Gastroenterology
Abstract
Acute high fat diet (HFD) exposure induces a brief period of hyperphagia before caloric balance is restored. Previous studies have demonstrated this period of regulation is associated with activation of synaptic NMDA receptors (NMDARs) on dorsal motor nucleus of the vagus (DMV) neurons, which increases vagal control of gastric functions. Our aim was to test the hypothesis that activation of DMV NMDARs occurs subsequent to activation of extrasynaptic NMDA receptors (NMDARex). Sprague-Dawley rats were fed control or HFD for 3-5 days prior to experimentation. Whole cell patch clamp recordings from gastric-projecting DMV neurons, in vivo recordings of gastric motility, tone, compliance, and emptying, as well as food intake studies were used to assess the effects of NMDAR antagonism on caloric regulation. Following acute HFD exposure, inhibition of NMDARex prevented the NMDARs-mediated increase in glutamatergic transmission to DMV neurons, as well as the increase in gastric tone and motility, while chronic NMDARex inhibition attenuated the regulation of caloric intake. Following acute HFD exposure, the regulation of food intake involves NMDARs-mediated currents, which occur in response to NMDARex activation. Understanding these events may provide a mechanistic basis for hyperphagia and identify potential novel therapeutic targets for the treatment of obesity.
Authors
Courtney Clyburn, R. Alberto Travagli, Amy C. Arnold, Kirsteen N. Browning
Abstract
Most colorectal cancers (CRCs) are moderately-differentiated or well-differentiated, a status that is preserved even in metastatic tumors. However, the molecular mechanisms underlying CRC differentiation remain to be elucidated. Herein, we unravel a novel post-transcriptional regulatory mechanism via a previously unappreciated LIN28B-CDX2 signaling axis that plays a critical role in mediating CRC differentiation. Owing to a large number of mRNA targets, the mRNA-binding protein LIN28B has diverse functions in development, metabolism, tissue regeneration and tumorigenesis. Our RNA-binding protein immunoprecipitation (RIP) assay revealed LIN28B directly binds CDX2 mRNA, which is a pivotal homeobox transcription factor in normal intestinal epithelial cell identity and differentiation. Furthermore, LIN28B overexpression results in enhanced CDX2 expression to promote both differentiation in subcutaneous xenograft tumors generated from CRC cells and metastatic tumor colonization through mesenchymal-epithelial transition in CRC liver metastasis mouse models. Chromatin immunoprecipitation (ChIP) sequence for CDX2 identified Alpha-Methylacyl-CoA racemase (AMACR) as a novel transcriptional target of CDX2 in the context of LIN28B overexpression. We also found AMACR enhances intestinal alkaline phosphatase (ALPi) activity, which is known as a key component of intestinal differentiation, through the upregulation of butyric acid. Overall, we demonstrate that LIN28B promotes CRC differentiation through CDX2-AMACR axis.
Authors
Kensuke Suzuki, Yasunori Masuike, Rei Mizuno, Uma M Sachdeva, Priya Chatterji, Sarah F. Andres, Wenping Sun, Andres J Klein-Szanto, Sepideh Besharati, Helen E Remotti, Michael P Verzi, Anil K. Rustgi
Abstract
Functional dyspepsia (FD) is associated with both chronic gastrointestinal distress and anxiety and depression. Here, we hypothesized that aberrant gastric signals, transmitted by the vagus nerve, may alter key brain regions modulating affective and pain behavior. Using a previously validated rat model of FD characterized by gastric hypersensitivity, depression- and anxiety-like behavior, we found that vagal activity in response to gastric distention was increased in FD rats. The FD phenotype was associated with gastric mast cell hyperplasia and increased expression of corticotrophin-releasing factor (CRF) and decreased brain-derived neurotrophic factor in the central amygdala. Subdiaphragmatic vagotomy reversed these changes and restored affective behavior to that of controls. Vagotomy partially attenuated pain responses to gastric distention, which may be mediated by central reflexes in the periaqueductal gray, as determined by local injection of lidocaine. Ketotifen, a mast cell stabilizer, reduced vagal hypersensitivity, normalized affective behavior and attenuated gastric hyperalgesia. In conclusion, vagal activity, partially driven by gastric mast cells, induces long-lasting changes in CRF signaling in the amygdala that may be responsible for enhanced pain and anxiety- and depression-like behaviors. Together, these results support a “bottom-up” pathway involving the gut-brain axis in the pathogenesis of both gastric pain and psychiatric co-morbidity in FD.
Authors
Zachary A. Cordner, Qian Li, Liansheng Liu, Kellie L. Tamashiro, Aditi Bhargava, Timothy H. Moran, Pankaj J. Pasricha
Abstract
Hindered by a limited understanding of the mechanisms responsible for diabetic gastroenteropathy (DGE), management is symptomatic. We investigated the duodenal mucosal expression of protein-coding genes and microRNAs (miRNA) in DGE and related them to clinical features. The diabetic phenotype, gastric emptying, mRNA, and miRNA expression and ultrastructure of duodenal mucosal biopsies were compared in 39 DGE patients and 21 controls. Among 3175 differentially expressed genes (FDR < 0.05), several mitochondrial DNA–encoded (mtDNA-encoded) genes (12 of 13 protein coding genes involved in oxidative phosphorylation [OXPHOS], both rRNAs and 9 of 22 transfer RNAs) were downregulated; conversely, nuclear DNA–encoded (nDNA-encoded) mitochondrial genes (OXPHOS) were upregulated in DGE. The promoters of differentially expressed genes were enriched in motifs for transcription factors (e.g., NRF1), which regulate mitochondrial biogenesis. Seventeen of 30 differentially expressed miRNAs targeted differentially expressed mitochondrial genes. Mitochondrial density was reduced and correlated with expression of 9 mtDNA OXPHOS genes. Uncovered by principal component (PC) analysis of 70 OXPHOS genes, PC1 was associated with neuropathy (P = 0.01) and delayed gastric emptying (P < 0.05). In DGE, mtDNA- and nDNA-encoded mitochondrial genes are reduced and increased — associated with reduced mitochondrial density, neuropathy, and delayed gastric emptying — and correlated with cognate miRNAs. These findings suggest that mitochondrial disturbances may contribute to delayed gastric emptying in DGE.
Authors
Susrutha Puthanmadhom Narayanan, Daniel O’Brien, Mayank Sharma, Karl Miller, Peter Adams, João F. Passos, Alfonso Eirin, Tamas Ordog, Adil E. Bharucha
Abstract
Severe acute pancreatitis (AP) is a life-threatening disease with up to 30% mortality. Therefore, prevention of AP aggravation and promotion of pancreatic regeneration are critical during the course and treatment of AP. Hypertriglyceridemia (HTG) is an established aggravating factor for AP that hinders pancreatic regeneration; however, its exact mechanism remains unclear. Using miRNA sequencing and further verification, we found that miRNA-153 (miR-153) was upregulated in the pancreas of HTG animal models and in the plasma of patients with HTG-AP. Increased miR-153 aggravated HTG-AP and delayed pancreatic repair via targeting TRAF3. Furthermore, miR-153 was transcriptionally suppressed by sterol regulatory element-binding transcription factor 1c (SREBP1c), which was suppressed by lipoprotein lipase malfunction-induced HTG. Overexpressing SREBP1c suppressed miR-153 expression, alleviated the severity of AP, and facilitated tissue regeneration in vivo. Finally, therapeutic administration of insulin also protected against HTG-AP via upregulating SREBP1c. Collectively, our results not only provide evidence that HTG leads to the development of more severe AP and hinders pancreatic regeneration via inducing persistent dysregulation of SREBP1c/miR-153 signaling, but also demonstrate that SREBP1c activators, including insulin, might be used to treat HTG-AP in patients.
Authors
Juanjuan Dai, Mingjie Jiang, Yangyang Hu, Jingbo Xiao, Bin Hu, Jiyao Xu, Xiao Han, Shuangjun Shen, Bin Li, Zengkai Wu, Yan He, Yingchun Ren, Li Wen, Xingpeng Wang, Guoyong Hu
Abstract
Diarrhea is a major cause of global mortality, and outbreaks of secretory diarrhea such as cholera remain an important problem in the developing world. Current treatment of secretory diarrhea primarily involves supportive measures such as fluid replacement. The calcium-sensing receptor (CaSR) regulates multiple biological activities in response to changes in extracellular Ca+2. The FDA-approved drug cinacalcet is an allosteric activator of CaSR used for treatment of hyperparathyroidism. Here, we found by short-circuit current measurements in human colonic T84 cells that CaSR activation by cinacalcet reduced forskolin-induced Cl- secretion by greater than 80%. Cinacalcet also reduced Cl- secretion induced by cholera toxin, heat-stable E. coli enterotoxin, and vasoactive intestinal peptide (VIP). The cinacalcet effect primarily involved indirect inhibition of cystic fibrosis transmembrane conductance regulator (CFTR)-mediated Cl- secretion following activation of CaSR, and downstream phospholipase C and phosphodiesterases. In mice, cinacalcet reduced fluid accumulation by more than 60% in intestinal closed-loop models of cholera and Traveler’s diarrhea. The cinacalcet effect involved both inhibition of CFTR-mediated secretion and stimulation of sodium-hydrogen exchanger 3 (NHE3)-mediated absorption. These findings support the therapeutic utility of the safe and commonly used drug cinacalcet in CFTR-dependent secretory diarrheas including cholera, Traveler’s diarrhea and VIPoma.
Authors
Apurva A. Oak, Parth D. Chhetri, Amber Rivera, Alan S. Verkman, Onur Cil
Abstract
Cantύ Syndrome (CS), caused by gain-of-function (GOF) mutations in pore-forming (Kir6.1, KCNJ8) and accessory (SUR2, ABCC9) ATP-sensitive potassium (KATP) channel subunit genes, is frequently accompanied by gastrointestinal (GI) dysmotility, and we describe one CS patient who required an implanted intestinal irrigation system for successful stooling. We used gene-modified mice to assess the underlying KATP channel subunits in gut smooth muscle, and to model the consequences of altered KATP channels in CS gut. We show that Kir6.1/SUR2 subunits underlie smooth muscle KATP channels throughout the small intestine and colon. Knock-in mice, carrying human KCNJ8 and ABCC9 CS mutations in the endogenous loci, exhibit reduced intrinsic contractility throughout the intestine, resulting in death when weaned onto solid food in the most severely affected animals. Death is avoided by weaning onto a liquid gel diet, implicating intestinal insufficiency and bowel impaction as the underlying cause, and GI transit is normalized by treatment with the KATP inhibitor glibenclamide. We thus define the molecular basis of intestinal KATP channel activity, the mechanism by which overactivity results in GI insufficiency, and a viable approach to therapy.
Authors
Nathaniel W. York, Helen Parker, Zili Xie, David Tyus, Maham A. Waheed, Zihan Yan, Dorothy K. Grange, Maria S. Remedi, Sarah K. England, Hongzhen Hu, Colin G. Nichols
Abstract
Loss of functional small bowel surface area following surgical resection for disorders such as Crohn’s disease, intestinal ischemic injury, radiation enteritis, and in children, necrotizing enterocolitis, atresia and gastroschisis, may result in short bowel syndrome (SBS) with attendant high morbidity, mortality and health care costs in the U.S. Following resection, the remaining small bowel epithelium mounts an adaptive response resulting in increased crypt cell proliferation, increased villus height, crypt depth and enhanced nutrient and electrolyte absorption. Although these morphologic and functional changes are well-described in animal models, the adaptive response in humans is less well understood, and clinically the response is unpredictable and often inadequate. Here we address the hypotheses that human intestinal stem cell populations are expanded and the stem cell niche is regulated following massive gut resection in short bowel syndrome. We use intestinal enteroid cultures from SBS patients to show that the magnitude and phenotype of the adaptive stem cell response is regulated by stromal niche cells including intestinal subepithelial myofibroblasts, which are activated by intestinal resection to enhance epithelial stem and proliferative cell responses. Our data suggest that myofibroblast regulation of bone morphogenetic protein signaling pathways plays a role in the gut adaptive response post resection.
Authors
Vered Gazit, Elzbieta A. Swietlicki, Miranda U. Liang, Adam Surti, Raechel McDaniel, Mackenzie Geisman, David M. Alvarado, Matthew A. Ciorba, Grant V. Bochicchio, Obeid Ilahi, John Kirby, William J. Symons, Nicholas O. Davidson, Marc S. Levin, Deborah C. Rubin
Abstract
High-fat feeding (HFF) leads to gut dysbiosis through unclear mechanisms. We hypothesize that bile acids secreted in response to high-fat diets (HFDs) may act on intestinal Paneth cells, leading to gut dysbiosis. We found that HFF resulted in widespread taxonomic shifts in the bacteria of the ileal mucosa, characterized by depletion of Lactobacillus and enrichment of Akkermansia muciniphila, Clostridium XIVa, Ruminococcaceae, and Lachnospiraceae, which were prevented by the bile acid binder cholestyramine. Immunohistochemistry and in situ hybridization studies showed that G protein–coupled bile acid receptor (TGR5) expressed in Paneth cells was upregulated in the rats fed HFD or normal chow supplemented with cholic acid. This was accompanied by decreased lysozyme+ Paneth cells and α-defensin 5 and 6 and increased expression of XBP-1. Pretreatment with ER stress inhibitor 4PBA or with cholestyramine prevented these changes. Ileal explants incubated with deoxycholic acid or cholic acid caused a decrease in α-defensin 5 and 6 and an increase in XBP-1, which was prevented by TGR5 antibody or 4PBA. In conclusion, this is the first demonstration to our knowledge that TGR5 is expressed in Paneth cells. HFF resulted in increased bile acid secretion and upregulation of TGR5 expression in Paneth cells. Bile acid toxicity in Paneth cells contributes to gut dysbiosis induced by HFF.
Authors
Hui Zhou, Shi-Yi Zhou, Merritt Gillilland III, Ji-Yao Li, Allen Lee, Jun Gao, Guanpo Zhang, Xianjun Xu, Chung Owyang
Abstract
Severe burn injury induces gut barrier dysfunction and subsequently a profound systemic inflammatory response. In the present study, we examined the role of the small intestinal brush border enzyme, intestinal alkaline phosphatase (IAP), in preserving gut barrier function and preventing systemic inflammation after burn wound infection in mice. Mice were subjected to a 30% total body surface area dorsal burn with or without intradermal injection of Pseudomonas aeruginosa. Mice were gavaged with 2000 units of IAP or vehicle at 3 and 12 hours after the insult. We found that both endogenously produced and exogenously supplemented IAP significantly reduced gut barrier damage, decreased bacterial translocation to the systemic organs, attenuated systemic inflammation, and improved survival in this burn wound infection model. IAP attenuated liver inflammation and reduced the proinflammatory characteristics of portal serum. Furthermore, we found that intestinal luminal contents of burn wound–infected mice negatively impacted the intestinal epithelial integrity compared with luminal contents of control mice and that IAP supplementation preserved monolayer integrity. These results indicate that oral IAP therapy may represent an approach to preserving gut barrier function, blocking proinflammatory triggers from entering the portal system, preventing gut-induced systemic inflammation, and improving survival after severe burn injuries.
Authors
Fatemeh Adiliaghdam, Paul Cavallaro, Vidisha Mohad, Marianna Almpani, Florian Kühn, Mohammad Hadi Gharedaghi, Mehran Najibi, Laurence G. Rahme, Richard A. Hodin
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