Crypt Epithelial Cell Proliferation Research Articles

Von Willebrand factor promotes radiation-induced intestinal injury (RIII) development and its cleavage enzyme rhADAMTS13 protects against RIII by reducing inflammation and oxidative stress

Patients with abdominopelvic cancer undergoing radiotherapy commonly develop radiation-induced intestinal injury (RIII); however, its underlying pathogenesis remains elusive. The von Willebrand factor (vWF)/a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 (ADAMTS13) axis has been implicated in thrombosis, inflammation, and oxidative stress. However, its role in RIII remains unclear. In this study, the effect of radiation on vWF and ADAMTS13 expression was firstly evaluated in patients with cervical cancer undergoing radiotherapy and C57BL/6J mice exposed to different doses of total abdominal irradiation. Then, mice with the specific deletion of vWF in the platelets and endothelium were established to demonstrate the contribution of vWF to RIII. Additionally, the radioprotective effect of recombinant human (rh) ADAMTS13 against RIII was assessed. Results showed that both the patients with cervical cancer undergoing radiotherapy and RIII mouse model exhibited increased vWF levels and decreased ADAMTS13 levels. The knockout of platelet- and endothelium-derived vWF rectified the vWF/ADAMTS13 axis imbalance; improved intestinal structural damage; increased crypt epithelial cell proliferation; and reduced radiation-induced apoptosis, inflammation, and oxidative stress, thereby alleviating RIII. Administration of rhADAMTS13 could equally alleviate RIII. Our results demonstrated that abdominal irradiation affected the balance of the vWF/ADAMTS13 axis. vWF exerted a deleterious role and ADAMTS13 exhibited a protective role in RIII progression. rhADAMTS13 has the potential to be developed into a radioprotective agent.

Survival of Stem Cells and Progenitors in the Intestine Is Regulated by LPA5-Dependent Signaling.

Background & AimsRegeneration of the epithelium by stem cells in the intestine is supported by intrinsic and extrinsic factors. Lysophosphatidic acid (LPA), a bioactive lipid mediator, regulates many cellular functions, including cell proliferation, survival, and cytokine secretion. Here, we identify LPA5 receptor as a potent regulator of the survival of stem cells and transit-amplifying cells in the intestine.MethodsWe have used genetic mouse models of conditional deletion of Lpar5, Lpar5f/f;Rosa-CreERT (Lpar5KO), and intestinal epithelial cell–specific Lpar5f/f;AhCre (Lpar5IECKO) mice. Mice were treated with tamoxifen or β-naphthoflavone to delete Lpar5 expression. Enteroids derived from these mice were used to determine the effect of Lpar5 loss on the apoptosis and proliferation of crypt epithelial cells.ResultsConditional loss of Lpar5 induced ablation of the intestinal mucosa, which increased morbidity of Lpar5KO mice. Epithelial regeneration was compromised with increased apoptosis and decreased proliferation of crypt epithelial cells by Lpar5 loss. Interestingly, intestinal epithelial cell–specific Lpar5 loss did not cause similar phenotypic defects in vivo. Lpar5 loss reduced intestinal stem cell marker gene expression and reduced lineage tracing from Lgr5+ ISCs. Lpar5 loss induced CXCL10 expression which exerts cytotoxic effects on intestinal stem cells and progenitors in the intestinal crypts. By co-culturing Lpar5KO enteroids with wild-type or Lpar5KO splenocytes, we demonstrated that lymphocytes protect the intestinal crypts via a LPA5-dependent suppression of CXCL10.ConclusionsLPA5 is essential for the regeneration of intestinal epithelium. Our findings reveal a new finding that LPA5 regulates survival of stem cells and transit-amplifying cells in the intestine.

The protective role of Chitooligosaccharides against chronic ulcerative colitis induced by dextran sulfate sodium in mice

• Protective effect of COS on the DSS-induced chronic experimental UC development. • Anti-inflammatory and anti-apoptotic activities of COS in chronic UC mice. • COS promoted the proliferation of crypt epithelial cells in chronic UC mice. • COS modulated gut microbiota in chronic UC mice. • Possible involvement of TLR4 signal pathway in COS-mediated protection in chronic UC. • Possible involvement of colonic Reg3g in COS-mediated protection in chronic UC. Chitooligosaccharides(COS) as potential functional food has been paid great attention because of their outstanding biological activities. Here, we found COS (250 and 500 mg/kg) protected against development of dextran sodium sulfate(DSS)- induced chronic experimental ulcerative colitis(UC) in mice. DSS-elevated serum and colonic tumor necrosis factor(TNF)-α, interleukin(IL)-1β and IL-6 levels were significantly supressed. TUNEL + apoptoic cells and Cleaved Caspase-9 expression were downregulated, Ki-67 + proliferative cells in colonic crypts and Bcl-2/Bax ratio were upregulated in COS-treated colons compared to DSS controls. COS enhanced gut microbiota diversity and restored community structure in chronic UC mice. Colonic expression of TLR4, NF-κB, STAT3 and pSTAT3 was significantly decreased, but that of regenerating islet derived 3 gamma(Reg3g) was increased by COS. Overall, COS represents a treatment alternative in chronic UC management via ameliorating inflammation, promoting mucosal repair and modulating gut microbiota. Inhibition of TLR4 signal and induction of colonic Reg3g might be involved in molecular mechanisms.

Condomless receptive anal intercourse is associated with markers of mucosal inflammation in a cohort of men who have sex with men in Atlanta, Georgia.

IntroductionWe previously showed that the rectal mucosal immune environment among men who have sex with men (MSM) engaging in condomless receptive anal intercourse (CRAI) is immunologically distinct from that of men who do not engage in anal intercourse (AI). Here, we further examined these differences with quantitative immunohistochemistry to better understand the geographic distribution of immune markers of interest.MethodsWe enrolled a cohort of MSM engaging in CRAI (n = 41) and men who do not engage in AI (n = 21) between October 2013 and April 2015. Participants were healthy, HIV‐negative men aged 18–45 from the metro Atlanta area. We performed rectal mucosal sampling via rigid sigmoidoscopy during two study visits separated by a median of nine weeks and timed with sexual activity for MSM engaging in CRAI. We used standardized, automated immunohistochemistry and quantitative image analysis to investigate the rectal mucosal distribution of neutrophils (MPO), IL‐17‐producing cells (IL‐17) and Tregs (FOXP3) in the lamina propria, and cellular proliferation (Ki67) and adherens junction protein (E‐cadherin) in the epithelium. We examined associations between biomarker expression and the rectal mucosal microbiota composition by 16s rRNA sequencing.ResultsRelative to the colonic crypt base, IL‐17, FOXP3, and MPO expression increased towards the rectal lumen, while Ki67 decreased and E‐cadherin was more uniformly distributed. Throughout the rectal mucosa distribution examined, MSM engaging in CRAI had higher mean lamina propria MPO expression (p = 0.04) and epithelial Ki67 (p = 0.04) compared to controls. There were no significant differences in IL‐17, FOXP3 or E‐cadherin expression. We found no significant associations of the five biomarkers with the global rectal microbiota composition or the individual taxa examined.ConclusionsUnderstanding the mucosal distribution of inflammatory mediators can enhance our knowledge of the earliest events in HIV transmission. Neutrophil enrichment and crypt epithelial cell proliferation likely represent sub‐clinical inflammation in response to CRAI in the rectal mucosa of MSM, which could increase the risk for HIV acquisition. However, the contributory role of the microbiota in mucosal inflammation among MSM remains unclear. HIV prevention may be enhanced by interventions that reduce inflammation or capitalize on the presence of specific inflammatory mechanisms during HIV exposure.

Sex-dependent lifespan extension of Apc Min/+ FAP mice by chronic mTOR inhibition.

Apc Min/+ mice model familial adenomatous polyposis (FAP), a disease that causes numerous colon polyps leading to colorectal cancer. We previously showed that chronic treatment of Apc Min/+ females with the anti-aging drug, rapamycin, restored a normal lifespan through reduced polyposis and anemia prevention. Lifespan extension by chronic rapamycin in wildtype UM-HET3 mice is sex-dependent with females gaining the most benefit. Whether Apc Min/+ mice have a similar sex-dependent response to chronic mTOR inhibition is not known. To address this knowledge gap and gain deeper insight into how chronic mTOR inhibition prevents intestinal polyposis, we compared male and female Apc Min/+ mice responses to chronic treatment with a rapamycin-containing diet. Animals were fed a diet containing either 42 ppm microencapsulate rapamycin or empty capsules, one group was used to determine lifespan and a second group with similar treatment was harvested at 16 weeks of age for cross-sectional studies. We found that the survival of males is greater than females in this setting (P < 0.0197). To explore the potential basis for this difference we analyzed factors affected by chronic rapamycin. Immunoblot assays showed that males and females exhibited approximately the same level of mTORC1 inhibition using phosphorylation of ribosomal protein S6 (rpS6) as an indirect measure. Immunohistochemistry assays of rpS6 phosphorylation showed that rapamycin reduction of mTORC1 activity was on the same level, with the most prominent difference being in intestinal crypt Paneth cells in both sexes. Chronic rapamycin also reduced crypt depths in both male and female Apc Min/+ mice (P < 0.0001), consistent with reduced crypt epithelial cell proliferation. Finally, chronic rapamycin prevented anemia equally in males and females. In males and females, these findings link rapamycin-mediated intestinal polyposis prevention with mTORC1 inhibition in Paneth cells and concomitant reduced epithelial cell proliferation.

Condomless Receptive Anal Intercourse is Associated with Markers of Mucosal Injury in a Cohort of Men Who Have Sex with Men

Background: The rectal mucosal immune environment among men who have sex with men (MSM) engaging in condomless receptive anal intercourse (CRAI) is immunologically distinct from that of men who do not engage in anal intercourse (AI). Methods: We enrolled a cohort of MSM engaging in CRAI (n=41) and men who do not engage in AI (controls; n=21) and used standardized, automated immunohistochemistry and quantitative image analysis to investigate the rectal mucosal distribution of neutrophils (MPO), IL-17-producing cells (IL-17), and Tregs (FOXP3) in the lamina propria, and cellullar proliferation (Ki67) and adherens junctions (E-cadherin) in the epithelium. We also examined associations between biomarker expression and the composition of the rectal mucosal microbiota. Findings: MSM engaging in CRAI had higher mean expression of MPO in the lamina propria (p=0.04) and Ki67 (p=0.04) in the epithelium relative to controls. There were no differences in IL-17, FOXP3, or E-cadherein expression. We found no statistically significant associations of the five biomarkers with the global rectal microbiota composition or with the individual taxa examined. Interpretation: Increased infiltration of neutrophils and proliferation of crypt epithelial cells may represent an injury response to frequent CRAI in the rectal mucosa of MSM; however, a contributory role of the microbiota to mucosal inflammation among MSM remains unclear. Prevention may be enhanced by interventions that reduce rectal mucosal inflammation or capitalize on the presence of a specific inflammatory mechanism at the time of HIV exposure. Funding Statement: This study was funded by the National Institutes of Health (K23 AI108335). Declaration of Interests: All authors declare no conflict of interest. Ethics Approval Statement: The Institutional Review Board (IRB) at Emory University approved this study.

Exosomes-mediated Transfer of miR-125a/b in Cell-to-cell Communication: A Novel Mechanism of Genetic Exchange in the Intestinal Microenvironment.

Glucagon-like peptide-2 (GLP-2), a key factor in intestinal rehabilitation therapy of short bowel syndrome (SBS), may require cell-to-cell communication to exert its biological functions. However, understanding of the mechanism remains elusive. Here, we report participation of exosomal miR-125a/b in GLP-2 mediated intestinal epithelial cells-myofibroblasts cross-talk in intestinal microenvironment.Methods: The effects of GLP-2 on the proliferation and apoptosis of intestinal epithelial cells in SBS rat models were evaluated. Exosomes were extracted from residual jejunum tissue of GLP-2 or vehicle treated SBS rats using ultracentrifugation method, and identified by nanoparticle trafficking analysis (NTA), transmission electron microscopy and western blotting. miRNA sequencing combined with qRT-PCR validation were used to identify differentially expressed miRNAs. miRNAs, which might be involved in proliferation and apoptosis of intestinal epithelial cells, were screened and further verified by miRNA functional experiments. Moreover, the proliferation-promoting and anti-apoptosis effects of GLP-2 on intestinal myofibroblasts, which expressing GLP-2 receptor, and whether GLP-2 could influence the content of miRNAs in the derived exosomes were studied. The downstream pathways were explored by miRNA function recovery experiment, luciferase reporter assay, pull down experiment, knockdown and overexpression of target gene and other experiments based on the bioinformatics prediction of miRNA target gene.Results: GLP-2 significantly promoted intestinal growth, facilitated the proliferation of intestinal crypt epithelial cells and inhibited the apoptosis of intestinal villi epithelial cells in type II SBS rats. GLP-2 significantly down-regulated exosomal miR-125a/b both in residual jejunums derived exosomes and in exosomes secreted by GLP-2R positive cells. Exosomal miR-125a/b was responsible for GLP-2 mediated intestinal epithelial cells proliferation promotion and apoptosis attenuation. miR-125a/b inhibited the proliferation and promotes apoptosis of intestinal epithelial cells by suppressing the myeloid cell leukemia-1 (MCL1).Conclusions: miR-125a/b shuttled by intestinal myofibroblasts derived exosomes regulate the proliferation and apoptosis of intestinal epithelial cells. GLP-2 treatment significantly decreases the level of miR-125a/b in the exosomes of intestinal myofibroblasts. miR-125a/b modulates the proliferation and apoptosis of intestinal epithelial cells by targeting the 3'UTR region of MCL1. Hence, this study indicates a novel mechanism of genetic exchange between cells in intestinal microenvironment.

Dual-specificity phosphatase 6 deletion protects the colonic epithelium against inflammation and promotes both proliferation and tumorigenesis.

The Ras/mitogen‐activated protein kinase (MAPK) pathway controls fundamental cellular processes such as proliferation, differentiation, and apoptosis. The dual‐specificity phosphatase 6 (DUSP6) regulates cytoplasmic MAPK signaling by dephosphorylating and inactivating extracellular signal‐regulated kinase (ERK1/2) MAPK. To determine the role of DUSP6 in the maintenance of intestinal homeostasis, we characterized the intestinal epithelial phenotype of Dusp6 knockout (KO) mice under normal, oncogenic, and proinflammatory conditions. Our results show that loss of Dusp6 increased crypt depth and epithelial cell proliferation without altering colonic architecture. Crypt regeneration capacity was also enhanced, as revealed by ex vivo Dusp6 KO organoid cultures. Additionally, loss of Dusp6 induced goblet cell expansion without affecting enteroendocrine and absorptive cell differentiation. Our data also demonstrate that Dusp6 KO mice were protected from acute dextran sulfate sodium‐induced colitis, as opposed to wild‐type mice. In addition, Dusp6 gene deletion markedly enhanced tumor load in Apc Min/+ mice. Decreased DUSP6 expression by RNA interference in HT29 colorectal cancer cells enhanced ERK1/2 activation levels and promoted both anchorage‐independent growth in soft agar as well as invasion through Matrigel. Finally, DUSP6 mRNA expression in human colorectal tumors was decreased in advanced stage tumors compared with paired normal tissues. These results demonstrate that DUSP6 phosphatase, by controlling ERK1/2 activation, regulates colonic inflammatory responses, and protects the intestinal epithelium against oncogenic stress.

Butyrate and bioactive proteolytic form of Wnt-5a regulate colonic epithelial proliferation and spatial development.

Proliferation and spatial development of colonic epithelial cells are highly regulated along the crypt vertical axis, which, when perturbed, can result in aberrant growth and carcinogenesis. In this study, two key factors were identified that have important and counterbalancing roles regulating these processes: pericrypt myofibroblast-derived Wnt-5a and the microbial metabolite butyrate. Cultured YAMC cell proliferation and heat shock protein induction were analzyed after butryate, conditioned medium with Wnt5a activity, and FrzB containing conditioned medium. In vivo studies to modulate Hsp25 employed intra-colonic wall Hsp25 encoding lentivirus. To silence Wnt-5a in vivo, intra-colonic wall Wnt-5a silencing RNA was used. Wnt-5a, secreted by stromal myofibroblasts of the lower crypt, promotes proliferation through canonical β-catenin activation. Essential to this are two key requirements: (1) proteolytic conversion of the highly insoluble ~40 kD Wnt-5a protein to a soluble 36 mer amino acid peptide that activates epithelial β-catenin and cellular proliferation, and (2) the simultaneous inhibition of butyrate-induced Hsp25 by Wnt-5a which is necessary to arrest the proliferative process in the upper colonic crypt. The interplay and spatial gradients of these factors insures that crypt epithelial cell proliferation and development proceed in an orderly fashion, but with sufficient plasticity to adapt to physiological perturbations including inflammation.

Protective Actions of Epithelial 5-Hydroxytryptamine 4 Receptors in Normal and Inflamed Colon

The 5-hydroxytryptamine receptor 4 (5-HT4R or HTR4) is expressed in the colonic epithelium but little is known about its functions there. We examined whether activation of colonic epithelial 5-HT4R protects colons of mice from inflammation. The 5-HT4R agonist tegaserod (1 mg/kg), the 5-HT4R antagonist GR113808 (1 mg/kg), or vehicle (control) were delivered by enema to wild-type or 5-HT4R knockout mice at the onset of, or during, active colitis, induced by administration of dextran sodium sulfate or trinitrobenzene sulfonic acid. Inflammation was measured using the colitis disease activity index and by histologic analysis of intestinal tissues. Epithelial proliferation, wound healing, and resistance to oxidative stress-induced apoptosis were assessed, as was colonic motility. Rectal administration of tegaserod reduced the severity of colitis compared with mice given vehicle, and accelerated recovery from active colitis. Rectal tegaserod did not improve colitis in 5-HT4R knockout mice, and intraperitoneally administered tegaserod did not protect wild-type mice from colitis. Tegaserod increased proliferation of crypt epithelial cells. Stimulation of 5-HT4R increased Caco-2 cell migration and reduced oxidative stress-induced apoptosis; these actions were blocked by co-administration of the 5-HT4R antagonist GR113808. In noninflamed colons of wild-type mice not receiving tegaserod, inhibition of 5-HT4Rs resulted in signs of colitis within 3 days. In these mice, epithelial proliferation decreased and bacterial translocation to the liver and spleen was detected. Daily administration of tegaserod increased motility in inflamed colons of guinea pigs and mice, whereas administration of GR113808 disrupted motility in animals without colitis. 5-HT4R activation maintains motility in healthy colons of mice and guinea pigs, and reduces inflammation in colons of mice with colitis. Agonists might be developed as treatments for patients with inflammatory bowel diseases.

Serotonin transporter variant drives preventable gastrointestinal abnormalities in development and function.

Autism spectrum disorder (ASD) is an increasingly common behavioral condition that frequently presents with gastrointestinal (GI) disturbances. It is not clear, however, how gut dysfunction relates to core ASD features. Multiple, rare hyperfunctional coding variants of the serotonin (5-HT) transporter (SERT, encoded by SLC6A4) have been identified in ASD. Expression of the most common SERT variant (Ala56) in mice increases 5-HT clearance and causes ASD-like behaviors. Here, we demonstrated that Ala56-expressing mice display GI defects that resemble those seen in mice lacking neuronal 5-HT. These defects included enteric nervous system hypoplasia, slow GI transit, diminished peristaltic reflex activity, and proliferation of crypt epithelial cells. An opposite phenotype was seen in SERT-deficient mice and in progeny of WT dams given the SERT antagonist fluoxetine. The reciprocal phenotypes that resulted from increased or decreased SERT activity support the idea that 5-HT signaling regulates enteric neuronal development and can, when disturbed, cause long-lasting abnormalities of GI function. Administration of a 5-HT4 agonist to Ala56 mice during development prevented Ala56-associated GI perturbations, suggesting that excessive SERT activity leads to inadequate 5-HT4-mediated neurogenesis. We propose that deficient 5-HT signaling during development may contribute to GI and behavioral features of ASD. The consequences of therapies targeting SERT during pregnancy warrant further evaluation.

Enteral Nutrition in the Management of Pediatric Intestinal Failure

Intestinal failure is an uncommon but devastating condition whose natural history has dramatically improved over the past two decades (1). Infants with intestinal failure due to severe short bowel syndrome or other diagnoses previously considered incompatible with life are now routinely being saved and cared for in cutting edge, multidisciplinary programs. Enteral nutrition plays a central role in the management of children with intestinal failure. This review provides an overview of enteral nutrition in pediatric intestinal failure, with specific emphasis on recent advances in clinical management, patient outcomes, and emerging therapies.

Loss of Sonic hedgehog leads to alterations in intestinal secretory cell maturation and autophagy.

BackgroundIntestinal epithelial cells express the Sonic and Indian hedgehog ligands. Despite the strong interest in gut hedgehog signaling in GI diseases, no studies have specifically addressed the singular role of intestinal epithelial cell Sonic hedgehog signaling. The aim of this study was to investigate the specific role of Sonic hedgehog in adult ileal epithelial homeostasis.Methodology/Principal FindingsA Sonic hedgehog intestinal epithelial conditional knockout mouse model was generated. Assessment of ileal histological abnormalities, crypt epithelial cell proliferation, epithelial cell fate, junctional proteins, signaling pathways, as well as ultrastructural analysis of intracellular organelles were performed in control and mutant mice. Mice lacking intestinal epithelial Sonic Hedgehog displayed decreased ileal crypt/villus length, decreased crypt proliferation as well as a decrease in the number of ileal mucin-secreting goblet cells and antimicrobial peptide-secreting Paneth cells during adult life. These secretory cells also exhibited disruption of their secretory products in mutant mice. Ultrastructural microscopy analysis revealed a dilated ER lumen in secretory cells. This phenotype was also associated with a decrease in autophagy.Conclusions/SignificanceAltogether, these findings indicate that the loss of Sonic hedgehog can lead to ileal secretory cell modifications indicative of endoplasmic reticulum stress, accompanied by a significant reduction in autophagy.

Down-regulation of mechanisms involved in cell transport and maintenance of mucosal integrity in pigs infected with Lawsonia intracellularis.

Lawsonia intracellularis is an obligate intracellular bacterium, responsible for the disease complex known as proliferative enteropathy (PE). L. intracellularis is associated with intestinal crypt epithelial cell proliferation but the mechanisms responsible are yet to be defined. Microarray analysis was used to investigate the host-pathogen interaction in experimentally infected pigs to identify pathways that may be involved. Ileal samples originating from twenty-eight weaner pigs experimentally challenged with a pure culture of L. intracellularis (strain LR189/5/83) were subjected to microarray analysis. Microarray transcriptional signatures were validated using immunohistochemistry and quantitative real time PCR of selected genes at various time points post challenge. At peak of infection (14 days post challenge) 86% of altered transcripts were down regulated, particularly those involved in maintenance of mucosal integrity and regulation of cell transport. Among the up-regulated transcripts, CD163 and CDK1 were novel findings and considered to be important, due to their respective roles in innate immunity and cellular proliferation. Overall, targeted cellular mechanisms included those that are important in epithelial restitution, migration and protection; maintenance of stable inter-epithelial cell relationships; cell transport of nutrients and electrolytes; innate immunity; and cell cycle.

The Jagged-2/Notch-1/Hes-1 Pathway Is Involved in Intestinal Epithelium Regeneration after Intestinal Ischemia-Reperfusion Injury

BackgroundNotch signaling plays a critical role in the maintenance of intestinal crypt epithelial cell proliferation. The aim of this study was to investigate the role of Notch signaling in the proliferation and regeneration of intestinal epithelium after intestinal ischemia reperfusion (I/R) injury.MethodsMale Sprague-Dawley rats were subjected to sham operation or I/R by occlusion of the superior mesenteric artery (SMA) for 20 min. Intestinal tissue samples were collected at 0, 1, 2, 4, and 6 h after reperfusion. Proliferation of the intestinal epithelium was evaluated by immunohistochemical staining of proliferating nuclear antigen (PCNA). The mRNA and protein expression levels of Notch signaling components were examined using Real-time PCR and Western blot analyses. Immunofluorescence was also performed to detect the expression and location of Jagged-2, cleaved Notch-1, and Hes-1 in the intestine. Finally, the γ-secretase inhibitor DAPT and the siRNA for Jagged-2 and Hes-1 were applied to investigate the functional role of Notch signaling in the proliferation of intestinal epithelial cells in an in vitro IEC-6 culture system.ResultsI/R injury caused increased intestinal crypt epithelial cell proliferation and increased mRNA and protein expression of Jagged-2, Notch-1, and Hes-1. The immunofluorescence results further confirmed increased protein expression of Jagged-2, cleaved Notch-1, and Hes-1 in the intestinal crypts. The inhibition of Notch signaling with DAPT and the suppression of Jagged-2 and Hes-1 expression using siRNA both significantly inhibited the proliferation of IEC-6 cells.ConclusionThe Jagged-2/Notch-1/Hes-1 signaling pathway is involved in intestinal epithelium regeneration early after I/R injury by increasing crypt epithelial cell proliferation.

The Jagged-1/Notch-1/Hes-1 Pathway Is Involved in Intestinal Adaptation in a Massive Small Bowel Resection Rat Model

Notch signaling is required for the maintenance of intestinal epithelial proliferation. Dysfunction of this signaling pathway is associated with the loss of proliferated crypt epithelial cells. The aim of this study was to investigate the role of Notch signaling in small bowel resection (SBR)-associated crypt epithelial cell proliferation. Male Sprague-Dawley rats were subjected to sham operation (bowel transection and reanastomosis) or 70% mid-SBR. Intestinal tissue samples were collected at 0.5, 1, 6, 12, 24, 72, and 168 h after operation. The expression of Notch pathway mRNAs and proteins was analyzed using RT-PCR and Western blot. The expression of the Notch pathway proteins Jagged-1, NICD and Hes-1 was also determined through immunohistochemical staining using day 3 postoperative intestinal tissues. The degree of crypt epithelial cell proliferation was evaluated using the immunohistochemical staining of proliferating cell nuclear antigen (PCNA). Furthermore, IEC-6 cells were used to examine the function of the Jagged-1 signaling system. SBR led to increased crypt epithelial cell proliferation and increased expression of Jagged-1 and Hes-1 mRNA and protein along with cleaved Notch-1. Immunohistochemical staining showed that Jagged-1, cleaved Notch-1 and Hes-1 colocalized in the same proliferated crypt epithelial cell population. Recombinant Jagged-1 significantly stimulated the proliferation of IEC-6 cells. Transient upregulation of Jagged-2 expression was found 1 h after SBR, and it was accompanied by cleaved Notch-1 and Hes-1 upregulation. The Jagged-1/Notch-1/Hes-1 signaling pathway is involved in intestinal adaptation through increasing crypt epithelial cell proliferation.

GM-CSF Produced by Nonhematopoietic Cells Is Required for Early Epithelial Cell Proliferation and Repair of Injured Colonic Mucosa

GM-CSF is a growth factor that promotes the survival and activation of macrophages and granulocytes, as well as dendritic cell differentiation and survival in vitro. The mechanism by which exogenous GM-CSF ameliorates the severity of Crohn's disease in humans and colitis in murine models has mainly been considered to reflect its activity on myeloid cells. We used GM-CSF-deficient (GM-CSF(-/-)) mice to probe the functional role of endogenous host-produced GM-CSF in a colitis model induced after injury to the colon epithelium. Dextran sodium sulfate (DSS), at doses that resulted in little epithelial damage and mucosal ulceration in wild type mice, caused marked colon ulceration and delayed ulcer healing in GM-CSF(-/-) mice. Colon crypt epithelial cell proliferation in vivo was significantly decreased in GM-CSF(-/-) mice at early times after DSS injury. This was paralleled by decreased expression of crypt epithelial cell genes involved in cell cycle, proliferation, and wound healing. Decreased crypt cell proliferation and delayed ulcer healing in GM-CSF(-/-) mice were rescued by exogenous GM-CSF, indicating the lack of a developmental abnormality in the epithelial cell proliferative response in those mice. Nonhematopoietic cells, and not myeloid cells, produced the GM-CSF important for colon epithelial proliferation after DSS-induced injury, as revealed by bone marrow chimera and dendritic cell-depletion experiments, with colon epithelial cells being the cellular source of GM-CSF. Endogenous epithelial cell-produced GM-CSF has a novel nonredundant role in facilitating epithelial cell proliferation and ulcer healing in response to injury of the colon crypt epithelium.

Mo2044 GM-CSF Facilitates Repair of Injured Colon Mucosa in Mice In Vivo by Promoting Crypt Epithelial Cell Proliferation

Mo2044 GM-CSF Facilitates Repair of Injured Colon Mucosa in Mice In Vivo by Promoting Crypt Epithelial Cell Proliferation

Perinatal undernutrition alters intestinal alkaline phosphatase and its main transcription factors KLF4 and Cdx1 in adult offspring fed a high-fat diet

Nutrient restriction during gestation and/or suckling is associated with an increased risk of developing inflammation, obesity and metabolic diseases in adulthood. However, the underlying mechanisms, including the role of the small intestine, are unclear. We hypothesized that intestinal adaptation to the diet in adulthood is modulated by perinatal nutrition. This hypothesis was tested using a split-plot design experiment with 20 controls and 20 intrauterine growth-retarded (IUGR) rats aged 240 days and randomly assigned to be fed a standard chow or a high-fat (HF) diet for 10 days. Jejunal tissue was collected at necropsy and analyzed for anatomy, digestive enzymes, goblet cells and mRNA levels. Cecal contents and blood serum were analyzed for alkaline phosphatase (AP). IUGR rats failed to adapt to HF by increasing AP activity in jejunal tissue and cecal content as observed in controls. mRNA levels of transcription factors KLF4 and Cdx1 were blunted in jejunal epithelial cell of IUGR rats fed HF. mRNA levels of TNF-α were lower in IUGR rats. They also displayed exacerbated aminopeptidase N response and reduced jejunal goblet cell density. Villus and crypt architecture and epithelial cell proliferation increased with HF in both control and IUGR rats. Serum AP tended to be lower, and serum levamisole inhibition-resistant AP fraction was lower, in IUGR than controls with HF. Serum fatty acids and triglycerides were higher in IUGR rats and higher with HF. In conclusion, the adult intestine adapts to an HF diet differentially depending on early nutrition, jejunal AP and transcription factors being blunted in IUGR individuals fed HF.

Administration of a dipeptidyl peptidase IV inhibitor enhances the intestinal adaptation in a mouse model of short bowel syndrome

Glucagon-like peptide-2 induces small intestine mucosal epithelial cell proliferation and may have benefit for patients who suffer from short bowel syndrome. However, glucagon-like peptide-2 is inactivated rapidly in vivo by dipeptidyl peptidase IV. Therefore, we hypothesized that selectively inhibiting dipeptidyl peptidase IV would prolong the circulating life of glucagon-like peptide-2 and lead to increased intestinal adaptation after development of short bowel syndrome. Eight-week old C57BL/6J mice underwent a 50% proximal small bowel resection and were treated with either sitagliptin, a dipeptidyl peptidase IV-inhibitor, starting 1 day before surgery versus placebo. The efficacy of dipeptidyl peptidase IV-inhibitor was assessed 3 days after resection, including intestinal morphology, epithelial cell apoptosis, and epithelial cell proliferation. Adaptive mechanisms were assessed with quantitative real-time polymerase chain reaction, and plasma bioactive glucagon-like peptide-2 was measured by radioimmunoassay. Body weight loss and peripheral blood glucose levels did not change compared with short bowel syndrome controls. Dipeptidyl peptidase IV-inhibitor treatment led to significant increases in villus height and crypt depth. Dipeptidyl peptidase IV-inhibitor treatment did not change EC apoptosis rates significantly, but it did increase crypt epithelial cell proliferation significantly versus placebo-short bowel syndrome controls. Dipeptidyl peptidase IV-inhibitor treatment markedly increased messenger RNA expression of β-catenin and c-myc in ileal mucosa. Plasma glucagon-like peptide-2 levels increased significantly (∼ 40.9%) in dipeptidyl peptidase IV-inhibitor short bowel syndrome mice. Dipeptidyl peptidase IV-inhibitor treatment increased short bowel syndrome adaptation and might potentially be useful for short bowel syndrome patients.