Effect of supplemental milk replacer and liquid starter diet for 4 and 11 days postweaning on intestinal parameters of weaned piglets and growth to slaughter.

Reduced piglet feed intake immediately postweaning (pw) leads to disruption of small intestine structure and function and reduced growth. Our objective was to evaluate the effect of providing supplemental milk or liquid starter diet for either 4 or 11 days pw, on intestinal parameters of newly weaned piglets and growth to slaughter. At weaning (28 ± 0.6 days old), five hundred and eighty-seven piglets ((Large White × Landrace) × Duroc) were divided into 59 pen groups, each containing 9-10 same sex (entire male or female) piglets. The pen groups were blocked by sex and weaning weight and provided with ad-libitum access to one of five dietary treatments: (1) Dry pelleted starter diet (control; CON); (2) CON+liquid milk replacer for 4 days pw (M4); (3) CON+liquid milk replacer for 11 days pw (M11); (4) CON+liquid starter diet for 4 days pw (S4) and (5) CON+liquid starter diet for 11 days pw (S11). Pen groups were weighed at weaning, days 11, 20, 28, and 47 pw and at target sale weight. Feed disappearance (on a DM basis) was recorded on each weighing day. On day 7 pw, 10 piglets per treatment were euthanised to collect small intestine tissue samples for determination of villus height (VH), crypt depth and brush-border membrane enzyme activity. Data were analysed using SAS-version 9.4. Between days 0 and 11 pw, M11 increased average daily feed intake by 48% and average daily gain (ADG) by 57% compared to CON (P < 0.05), and increased ADG by 54% (P < 0.05) compared to S4. Piglets on M11 also had improved feed conversion efficiency compared with CON piglets between days 0 and 11 pw. Treatment did not affect growth performance after day 28 pw, or carcass parameters at slaughter. At day 7 pw, M11 piglets had 37% higher jejunal VH than CON piglets (P < 0.05) and S11 piglets had 28% higher ileal VH than S4 piglets (P < 0.05). M11 piglets had up to 150% higher ileal sucrase activity than M4, S4 and S11 piglets (P < 0.05) and 180% higher ileal maltase activity than S4 piglets (P < 0.05). In conclusion, M11 reduced the immediate negative effects of weaning, as it was associated with increased feed intake, growth, brush-border membrane enzyme activity and improved intestinal structure early pw. However, there were no carryover effects of any of the liquid supplements on growth or feed efficiency or carcass weight at slaughter.

Comparative anatomy, pre- and postnatal changes during the development and maturation of the small intestine: Life-stage influences on exposure.

The gastrointestinal (GI) system absorbs nutrients and xenobiotics, excretes waste, and performs immunologic and endocrine functions. The subdivisions of the mature gut and the complexity of their corrugated, absorptive luminal surfaces differ greatly among mammals. Regardless, the embryonic gut tube in all mammalian species arises when cephalocaudal folding incorporates the roof of the yolk sac into the embryo. The gut tube quickly lengthens and bulges into the umbilical cord. Upon reentry into the abdominal cavity, the gut tube begins to differentiate-a process that continues until well into the lactation period. Differentiation of the small intestine involves (1) increasing the absorptive surface area of the lumen; (2) establishing mechanisms to control the pH of luminal contents; (3) forming a hierarchical vascular system for distribution of absorbed nutrients; (4) developing a complex enteric nervous system to control motility; (5) providing a system for replenishment of cells; and (6) contributing to the immunity of the organism. Because the length of gestation varies among species typically used in safety tests and is much shorter than human gestation, the state of GI maturation at the time of parturition differs significantly. Differences in GI maturation can contribute to species differences in the rate and extent of absorption; these differences must be considered when designing and interpreting pharmacological/toxicological studies and extrapolating safety test results to humans.

Intra-amniotic administration of l-glutamine promotes intestinal maturation and enteroendocrine stimulation in chick embryos.

Initial nutritional stimulation is a key driving force for small intestinal maturation. In chick embryos, administration of l-glutamine (Gln) into the amniotic fluid stimulates early development of the small intestinal epithelium by promoting enterocyte differentiation. In this study, we evaluated the effects of intra-amniotic administration of Gln on enterocyte morphology and function, and elucidated a potential enteroendocrine pathway through which Gln stimulates small intestinal maturation. Our results show that Gln stimulation at embryonic day 17 significantly increased enterocyte and microvilli dimensions by 10 and 20%, respectively, within 48 h. Post-hatch, enterocytes and microvilli were 20% longer in Gln-treated chicks. Correspondingly, Gln stimulation significantly upregulated mRNA expression of brush border nutrient transporters PepT-1 and SGLT-1 and tight junction proteins TJP-1 and TJP-2, before and after hatch (P < 0.05). Since GLP-2 signaling from intestinal L-cells is associated with enterocyte growth, functionality and integrity, we examined the effects of Gln stimulation on mRNA expression of key hormones and receptors within this enteroendocrine pathway and found significant increases in GLP-2R, IGF-1 and IGF-1R expression before and after hatch (P < 0.05). In conclusion, our findings link primary nutrient stimulation in the developing small intestine with enterocyte morphological and functional maturation and enteroendocrine signaling.

Characterization of an in vitro 3D intestinal organoid model by using massive RNAseq-based transcriptome profiling.

Organoids culture provides unique opportunities to study human diseases and to complement animal models. Several organs and tissues can be in vitro cultured in 3D structures resembling in vivo tissue organization. Organoids culture contains most of the cell types of the original tissue and are maintained by growth factors mimicking the in vivo state. However, the system is yet not fully understood, and specific in vivo features especially those driven by cell-extrinsic factors may be lost in culture. Here we show a comprehensive transcriptome-wide characterization of mouse gut organoids derived from different intestinal compartments and from mice of different gender and age. RNA-seq analysis showed that the in vitro culture strongly influences the global transcriptome of the intestinal epithelial cells (~ 60% of the total variance). Several compartment-, age- and gender-related transcriptome features are lost after culturing indicating that they are driven by niche or systemic factors. However, certain intrinsic transcriptional programs, for example, some compartment-related features and a minority of gender- and aging- related features are maintained in vitro which suggested possibilities for these features to be studied in this system. Moreover, our study provides knowledge about the cell-extrinsic or cell-intrinsic origin of intestinal epithelial transcriptional programs. We anticipated that our characterization of this in vitro system is an important reference for scientists and clinicians using intestinal organoids as a research model.

Dietary Tryptophan Supplementation Alters Fat and Glucose Metabolism in a Low-Birthweight Piglet Model.

Low birthweight (LBW) is associated with metabolic complications, such as glucose and lipid metabolism disturbances in early life. The objective of this study was to assess: (1) the effect of dietary tryptophan (Trp) on glucose and fat metabolism in an LBW piglet model, and (2) the role peripheral 5-hydroxytryptamine type 3 (5HT3) receptors in regulating the feeding behavior in LBW piglets fed with Trp-supplemented diets. Seven-day-old piglets were assigned to 4 treatments: normal birthweight-0%Trp (NBW-T0), LBW-0%Trp (LBW-T0), LBW-0.4%Trp (LBW-T0.4), and LBW-0.8%Trp (LBW-T0.8) for 3 weeks. Compared to LBW-T0, the blood glucose was decreased in LBW-T0.8 at 60 min following the meal test, and the triglycerides were lower in LBW-T0.4 and LBW-T0.8. Relative to LBW-T0, LBW-T0.8 had a lower transcript and protein abundance of hepatic glucose transporter-2, a higher mRNA abundance of glucokinase, and a lower transcript of phosphoenolpyruvate carboxykinase. LBW-T0.4 tended to have a lower protein abundance of sodium-glucose co-transporter 1 in the jejunum. In comparison with LBW-T0, LBW-T0.4 and LBW-T0.8 had a lower transcript of hepatic acetyl-CoA carboxylase, and LBW-T0.4 had a higher transcript of 3-hydroxyacyl-CoA dehydrogenase. Blocking 5-HT3 receptors with ondansetron reduced the feed intake in all groups, with a transient effect on LBW-T0, but more persistent effect on LBW-T0.8 and NBW-T0. In conclusion, Trp supplementation reduced the hepatic lipogenesis and gluconeogenesis, but increased the glycolysis in LBW piglets. Peripheral serotonin is likely involved in the regulation of feeding behavior, particularly in LBW piglets fed diets supplemented with a higher dose of Trp.

Acceleration of Small Intestine Development and Remodeling of the Microbiome Following Hyaluronan 35 kDa Treatment in Neonatal Mice.

The beneficial effects of human milk suppressing the development of intestinal pathologies such as necrotizing enterocolitis in preterm infants are widely known. Human milk (HM) is rich in a multitude of bioactive factors that play major roles in promoting postnatal maturation, differentiation, and the development of the microbiome. Previous studies showed that HM is rich in hyaluronan (HA) especially in colostrum and early milk. This study aims to determine the role of HA 35 KDa, a HM HA mimic, on intestinal proliferation, differentiation, and the development of the intestinal microbiome. We show that oral HA 35 KDa supplementation for 7 days in mouse pups leads to increased villus length and crypt depth, and increased goblet and Paneth cells, compared to controls. We also show that HA 35 KDa leads to an increased predominance of Clostridiales Ruminococcaceae, Lactobacillales Lactobacillaceae, and Clostridiales Lachnospiraceae. In seeking the mechanisms involved in the changes, bulk RNA seq was performed on samples from the terminal ileum and identified upregulation in several genes essential for cellular growth, proliferation, and survival. Taken together, this study shows that HA 35 KDa supplemented to mouse pups promotes intestinal epithelial cell proliferation, as well as the development of Paneth cells and goblet cell subsets. HA 35 KDa also impacted the intestinal microbiota; the implications of these responses need to be determined.

How to Obtain a Mega-Intestine with Normal Morphology: In Silico Modelling of Postnatal Intestinal Growth in a Cd97-Transgenic Mouse.

Intestinal cylindrical growth peaks in mice a few weeks after birth, simultaneously with crypt fission activity. It nearly stops after weaning and cannot be reactivated later. Transgenic mice expressing Cd97/Adgre5 in the intestinal epithelium develop a mega-intestine with normal microscopic morphology in adult mice. Here, we demonstrate premature intestinal differentiation in Cd97/Adgre5 transgenic mice at both the cellular and molecular levels until postnatal day 14. Subsequently, the growth of the intestinal epithelium becomes activated and its maturation suppressed. These changes are paralleled by postnatal regulation of growth factors and by an increased expression of secretory cell markers, suggesting growth activation of non-epithelial tissue layers as the origin of enforced tissue growth. To understand postnatal intestinal growth mechanistically, we study epithelial fate decisions during this period with the use of a 3D individual cell-based computer model. In the model, the expansion of the intestinal stem cell (SC) population, a prerequisite for crypt fission, is largely independent of the tissue growth rate and is therefore not spontaneously adaptive. Accordingly, the model suggests that, besides the growth activation of non-epithelial tissue layers, the formation of a mega-intestine requires a released growth control in the epithelium, enabling accelerated SC expansion. The similar intestinal morphology in Cd97/Adgre5 transgenic and wild type mice indicates a synchronization of tissue growth and SC expansion, likely by a crypt density-controlled contact inhibition of growth of intestinal SC proliferation. The formation of a mega-intestine with normal microscopic morphology turns out to originate in changes of autonomous and conditional specification of the intestinal cell fate induced by the activation of Cd97/Adgre5.

Low birth weight female piglets show altered intestinal development, gene expression, and epigenetic changes at key developmental loci.

Intestinal development is compromised in low birth weight (LBW) pigs, negatively impacting their growth, health, and resilience. We investigated the molecular mechanisms of the altered intestinal maturation observed in neonatal and juvenile LBW female piglets by comparing the changes in intestinal morphology, gene expression, and methylation in LBW versus normal birth weight (NBW) female piglets. A total of 16 LBW/NBW sibling pairs were sacrificed at 0 hours, 8 hours, 10 days, and 8 weeks of age. The gastrointestinal tract was weighed, measured, and the small intestine was sampled for histomorphology, gene expression, and methylation analyses. Impaired intestinal development, with shorter villi and shallower crypts, was observed in LBW female piglets. The expression of intestinal development markers (ALPI and OLFM) rapidly peaked after birth in NBW but not in LBW female piglets. The lower expression of genes involved in nutrient digestion (ANPEP and SI) and barrier function (OCLN and CLDN4) in LBW, together with their delayed development of intestinal villi and crypts could help to explain the compromised health and growth potential of LBW female piglets. The changes in methylation observed in LBW in key regulators of intestinal development (OLFM4 and FZD5) suggest long-term effects of BW on intestinal gene expression, development, and function. Accordingly, experimental demethylation induced in IPEC-J2 cells led to increased expression of intestinal genes (MGA, DPP4, and GLUT2). Overall, we have identified the alterations in transcription or epigenetic marking at a number of genes critical to intestinal development, which may contribute to both the short- and long-term failure of LBW female piglets to thrive.

Impaired Performance of Broiler Chickens Fed Diets Naturally Contaminated with Moderate Levels of Deoxynivalenol.

Mycotoxin exposure is common in the poultry industry. Deoxynivalenol (DON) is usually detected at levels below the maximum threshold (5000 ppb), but depending on diet and age, broiler performance can be affected. We evaluated the effects of 900 ppb and 2300 ppb DON on the performance, intestinal morphometry, and lesion scores of broiler chickens. One-day-old male Ross broilers (n = 736) were divided into 4 treatments with 8 replicates each, and a pen containing 23 birds was the experimental unit. The animals were fed diets naturally contaminated with two levels of DON: 900 (Low DON-LD) or 2300 (Moderate DON-MD) ppb, with or without activated charcoal, over 28 days. After this, all birds were fed a marginally DON-contaminated diet without charcoal. During the first 28 days, body weight gain (BWG) and feed conversion ratio (FCR) were significantly impaired when broilers were fed a MD diet without activated charcoal. Even after feeding a marginally contaminated diet from D28-35, birds previously fed the MD diet presented a significantly lower performance. The villus height:crypt depth (VH:CD) ratio was significantly higher in the ileum from 14-day-old broilers fed the MD when compared with the LD diet. At D28, the MD diet caused decreased villus height (VH) and increased crypt depth (CD), affecting VH:CD ratio in both intestinal segments, with higher levels in the jejunum from 28-day-old broilers fed a non-supplemented LD diet. Broiler production was negatively affected by DON, even at moderate levels (2300 ppb).

The differences in intestinal growth and microorganisms between male and female ducks.

There are great differences in physiological and biological functions between animals of different sexes. However, whether there is a consensus between sexes in duck intestinal development and microorganisms is still unknown. The current study used Nonghua ducks to estimate the effect of sex on the intestine by evaluating differences in intestinal growth indexes and microorganisms. The intestines of male and female ducks were sampled at 2, 5, and 10 wk from the duodenum, jejunum, ileum, and cecum. Then, the intestinal length and weight were measured, the morphology was observed with HE staining, and the intestinal content was analyzed by 16S rRNA sequencing. The results showed that male ducks have shorter intestinal lengths with higher relative weights/relative lengths. The values of jejunal villus height (VH)/crypt depth (CD) of female ducks were significantly higher at 2 wk, whereas the jejunal VH/CD was significantly lower at 10 wk. There was obvious separation of microorganisms in each intestinal segment of ducks of different sexes at the 3 time periods. The dominant phyla at different stages were Firmicutea, Proteobacteria, Bacteroidetes, and Actinobacteria. The duodenal Chao index at the genus level of male ducks was significantly higher at 10 wk than that of female ducks. Significantly different genera were found only in the jejunum, and the abundances of Escherichia_Shigella, Pseudomonas, Clostridium_sensu_stricto_1, Sphingomonas, and Desulfovibrio in male ducks were higher than those in female ducks, whereas the abundance of Rothia was lower, and the abundance of viral infectious diseases, lipid metabolism, metabolism of terpenoids and polyketides, parasitic infectious diseases, xenobiotic biodegradation and metabolism, cardiovascular disease, and metabolism of other amino acids in male ducks were higher than that in female ducks, whereas gene folding, sorting and degradation pathways, and nucleotide metabolism were lower. This study provides a basic reference for the intestinal development and microbial symbiosis of ducks of different sexes.

Postnatal growth retardation is associated with deteriorated intestinal mucosal barrier function using a porcine model.

Individuals with postnatal growth retardation (PGR) are prone to developing chronic diseases. Abnormal development in small intestine is casually implicated in impaired growth. However, the exact mechanism is still implausible. In this present study, PGR piglets (aged 42 days) were employed as a good model to analyze developmental changes in intestinal mucosal barrier function. Our data demonstrated that PGR piglets exhibited impaired jejunal and ileal epithelial villous morphology and permeability, accompanied by decreased cell proliferation ability and increased apoptosis rate. In addition, the expression of tight junction proteins (ZO-1, claudin 1, and occludin) and E-cadherin was markedly inhibited by PGR. The expression of P-glycoprotein was significantly reduced in PGR piglets, as well as decreased activity of lysozyme. Moreover, the mRNA abundance and content of inflammatory cytokines were significantly increased in the intestinal mucosa and plasma of PGR piglets, respectively. PGR also contributed to lower level of sIgA, and higher level of CD68-positive rate, ß-defensins, and protein expression involved p38 MAPK/NF-κB pathway. Furthermore, PGR altered the intestinal microbial community such as decreased genus Alloprevotella and Oscillospira abundances, and led to lower microbial-derived butyrate production, which may be potential targets for treatment. Collectively, our findings indicated that the intestinal mucosal barrier function of PGR piglets could develop the nutritional intervention strategies in prevention and treatment of the intestinal mucosal barrier dysfunction in piglets and humans.

Dietary 25-hydroxycholecalciferol supplementation improves performance, immunity, antioxidant status, intestinal morphology, and bone quality in weaned piglets.

BACKGROUND: 25-Hydroxycholecalciferol (25OHD3 ) is a new feed additive, which is a potential alternative to vitamin D3 in swine nutrition. The objective of this study was to determine the effects of different doses of 25OHD3 supplementation on performance, immunity, antioxidant capacity, intestinal morphology and bone quality in piglets. RESULTS: As dietary 25OHD3 supplementation increased, the average daily gain (ADG) improved (P < 0.05) quadratically during days 1-14, and tended to increase (P = 0.06) quadratically during the overall period of the experiment. Increasing 25OHD3 supplementation increased (linear effect, P < 0.05) the serum 25OHD3 level and serum glutathione peroxidase (GSH-Px) activity. On day 14, serum immunoglobulin A (IgA) was increased (linear and quadratic effects, P < 0.05) as dietary 25OHD3 supplementation increased. On day 28, serum IgA level was higher (P < 0.05) linearly and the complement 3 (C3) level was reduced (P < 0.05) linearly as dietary supplementation of 25OHD3 increased. The mucosal GSH-Px activity of the small intestine was higher (quadratic effect, P < 0.05) with increasing 25OHD3 supplementation. Jejunal villus height (P = 0.06) and villus height to crypt depth ratio (P = 0.07) tended to increase quadratically, and the villus height to crypt-depth ratio of the ileum increased (P < 0.05) linearly and quadratically with increasing 25OHD3 supplementation. Dietary supplementation with an increasing level of 25OHD3 increased breaking strength of tibias and femurs (quadratic effect, P < 0.05). CONCLUSION: Increasing dietary 25OHD3 supplementation partly improved performance, immunity, antioxidant status, intestinal morphology, and bone properties of weaned piglets. © 2020 Society of Chemical Industry.

Constitutive Activation of Nrf2 in Mice Expands Enterogenesis in Small Intestine Through Negative Regulation of Math1.

BACKGROUND & AIMS: Notch signaling coordinates cell differentiation processes in the intestinal epithelium. The transcription factor Nrf2 orchestrates defense mechanisms by regulating cellular redox homeostasis, which, as shown previously in murine liver, can be amplified through signaling crosstalk with the Notch pathway. However, interplay between these 2 signaling pathways in the gut is unknown. METHODS: Mice modified genetically to amplify Nrf2 in the intestinal epithelium (Keap1f/f::VilCre) were generated as well as pharmacological activation of Nrf2 and subjected to phenotypic and cell lineage analyses. Cell lines were used for reporter gene assays together with Nrf2 overexpression to study transcriptional regulation of the Notch downstream effector. RESULTS: Constitutive activation of Nrf2 signaling caused increased intestinal length along with expanded cell number and thickness of enterocytes without any alterations of secretory lineage, outcomes abrogated by concomitant disruption of Nrf2. The Nrf2 and Notch pathways in epithelium showed inverse spatial profiles, where Nrf2 activity in crypts was lower than villi. In progenitor cells of Keap1f/f::VilCre mice, Notch downstream effector Math1, which regulates a differentiation balance of cell lineage through lateral inhibition, showed suppressed expression. In vitro results demonstrated Nrf2 negatively regulated Math1, where 6 antioxidant response elements located in the regulatory regions contributed to this repression. CONCLUSIONS: Activation of Nrf2 perturbed the dialog of the Notch cascade though negative regulation of Math1 in progenitor cells, leading to enhanced enterogenesis. The crosstalk between the Nrf2 and Notch pathways could be critical for fine-tuning intestinal homeostasis and point to new approaches for the pharmacological management of absorptive deficiencies.

Short communication: Effect of delaying the first colostrum feeding on small intestinal histomorphology and serum insulin-like growth factor-1 concentrations in neonatal male Holstein calves.

The primary objective of this study was to determine the effect of delaying the first colostrum feeding on small intestinal histomorphology and serum insulin-like growth factor-1 (IGF-1) concentrations, and the secondary objective was to characterize the ultrastructure of the small intestine of neonatal calves at 51 h of life. Twenty-seven male Holstein calves were fed pooled, pasteurized colostrum (7.5% of birth body weight; 62 g of IgG/L) at 45 min (0H, n = 9), 6 h (6H, n = 9), or 12 h (12H, n = 9) after birth. At 12 h after their respective colostrum feeding, calves were fed milk replacer at 2.5% of birth body weight per meal and every 6 h thereafter. Blood samples were collected every 6 h using a jugular catheter and analyzed for serum IGF-1 concentrations using an automated solid-phase chemiluminescent immunoassay. At 51 h of life, calves were euthanized to facilitate collection of the duodenum, proximal and distal jejunum, and ileum. All segments of the small intestine were assessed for histomorphology, whereas scanning electron and transmission electron microscopy analyses were conducted only for the proximal jejunum and ileum. The results revealed that there was no overall effect of colostrum feeding time on serum IGF-1 concentrations; however, serum IGF-1 concentrations were influenced by time. Specifically, concentrations of serum IGF-1 at 48 h were 29% greater than concentrations at 0 h of life (312.8 ± 14.85 vs. 241.9 ± 14.06 ng/mL). Although there was no overall effect of colostrum feeding time on all histomorphological measures assessed, treatment × segment interactions were observed. Villi height was 1.4 times greater in the distal jejunum of 0H calves than in 6H and 12H calves, and 0H calves tended to have 1.2 times greater ileal villus height than 12H calves. In addition, 0H calves had 1.2 and 1.3 times greater ileal crypt depth than 6H and 12H calves, respectively, and 1.3 times greater surface area index than 12H calves in the distal jejunum. Qualitative ultrastructural evaluation of small intestinal enterocytes conducted irrespective of colostrum treatment revealed the presence of large vacuoles of electron-dense material, apical mitochondria, and apical canalicular systems in the jejunum and ileum. These results indicate that the calf intestine at 51 h of life contains unique enterocyte characteristics similar to fetal enterocytes and that delaying colostrum feeding may negatively influence intestinal growth and development.

Mucosal characteristics vary across developmental stages in the small intestine of C57BL/6J mice.

AIMS: The benefits of utilizing laboratory mice include low cost, ease of maintenance, and accessibility of molecular tools. However, the ages of experimental mice in the literature vary drastically. We hypothesized that there exists age-related variation in the murine small intestine across developmental stages. MATERIALS AND METHODS: Segments of small intestine were harvested from C57BL/6J mice of varying ages (E17 to 24 weeks; n = 3-4/group). Slides were analyzed for morphometric parameters, cell types, and crypt proliferation index (CPI). Secondary analysis comparing age-matched males and females (n = 4/group) was performed. Means were compared with Student's t-test and variance of proportions with the Chi-squared test to a significance of p < 0.05. KEY FINDINGS: There were small but significant differences including regional variation in villus height, which abolished when examining the small intestine as a whole. Sexually immature mice had increased CPI compared to mature animals. The most dramatic differences were seen in mice at weaning, which demonstrated shallower crypts, increased CPI, fewer Paneth and goblet cells, and more enterochromaffin cells. Examination of embryonic intestine revealed an underdeveloped mucosa lacking differentiated cells. There were minimal differences when comparing age-matched males and females. SIGNIFICANCE: Small, but statistically significant differences in villus height, crypt depth, and crypt proliferation are present in mice across early developmental stages. Mice at weaning exhibit variation in crypt-villus cell composition compared to older animals, which may explain the propensity for certain intestinal conditions in the very young. Investigators studying the GI mucosa should employ consistent age-matching in order to allow direct comparison between studies.

LIF is essential for ISC function and protects against radiation-induced gastrointestinal syndrome.

Leukemia inhibitory factor (LIF) is a cytokine essential for maintaining pluripotency of mouse embryonic stem cells. However, its role in adult intestinal stem cells (ISCs) is unclear. The adult intestinal epithelium has a high self-renewal rate driven by ISCs in crypts. Here, we find that LIF is present in the ISC niche in crypts and critical for the function of ISCs in maintaining the intestinal epithelial homeostasis and regeneration. Mechanistically, LIF maintains ß-catenin activity through the AKT/GSK3ß signaling to regulate ISC functions. LIF deficiency in mice impairs the renewal of the intestinal epithelium under the physiological condition. Further, LIF deficiency in mice impairs the regeneration of intestinal epithelium in response to radiation and shortens the lifespan of mice after high doses of radiation due to gastrointestinal (GI) syndrome, which can be rescued by administering recombinant LIF (rLIF). Importantly, LIF exhibits a radioprotective role in wild-type (WT) mice by protecting mice from lethal radiation-induced GI syndrome; administering rLIF promotes intestinal epithelial regeneration and prolongs survival in WT mice after radiation. These results reveal a previously unidentified and a crucial role of LIF in ensuring ISC function, promoting regeneration of the intestinal epithelium in response to radiation and protecting against radiation-induced GI syndrome.

Intestinal stem cells promote crypt fission during postnatal growth of the small intestine.

OBJECTIVE: Wnt-ß-catenin signalling is essential for intestinal stem cells. Our aim was to investigate the relationship between intestinal stem cells and crypt fission which peaks during infancy. DESIGN: Duodenal biopsies were obtained during endoscopy to assess the severity of reflux oesophagitis of 15 infants, children and teenagers, which would not affect the duodenum. Samples of small intestine were also obtained from rats 7-72 days of life. Crypt fission was assessed using microdissection of 100 whole crypts and recording the percentage of bifid crypts. Intestinal LGR5+ stem cells were identified by in situ hybridisation. Rats were treated with Dickkopf to block Wnt-ß-catenin signalling. RESULTS: Crypt fission peaked during infancy before declining after 3-4 years in humans and after 21 days of life in rats. Occasional mitotic figures were seen in bifid crypts during early fission. Stem cells were elevated for a greater period during infancy and childhood in humans. Clustering of Paneth cells was present around the stem cells at the crypt base. Dickkopf reduced the number of stem cells and crypt fission to 45% and 29%, respectively, of control values, showing dependence of both crypt fission and Lgr5+ stem cells on Wnt signalling. However, Dickkopf did not decrease mitotic count per crypt, indicating a difference in signalling between stem cells and their progeny in the transit amplifying zone. CONCLUSION: Crypt fission peaks during infancy and is dependent on intestinal stem cells. This is relatively hidden by 'a cloak of invisibility' due to the low proliferation of stem cells.

Prenatal over- and undernutrition differentially program small intestinal growth, angiogenesis, absorptive capacity, and endocrine function in sheep.

The aim was to test the hypothesis that prenatal under- and overnutrition in late gestation can program small intestinal (SI) growth, angiogenesis, and endocrine function to predispose for a hyperabsorptive state, thereby increasing the susceptibility to the adverse effects of an early postnatal obesogenic diet. Twin-pregnant ewes were exposed to adequate (NORM), LOW (50% of NORM), or HIGH (150% energy and 110% protein of NORM) diets through the last trimester (term ~147 days). From 3 days to 6 months of age, their lambs were fed either a moderate (CONV) or a high-carbohydrate high-fat (HCHF) diet. At 6 months of age, responses in plasma metabolites and insulin to refeeding after fasting were determined and then different segments of the SI were sampled at autopsy. Prenatal overnutrition impacts were most abundant in the duodenum where HIGH had increased villus amplification factor and lowered villi thickness with increased IRS-1 and reduced GH-R expressions. In jejunum, HIGH lambs had an increased expression of Lactate gene and amplified when exposed to HCHF postnatally. Specifically, in LOW, sensitivity to HCHF was affected in ileum. Thus, the mismatching LOW-HCHF nutrition increased expressions of angiogenic genes (VEGF, VEGF-R1, ANGPT1, RTK) and increased mucosa layer (tunica mucosa) thickness but reduced muscle layer (Tunica muscularis) thickness. The SI is a target of prenatal nutritional programming, where late gestation overnutrition increased and shifted digestive capacity for carbohydrates toward the jejunum, whereas late gestation undernutrition predisposed for ileal angiogenesis and carbohydrate and fat hyperabsorptive capacity upon subsequent exposure to postnatal obesogenic diet.

Pharmacological activation of TGR5 promotes intestinal growth via a GLP-2-dependent pathway in mice.

Glucagon-like peptide (GLP)-1 and -2-secreting L cells have been shown to express the bile acid receptor Takeda G protein-receptor-5 (TGR5) and increase secretion upon receptor activation. Previous studies have explored GLP-1 secretion following acute TGR5 activation, but chronic activation and GLP-2 responses have not been characterized. In this study, we aimed to investigate the consequences of pharmacological TGR5 receptor activation on L cell hormone production in vivo using the specific TGR5 agonist RO5527239 and the GLP-2 receptor knockout mouse. Here, we show that 1) TGR5 receptor activation led to increased GLP-1 and GLP-2 content in the colon, which 2) was associated with an increased small intestinal weight that 3) was GLP-2 dependent. Additionally, we report that TGR5-mediated gallbladder filling occurred independently of GLP-2 signaling. In conclusion, we demonstrate that pharmacological TGR5 receptor activation stimulates L cells, triggering GLP-2-dependent intestinal adaption in mice.NEW & NOTEWORTHY Using the specific Takeda G protein-receptor-5 (TGR5) agonist RO5527239 and GLP-2 receptor knockout mice, we show that activation of TGR5 led to the increase in colonic GLP-1 and GLP-2 concomitant with a GLP-2 dependent growth response in the proximal portion of the small intestine.

Effect of arginine or glutamine supplementation and milk feeding allowance on small intestine development in calves.

The development of the small intestine (SI) is important for the health and growth of neonatal calves. This study evaluated the effect of arginine (Arg) and glutamine (Gln) supplementation and 2 levels of milk allowance on the histomorphological development of the SI in preweaning calves. Sixty mixed-sex Friesian × Jersey calves (3-5 d of age) were offered reconstituted whole milk (125 g/L, 26% fat, 26% protein) at either high (20% of arrival body weight/d; HM) or low (10% of arrival body weight/d; LM) milk allowance without (Ctrl) or with supplementary Arg or Gln (at 1% of milk dry matter) in a 2 × 3 factorial design (n = 10/treatment). After 35 d on the diets, all calves were slaughtered to collect tissues for examination of SI development. Calves in the HM group had higher milk intake, total weight gain, and average daily gain compared with LM calves, but no effect of AA supplementation nor an interaction between milk allowance and AA supplementation was observed. For the duodenum, we observed an AA by milk allowance interaction for villus height and width, and goblet cell number per villus (HM-Arg > HM-Gln > HM-Ctrl), and villus height to crypt depth ratio (HM-Arg > HM-Gln = HM-Ctrl), but no effect of AA supplementation in the LM group. Goblet cell numbers per 100 µm of SI were greater in Arg-supplemented calves than in unsupplemented controls, with Gln-supplemented calves intermediate to but not different from the other groups. Epithelium thickness was greater in LM than in HM calves. Villus density, crypt depth, and muscle thickness did not differ between groups. For the jejunum, there was an AA by milk allowance interaction for villus height, villus surface area, and villus height to crypt depth ratio (HM-Arg = HM-Gln > HM-Ctrl), with no effect of AA supplementation in the LM groups. Amino acid supplementation affected goblet cell number per villus (HM-Gln > HM-Ctrl calves, HM-Arg intermediate), and both LM-Arg and LM-Gln calves had greater numbers than LM-Ctrl calves. Villus width, crypt depth, and muscle thickness were greater in HM than LM calves but there was no effect of AA supplementation. Villus density, goblet cell number per 100 µm of SI, and epithelium thickness were unaffected by AA supplementation and milk allowance. Milk allowance and AA supplementation had no effect on SI morphology in the ileum. Increasing milk allowance improved villus height, width, and surface area but only in Arg- or Gln-supplemented calves, not in control calves. The observed changes in development may be important for intestinal functionality, integrity, and barrier function in preweaning calves, potentially through increased cell growth and proliferation or reduced levels of cellular atrophy.