Intestinal Epithelial Cell Apoptosis Research Articles

Olfactomedin-4 deletion exacerbates DSS-induced colitis through a matrix metalloproteinase-9-dependent mechanism.

Background and Aims: Olfactomedin-4 is a glycoprotein that is upregulated in inflamed gastrointestinal tissues. This study aimed to investigate the role and underlying mechanisms of olfactomedin-4 in ulcerative colitis. Methods: C57BL/6 mice and olfactomedin-4 knockout mice were fed dextran sulfate sodium in drinking water to establish a colitis model. An in vitro inflammation model was constructed in HCT116 and NCM460 cells stimulated with lipopolysaccharide. The expression of olfactomedin-4 was detected by Western blotting, immunohistochemistry staining, and qRT‒PCR. The differences in the severity of colitis between olfactomedin-4 knockout mice and wild-type mice were compared, and the underlying mechanisms were explored. Results: Olfactomedin-4 expression was significantly upregulated in colonic tissues of active ulcerative colitis patients and in cellular and mouse models of colitis. Compared with wild-type littermates, olfactomedin-4 knockout mice were more susceptible to dextran sulfate sodium-induced colitis and produced higher levels of proinflammatory cytokines and chemokines. In addition, olfactomedin-4 deficiency significantly promoted intestinal epithelial cell apoptosis and increased intestinal permeability, which was mediated by the p53 pathway. Moreover, olfactomedin-4 directly interacted with and negatively regulated matrix metalloproteinase-9. Inhibiting matrix metalloproteinase-9 significantly decreased colonic p53 expression and ameliorated experimental colitis in olfactomedin-4 knockout mice, while overexpression of matrix metalloproteinase-9 aggravated colitis. Further experiments showed that matrix metalloproteinase-9 regulated p53 through the Notch1 signaling pathway to promote ulcerative colitis progression. Conclusions: Olfactomedin-4 is significantly upregulated in ulcerative colitis and may protect against colitis by directly inhibiting matrix metalloproteinase-9 and further decreasing p53-mediated apoptosis via Notch1 signaling.

Dexmedetomidine alleviates intestinal barrier dysfunction and inflammatory response in mice via suppressing TLR4/MyD88/NF-κB signaling in an experimental model of ulcerative colitis.

Ulcerative colitis (UC) is a nonspecific intestinal inflammatory disease. Dexmedetomidine (DEX) is a selective alpha 2-adrenergic receptor agonist commonly used for analgesia and sedation in intensive care units. Herein, the role and mechanism of DEX in dextran sulfate sodium (DSS)-induced colitis was explored. A murine model of DSS-induced colitis was established by adding 3.5% (w/v) DSS in drinking water to C57BL/6J female mice. The severity of colitis was measured by the disease activity index (DAI) score, colon length and body weight of mice. The serum concentration and mRNA levels of inflammatory cytokines in colon tissues were assessed by ELISA and RT-qPCR, respectively. Protein levels of apoptotic markers, tight junction proteins and genes involved in the TLR4/MyD88/NF-κB signaling were quantified utilizing Western blotting. The pathological changes of colon tissues were evaluated by hematoxylin-eosin (HE) staining and histological score. Intestinal permeability in vivo was assessed by fluorescein isothiocyanate (FITC)-dextran (FITC-D) administration. TUNEL assay was used to determine cell apoptosis in the intestinal epithelium. DSS administration resulted in weight loss, shortening of the colon, increased DAI score, histological abnormalities, and increased serum FITC-D levels in mice, all of which were reversed by DEX injection. Moreover, DEX attenuated DSS-triggered inflammatory response, intestinal barrier injury and intestinal epithelial cell apoptosis. Mechanically, DEX inactivated the TLR4/MyD88/NF-κB signaling in the colon tissues. DEX exerts beneficial effects against the intestinal barrier dysfunction, inflammatory response, and apoptosis of intestinal epithelial cells via inactivation of the TLR4/MyD88/NF-κB signaling in mice with DSS-induced colitis.

Glutathione Protects against Paraquat-Induced Oxidative Stress by Regulating Intestinal Barrier, Antioxidant Capacity, and CAR Signaling Pathway in Weaned Piglets.

Endogenous glutathione (GSH) effectively regulates redox homeostasis in the body. This study aimed to investigate the regulatory mechanism of different dietary levels of GSH supplementation on the intestinal barrier and antioxidant function in a paraquat-induced stress-weaned piglet model. Our results showed that dietary 0.06% GSH supplementation improved the growth performance of weaned piglets under normal and stressful conditions to some degree and decreased the diarrhea rate throughout. Exogenous GSH improved paraquat-induced changes in intestinal morphology, organelle, and permeability and reduced intestinal epithelial cell apoptosis. Moreover, GSH treatment alleviated intestinal oxidative stress damage by upregulating antioxidant (GPX4, CnZnSOD, GCLC, and GCLM) and anti-inflammatory (IL-10) gene expression and downregulating inflammatory cytokines (IFN-γ and IL-12) gene expression. Furthermore, GSH significantly reduced the expression levels of constitutive androstane receptor (CAR), RXRα, HSP90, PP2Ac, CYP2B22, and CYP3A29, and increased the expression levels of GSTA1 and GSTA2 in the jejunum and ileum of paraquat-induced piglets. We conclude that exogenous GSH protects against oxidative stress damage by regulating the intestinal barrier, antioxidant capacity, and CAR signaling pathway.

Deoxynivalenol hijacks the pathway of Janus kinase 2/signal transducers and activators of transcription 3 (JAK2/STAT-3) to drive caspase-3-mediated apoptosis in intestinal porcine epithelial cells

Deoxynivalenol (DON) can easily injure the intestinal tract, which represents the first barrier against food contaminants. The intestinal toxicity induced by DON was mainly focused on mitogen-activated protein kinase (MAPK) activation, however, the underlying mechanisms by which DON triggers apoptosis by other pathways remain poorly understood. In this study, the Janus kinase 2/signal transducers and activators of transcription 3 (JAK2/STAT-3) pathway was proposed to regulate the intrinsic apoptosis induced by DON and thoroughly investigated in intestinal porcine epithelial cells (IPEC-J2). First, DON was found to be able to efficiently inhibit cell viability and increase the release of lactate dehydrogenase. It could also enhance the activity of the cleaved caspase-3 in a time-dependent manner, accompanied by a loss of mitochondrial membrane potential and an up-regulation of the apoptosis rate. Then, the expression of genes associated with inflammation and apoptosis were investigated. DON increased the expression of IL-6, IL-1β, TNF-α, SOCS3 and Bax, but decreased the expression of Bcl-2 and Bcl-xL. Moreover, we discovered that DON robustly inhibited STAT-3 activity together with the down-regulation of JAK2, Bcl-2 and Bcl-xL, paralleling the increase in p38 phosphorylation. Furthermore, a pharmacological activation of JAK2/STAT-3 alleviated DON induced-apoptosis. Concurrent with the apoptotic pathway, during the initial exposure to DON (first 4 h), a survival pathway involving phosphorylated Erk1/2, Akt, and FoxO1 was also observed. Thus, apoptosis induced by DON was Janus faced: although the survival pathway was activated, the DON-induced apoptotic JAK2/STAT-3/caspase-3 pathway dominated, leading to an imbalance in cell homeostasis. This study provides a novel avenue to comprehensively reveal the pathological mechanisms of DON-induced intestinal disorders, which is promising for future applications to other contaminants in food and feed.

Fusobacterium nucleatum aggravates ulcerative colitis through promoting gut microbiota dysbiosis and dysmetabolism.

The correlation between periodontitis and ulcerative colitis (UC) has drawn widespread attention recently. Fusobacterium nucleatum (F. nucleatum) as a periodontal pathogen also has reservoirs in gut and may play a role in intestinal diseases. However, its role in the pathogenesis of UC is unclear. Mice were orally given dextran sulphate sodium (DSS) solution and F. nucleatum to construct experimental models. The survival rate, weight, and disease activity index (DAI) of mice were monitored. Alveolar bone loss, abundance of F. nucleatum in colon, colon length, histopathological assessment, and inflammatory cytokines were detected. Apoptosis of intestinal epithelial cells (IECs) were evaluated by TUNEL assay and pro-apoptotic gene Bax. The epithelial barrier function was assessed by tight junction proteins. By 16S rRNA gene sequencing and LC-MS-based methods, the composition of the intestinal microbiota and metabolites in mice were analyzed. F. nucleatum facilitated alveolar bone loss and colonized only in infected colon tissue. Mice fed with DSS showed destruction of gut structure, increased expressions of interleukin one-beta (IL-1β) and tumor necrosis factor alpha (TNF-α), decreased expression of IL-10, higher apoptosis of IECs, microbiota dysbiosis and bile acid dysmetabolism compared to healthy ones. F. nucleatum further aggravated intestinal inflammation and epithelial barrier damage. Probiotics such as Bifidobacterium and Faecalibacterium decreased, opportunistic pathogensEscherichia-Shigella increased and the differential microorganisms highly associated with inflammatory parameters and metabolites. Meanwhile, level of uric acid involving in the purine metabolism significantly elevated compared to UC mice. F. nucleatum promotes gut inflammation, epithelial barrier dysfunction, microbiota dysbiosis and dysmetabolism to aggravate UC.

The Polo-Like Kinase 1–Mammalian Target of Rapamycin Axis Regulates Autophagy to Prevent Intestinal Barrier Dysfunction During Sepsis

The intestines play a crucial role in the development of sepsis. The balance between autophagy and apoptosis in intestinal epithelial cells is dynamic and determines intestinal permeability. The present study focused on the potential role of autophagy in sepsis-induced intestinal barrier dysfunction and explored the mechanisms invivo and invitro. Excessive apoptosis in intestinal epithelia and a disrupted intestinal barrier were observed in septic mice. Promoting autophagy with rapamycin reduced intestinal epithelial apoptosis and restored intestinal barrier function, presenting as decreased serum diamine oxidase (DAO) and fluorescein isothiocyanate-dextran 40 (FD40) levels and increased expression of zonula occludens-1 (ZO-1) and Occludin. Polo-like kinase 1 (PLK1) knockdown in mice ameliorated intestinal epithelial apoptosis and the intestinal barrier during sepsis, whereas these effects were reduced with chloroquine and enhanced with rapamycin. PLK1 also promoted cell autophagy and improved lipopolysaccharide-induced apoptosis and high permeability invitro. Moreover, PLK1 physically interacted with mammalian target of rapamycin (mTOR) and participated in reciprocal regulatory crosstalk in intestinal epithelial cells during sepsis. This study provides novel insight into the role of autophagy in sepsis-induced intestinal barrier dysfunction and indicates that the PLK1-mTOR axis may be a promising therapeutic target for sepsis.

Hypoxic mesenchymal stem cell-derived exosomes alleviate ulcerative colitis injury by limiting intestinal epithelial cells reactive oxygen species accumulation and DNA damage through HIF-1α.

Ulcerative colitis (UC) is an inflammatory disease with a complex pathogenic mechanism. Mounting evidence suggests that UC pathogenesis is linked to excessive production of reactive oxygen species (ROS) and cellular DNA damage. Recent studies have shown that bone mesenchymal stem cells (BMSCs) mainly exert their therapeutic effects through paracrine exosomes, and oxygen concentration is extremely important to BMSCs and exosomes. The main objective of this study was to determine whether exosomes from BMSCs under hypoxic conditions (HP-Exos) exhibit a greater therapeutic effect on UC compared to exosomes under normoxic conditions (Exos) and to resolve the mechanism of HP-Exos. We observed that hypoxia enhances the activity and migration of BMSCs and inhibits BMSC apoptosis without changing their morphological characteristics. Furthermore, HP-Exos significantly relieved UC symptoms and pathological damage. In order to further understand the mechanism of HP-Exos in UC, findings from in vivo experiments demonstrated that HP-Exos reduces ROS production, DNA damage and apoptosis in intestinal epithelial cells. As hypoxia-inducible factor 1α (HIF-1α) plays an important role in hypoxia, we knocked down HIF-1α in BMSCs. HIF-1α knockout reversed the effects of hypoxia on the activity, migration and apoptosis of BMSCs. Moreover, inhibition of HIF-1α expression also reversed the regulation of UC by HP-Exos. Therefore, we conclude that HP-Exos regulates ROS accumulation, DNA damage and immune homeostasis in intestinal epithelial cells via HIF-1α.

Modulation of intestinal epithelial cell proliferation and apoptosis by Lactobacillus gasseri SF1183

The gut microbiota exerts a variety of positive effects on the intestinal homeostasis, including the production of beneficial molecules, control of the epithelial barrier integrity and the regulation of the balance between host’s cell death and proliferation. The interactions between commensal bacteria and intestinal cells are still under-investigated and is then of paramount importance to address such interactions at the molecular and cellular levels. We report an in vitro analysis of the effects of molecules secreted by Lactobacillus gasseri SF1183 on HCT116 cells, selected as a model of intestinal epithelial cells. SF1183 is a L. gasseri strain isolated from an ileal biopsy of a human healthy volunteer, able to prevent colitis symptoms in vivo. Expanding previous findings, we show that bioactive molecules secreted by SF1183 reduce the proliferation of HCT116 cells in a reversible manner determining a variation in cell cycle markers (p21WAF, p53, cyclin D1) and resulting in the protection of HCT116 cells from TNF-alfa induced apoptosis, an effect potentially relevant for the protection of the epithelial barrier integrity and reconstitution of tissue homeostasis. Consistently, SF1183 secreted molecules increase the recruitment of occludin, a major component of TJ, at the cell–cell contacts, suggesting a reinforcement of the barrier function.

Postbiotic muramyl dipeptide alleviates colitis via activating autophagy in intestinal epithelial cells.

The pathogenesis of IBD is complicated and still unclear. Nucleotide-binding oligomerization domain 2 (NOD2) plays a significant role in regulating gut inflammation under the activation of muramyl dipeptide (MDP), which is used as a postbiotic. The study aimed to investigate the effect of MDP on the intestinal barrier in colitis and the mechanism involved. In this study, C57BL/6 mice were challenged with dextran sodium sulfate (DSS) for establishing a colitis model with the pre-treatment of MDP in vivo. Intestinal permeability was reflected by detecting the serum concentration of 4kDa Fluorescein Isothiocyanate-Dextran. The expression of inflammation, barrier-related proteins, and autophagy was tested by Western Blotting. Proliferation and apoptosis in intestinal epithelial cells were detected by immunohistochemistry. Caco-2 cells were exposed to lipopolysaccharide for imitating inflammation in vitro. The findings showed that administration of MDP ameliorated losses of body weight loss, gross injury, and histology score of the colon in the DSS-induced colitis mice. MDP significantly ameliorated the condition of gut permeability, and promoted intestinal barrier repair by increasing the expression of Zonula occludens-1 and E-cadherin. Meanwhile, MDP promoted proliferation and reduced apoptosis of intestinal epithelial cells. In the experiment group treated with MDP, LC3 was upregulated, and p62 was downregulated, respectively. These results suggested that MDP stimulation attenuates intestinal inflammation both in vivo and in vitro. Potentially, MDP reduced the intestinal barrier damage by regulating autophagy in intestinal epithelial cells. Future trials investigating the effects of MDP-based postbiotics on IBD may be promising.

Fermented Astragalus and its metabolites regulate inflammatory status and gut microbiota to repair intestinal barrier damage in dextran sulfate sodium-induced ulcerative colitis

Fermentation represents an efficient biotechnological approach to increase the nutritional and functional potential of traditional Chinese medicine. In this study, Lactobacillus plantarum was used to ferment traditional Chinese medicine Astragalus, the differential metabolites in the fermented Astragalus (FA) were identified by ultra-performance liquid chromatography-Q Exactive hybrid quadrupole-Orbitrap mass spectrometry (UPLC-Q-Exactive-MS), and the ameliorating effect of FA on dextran sulfate sodium (DSS)-induced colitis in mice were further explored. The results showed that 11 differential metabolites such as raffinose, progesterone and uridine were identified in FA, which may help improve the ability of FA to alleviate colitis. Prophylactic FA supplementation effectively improved DAI score, colon length and histopathological lesion in DSS-treated mice. The abnormal activation of the intestinal immune barrier in mice was controlled after FA supplementation, the contents of myeloperoxidase (MPO) and IgE were reduced and the contents of IgA were increased. The intestinal pro-inflammatory factors TNF-α, IL-1β, IL-6, and IL-17 were down-regulated and the anti-inflammatory factors IL-10 and TGF-β were up-regulated, suggesting that FA can intervene in inflammatory status by regulating the balance of Th1/Th2/Th17/Treg related cytokines. In addition, FA supplementation modified the structure of the intestinal microbiota and enriched the abundance of Akkermansia and Alistipes, which were positively associated with the production of short-chain fatty acids. These microbes and their metabolites induced by FA also be involved in maintaining the intestinal mucosal barrier integrity by affecting mucosal immunity. We observed that intestinal tight junction protein and mucous secreting protein ZO-1, occludin, and MUC2 genes expression were more pronounced in mice supplemented with FA compared to unfermented Astragalus, along with modulation of intestinal epithelial cells (IECs) apoptosis, verifying the intestinal mucosal barrier repaired by FA. This study is the first to suggest that FA as a potential modulator can more effectively regulate the inflammatory status and gut microbiota to repair the intestinal barrier damage caused by colitis.

Acute cytomegalovirus infection modulates the intestinal microbiota and targets intestinal epithelial cells.

Primary and recurrent cytomegalovirus (CMV) infections frequently cause CMV colitis in immunocompromised as well as inflammatory bowel disease (IBD) patients. Additionally, colitis occasionally occurs upon primary CMV infection in patients who are apparently immunocompetent. In both cases, the underlying pathophysiologic mechanisms are largely elusive - in part due to the lack of adequate access to specimens. We employed the mouse cytomegalovirus (MCMV) model to assess the association between CMV and colitis. During acute primary MCMV infection of immunocompetent mice, the gut microbial composition was affected as manifested by an altered ratio of the Firmicutes to Bacteroidetes phyla. Interestingly, these microbial changes coincided with high-titer MCMV replication in the colon, crypt hyperplasia, increased colonic pro-inflammatory cytokine levels, and a transient increase in the expression of the antimicrobial protein Regenerating islet-derived protein 3 gamma (Reg3γ). Further analyses revealed that murine and human intestinal epithelial cell lines, as well as primary intestinal crypt cells and organoids represent direct targets of CMV infection causing increased cell death. Accordingly, in vivo MCMV infection disrupted the intestinal epithelial barrier and increased apoptosis of intestinal epithelial cells. In summary, our data show that CMV transiently induces colitis in immunocompetent hosts by altering the intestinal homeostasis.

Mechanism of HSP90 Inhibitor in the Treatment of DSS-induced Colitis in Mice by Inhibiting MAPK Pathway and Synergistic Effect of Compound Sophorae Decoction.

The mechanism of Heat Shock Protein 90 (HSP90) in Ulcerative Colitis (UC) has been studied, and mitogenic-activated protein kinases (MAPK) also contribute to the pathogenesis of UC. However, the effect of the HSP90/MAPK pathway in UC is still unclear. Therefore, the mainstay of this research is to explore the mechanism of action of this pathway in UC. Compound sophorae decoction (CSD), as a Chinese herbal decoction, can synergistically affect the above process. This study aimed to uncover the synergistic effects of HSP90 inhibitors regulating the MAPK pathway for treating DSS-induced colitis in mice and the synergistic effects of CSD. This experiment used oral administration of standard diets containing 3% dextran sodium sulfate (DSS) to establish an experimental colitis model in mice. The model was treated with HSP90 inhibitor, CSD, or dexamethasone. Mouse feces, mobility, body weight, colon length, and colon histopathology scores were recorded daily to assess the degree of colitis inflammation. Expression levels of HSP90 and MAPK pathway-related genes and proteins were evaluated by Western blot and qPCR. The evaluation of intestinal mucosal permeability was measured by enzyme-linked immunosorbent assay (ELISA), which could detect the protein level of D-Amino Acid Oxidase (DAO) and D-lactic acid (D-LA). The same went for downstream molecules AFT-2, p53, and apoptosis-related proteins BAX, BCL-2, Caspase3, and survivin in the MAPK pathway. Immunohistochemical measured p-38, p-JNK, and p-ERK expressions. JAM-A and claudin-1 connexin were tested by immunofluorescence staining. The TUNEL method was for measuring the apoptosis rate of colonic epithelial cells. CBA kit determined the level of inflammatory factors of colons. HSP90 inhibitor can improve the degree of pathological damage in the colon of mice treated with DSS, increase the mice's weight and the length of the colon, and significantly reduce the disease activity index (DAI) score. Intraperitoneal injection of HSP90 inhibitor can reduce the expression of MAPK pathway markers P38, JNK, ERK, and their phosphorylation and decrease the content of AFT-2 and p53, which is downstream of the MAPK pathway. In addition, treatment of the HSP90 inhibitor up-regulated the expression of anti-apoptotic proteins BCL-2 and survivin, as well as down-regulated apoptotic protein caspase3, BAX in the colon of mice with colitis. Lower levels of inflammatory factors such as IL-6, MCP-1, IFN-γ, TNF, IL-12p70, and increased IL-10 were observed after HSP90 inhibitor therapy. Furthermore, the combination treatment of CSD can enhance the effect of the single HSP90 inhibitor treatment and play a synergistic effect. These data suggest that an HSP90 inhibitor is available to treat UC by inhibiting the MAPK signaling pathway. This axis can restore the intestinal mucosa barrier's function by reducing intestinal mucosa's permeability and inhibiting apoptosis of intestinal epithelial cells. The specific mechanism is that HSP90 inhibitor can reduce the pathological damage and inflammation levels of colitis mice, and reduce the apoptosis rate of colonic epithelial cells and the mucosal permeability, thereby restoring the mucosal barrier function. During this process, CSD works synergistically to improve the therapeutic effect of the HSP90 inhibitor.

Curcumin analog C66 alleviates inflammatory colitis by inhibiting the activation of NF-κB.

The pathogenesis of inflammatory bowel disease (IBD) remains unclear.C66, a derivative of curcumin, reportedly exerts anti-inflammatory, antifibrotic and anti-apoptotic effects by targeting the JNK pathway. However, the effect of C66 against IBD is not clear. In this study, we aimed to investigate the effect of C66 against IBD. C57BL/6J mice were treated with 2.5% DSS for 7days, and then administered water for 3days to develop the IBD mouse model. A mouse intestinal epithelial cell line, MODE-K, stimulated by lipopolysaccharide (LPS) was used as the in vitro model. The therapeutic effects of C66 were evaluated and the pharmacological mechanisms were explored. Compared to the model group, C66 treatment significantly reduced colitis-associated damage, including a decrease in disease activity index (DAI), a higher body weight and longer colon. In addition, the infiltration of distal inflammatory cells, loss of crypt tissues, and destruction of epithelial cells were reduced in C66-treated group. In addition, C66 treatment reduced fibrotic areas and inflammatory responses in the colon tissues, leading to increased epithelial cell proliferation and decreased apoptosis in colon. Furthermore, C66 treatment decreased the levels of p-JNK and p-P65, indicating that C66 inhibits the activation of the JNK and NF-κB signaling pathways induced by DSS in colon tissues.Finally, in vitro data show that C66 inhibited LPS-induced inflammation and apoptosis in small intestinal epithelial cells. CONCLUSIONS: The curcumin analog C66 exhibits its anti-inflammatory effect by inhibiting the DSS-induced activation of JNK/NF-κB signaling pathways. C66 may be a potential candidate for the treatment of IBD.

Toll-Like Receptor 9 Signaling Pathway Contributes to Intestinal Mucosal Barrier Injury in Mice With Severe Acute Pancreatitis.

The purpose of this study was to investigate the role and mechanism of toll-like receptor 9 (TLR9) in intestinal mucosal barrier injury in mice with severe acute pancreatitis (SAP). The mice were randomly divided into 3 groups: control group, SAP group, and TLR9 antagonist-treated group. The expression of tumor necrosis factor-α, interleukin-1β, interleukin-6, diamine oxidase, and endotoxin core antibodies were detected by enzyme-linked immunosorbent assay. The protein expression of zonula occluden-1 (ZO)-1, occludin, TLR9, myeloid differentiation factor 88 (MyD88), tumor necrosis factor receptor-associated factor 6 (TRAF6), p-nuclear factor (NF)-κB p65, and NF-κB p65 were detected by Western blot. TdT-mediated dUTP nick-end labeling staining was used for detecting intestinal epithelial cell apoptosis. The expression of TLR9 and its related pathway proteins MyD88, TRAF6, and p-NF-κB p65 in the intestinal tract of SAP mice were significantly increased compared with that of control mice. Inhibition of the TLR9 expression could reduce the level of serum proinflammatory cytokines, reduce the apoptosis of intestinal epithelial cells, improve intestinal permeability, and ultimately reduce the damage of intestinal mucosal barrier function in SAP. Toll-like receptor 9/MyD88/TRAF6/NF-κB signaling pathway plays an important role in intestinal mucosal barrier injury of SAP.

The Pathogenicity and Synergistic Action of Th1 and Th17 Cells in Inflammatory Bowel Diseases.

Inflammatory bowel diseases (IBDs), including ulcerative colitis and Crohn's disease, are characterized by chronic idiopathic inflammation of gastrointestinal tract. Although the pathogenesis of IBD remains unknown, intestinal immune dysfunction has been considered as the core pathogenesis. In the intestinal immune system, T helper 1 (Th1) and Th17 cells are indispensable for intestine homeostasis via preventing pathogenic bacteria invasion, regulating metabolism and functions of intestinal epithelial cells (IECs), and promoting IEC self-renewal. However, during the development of IBD, Th1 and Th17 cells acquire the pathogenicity and change from the maintainer of intestinal homeostasis to the destroyer of intestinal mucosa. Because of coexpressing interferon-γ and interleukin-17A, Th17 cells with pathogenicity are named as pathogenic Th17 cells. In disease states, Th1 cells impair IEC programs by inducing IEC apoptosis, recruiting immune cells, promoting adhesion molecules expression of IECs, and differentiating to epithelial cell adhesion molecule-specific interferon γ-positive Th1 cells. Pathogenic Th17 cells induce IEC injury by triggering IBD susceptibility genes expression of IECs and specifically killing IECs. In addition, Th1 and pathogenic Th17 cells could cooperate to induce colitis. The evidences from IBD patients and animal models demonstrate that synergistic action of Th1 and pathogenic Th17 cells occurs in the diseases development and aggravates the mucosal inflammation. In this review, we focused on Th1 and Th17 cell programs in homeostasis and intestine inflammation and specifically discussed the impact of Th1 and Th17 cell pathogenicity and their synergistic action on the onset and the development of IBD. We hoped to provide some clues for treating IBD.

Deoxynivalenol Induces Apoptosis via FOXO3a-Signaling Pathway in Small-Intestinal Cells in Pig.

Deoxynivalenol (DON) is a mycotoxin that is found in feed ingredients derived from grains such as corn and wheat. Consumption of DON-contaminated feed has been shown to cause damage to the intestine, kidneys, and liver. However, the molecular mechanism by which DON exerts its effect in the small intestine is not completely understood. As a result, we profiled gene expression in intestinal epithelial cells treated with DON and examined the molecular function in vitro. We hypothesized that DON could induce apoptosis via the FOXO3a-signaling pathway in intestinal epithelial cells based on these findings. DON induced the apoptosis and the translocation of FOXO3a into the nucleus. Moreover, the inhibiting of FOXO3a alleviated the apoptosis and expression of apoptosis-related genes (TRAL, BCL-6, CASP8, and CASP3). ERK1/2 inhibitor treatment suppressed the translocation of FOXO3a into the nucleus. Our discovery suggests that DON induces apoptosis in intestinal epithelial cells through the FOXO3a-signaling pathway.

MPST deficiency promotes intestinal epithelial cell apoptosis and aggravates inflammatory bowel disease via AKT

Background & aimsExcessive inflammatory responses and oxidative stress are considered the main characteristics of inflammatory bowel disease (IBD). Endogenous hydrogen sulfide (H2S) has been reported to show anti-inflammatory activity in IBD. The main aim of this study was to explore the role of 3-mercaptopyruvate sulfurtransferase (MPST), a key enzyme that regulates endogenous H2S biosynthesis, in IBD. MethodsColonic MPST expression was evaluated in mice and patients with IBD. Various approaches were used to explore the concrete mechanism underlying MPST regulation of the progression of colitis through in vivo and in vitro models. ResultsMPST expression was markedly decreased in colonic samples from patients with ulcerative colitis (UC) or Crohn's disease (CD) and from mice treated with DSS. MPST deficiency significantly aggravated the symptoms of murine colitis, exacerbated inflammatory responses and apoptosis, and inhibited epithelium stem cell-derived organoid formation in an H2S-independent manner. Consistently, when HT29 cells were treated with TNF-α, inhibition of MPST significantly increased the expression of proinflammatory cytokines, the amount of ROS and the prevalence of apoptosis, whereas overexpression of MPST markedly improved these effects. RNA-seq analysis showed that MPST might play a role in regulating apoptosis through AKT signaling. Mechanistically, MPST directly interacted with AKT and reduced the phosphorylation of AKT. Additionally, MPST expression was positively correlated with AKT expression in human IBD samples. In addition, overexpression of AKT rescued IEC apoptosis caused by MPST deficiency, while inhibition of AKT significantly aggravated it. ConclusionsMPST protects the intestines from inflammation most likely by regulating the AKT/apoptosis axis in IECs. Our results may provide a novel therapeutic strategy for the treatment of colitis.

Transcriptome Profile Analysis of Intestinal Upper Villus Epithelial Cells and Crypt Epithelial Cells of Suckling Piglets.

Simple SummaryTo investigate the genetic reprogramming that drives intestinal epithelial cells (IECs) maturation along the crypt-villus axis, entCerocytes were sequentially isolated from the villus tip to the crypts of porcine small intestine. The present study obtained the intestinal upper villus epithelial cells (F1) and crypt epithelial cells (F3) of 21-day suckling piglets using the divalent chelation and precipitation technique. By mRNA high-throughput sequencing analysis of F1 and F3, it was shown that a total of 672 unigenes were differentially expressed between F1 and F3, including 224 highly expressed and 448 minimally expressed unigenes. The greatest number of differentially expressed genes enriched in signal transduction, e.g., Wnt, Hippo, TGF-beta, mTOR, PI3K-Akt, and MAPK signaling pathways, were closely related to the differentiation, proliferation, maturation and apoptosis of IECs. Our results provide important information for identifying new regulators of IECs differentiation and maturation. Moreover, they can also provide insights into the regulatory mechanisms underlying intestinal epithelial cell renewal and the rapid repair of intestinal mucosal oxidative damage.It is well known that the small intestinal epithelial cells of mammals rapidly undergo differentiation, maturation, and apoptosis. However, few studies have defined the physiological state and gene expression changes of enterocytes along the crypt-villus axis in suckling piglets. In the present study, we obtained the intestinal upper villus epithelial cells (F1) and crypt epithelial cells (F3) of 21-day suckling piglets using the divalent chelation and precipitation technique. The activities of alkaline phosphatase, sucrase, and lactase of F1 were significantly higher (p < 0.05) than those of F3. To explore the differences at the gene transcription level, we compared the global transcriptional profiles of F1 and F3 using RNA-seq analysis technology. A total of 672 differentially expressed genes (DEGs) were identified between F1 and F3, including 224 highly expressed and 448 minimally expressed unigenes. Functional analyses indicated that some DEGs were involved in the transcriptional regulation of nutrient transportation (SLC15A1, SLC5A1, and SLC3A1), cell differentiation (LGR5, HOXA5 and KLF4), cell proliferation (PLK2 and TGFB3), transcriptional regulation (JUN, FOS and ATF3), and signaling transduction (WNT10B and BMP1), suggesting that these genes were related to intestinal epithelial cell maturation and cell renewal. Gene Ontology (GO) enrichment analysis showed that the DEGs were mainly associated with binding, catalytic activity, enzyme regulator activity, and molecular transducer activity. Furthermore, KEGG pathway analysis revealed that the DGEs were categorized into 284 significantly enriched pathways. The greatest number of DEGs enriched in signal transduction, some of which (Wnt, Hippo, TGF-beta, mTOR, PI3K-Akt, and MAPK signaling pathways) were closely related to the differentiation, proliferation, maturation and apoptosis of intestinal epithelial cells. We validated the expression levels of eight DEGs in F1 and F3 using qRT-PCR. The present study revealed temporal and regional changes in mRNA expression between F1 and F3 of suckling piglets, which provides insights into the regulatory mechanisms underlying intestinal epithelial cell renewal and the rapid repair of intestinal mucosal damage.

MicroRNA-16 inhibits the TLR4/NF-κB pathway and maintains tight junction integrity in irritable bowel syndrome with diarrhea

Irritable bowel syndrome with diarrhea (IBS-D) is a chronic and relapsing inflammatory disorder in which pathogenesis has been shown to be in part the result of miRNA-mediated signaling. Here, we investigated the alleviatory role of miR-16 in IBS-D. First, we established an IBS-D mouse model using colonic instillation of acetic acid and developed an IBS-D cell model using lipopolysaccharide exposure. The experimental data demonstrated that miR-16 was underexpressed in the serum of IBS-D patients, as well as in the colorectal tissues of IBS-D mouse models and lipopolysaccharide-exposed intestinal epithelial cells. Next, miR-16 and TLR4 were overexpressed or inhibited to characterize their roles in the viability and apoptosis of intestinal epithelial cells, inflammation, and epithelial tight junction. We found that miR-16 overexpression increased the viability of intestinal epithelial cells, maintained tight junction integrity, and inhibited cell apoptosis and inflammation. We showed that miR-16 targeted TLR4 and inhibited the TLR4/NF-κB signaling pathway. Additionally, inhibition of NF-κB suppressed the long noncoding RNA XIST, thereby promoting enterocyte viability, inhibiting apoptosis and cytokine production, and maintaining tight junction integrity. Invivo experiments further verified the alleviatory effect of miR-16 on IBS-D symptoms in mice. Taken together, we conclude that miR-16 downregulates XIST through the TLR4/NF-κB pathway, thereby relieving IBS-D. This study suggests that miR-16 may represent a potential target for therapeutic intervention against IBS-D.

Plakoglobin and High-Mobility Group Box 1 Mediate Intestinal Epithelial Cell Apoptosis Induced by Clostridioides difficile TcdB.

ABSTRACTClostridioides difficile infection (CDI) is the leading cause of antibiotic-associated intestinal disease, resulting in severe diarrhea and fatal pseudomembranous colitis. TcdB, one of the essential virulence factors secreted by this bacterium, induces host cell apoptosis through a poorly understood mechanism. Here, we performed an RNA interference (RNAi) screen customized to Caco-2 cells, a cell line model of the intestinal epithelium, to discover host factors involved in TcdB-induced apoptosis. We identified plakoglobin, also known as junction plakoglobin (JUP) or γ-catenin, a member of the catenin family, as a novel host factor and a previously known cell death-related chromatin factor, high-mobility group box 1 (HMGB1). Disruption of those host factors by RNAi and CRISPR resulted in resistance of cells to TcdB-mediated and mitochondrion-dependent apoptosis. JUP was redistributed from adherens junctions to the mitochondria and colocalized with the antiapoptotic factor Bcl-XL. JUP proteins could permeabilize the mitochondrial membrane, resulting in the release of cytochrome c. Our results reveal a novel role of JUP in targeting the mitochondria to promote the mitochondrial apoptotic pathway. Treatment with glycyrrhizin, an HMGB1 inhibitor, resulted in significantly increased resistance to TcdB-induced epithelial damage in cultured cells and a mouse ligated colon loop model. These findings demonstrate the critical roles of JUP and HMGB1 in TcdB-induced epithelial cell apoptosis.