASC) (not T cells) from the colonic LP, leading to a fecal pan-immunoglobulin deficit, severe colitis, and alterations of microbiota composition. Colitis was not due to defective regulation, as dendritic cells (DC), regulatory T cells, retinaldehyde dehydrogenase (RALDH) expression, activity, and regulatory T/B-cell cytokines were all comparable between the strains/treatment. Finally, an IgA deficit closely recapitulated the clinical phenotype and altered microbiota composition of β7-deficient IL-10−/− mice. Thus, a luminal IgA deficit contributes to accelerated colitis in the β7-deficient state. Given the critical/nonredundant dependence of IgA ASC on α4β7:MAdCAM-1 for intestinal homing, B cells may represent unappreciated targets of anti-integrin therapies.

Dissecting Common and Unique Effects of Anti-α4β7 and Anti-Tumor Necrosis Factor Treatment in Ulcerative Colitis.

Background and AimsVedolizumab is an anti-α4β7 antibody approved for the treatment of ulcerative colitis [UC]. Although it is assumed that vedolizumab blocks intestinal homing of lymphocytes, its effects on different intestinal cell populations are not fully stablished. In order to establish the unique mechanisms of action of vedolizumab in UC patients, we compared its effects to those induced by anti-tumour necrosis factor [TNF].MethodsPatients with active UC [endoscopic Mayo score >1] starting vedolizumab [n = 33] or anti-TNF [n = 45] and controls [n = 22] were included. Colon biopsies [at weeks 0, 14 and 46] and blood samples [at weeks 0, 2, 6, 14, 30 and 46] were used for cell phenotyping, transcriptional analysis [qPCR], and to measure receptor occupancy.ResultsVedolizumab, in contrast to anti-TNF, significantly reduced the proportion of α4β7+ cells within intestinal T subsets while preserving the percentage of α4β7+ plasma cells. The marked decrease in α4β7 did not change the percentage of colonic αEβ7+ cells [at 46 weeks]. Both vedolizumab and anti-TNF significantly downregulated inflammation-related genes in the colon of responders [Mayo score < 2]. Moreover, both treatments significantly decreased the percentage of intestinal, but not blood, total lymphocytes [T and plasma cells], as well as the proportion of α4β1+ cells within intestinal T lymphocytes.ConclusionsOur data show that while vedolizumab and anti-TNF block two unrelated targets, they induce remarkably similar effects. On the other hand, vedolizumab’s unique mechanism of action relies on blocking intestinal trafficking of α4β7 T cells, despite effectively binding to B and plasma cells that express α4β7.

Ahr-Foxp3-RORγt axis controls gut homing of CD4+ T cells by regulating GPR15.

The orphan chemoattractant receptor GPR15 is important for homing T lymphocytes to the large intestine, thereby maintaining intestinal immune homeostasis. However, the molecular mechanisms underlying the regulation of GPR15 expression remain elusive. Here, we show a central role of the aryl hydrocarbon receptor (Ahr) in promoting GPR15 expression in both mice and human, thus gut homing of T lymphocytes. Mechanistically, Ahr directly binds to open chromatin regions of the Gpr15 locus to enhance its expression. Ahr transcriptional activity in directing GPR15 expression was modulated by two transcription factors, Foxp3 and RORγt, both of which are expressed preferentially by gut regulatory T cells (Tregs) in vivo. Specifically, Foxp3 interacted with Ahr and enhanced Ahr DNA binding at the Gpr15 locus, thereby promoting GPR15 expression. In contrast, RORγt plays an inhibitory role, at least in part, by competing with Ahr binding to the Gpr15 locus. Our findings thus demonstrate a key role for Ahr in regulating Treg intestinal homing under the steady state and during inflammation and the importance of Ahr-RORγt-Foxp3 axis in regulating gut homing receptor GPR15 expression by lymphocytes.

T cell-intrinsic expression of HIC1 links retinoic acid to tissue residency

Abstract The interaction between micronutrients and the immune system is emerging as a key control point in immune cell function in homeostasis and disease. For example, the vitamin A metabolite all-trans retinoic acid (atRA) has established roles in immune cell migration and effector function. Vitamin A deficiency results in a significant reduction in intestinal immune cells, which is thought to be associated with a lack of intestinal homing receptor expression and reduced migration to the intestine. However, the precise molecular pathways controlling intestinal immune cell homeostasis remains unclear. Here, we show that the atRA-inducible BTB-POZ transcriptional repressor Hypermethylated in cancer 1 (HIC1, ZBTB29) controls intestinal T cell residency. Mice with a T cell-intrinsic deletion of HIC1 (Hic1ΔT mice) have reduced numbers of CD69+ CD103+ tissue resident memory T (TRM) cells in the intestine, demonstrating that the atRA/HIC1 axis is critical for establishing and maintaining TRM cells. Further, following influenza A virus infection, numbers of TRM cells in the lungs are significantly reduced in Hic1ΔT mice, as well as in mice treated with the pan-retinoic acid receptor inhibitor BMS493, or in mice with a T cell-specific deletion of RARa (RaraΔT mice). Consistent with this, activation of naive CD8+ T cells from mice or humans in the presence of IL-2, IL-15, TGFβ and atRA results in the generation of CD69+ CD103+ ‘TRM-like’ cells that share the transcriptional profile of bona fide TRM cells. Taken together, these results identify atRA as a critical regulator of T cell tissue residency, acting by inducing expression of HIC1 and promoting the tissue residency gene program.

Abnormal Peyer patch development and B-cell gut homing drive IgA deficiency in Kabuki syndrome

Kabuki syndrome (KS) is commonly caused by mutations in the histone-modifying enzyme lysine methyltransferase 2D (KMT2D). Immune dysfunction is frequently observed in individuals with KS, but the role of KMT2D in immune system function has not been identified. We sought to understand the mechanisms driving KS-associated immune deficiency (hypogammaglobulinemia [low IgA], splenomegaly, and diminished immunization responses). We performed a comprehensive evaluation of humoral immunity and secondary lymphoid tissues in an established KS (Kmt2d+/βGeo) mouse model and validated select findings in a patient with KS. Compared with wild-type littermates, Kmt2d+/βGeo mice demonstrated deficiencies in multiple B-cell lineages and reduced serum IgA and elevated IgM levels across multiple ages. The bone marrow, spleen, and intestine of Kmt2d+/βGeo mice contained diminished numbers of IgA-secreting cells, while elevated germinal center B cells were found in the mesenteric lymph node and Peyer patches. Kmt2d+/βGeo mice have decreased size and numbers of Peyer patches, a finding confirmed in human samples. We identified deficiency of Itgb7 RNA and protein expression, a gene encoding an adhesion protein that mediates intestinal homing, and we demonstrated KMT2D-dependent control of ITGB7 expression in a human cell line. Kmt2d haploinsufficiency has broad deleterious effects on B-cell differentiation, specifically hampering gut lymphocyte homing and IgA+ plasma cell differentiation. Intestinal lymphoid defects caused by ITGB7 deficiency have not previously been recognized in KS, and these results provide new mechanistic insights into the pathogenesis of KS-associated immune deficiency.

Donor mesenchymal stem cell kinetics after transamniotic stem cell therapy (TRASCET) in a rodent model of gastroschisis

PurposeWe sought to comprehensively scrutinize donor mesenchymal stem cell kinetics following transamniotic stem cell therapy (TRASCET) in experimental gastroschisis. MethodsA gastroschisis was surgically created in 102 rat fetuses at gestation day 18 (term = 22 days), immediately followed by volume-matched amniotic injections of either amniotic fluid mesenchymal stem cells (afMSCs) labeled with a luciferase reporter gene (n = 58), or luciferase protein alone (n = 44). Samples from multiple anatomical sites from survivors were screened for luciferase activity via microplate luminometry at term. Statistical analysis included Mann–Whitney U-test, Wald test, and kappa coefficient (p < 0.05). ResultsOverall survival was 42% (43/102), with no significant difference between the two groups (p = 0.82). When controlled by acellular luciferase, donor afMSCs were identified selectively in the placenta (p < 0.001) and bowel (p = 0.005), independently of the dams (respectively, p < 0.001 and p = 0.041). Bowel homing was documented exclusively in areas exposed to the amniotic cavity. There was no mutual correlation between placental and bowel homing (kappa = −0.02; p = 0.91). ConclusionsAmniotic mesenchymal stem cells home to specific sites after TRASCET in the setting of gastroschisis. Placental homing and intestinal homing are central yet seemingly independent constituents of cell trafficking, suggesting that both direct amniotic seeding and hematogenous routing take place. Level of evidenceN/A (animal and laboratory study).

The Compromised Mucosal Immune System of β7 Integrin-Deficient Mice Has Only Minor Effects on the Fecal Microbiota in Homeostasis.

The gastrointestinal tract is an ideal habitat for diverse bacterial species that reside in a homeostatic balance with local tissue and significantly contribute to host health. Negative shifts in gut microbiota profiles, also known as dysbiosis, may be implicated in the development of chronic disorders such as inflammatory bowel diseases (IBD). Adhesion molecule-dependent recruitment of immune cells to the gut is an important step in IBD pathogenesis. The adhesion molecule β7 integrin contributes to the development of the gut-associated lymphoid tissue (GALT), intestinal immune cell homing, and immune responses and is known to promote intestinal inflammation. Although many studies underlined the role of the gut microbiota in shaping the mucosal immune system, studies on the influence of the host immune system on the microbiota are rare, especially in homeostasis. We addressed this question via comparative 16S rRNA gene amplicon analysis of fecal microbial communities from wild-type and β7 integrin-deficient mice, the latter being characterized by a compromised GALT. Besides subtle changes in relative abundances of Muribaculaceae spp. and unknown members of the families Ruminococcaceae and Lachnospiraceae, there was altogether no major difference in microbiota profiles in β7 integrin-deficient mice vs. wild-type littermates. This indicates that, in conditions of homeostasis, there is only a minor influence of the host immune system on the fecal microbiota in our mouse model, stressing the potential importance of pathological factors for dysbiosis development.

Disrupting Myelin-Specific Th17 Cell Gut Homing Confers Protection in an Adoptive Transfer Experimental Autoimmune Encephalomyelitis

Multiple sclerosis (MS) is a common autoimmune disease of the CNS. Although an association between MS and inflammatory bowel diseases is observed, the link connecting intestinal immune responses and neuroinflammation remains unclear. Here we show that encephalitogenic Th17 cells infiltrate the colonic lamina propria before neurological symptom development in two murine MS models, active and adoptive transfer experimental autoimmune encephalomyelitis (EAE). Specifically targeting Th17 cell intestinal homing by blocking the α4β7-integrin and its ligand MAdCAM-1 pathway impairs Tcell migration to the large intestine and dampens EAE severity in the Th17 cell adoptive transfer model. Mechanistically, myelin-specific Th17 cells proliferate in the colon and affect gut microbiota composition. The beneficial effect of blocking the α4β7-integrin and its ligand MAdCAM-1 pathway on EAE is interdependent with gut microbiota. Those results show that disrupting myelin-specific Th17 cell trafficking to the large intestine harnesses neuroinflammation and suggests that the gut environment and microbiota catalyze the encephalitogenic properties of Th17 cells.

MAdCAM-1-Mediated Intestinal Lymphocyte Homing Is Critical for the Development of Active Experimental Autoimmune Encephalomyelitis.

Lymphocyte homing into the intestine is mediated by binding of leukocytes to mucosal addressin cell adhesion molecule 1 (MAdCAM-1), expressed on endothelial cells. Currently, the immune system of the gut is considered a major modulator not only of inflammatory bowel disease, but also of extra-intestinal autoimmune disorders, including multiple sclerosis (MS). Despite intense research in this field, the exact role of the intestine in the pathogenesis of (neuro-)inflammatory disease conditions remains to be clarified. This prompted us to investigate the role of MAdCAM-1 in immunological processes in the intestine during T cell-mediated autoimmunity of the central nervous system (CNS). Using the experimental autoimmune encephalomyelitis model of MS, we show that MAdCAM-1-deficient (MAdCAM-1-KO) mice are less susceptible to actively MOG35−55-induced disease. Protection from disease was accompanied by decreased numbers of immune cells in the lamina propria and Peyer's patches as well as reduced immune cell infiltration into the spinal cord. MOG35−55-recall responses were intact in other secondary lymphoid organs of MAdCAM-1-KO mice. The composition of specific bacterial groups within the microbiome did not differ between MAdCAM-1-KO mice and controls, while MAdCAM-1-deficiency severely impaired migration of MOG35−55-activated lymphocytes to the gut. Our data indicate a critical role of MAdCAM-1 in the development of CNS inflammation by regulating lymphocyte homing to the intestine, and may suggest a role for the intestinal tract in educating lymphocytes to become encephalitogenic.

Differential Effects of the Absence of Nkx2-3 and MAdCAM-1 on the Distribution of Intestinal Type 3 Innate Lymphoid Cells and Postnatal SILT Formation in Mice.

Seeding of leukocytes to developing lymphoid tissues in embryonic and early postnatal age and to the mucosa throughout adulthood depends on the interaction between endothelial MAdCAM-1 addressin and its cognate ligand α4β7 integrin. Nkx2-3 as a transcriptional regulator of MAdCAM-1 controls vascular patterning in visceral lymphoid tissues in mice, and has been identified as a susceptibility factor for inflammatory bowel diseases in humans, associated with lymphoid neogenesis in the inflamed intestines. The role of Nkx2-3 in the organogenesis of the solitary intestinal lymphoid tissues (SILTs) involving type 3 innate lymphoid cells (ILC3) is still unknown. Here we investigated the effect of Nkx2-3 on the postnatal distribution of intestinal ILC3s and the development of SILTs, comparing these to mice lacking MAdCAM-1, but preserving Nkx2-3. At 1 week of age small intestines (SI) contained significantly higher number of ILC3s relative to the colon, with a substantial reduction in MAdCAM-1−/− mice compared to C57BL/6 controls. One week later SI ILC3 number decreased in all genotypes, the number of colonic ILC3 of both Nkx2-3-deficient and Nkx2-3-heterozygous mice significantly increased. On the fourth postnatal week a further reduction of SI ILC3s was observed in both Nkx2-3-deficient and Nkx2-3-heterozygous mice, while in the colon the number of ILC3s showed a significant reduction in all genotypes. At 1 week of age only sporadic SILT components were present in all genotypes. By the second week mice deficient for either Nkx2-3 or MAdCAM-1 showed absence of SILT maturation compared to their relevant controls, lacking mature isolated lymphoid follicles (ILF). By the fourth week both Nkx2-3-deficient and Nkx2-3-heterozygous mice showed a similar distribution of ILFs relative to cryptopatches (CP), whereas in MAdCAM-1−/− mice CPs and immature ILFs were present, mature ILFs were scarce. Our data demonstrate that the complete absence of MAdCAM-1 partially impairs intestinal seeding of ILC3s and causes partial blockade of SILT maturation, without affecting peripheral lymph node development. In contrast, the inactivation of Nkx2-3 permits postnatal seeding, and its blocking effect on SILT maturation prevails at later stage, thus other adhesion molecules may compensate for the intestinal homing of ILC3s in the absence of MAdCAM-1.