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Intelectins (intestinal lectins) are highly conserved across chordate evolution and have been implicated in various human diseases, including Crohn's disease (CD). The human genome encodes two intelectin genes, intelectin-1 (ITLN1) and intelectin-2 (ITLN2). Other than its high sequence similarity with ITLN1, little is known about ITLN2. To address this void in knowledge, we report that ITLN2 exhibits discrete, yet notable differences from ITLN1 in primary structure, including a unique amino terminus, as well as changes in amino acid residues associated with the glycan-binding activity of ITLN1. We identified that ITLN2 is a highly abundant Paneth cell-specific product, which localizes to secretory granules, and is expressed as a multimeric protein in the small intestine. In surgical specimens of ileal CD, ITLN2 mRNA levels were reduced approximately five-fold compared to control specimens. The ileal expression of ITLN2 was unaffected by previously reported disease-associated variants in ITLN2 and CD-associated variants in neighboring ITLN1 as well as NOD2 and ATG16L1. ITLN2 mRNA expression was undetectable in control colon tissue; however, in both ulcerative colitis (UC) and colonic CD, metaplastic Paneth cells were found to express ITLN2. Together, the data reported establish the groundwork for understanding ITLN2 function(s) in the intestine, including its possible role in CD.

Intelectins are carbohydrate-binding proteins implicated in innate immunity and highly conserved across chordate evolution, including both ascidians and humans. Human intelectin-1 (ITLN1) is highly abundant within the intestinal mucosa and binds microbial but not host glycans. Genome-wide association studies identified SNPs in ITLN1 that are linked to susceptibility for Crohn's disease. Moreover, ITLN1 has been implicated in the pathophysiology of obesity and associated metabolic disease. To gain insight on biological activities of human ITLN1 in vivo, we developed a C57BL/6 mouse model genetically targeting the gene encoding the functional mouse ortholog. In wild-type C57BL/6 mice, both mRNA and protein analysis showed high expression of Itln1 in the small intestine, but manifold lower levels in colon and other extraintestinal tissues. Whereas intestinal expression of human ITLN1 localizes to goblet cells, our data confirm that mouse Itln1 is expressed in Paneth cells. Compared to wild-type littermate controls, mice homozygous for the Itln1 hypomorphic trapping allele had reduced expression levels of Itln1 expression (~10,000-fold). The knockout mice exhibited increased susceptibility in an acute model of experimentally induced colitis with 2% w/v dextran sulfate sodium (DSS). In a model of chronic colitis using a lower dose of DSS (1.5% w/v), which enabled a detailed view of disease activity across a protracted period, no differences were observed in body weight, fecal texture, hemoccult scores, food/water intake, or colon length at necropsy, but there was a statistically significant genotype over time effect for the combined fecal scores of disease activity. In model of diet-induced obesity, using two western-style diets, which varied in amounts of sugar (as sucrose) and saturated fat (as lard), mice with Itln1 expression ablated showed no increased susceptibility, in terms of weight gain, food intake, plasma markers of obesity compared to wildtype littermates. W

The microbiome modulates host immunity and aids the maintenance of tolerance in the gut, where microbial and food-derived antigens are abundant. Yet modern dietary factors and the excessive use of antibiotics have contributed to the rising incidence of food allergies, inflammatory bowel disease and other non-communicable chronic diseases associated with the depletion of beneficial taxa, including butyrate-producing Clostridia. Here we show that intragastrically delivered neutral and negatively charged polymeric micelles releasing butyrate in different regions of the intestinal tract restore barrier-protective responses in mouse models of colitis and of peanut allergy. Treatment with the butyrate-releasing micelles increased the abundance of butyrate-producing taxa in Clostridium cluster XIVa, protected mice from an anaphylactic reaction to a peanut challenge and reduced disease severity in a T-cell-transfer model of colitis. By restoring microbial and mucosal homoeostasis, butyrate-releasing micelles may function as an antigen-agnostic approach for the treatment of allergic and inflammatory diseases.

The gut microbiota is implicated in the adverse developmental outcomes of postnatal iron supplementation. To generate hypotheses on how changes to the gut microbiota by iron adversely affect development, and to determine whether the form of iron influences microbiota outcomes, we characterized gut microbiome and metabolome changes in Sprague-Dawley rat pups given oral supplements of ferrous sulfate (FS), ferrous bis-glycinate chelate (FC), or vehicle control (CON) on postnatal day (PD) 2-14. Iron supplementation reduced microbiome alpha-diversity (p < 0.0001) and altered short-chain fatty acids (SCFAs) and trimethylamine (TMA) in a form-dependent manner. To investigate the long-term effects of iron provision in early life, an additional cohort was supplemented with FS, FC, or CON until PD 21 and then weaned onto standard chow. At ~8 weeks of age, young adult (YA) rats that received FS exhibited more diverse microbiomes compared to CON (p < 0.05), whereas FC microbiomes were less diverse (p < 0.05). Iron provision resulted in 10,000-fold reduced abundance of Lactobacilli in pre-weanling and YA animals provided iron in early life (p < 0.0001). Our results suggest that in pre-weanling rats, supplemental iron form can generate differential effects on the gut microbiota and microbial metabolism that persist into adulthood.

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Iron supplements are frequently provided to infants in high-income countries despite low incidence of iron deficiency. There is growing concern regarding adverse health and development outcomes of excess iron provision in early life. Excess iron may directly damage developing organs through the formation of reactive oxygen species, alter systemic inflammatory signaling, and/or dysregulate trace mineral metabolism. To better characterize the in vivo effects of excess iron on development, we utilized a pre-weanling rat pup model. Lewis rat litters were culled to eight pups (four males and four females) and randomly assigned to daily supplementation groups receiving either vehicle control (CON; 10% w/v sucrose solution) or ferrous sulfate (FS) iron at one of the following doses: 10, 30, or 90 mg iron/kg body weight-FS-10, FS-30, and FS-90, respectively-from postnatal day (PD) 2 through 9. FS-90 litters, but not FS-30 or FS-10, failed to thrive compared to CON litters and had smaller brains on PD 10. Among the groups, FS-90 liver iron levels were highest, as were white blood cell counts. Compared to CON, circulating MCP-1 and liver zinc were increased in FS-90 pups, whereas liver copper was decreased. Growth defects due to excess FS provision in pre-weanling rats may be related to liver injury, inflammation, and altered trace mineral metabolism.

Intelectins are a family of multimeric secreted proteins that bind microbe-specific glycans. Both genetic and functional studies have suggested that intelectins have an important role in innate immunity and are involved in the etiology of various human diseases, including inflammatory bowel disease. Experiments investigating the role of intelectins in human disease using mouse models are limited by the fact that there is not a clear one-to-one relationship between intelectin genes in humans and mice, and that the number of intelectin genes varies between different mouse strains. In this study we show by gene sequence and gene expression analysis that human intelectin-1 (ITLN1) has multiple orthologues in mice, including a functional homologue Itln1; however, human intelectin-2 has no such orthologue or homologue. We confirm that all sub-strains of the C57 mouse strain have a large deletion resulting in retention of only one intelectin gene, Itln1. The majority of laboratory strains have a full complement of six intelectin genes, except CAST, SPRET, SKIVE, MOLF and PANCEVO strains, which are derived from different mouse species/subspecies and encode different complements of intelectin genes. In wild mice, intelectin deletions are polymorphic in Mus musculus castaneus and Mus musculus domesticus. Further sequence analysis shows that Itln3 and Itln5 are polymorphic pseudogenes due to premature truncating mutations, and that mouse Itln1 has undergone recent adaptive evolution. Taken together, our study shows extensive diversity in intelectin genes in both laboratory and wild-mice, suggesting a pattern of birth-and-death evolution. In addition, our data provide a foundation for further experimental investigation of the role of intelectins in disease.

Intelectins are a family of multimeric secreted proteins that bind microbe-specific glycans. Both genetic and functional studies have suggested that intelectins have an important role in innate immunity and are involved in the etiology of various human diseases, including inflammatory bowel disease. Experiments investigating the role of intelectins in human disease using mouse models are limited by the fact that there is not a clear one-to-one relationship between intelectin genes in humans and mice, and that the number of intelectin genes varies between different mouse strains. In this study we show by gene sequence and gene expression analysis that human intelectin-1 (ITLN1) has multiple orthologues in mice, including a functional homologue Itln1; however, human intelectin-2 has no such orthologue or homologue. We confirm that all sub-strains of the C57 mouse strain have a large deletion resulting in retention of only one intelectin gene, Itln1. The majority of laboratory strains have a full complement of six intelectin genes, except CAST, SPRET, SKIVE, MOLF and PANCEVO strains, which are derived from different mouse species/subspecies and encode different complements of intelectin genes. In wild mice, intelectin deletions are polymorphic in Mus musculus castaneus and Mus musculus domesticus. Further sequence analysis shows that Itln3 and Itln5 are polymorphic pseudogenes due to premature truncating mutations, and that mouse Itln1 has undergone recent adaptive evolution. Taken together, our study shows extensive diversity in intelectin genes in both laboratory and wild-mice, suggesting a pattern of birth-and-death evolution. In addition, our data provide a foundation for further experimental investigation of the role of intelectins in disease.

The intestinal mucus layer, an important element of epithelial protection, is produced by goblet cells. Intestinal goblet cells are assumed to be a homogeneous cell type. In this study, however, we delineated their specific gene and protein expression profiles and identified several distinct goblet cell populations that form two differentiation trajectories. One distinct subtype, the intercrypt goblet cells (icGCs), located at the colonic luminal surface, produced mucus with properties that differed from the mucus secreted by crypt-residing goblet cells. Mice with defective icGCs had increased sensitivity to chemically induced colitis and manifested spontaneous colitis with age. Furthermore, alterations in mucus and reduced numbers of icGCs were observed in patients with both active and remissive ulcerative colitis, which highlights the importance of icGCs in maintaining functional protection of the epithelium.

Obesity and accompanying metabolic disease is negatively correlated with lung health yet the exact mechanisms by which obesity affects the lung are not well characterized. Since obesity is associated with lung diseases as chronic bronchitis and asthma, we designed a series of experiments to measure changes in lung metabolism in mice fed obesogenic diets. Mice were fed either control or high fat/sugar diet (45%kcal fat/17%kcal sucrose), or very high fat diet (60%kcal fat/7% sucrose) for 150 days. We performed untargeted metabolomics by GC-TOFMS and HILIC-QTOFMS and lipidomics by RPLC-QTOFMS to reveal global changes in lung metabolism resulting from obesity and diet composition. From a total of 447 detected metabolites, we found 91 metabolite and lipid species significantly altered in mouse lung tissues upon dietary treatments. Significantly altered metabolites included complex lipids, free fatty acids, energy metabolites, amino acids and adenosine and NAD pathway members. While some metabolites were altered in both obese groups compared to control, others were different between obesogenic diet groups. Furthermore, a comparison of changes between lung, kidney and liver tissues indicated few metabolic changes were shared across organs, suggesting the lung is an independent metabolic organ. These results indicate obesity and diet composition have direct mechanistic effects on composition of the lung metabolome, which may contribute to disease progression by lung-specific pathways.

Obesity and accompanying metabolic disease is negatively correlated with lung health yet the exact mechanisms by which obesity affects the lung are not well characterized. Since obesity is associated with lung diseases as chronic bronchitis and asthma, we designed a series of experiments to measure changes in lung metabolism in mice fed obesogenic diets. Mice were fed either control or high fat/sugar diet (45%kcal fat/17%kcal sucrose), or very high fat diet (60%kcal fat/7% sucrose) for 150 days. We performed untargeted metabolomics by GC-TOFMS and HILIC-QTOFMS and lipidomics by RPLC-QTOFMS to reveal global changes in lung metabolism resulting from obesity and diet composition. From a total of 447 detected metabolites, we found 91 metabolite and lipid species significantly altered in mouse lung tissues upon dietary treatments. Significantly altered metabolites included complex lipids, free fatty acids, energy metabolites, amino acids and adenosine and NAD pathway members. While some metabolites were altered in both obese groups compared to control, others were different between obesogenic diet groups. Furthermore, a comparison of changes between lung, kidney and liver tissues indicated few metabolic changes were shared across organs, suggesting the lung is an independent metabolic organ. These results indicate obesity and diet composition have direct mechanistic effects on composition of the lung metabolome, which may contribute to disease progression by lung-specific pathways.