Intestinal Epithelial Proliferation Research Articles

MiR-495 promotes intestinal epithelial cell apoptosis through downregulation of Sphingosine-1-phosphate.

Many pathological conditions lead to defects in intestinal epithelial integrity and loss of barrier function; Sphingosine-1-phosphate (S1P) has been shown to augment intestinal barrier integrity, though the exact mechanisms are not completely understood. We have previously shown that overexpression of Sphingosine Kinase 1 (SphK1), the rate limiting enzyme for S1P synthesis, significantly increased S1P production and cell proliferation. Here we show that microRNA 495 (miR-495) upregulation led to decreased levels of SphK1 resultant from a direct effect at the SphK1 mRNA. Increasing expression of miR-495 in intestinal epithelial cells resulted in decreased proliferation and increased susceptibility to apoptosis. Transgenic expression of miR-495 inhibited mucosal growth, as well as decreased proliferation in the crypts. The intestinal villi also expressed decreased levels of barrier proteins and exaggerated damage upon exposure to cecal ligation-puncture. These results implicate miR-495 as a critical negative regulator of intestinal epithelial protection and proliferation through direct regulation of SphK1, the rate limiting enzyme critical for production of S1P.

Increased activity of epithelial Cdc42 Rho GTPase and tight junction permeability in the Cftr knockout intestine.

Increased intestinal permeability is a manifestation of cystic fibrosis (CF) in people with CF (pwCF) and in CF mouse models. CF transmembrane conductance regulator knockout (Cftr KO) mouse intestine exhibits increased proliferation and Wnt/β-catenin signaling relative to wild-type mice (WT). Since the Rho GTPase Cdc42 plays a central role in intestinal epithelial proliferation and tight junction remodeling, we hypothesized that Cdc42 may be altered in the Cftr KO crypts. Immunofluorescence showed distinct tight junction localization of Cdc42 in Cftr KO fresh crypts and enteroids, the latter indicating an epithelial-autonomous feature. Quantitative PCR and immunoblots revealed similar expression of Cdc42 in the Cftr KO crypts/enteroids relative to WT, whereas pulldown assays showed increased GTP-bound (active) Cdc42 in proportion to total Cdc42 in Cftr KO enteroids. Cdc42 activity in the Cftr KO and WT enteroids could be reduced by inhibition of the Wnt transducer Disheveled. With the use of a dye permeability assay, Cftr KO enteroids exhibited increased paracellular permeability to 3 kDa dextran relative to WT. Leak permeability and Cdc42 tight junction localization were reduced to a greater extent by inhibition of Wnt/β-catenin signaling with endo-IWR1 in Cftr KO relative to WT enteroids. Increased proliferation or inhibition of Cdc42 activity with ML141 in WT enteroids had no effect on permeability. In contrast, inhibition of Cdc42 with ML141 increased permeability to both 3 kDa dextran and tight junction impermeant 500 kDa dextran in Cftr KO enteroids. These data suggest that increased constitutive Cdc42 activity may alter the stability of paracellular permeability in Cftr KO crypt epithelium.NEW & NOTEWORTHY Increased tight junction localization and GTP-bound activity of the Rho GTPase Cdc42 was identified in small intestinal crypts and enteroids of cystic fibrosis (CF) transmembrane conductance regulator knockout (Cftr KO) mice. The increase in epithelial Cdc42 activity was associated with increased Wnt signaling. Paracellular flux of an uncharged solute (3 kDa dextran) in Cftr KO enteroids indicated a moderate leak permeability under basal conditions that was strongly exacerbated by Cdc42 inhibition. These findings suggest increased activity of Cdc42 in the Cftr KO intestine underlies alterations in intestinal permeability.

Human Milk Supports Robust Intestinal Organoid Growth, Differentiation, and Homeostatic Cytokine Production

Background and AimsNecrotizing enterocolitis (NEC) is a severe gastrointestinal complication of prematurity. Using small intestinal organoids derived from fetal tissue of a gestational age similar to an extremely preterm infant, this study aims to assess the effect of diet on intestinal epithelial growth and differentiation to elucidate the role nutrition type plays in intestinal development and modifies the risk for NEC. MethodsOrganoids were cultured for 5 days in growth media and 5 days in differentiation media supplemented 1:40 with four different diets: parental milk (PM), donor human milk (DHM), standard formula, or extensively hydrolyzed formula. Images were captured daily and organoids were quantified. Organoids were preserved for RNA sequencing and immunofluorescence staining with Ki67, cleaved caspase 3, and chromogranin-A. Media was saved for cytokine/chemokine and growth factor analysis. ResultsHuman milk supplementation improved growth and differentiation of intestinal organoids generating larger organoids during the growth phase and organoids with longer and wider buds during differentiation compared to formula. Ki67 staining confirmed the proliferative nature of milk-supplemented organoids and chromogranin A staining proved that MM-supplemented organoids induced highest enteroendocrine differentiation. Human milk supplementation also upregulated genes involved in Wnt signaling and fatty acid metabolism pathways and promoted a homeostatic immune landscape, including via increased secretion of TNF-related apoptosis inducing ligand (TRAIL) among other cytokines. Conversely, organoids supplemented with formula had a downregulation of cell-cycle-promoting genes and a more inflammatory immune signature, including a reduced levels of leukemia inhibitory factor (LIF). ConclusionsOur results demonstrate that PM, and to a lesser extent DHM, support robust intestinal epithelial proliferation, differentiation, and homeostatic cytokine production, suggesting a critical role for factors enriched in human milk in intestinal epithelial health.

Secondary bile acids mediate high-fat diet-induced upregulation of R-spondin 3 and intestinal epithelial proliferation.

Secondary bile acids mediate high-fat diet-induced upregulation of R-spondin 3 and intestinal epithelial proliferation.

Inhibition of miR-142-3p promotes intestinal epithelial proliferation and barrier function after ischemia/reperfusion injury by targeting FoxM1.

Damage of intestinal barrier function (BF) after ischemia/reperfusion (I/R) injury can induce serious complications and high mortality. MicroRNAs (miRNAs) are involved in intestinal mucosal BF and epithelial proliferation after I/R injury have been reported. We aimed to investigate the role and regulatory mechanism of miR-142-3p (miR-142) in intestinal epithelial proliferation and BF after I/R injury. We detected the proliferation, barrier function and miR-142 expression in clinical ischemic intestinal tissues. Furthermore, we induced an in vivo intestinal I/R injury mouse model and in vitro IEC-6 cells hypoxia/reoxygenation (H/R) injury model. After increasing and decreasing expression of miR-142, we detected the proliferation and barrier function of intestinal epithelial cells after I/R or H/R injury. We found that miR-142 expression was significantly increased in clinical ischemic intestinal mucosa and mouse intestinal mucosa exposed to I/R injury, and there was an inverse relationship between miR-142 and proliferation/BF. Inhibition of miR-142 significant promoted intestinal epithelial proliferation and BF after I/R injury. Furthermore, inhibition of miR-142 improved overall survival rate of mice after I/R injury. MiR-142 directly targeted FoxM1 which was identified by bioinformatics analysis and luciferase activity assay in IEC-6 cells. Inhibition of miR-142 promotes intestinal epithelial proliferation and BF after I/R injury in a FoxM1-mediated manner.

Microbiota-derived I3A protects the intestine against radiation injury by activating AhR/IL-10/Wnt signaling and enhancing the abundance of probiotics

ABSTRACT The intestine is prone to radiation damage in patients undergoing radiotherapy for pelvic tumors. However, there are currently no effective drugs available for the prevention or treatment of radiation-induced enteropathy (RIE). In this study, we aimed at investigating the impact of indole-3-carboxaldehyde (I3A) derived from the intestinal microbiota on RIE. Intestinal organoids were isolated and cultivated for screening radioprotective tryptophan metabolites. A RIE model was established using 13 Gy whole-abdominal irradiation in male C57BL/6J mice. After oral administration of I3A, its radioprotective ability was assessed through the observation of survival rates, clinical scores, and pathological analysis. Intestinal stem cell survival and changes in the intestinal barrier were observed through immunofluorescence and immunohistochemistry. Subsequently, the radioprotective mechanisms of I3A was investigated through 16S rRNA and transcriptome sequencing, respectively. Finally, human colon cancer cells and organoids were cultured to assess the influence of I3A on tumor radiotherapy. I3A exhibited the most potent radioprotective effect on intestinal organoids. Oral administration of I3A treatment significantly increased the survival rate in irradiated mice, improved clinical and histological scores, mitigated mucosal damage, enhanced the proliferation and differentiation of Lgr5+ intestinal stem cells, and maintained intestinal barrier integrity. Furthermore, I3A enhanced the abundance of probiotics, and activated the AhR/IL-10/Wnt signaling pathway to promote intestinal epithelial proliferation. As a crucial tryptophan metabolite, I3A promotes intestinal epithelial cell proliferation through the AhR/IL-10/Wnt signaling pathway and upregulates the abundance of probiotics to treat RIE. Microbiota-derived I3A demonstrates potential clinical application value for the treatment of RIE.

Bacillus subtilis alleviates excessive apoptosis of intestinal epithelial cells in intrauterine growth restriction suckling piglets via the members of Bcl-2 and caspase families.

The intestine is a barrier resisting various stress responses. Intrauterine growth restriction (IUGR) can cause damage to the intestinal barrier via destroying the balance of intestinal epithelial cells' proliferation and apoptosis. Bacillus subtilis has been reported to regulate intestinal epithelial cells' proliferation and apoptosis. Thus, the purpose of this study was to determine if B. subtilis could regulate intestinal epithelial cells' proliferation and apoptosis in intrauterine growth restriction suckling piglets. Compared with the normal birth weight group, the IUGR group showed greater mean optical density values of Ki-67-positive cells in the ileal crypt (P < 0.05). IUGR resulted in higher ability of proliferation and apoptosis of intestinal epithelial cells, by upregulation of the messenger RNA (mRNA) or proteins expression of leucine rich repeat containing G protein coupled receptor 5, Caspase-3, Caspase-7, β-catenin, cyclinD1, B-cell lymphoma-2 associated agonist of cell death, and BCL2 associated X (P < 0.05), and downregulation of the mRNA or protein expression of B-cell lymphoma-2 and B-cell lymphoma-2-like 1 (P < 0.05). However, B. subtilis supplementation decreased the mRNA or proteins expression of leucine rich repeat containing G protein coupled receptor 5, SPARC related modular calcium binding 2, tumor necrosis factor receptor superfamily member 19, cyclinD1, Caspase-7, β-catenin, B-cell lymphoma-2 associated agonist of cell death, and Caspase-3 (P < 0.05), and increased the mRNA expression of B-cell lymphoma-2 (P < 0.05). IUGR led to excessive apoptosis of intestinal epithelial cells, which induced compensatory proliferation. However, B. subtilis treatment prevented intestinal epithelial cells of IUGR suckling piglets from excessive apoptosis. © 2024 Society of Chemical Industry.

The road to refractory graft-versus-host disease is paved with good intentions.

Refractory acute graft-versus-host disease (GVHD) occurs when the immune injury exceeds the capacity of injured tissues to regenerate and repair. While glucocorticoids have been used for decades to treat GVHD, Arnhold, Chang, and colleagues in this issue of the JCI question whether this approach can in fact be counterproductive. Using in vivo experimental models of GVHD and in vitro intestinal organoids, the study authors show that glucocorticoid exposure directly impeded small intestinal epithelial proliferation and survival, thus preventing the resolution of injury. These findings suggest that future treatment approaches for acute GVHD should include measures to reduce immune reactivity as well as interventions to actively promote tissue resilience.

In-Ovo Glutamine Administration Enhances Intestinal Development and Functions in Broiler Chickens: Insights from Enteroid Models

BackgroundEarly life events play significant roles in tissue development and animal health in their later life. Early nutrition, through in-ovo delivery, has shown beneficial effects on improving intestinal health in broiler chickens. However, the underlying mechanism is not fully investigated. A recently developed enteroid culture technique allows investigations on intestinal epithelial functions that are close to physiologic conditions. ObjectivesIn this study, we evaluated the short- and long-term effects of in-ovo administration of glutamine (Gln) on intestinal epithelial development and functions by using intestinal enteroid culture and tissue electrophysiologic analysis. MethodsA hundred eggs of commercial Cobb500 broilers were in-ovo injected with 0.2 mL of either phosphate-buffered saline (PBS) or 3% Gln at embryonic day 18 (E18). Chicks were killed on the day of hatch, and at 3- and 14-d posthatch. Enteroids were generated from the small intestine. After 4 d of culture, enteroids were harvested for 5-ethynyl-2′-deoxyuridine proliferation, fluorescein isothiocyanate-4 kDa dextran permeability, and glucose absorption assays. At day 3 (d3) and day 14 (d14), intestinal barrier and nutrient transport functions were measured by the Ussing chamber. The gene expression of epithelial cell markers, nutrient transporters, and tight-junction proteins were analyzed in both intestinal tissues and enteroids. ResultsIn comparison with the PBS control group, in-ovo Gln increased intestinal villus morphology, epithelial cell proliferation, and differentiation, and altered epithelial cell population toward increased number of enteroendocrine and goblet cells while decreasing Paneth cells. Enteroids gene expression of nutrient transporters (B0AT1, SGLT1, and EAAT3), tight junction (ZO2), glucose absorption, and barrier functions were enhanced on the day of hatch. Long-term increases of intestinal di-peptide and alanine transport were observed at day 14 posthatch. ConclusionsTogether our results suggested that the in-ovo injection of Gln stimulated intestinal epithelium proliferation and programmed the epithelial cell differentiation toward absorptive cells.

Spatial Variation of the Gastrointestinal Microbiota in Response to Long-Term Administration of Vonoprazan in Mice With High Risk of Gastric Cancer.

Vonoprazan, a potassium-competitive acid blocker, is superior to traditional proton pump inhibitor (PPI) in acid suppression and has been approved in the treatment of acid-related disorders. Accumulating evidence suggest associations between PPI use and gut microbiota, yet the effect of vonoprazan on GI microbiota is obscure. Transgenic FVB/N insulin-gastrin (INS-GAS) mice as a model of gastric cancer (GC) were administered vonoprazan by gavage every other day for 12 weeks. Stomachs were evaluated by histopathology, Ki-67 proliferation index, and inflammatory cytokines. The mucosal and lumen microbiota from stomach, jejunum, ileum, cecum, and feces were detected using 16S rRNA gene sequencing. Higher incidence of intestinal metaplasia and epithelial proliferation were observed in the vonoprazan group than that in the control mice. Vonoprazan also elevated the gastric expression of proinflammatory cytokines, including TNF-α, IL-1β, and IL-6. Each mice comprised a unique microbiota composition that was consistent across different niches. The structure of GI microbiota changed dramatically after vonoprazan treatment with the stomach being the most disturbed segment. Vonoprazan administration shifted the gut microbiota toward the enrichment of pathogenic Streptococcus, Staphylococcus, Bilophila, and the loss of commensal Prevotella, Bifidobacterium, and Faecalibacterium. Interestingly, compared to the controls, microbial interactions were weaker in the stomach while stronger in the jejunum of the vonoprazan group. Long-term vonoprazan treatment promoted gastric lesions in male INS-GAS mice, with the disequilibrium of GI microbiome. The clinical application of vonoprazan needs to be judicious particularly among those with high risk of GC.

Maternal resveratrol improves the intestinal health and weight gain of suckling piglets during high summer temperatures: The involvement of exosome-derived microRNAs and immunoglobin in colostrum

Previous studies have shown that maternal resveratrol improved growth performance and altered the microbial composition of suckling piglets under hot summer conditions. However, it remains unclear how maternal resveratrol improves growth performance of suckling piglets during high summer temperatures. A total of 20 sows (Landrace × Large White; three parity) were randomly assigned to 2 groups (with or without 300 mg/kg resveratrol) from d 75 of gestation to d 21 of lactation during high ambient temperatures (from 27 to 30 °C). The results showed that maternal resveratrol supplementation increased total daily weight gain of piglets under hot summer conditions, which is consistent with previous studies. Furthermore, we found that maternal resveratrol improved the intestinal morphology and intestinal epithelial proliferation in suckling piglets. Dietary resveratrol supplementation affected the characteristics of exosome-derived microRNAs (miRNAs) in sow colostrum, as well as the genes targeted by differentially produced miRNAs. MiRNAs are concentrated in the tight junction pathway. As a result, the expression of intestinal tight junction proteins was increased in suckling piglets (P < 0.05). Notably, maternal resveratrol increased the intestinal secretory immunoglobulin A (sIgA) levels of suckling piglets via colostrum immunoglobin (P < 0.05), which could increase the abundance of beneficial microbiota to further increase the concentration of short chain fatty acids (SCFA) in suckling piglets' intestine (P < 0.05). Finally, our correlation analysis further demonstrated the positive associations between significantly differential intestinal microbiota, intestinal sIgA production and SCFA concentrations, as well as the positive relation between total daily weight gain and intestinal health of suckling piglets. Taken together, our findings suggested that maternal resveratrol could promote intestinal health to improve piglet growth during high summer temperatures, which might be associated with the immunoglobin and exosome-derived miRNAs in sows' colostrum.

A purified diet affects intestinal epithelial proliferation and barrier functions through gut microbial alterations.

The gut microbiota plays a crucial role in maintaining epithelial barrier function. Although multiple studies have demonstrated the significance of dietary factors on the gut microbiota and mucosal barrier function, the impact of a purified diet, which has long been used in various animal experiments, on intestinal homeostasis remains to be elucidated. Here, we compared the impact of two different types of diets, a crude diet and an AIN-93G-formula purified diet, on epithelial integrity and the gut microbiota. Purified diet-fed mice exhibited shorter villi and crypt lengths and slower epithelial turnover, particularly in the ileum. In addition, antimicrobial products, including REG3γ, were substantially decreased in purified diet-fed mice. Purified diet feeding also suppressed α1,2-fucosylation on the epithelial surface. Furthermore, the purified diet induced metabolic rewiring to fatty acid oxidation and ketogenesis. 16S ribosomal RNA gene sequencing of the ileal contents and mucus layer revealed distinct gut microbiota compositions between the purified and crude diet-fed mice. Purified diet feeding reduced the abundance of segmented filamentous bacteria (SFB), which potently upregulate REG3γ and fucosyltransferase 2 (Fut2) by stimulating group 3 innate lymphoid cells (ILC3s) to produce IL-22. These observations illustrate that the intake of a crude diet secures epithelial barrier function by facilitating SFB colonization, whereas a purified diet insufficiently establishes the epithelial barrier, at least partly owing to the loss of SFB. Our data suggest that the influence of purified diets on the epithelial barrier integrity should be considered in experiments using purified diets.

Dietary sodium butyrate and forskolin promote cell proliferation to resist Citrobacter rodentium infection by lysozyme upregulation

The invasion of pathogens derived from diet or the environment triggers intestinal inflammation, which is a result of the host intestinal barrier damage and immune dysfunction, such as the decrease of host defense peptides (HDPs) and inhibition of cell proliferation. Herein, the siRNA interference demonstrated that a critical lysozyme, which was mainly induced by dietary supplementations of sodium butyrate and forskolin, improved cell proliferation by inhibiting p38 MAPK and JNK signaling pathways and reinforced the intestinal barrier. Moreover, we established a mouse model of intestinal inflammation with Citrobacter rodentium (C. rodentium). It was found that NaBu and FSK stimulated intestinal epithelial proliferation to repair barrier damage, and performed stronger disease resistance to C. rodentium infection on account of the lysozyme upregulation in colon. Therefore, this study revealed that NaBu and FSK prevented or alleviated the development of foodborne infections by promoting lysozyme expression, which may be mediated by the effect of lysozyme on promoting cell proliferation and improving barrier function.

Quantitative Analysis of NKX2-3 Expression in Human Colon: An Immunohistochemical Study.

In mice, Nkx2-3 homeodomain transcription factor defines the vascular specification of secondary and tertiary lymphoid tissues of the intestines. In human studies, polymorphisms in NKX2-3 have been identified as a susceptibility factor in inflammatory bowel diseases, whereas in mice, its absence is associated with protection against experimental colitis and enhanced intestinal epithelial proliferation. Here, we investigated the expression of NKX2-3 in normal, polyp, and adenocarcinoma human colon samples using immunohistochemistry and quantitative morphometry, correlating its expression with endothelial and mesenchymal stromal markers. Our results revealed that the expression of NKX2-3 is regionally confined to the lamina propria and lamina muscularis mucosae, and its production is restricted mostly to endothelial cells and smooth muscle cells with variable co-expression of CD34, alpha smooth muscle antigen (αSMA), and vascular adhesion protein-1 (VAP-1). The frequency of NKX2-3-positive cells and intensity of expression correlated inversely with aging. Furthermore, in most colorectal carcinoma samples, we observed a significant reduction of NKX2-3 expression. These findings indicate that the NKX2-3 transcription factor is produced by both endothelial and non-endothelial tissue constituents in the colon, and its expression changes during aging and in colorectal malignancies. (J Histochem Cytochem XX: XXX-XXX, XXXX).

Human intestinal organoids from Cronkhite-Canada syndrome patients reveal link between serotonin and proliferation.

Cronkhite-Canada Syndrome (CCS) is a rare, noninherited polyposis syndrome affecting 1 in every million individuals. Despite over 50 years of CCS cases, the etiopathogenesis and optimal treatment for CCS remains unknown due to the rarity of the disease and lack of model systems. To better understand the etiology of CCS, we generated human intestinal organoids (HIOs) from intestinal stem cells isolated from 2 patients. We discovered that CCS HIOs are highly proliferative and have increased numbers of enteroendocrine cells producing serotonin (also known as 5-hydroxytryptamine or 5HT). These features were also confirmed in patient tissue biopsies. Recombinant 5HT increased proliferation of non-CCS donor HIOs and inhibition of 5HT production in the CCS HIOs resulted in decreased proliferation, suggesting a link between local epithelial 5HT production and control of epithelial stem cell proliferation. This link was confirmed in genetically engineered HIOs with an increased number of enteroendocrine cells. This work provides a new mechanism to explain the pathogenesis of CCS and illustrates the important contribution of HIO cultures to understanding disease etiology and in the identification of novel therapies. Our work demonstrates the principle of using organoids for personalized medicine and sheds light on how intestinal hormones can play a role in intestinal epithelial proliferation.

Mmp17-deficient mice exhibit heightened goblet cell effector expression in the colon and increased resistance to chronic Trichuris muris infection.

Intestinal epithelial homeostasis is maintained by intrinsic and extrinsic signals. The extrinsic signals include those provided by mesenchymal cell populations that surround intestinal crypts and is further facilitated by the extracellular matrix (ECM), which is modulated by proteases such as matrix metalloproteinases (MMPs). Extrinsic signals ensure an appropriate balance between intestinal epithelial proliferation and differentiation. This study explores the role of MMP17, which is preferentially expressed by smooth muscle cells in the intestine, in intestinal homeostasis and during immunity to infection. Mice lacking MMP17 expressed high levels of goblet-cell associated genes and proteins, such as CLCA1 and RELM-β, which are normally associated with immune responses to infection. Nevertheless, Mmp17 KO mice did not have altered resistance during a bacterial Citrobacter rodentium infection. However, when challenged with a low dose of the helminth Trichuris muris, Mmp17 KO mice had increased resistance, without a clear role for an altered immune response during infection. Mechanistically, we did not find changes in traditional modulators of goblet cell effectors such as the NOTCH pathway or specific cytokines. We found MMP17 expression in smooth muscle cells as well as lamina propria cells such as macrophages. Together, our data suggest that MMP17 extrinsically alters goblet cell maturation which is sufficient to alter clearance in a helminth infection model.

(-)-Epigallocatechin-3-gallate promotes intestinal epithelial proliferation and barrier function after ischemia/reperfusion injury via activation of Nurr1

Context (-)-Epigallocatechin-3-gallate (EGCG) is involved in cell proliferation and ischemia/reperfusion (I/R) injury of several organs. Objective To identify the role of EGCG in intestinal epithelial proliferation and barrier exposed to I/R injury. Material and methods Fifty Sprague-Dawley rats were divided into sham, I/R, I/R + EGCG (12.5 mg/kg), I/R + EGCG (25 mg/kg) and I/R + EGCG (50 mg/kg). I/R group rats were subjected to intestinal ischemia for 1 h and 6 h reperfusion. The rats were supplemented with EGCG 12.5, 25 and 50 mg/kg daily for 3 days via intraperitoneal injection before surgery. We used IEC-6 to expose to hypoxia/reoxygenation (H/R) injury to mimic I/R in vivo. IEC-6 cells were divided into control, H/R and H/R + EGCG (40 μmol/L). The effects of EGCG and its mechanism was explored. Results Pharmacological treatment with EGCG notably improves intestinal epithelial proliferation (12.5 mg/kg, 1.74-fold; 25 mg/kg, 2.93-fold, and 50 mg/kg, 4.33-fold) and barrier function after I/R injury. EGCG promoted cell proliferation (2.99-fold) and increased the expression of occludin (2.36-fold) and ZO-1 (1.64-fold) in IEC-6 cells after H/R injury. EGCG promoted proliferation of IEC-6 cells with ED50 values of 18.16 μmol/L. Further investigations indicated that EGCG activated Nurr1 expression in intestine after I/R injury. EGCG promote cell proliferation and increased the expression of occludin and ZO-1 in IEC-6 cells after H/R injury were abrogated in the knockdown of Nurr1 by siRNA. Discussion and conclusion Our findings indicate that EGCG promotes intestinal epithelial cell proliferation and barrier function after I/R injury in vitro and in vivo via activation of Nurr1.

MiR-379-5P INHIBITION ENHANCES INTESTINAL EPITHELIAL PROLIFERATION AND BARRIER FUNCTION RECOVERY AFTER ISCHEMIA/REPERFUSION BY TARGETING EIF4G2.

Gut barrier dysfunction caused by intestinal ischemia/reperfusion (I/R) injury is associated with substantial death and morbidity. In this research, the role of microRNAs (miRNAs) in regulating intestinal I/R injury was investigated. We used miRNA sequencing to analyze clinical ischemic and normal intestinal samples. Through bioinformatics analysis based on sequencing results, we found that upregulated miRNAs inhibited epithelial barrier function and cell proliferation, with miR-379-5p being the most significantly upregulated in the ischemic intestine. Further studies confirmed the role of miR-379-5p through experiments in the human ischemic intestine, the mouse I/R injury model in vivo , and cell hypoxia/reoxygenation models in vitro . Inhibiting miR-379-5p increased epithelial cell proliferation and improved barrier function after I/R injury. We also identified eukaryotic translation initiation factor 4 gamma 2 (EIF4G2) as a downstream target gene of miR-379-5p through bioinformatics prediction and experimental verification. The findings suggest that inhibiting miR-379-5p could improve intestinal epithelial cell proliferation and barrier function by targeting EIF4G2. The goal of this study was to find a potential target for treating I/R injury in the intestine, as well as to prevent and mitigate the damage caused.

Total ginsenosides promotes intestinal epithelial proliferation via affecting polyamine-mediated HuR on post-transcriptional control

Ginseng, as a functional food, is widely used worldwide because of its various benefits. This study aims to elucidate the potential effects of total ginsenosides (TG) in intestinal mucosal restitution. Establishing IEC-6 cell proliferation inhibition model by culturing with 5 mmol/L DL-α-difluoromethylornithine (DFMO, a polyamine inhibitor) for 48 h. The intestinal injury mice model was induced by 3% Dextran sodium sulfate (DSS) for 7 d. TG reversed the cell growth inhibition, regulated cell cycle and the decrease in c-Myc protein expression caused by DFMO, without affecting mRNA expression; TG increased translation efficiency and half-life of c-Myc mRNA, suggesting that TG promoted cell proliferation through affecting translation of c-Myc. Furthermore, TG could prevent the decrease of human antigen R (HuR)/c-Myc mRNA, p-HuR and p-Chk2 expression caused by DFMO, as well as reduce the expression of HuR in cytoplasm while increasing the expression in nucleus, indicating that TG influenced post-transcriptional control of c-Myc through polyamine-mediated HuR pathway for intestinal mucosal restitution. Panax ginseng aqueous extract (PGE) prevented intestinal mucosal injury induced by DSS(improved clinical symptoms and restored colon integrity) through enhancing HuR and c-Myc levels. Our results may provide a scientific basis for ginseng as a potential functional food ingredient and assistance therapeutic agent to prevent intestinal injury.

HucMSC-Exo Promote Mucosal Healing in Experimental Colitis by Accelerating Intestinal Stem Cells and Epithelium Regeneration via Wnt Signaling Pathway.

Mucosal healing has emerged as a crucial therapeutic goal for inflammatory bowel diseases (IBD). Exosomes (Exo) as a potential acellular candidate for stem cell therapy might be competent to promote mucosal healing, while its mechanism remains unexplored. Exosomes derived from human umbilical cord mesenchymal stem cells (hucMSCs) were subjected to experimental colitis mice intraperitoneally to estimate the role in mucosal healing and the regeneration of intestinal stem cells (ISCs) and epithelium. The intestinal organoid model of IBD was constructed utilizing tumor necrosis factor (TNF)-α for subsequent function analysis in vitro. Transcriptome sequencing was performed to decipher the underlying mechanism and Wnt-C59, an oral Wnt inhibitor, was used to confirm that further. Finally, the potential specific components of hucMSC‑exo were investigated based on several existing miRNA expression datasets. HucMSC-exo showed striking potential for mucosal healing in colitis mice, characterized by decreased histopathological injuries and neutrophil infiltration as well as improved epithelial integrity. HucMSC-exo up-regulated the expression of leucine-rich repeat-containing G-protein coupled receptor 5 (Lgr5), a specific marker for ISCs and accelerated the proliferation of intestinal epithelium. HucMSC-exo endowed intestinal organoids with more excellent capacity to grow and bud under TNF-α stimulation. More than that, the fact that hucMSC-exo activated the canonical Wnt signaling pathway to promote mucosal healing was uncovered by not only RNA-sequencing but also relevant experimental data. Finally, bioinformatics analysis of the existing miRNA expression datasets indicated that several miRNAs abundant in hucMSC-exo involved widely in regeneration or repair related biological processes and Wnt signaling pathway might be one of the most important signal transduction pathways. Our results suggested that hucMSC-exo could facilitate mucosal healing in experimental colitis by accelerating ISCs and intestinal epithelium regeneration via transferring key miRNAs, which was dependent on the activation of Wnt/β-catenin signaling pathway.