Intestinal Epithelial Turnover Research Articles

Propionate Enhances Cell Speed and Persistence to Promote Intestinal Epithelial Turnover and Repair.

Background and AimsGut bacteria-derived short-chain fatty acids (SCFAs) play crucial roles in the maintenance of intestinal homeostasis. However, how SCFAs regulate epithelial turnover and tissue repair remain incompletely understood. In this study, we investigated how the SCFA propionate regulates cell migration to promote epithelial renewal and repair.MethodsMouse small intestinal epithelial cells (MSIE) and human Caco-2 cells were used to determine the effects of SCFAs on gene expression, proliferation, migration, and cell spreading in vitro. Video microscopy and single cell tracking were used to assess cell migration kinetically. 5-bromo-2’-deoxyuridine (BrdU) and hydroxyurea were used to assess the effects of SCFAs on migration in vivo. Lastly, an acute colitis model using dextran sulfate sodium (DSS) was used to examine the effects of SCFAs in vivo.ResultsUsing video microscopy and single cell tracking, we found that propionate promoted intestinal epithelial cell migration by enhancing cell spreading and polarization, which led to increases in both cell speed and persistence. This novel function of propionate was dependent on inhibition of class I histone deacetylases (HDAC) and GPR43 and required signal transducer and activator of transcription 3 (STAT3). Furthermore, using 5-bromo-2’-deoxyuridine (BrdU) and hydroxyurea in vivo, we found that propionate enhanced cell migration up the crypt-villus axis under homeostatic conditions, while also protecting against ulcer formation in experimental colitis.ConclusionOur results demonstrate a mechanism by which propionate stimulates cell migration in an HDAC inhibition, GPR43, and STAT3 dependent manner, and suggest that propionate plays an important role in epithelial migration independent of proliferation.

The role of single cell mechanical behavior and polarity in driving collective cell migration.

The directed migration of cell collectives is essential in various physiological processes, such as epiboly, intestinal epithelial turnover, and convergent extension during morphogenesis as well as during pathological events like wound healing and cancer metastasis. Collective cell migration leads to the emergence of coordinated movements over multiple cells. Our current understanding emphasizes that these movements are mainly driven by large-scale transmission of signals through adherens junctions. In this study, we show that collective movements of epithelial cells can be triggered by polarity signals at the single cell level through the establishment of coordinated lamellipodial protrusions. We designed a minimalistic model system to generate one-dimensional epithelial trains confined in ring shaped patterns that recapitulate rotational movements observed in vitro in cellular monolayers and in vivo in genitalia or follicular cell rotation. Using our system, we demonstrated that cells follow coordinated rotational movements after the establishment of directed Rac1-dependent polarity over the entire monolayer. Our experimental and numerical approaches show that the maintenance of coordinated migration requires the acquisition of a front-back polarity within each single cell but does not require the maintenance of cell-cell junctions. Taken together, these unexpected findings demonstrate that collective cell dynamics in closed environments as observed in multiple in vitro and in vivo situations can arise from single cell behavior through a sustained memory of cell polarity.

Pasteurized Akkermansia muciniphila increases whole-body energy expenditure and fecal energy excretion in diet-induced obese mice

ABSTRACT Accumulating evidence points to Akkermansia muciniphila as a novel candidate to prevent or treat obesity-related metabolic disorders. We recently observed, in mice and in humans, that pasteurization of A. muciniphila increases its beneficial effects on metabolism. However, it is currently unknown if the observed beneficial effects on body weight and fat mass gain are due to specific changes in energy expenditure. Therefore, we investigated the effects of pasteurized A. muciniphila on whole-body energy metabolism during high-fat diet feeding by using metabolic chambers. We confirmed that daily oral administration of pasteurized A. muciniphila alleviated diet-induced obesity and decreased food energy efficiency. We found that this effect was associated with an increase in energy expenditure and spontaneous physical activity. Strikingly, we discovered that energy expenditure was enhanced independently from changes in markers of thermogenesis or beiging of the white adipose tissue. However, we found in brown and white adipose tissues that perilipin2, a factor associated with lipid droplet and known to be altered in obesity, was decreased in expression by pasteurized A. muciniphila. Finally, we observed that treatment with pasteurized A. muciniphila increased energy excretion in the feces. Interestingly, we demonstrated that this effect was not due to the modulation of intestinal lipid absorption or chylomicron synthesis but likely involved a reduction of carbohydrates absorption and enhanced intestinal epithelial turnover. In conclusion, this study further dissects the mechanisms by which pasteurized A. muciniphila reduces body weight and fat mass gain. These data also further support the impact of targeting the gut microbiota by using specific bacteria to control whole-body energy metabolism.

Zinc L-Aspartate enhances intestinal stem cell activity to protect the integrity of the intestinal mucosa against deoxynivalenol through activation of the Wnt/β-catenin signaling pathway

The micronutrient, zinc, plays a vital role in modulating cellular signaling recognition and enhancing intestinal barrier function. However, the precise mechanisms underlying the zinc regulation of intestinal stem cell (ISC) renewal and regeneration ability, which drive intestinal epithelial turnover to maintain the intestinal barrier, under physiological and pathological conditions are unknown. In this study, we used in vivo mouse plus ex vivo enteroid model to investigate thoroughly the protection efficacy of zinc L-aspartate (Zn-Asp) on intestinal mucosal integrity exposed to deoxynivalenol (DON). The results showed that 10 rather than 20 mg/kg body weight (BW) Zn-Asp (calculation in zinc) significantly increased the jejunum mass and ameliorated mucosa injury caused by 2 mg/kg BW DON treatment, including improvement of the intestinal morphology and barrier, as well as enteroid-forming and -budding efficiency, which was expanded from crypt cells isolated from jejunum of mice in each group. The repair process stimulated by Zn-Asp was also accompanied by increased fluorescence signal intensity of KRT20 and Villin; increased numbers of MUC2+, CAG+, LYZ+, BrdU+ and Ki67+ cells in mouse jejunum; and protein expression of Ki67 and PCNA in the jejunum, crypt and enteroid. Simultaneously, Zn-Asp increased ISC activity to promote intestinal epithelial renewal even under physiological conditions. These results were further verified in ex vivo enteroid culture experiments, which were treated with 100 μmol/L Zn-Asp (calculation in zinc) and 100 ng/mL DON for 72 h. Furthermore, we demonstrated that Zn-Asp improved intestinal integrity or accelerated wound healing along with Wnt/β-catenin signaling upregulation or reactivation. Our findings indicate Zn-Asp, especially Zn, enhances ISC activity to maintain the intestinal integrity by activating the Wnt/β-catenin signaling, which sheds some light upon effective preventive strategies for intestinal injury induced by mycotoxin based on ISCs with exogenous zinc preparations in the proper drugs, health foods or qualified feed.

Atg14 protects the intestinal epithelium from TNF-triggered villus atrophy

ABSTRACTRegulation of intestinal epithelial turnover is a key component of villus maintenance in the intestine. The balance of cell turnover can be perturbed by various extrinsic factors including the cytokine TNF, a cell signaling protein that mediates both proliferative and cytotoxic outcomes. Under conditions of infection and damage, defects in autophagy are associated with TNF-mediated cell death and tissue damage in the intestinal epithelium. However, a direct role of autophagy within the context of enterocyte cell death during homeostasis is lacking. Here, we generated mice lacking ATG14 (autophagy related 14) within the intestinal epithelium [Atg14f/f Vil1-Cre (VC)+]. These mice developed spontaneous villus loss and intestinal epithelial cell death within the small intestine. Based on marker studies, the increased cell death in these mice was due to apoptosis. Atg14f/f VC+ intestinal epithelial cells demonstrated sensitivity to TNF-triggered apoptosis. Correspondingly, both TNF blocking antibody and genetic deletion of Tnfrsf1a/Tnfr1 rescued villus loss and cell death phenotype in Atg14f/f VC+ mice. Lastly, we identified a similar pattern of spontaneous villus atrophy and cell death when Rb1cc1/Fip200 was conditionally deleted from the intestinal epithelium (Rb1cc1f/f VC+). Overall, these findings are consistent with the hypothesis that factors that control entry into the autophagy pathway are also required during homeostasis to prevent TNF triggered death in the intestine.Abbreviations: ANOVA: analysis of variance; Atg14: autophagy related 14; Atg16l1: autophagy related 16-like 1 (S. cerevisiae); Atg5: autophagy related 5; cCASP3: cleaved CASP3/caspase-3; cCASP8: cleaved CASP8/caspase-8; CHX: cycloheximide; EdU: 5-ethynyl-2´-deoxyuridine thymidine; f/f: flox/flox; H&E: hematoxylin and eosin; MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; Nec-1: necrostatin-1; Rb1cc1/Fip200: RB1-inducible coiled-coil 1; Ripk1: receptor (TNFRSF)-interacting serine-threonine kinase 1; Ripk3: receptor (TNFRSF)-interacting serine-threonine kinase 3; Tnfrsf1a/Tnfr1: tumor necrosis factor receptor superfamily, member 1a; Tnf/ Tnfsf1a: tumor necrosis factor; VC: Vil1/villin 1-Cre

Gastrointestinal iron excretion and reversal of iron excess in a mouse model of inherited iron excess.

The current paradigm in the field of mammalian iron biology states that body iron levels are determined by dietary iron absorption, not by iron excretion. Iron absorption is a highly regulated process influenced by iron levels and other factors. Iron excretion is believed to occur at a basal rate irrespective of iron levels and is associated with processes such as turnover of intestinal epithelium, blood loss, and exfoliation of dead skin. Here we explore iron excretion in a mouse model of iron excess due to inherited transferrin deficiency. Iron excess in this model is attributed to impaired regulation of iron absorption leading to excessive dietary iron uptake. Pharmacological correction of transferrin deficiency not only normalized iron absorption rates and halted progression of iron excess but also reversed body iron excess. Transferrin treatment did not alter the half-life of 59Fe in mutant mice. 59Fe-based studies indicated that most iron was excreted via the gastrointestinal tract and suggested that iron-loaded mutant mice had increased rates of iron excretion. Direct measurement of urinary iron levels agreed with 59Fe-based predictions that urinary iron levels were increased in untreated mutant mice. Fecal ferritin levels were also increased in mutant mice relative to wild-type mice. Overall, these data suggest that mice have a significant capacity for iron excretion. We propose that further investigation into iron excretion is warranted in this and other models of perturbed iron homeostasis, as pharmacological targeting of iron excretion may represent a novel means of treatment for diseases of iron excess.

Coinfection with Eimeria spp. decreases bacteremia and spinal lesions caused by pathogenic Enterococcus cecorum

Pathogenic strains of Enterococcus cecorum (EC) escape the gut niche to infect the spine of broilers at the free thoracic vertebra (FTV) causing the disease enterococcal spondylitis or ‘kinky-back’. Intestinal barrier damage caused by coinfection with Eimeria spp. has been suggested to play a role in potentiating EC bacteremia and FTV lesion development. To test this hypothesis, 1440 broilers were experimentally infected with EC only, EC and a coinfection of E. acervulina, E. maxima and E. tenella (EC:Cocci), and a saline only control (Sham). Birds were grown for 35 days, spleen cultures, histologic lesions in the FTV and live performance parameters were compared among groups. Coccidian coinfection significantly decreased the prevalence of EC bacteremia and histologic lesions in the FTV. Histologic evaluation of the ceca revealed significantly increased cecal mucosal height and mean inflammatory scores in the EC:Cocci group compared to EC only and sham inoculated controls. These findings indicate that the decrease in pathogenic EC bacteremia observed with coccidia coinfection may be due to increased intestinal epithelial turnover or increased immune surveillance of the intestine. In both infection groups, body weights, body weight gain and feed intake were significantly decreased and feed conversion ratios were significantly increased. These undesirable alterations in live performance parameters were exacerbated by nicarbazin treatment but not zoalene or bacitracin treatment. Further work is needed to determine the mechanism for the observed benefit of coccidian coinfection in decreasing bacteremia and FTV lesions due to pathogenic EC.

Vitamin D Improves Intestinal Barrier Function in Cirrhosis Rats by Upregulating Heme Oxygenase-1 Expression.

Intestinal barrier dysfunction always accompanies cirrhosis in patients with advanced liver disease and is an important contributor facilitating bacterial translocation (BT), which has been involved in the pathogenesis of cirrhosis and its complications. Several studies have demonstrated the protective effect of Vitamin D on intestinal barrier function. However, severe cholestasis leads to vitamin D depletion. This study was designed to test whether vitamin D therapy improves intestinal dysfunction in cirrhosis. Rats were subcutaneously injected with 50% sterile CCl4 (a mixture of pure CCl4 and olive oil, 0.3 mL/100 g) twice a week for 6 weeks. Next, 1,25(OH)2D3 (0.5 µg/100 g) and the vehicle were administered simultaneously with CCl4 to compare the extent of intestinal histologic damage, tight junction protein expression, intestinal barrier function, BT, intestinal proliferation, apoptosis, and enterocyte turnover. Intestinal heme oxygenase-1 (HO-1) expression and oxidative stress were also assessed. We found that vitamin D could maintain intestinal epithelial proliferation and turnover, inhibit intestinal epithelial apoptosis, alleviate structural damage, and prevent BT and intestinal barrier dysfunction. These were achieved partly through restoration of HO-1 and inhibition of oxidative stress. Taken together, our results suggest that vitamin D ameliorated intestinal epithelial turnover and improved the integrity and function of intestinal barrier in CCl4-induced liver cirrhotic rats. HO-1 signaling activation was involved in these above beneficial effects.

Long-term effect of parasympathetic or sympathetic denervation on intestinal epithelial cell proliferation and apoptosis

Intestinal epithelial tissue is constantly regenerated as a means to maintain proper tissue function. Previous studies have demonstrated that denervation of the parasympathetic or sympathetic nervous system to the intestine alters this process. However, results are inconsistent between studies, showing both increases and decreases in proliferation after denervation of the parasympathetic or sympathetic. The effect appears to correlate with (1) the timing post-denervation, (2) denervation-induced changes in food intake, (3) the denervation technique used, and (4) which intestinal segment is investigated. Thus, we proposed that parasympathetic or sympathetic denervation does not have an effect on intestinal epithelial regeneration when you (1) evaluate denervation after long-term denervation, (2) control for post-surgical changes in food intake, (3) use minimally invasive surgical techniques and (4) include a segmental analysis. To test this, adult male Sprague Dawley rats underwent parasympathetic denervation via subdiaphragmatic vagotomy, sympathetic denervation via celiacomesenteric ganglionectomy, a parasympathetic denervation sham surgery, or a sympathetic denervation sham surgery. Sham surgery ad libitum-fed groups and sham surgery pair-fed groups were used to control for surgically induced changes in food intake. Three weeks post-surgery, animals were sacrificed and tissue from the duodenum, jejunum, and ileum was excised and immunohistochemically processed to visualize indicators of proliferation (bromodeoxyuridine-positive cells) and apoptosis (caspase-3-positive cells). Results showed no differences between groups in proliferation, apoptosis, or total cell number in any intestinal segment. These results suggest that parasympathetic or sympathetic denervation does not have a significant long-term effect on intestinal epithelial turnover. Thus, intestinal epithelial regeneration is able to recover after autonomic nervous system injury. Impact statement This study investigates the long-term effect of autonomic denervation on intestinal epithelial cell turnover, as measured by proliferation, apoptosis, and total cell number. Although previous research has established that autonomic denervation can alter intestinal epithelial turnover under short-term conditions, here we establish for the first time that these changes do not persist long-term when you control for surgical-induced changes in food intake and use targeted denervation procedures. These findings add to the base of knowledge on autonomic control of tissue turnover, highlight the ability of the intestinal epithelium to recover after autonomic injury and reveal possible implications of the use of ANS denervation for disease treatment in humans.

TWEAK/Fn14 Pathway: A Nonredundant Role in Intestinal Damage in Mice Through a TWEAK/Intestinal Epithelial Cell Axis

Tumor necrosis factor (TNF) superfamily members have attracted attention as new therapeutic targets for treating inflammatory disease. TNF-like weak inducer of apoptosis (TWEAK) is a unique, multifunctional TNF family cytokine that signals through its receptor, fibroblast growth factor-inducible molecule 14 (Fn14). The role of this pathway in the intestine has not been previously reported. The 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis model was conducted in TWEAK- or Fn14-deficient mice or in normal mice treated with a TWEAK-blocking monoclonal antibody, and clinical severity, histopathology, immunohistochemistry for cell infiltrates, TWEAK and Fn14, gene expression profiling in the colon, and systemic adaptive immunity were assessed. The effect of TWEAK on colon epithelial cell production of inflammatory mediators was analyzed in vitro. The gamma-irradiation injury model was conducted in TWEAK- or Fn14-deficient mice, and crypt epithelial death was assessed. Colitis severity and histologic scores were significantly reduced by TWEAK pathway deficiency or TWEAK-blocking monoclonal antibody. Neutrophil and macrophage infiltrates, chemokines, cytokines, and matrix metalloproteinase expression were reduced in the TWEAK-deficient colon after TNBS administration; however, systemic adaptive immune responses to trinitrophenyl were not altered. Fn14 is expressed on colon epithelial cells in TNBS colitis, and TWEAK induces epithelial production of pathogenic mediators. TWEAK also regulates intestinal epithelial turnover, as evidenced by reduced epithelial cell death after gamma-irradiation injury in TWEAK and Fn14 knockout mice. Our studies elucidate a nonredundant TWEAK-intestinal epithelial cell axis and suggest that blocking TWEAK may dampen chronic intestinal inflammation and allow normal epithelial repair.

In vivo effects of thyroid hormone, corticosteroids and prolactin on cell proliferation and apoptosis in the anterior intestine of the euryhaline mudskipper ( Periophthalmus modestus)

We have previously shown that anterior intestinal epithelium of the euryhaline mudskipper ( Periophthalmus modestus) undergoes apoptosis during seawater (SW) acclimation, whereas elevated cell proliferation was observed in freshwater (FW)-acclimated fish. To understand the possible endocrine regulation of the gastrointestinal cell turnover during salinity acclimation, we examined the ratios of apoptotic and proliferating cells in the anterior intestine of one-third SW-acclimated mudskipper treated with triiodothyronine (T 3), cortisol, 11-deoxycorticosterone (DOC, the putative teleostean mineralocorticoid), or prolactin (PRL). In situ nick end labeling of genomic DNA (TUNEL) and immunohistochemistry of proliferating cells nuclear antigen (PCNA) were used as indicators of apoptosis and cell proliferations, respectively. Cortisol significantly elevated apoptosis ( P < 0.05) in the epithelia and connective tissues and also stimulated the epithelial cell proliferation ( P < 0.05). PRL induced epithelial cell proliferation ( P < 0.05), but did not affect apoptotic status of the intestinal epithelium. Neither T 3 nor DOC had any impact on cell proliferation or apoptosis. Together, our results suggest a role for cortisol and PRL in the regulation of anterior intestinal epithelial turnover during salinity acclimation in this species.

A Role for Membrane-Type Serine Protease (MT-SP1) in Intestinal Epithelial Turnover

Membrane type-serine protease 1 (MT-SP1) plays potential roles in the process of invasion and metastasis of carcinomas. In the present study, we cloned a rat MT-SP1 cDNA and investigated the intestinal distribution and proteolytic properties of the enzyme. By in situ hybridization we found the prominent expression of the mRNA in the epithelial layer of the small intestinal upper villi and of the colon, where cells are loosely attached to the basement membrane. When MT-SP1 was expressed in Caco-2, a colonic carcinoma cell line, the protein was localized exclusively on the basolateral side. A secreted form of the enzyme produced in COS-1 cells digested fibronectin and laminin. These findings suggest that MT-SP1 participates in the control of intestinal epithelial turnover by regulating the cell-substratum adhesion.

The Turnover of Intestinal Epithelium in the Chick

THE cells of the intestinal epithelium originate in the crypts of Lieberkuhn and migrate up the sides of the villi. When they reach the tips (extrusion zone) of the villi the cells are extruded into the lumen of the intestine. The mechanism of migration is not fully understood, but the movement has been shown to be rapid. Leblond and Stevens (1948) reported that replacement of the epithelium in rats took 1.57 days in the duodenum and 1.35 days in the ileum. In cats, the renewal time was found to be 2.25 and 2.75 days for the duodenum and ileum, respectively (McMinn, 1954). In human duodenum, a renewal time of 1.8 days was reported by Bertalanffy and Nagy (1958).As part of investigations on avian digestion and absorption, the turnover of intestinal epithelium in chicks was studied. The movement of intestinal mucosal cells was followed using microradioautography after injecting chicks with …