American journal of physiology. Gastrointestinal and liver physiology最新文献

The global population is aging, with one in six people projected to be 65 yr or older by 2050. Since people aged 65 and older experience higher rates of morbidity and mortality after burn injury, there is an increased need to develop effective burn treatments in this age group. Heightened morbidity and risk of mortality may stem from increased gut leakiness and death of intestinal epithelial cells of aged individuals. Herein, we used our clinically relevant model of scald burn injury in young and aged mice to ascertain whether the colon, isolated colonic epithelium, and organoids grown from the colon have deficiencies in cell growth, senescence, and apoptosis pathways. Aged, burn-injured mice displayed increased senescence marker cdkn2a in the colon and isolated epithelium, and displayed a reduction in proliferation marker cdk4 in the colon when compared with young mice. Changes in senescence and proliferation coincided with a reduction in stem cell marker lgr5 in the colon and colonic epithelium, suggesting a burn-related reduction in the stemness of the epithelial crypt. Although we failed to see histological changes in the colonic epithelium, we observed an increase in proapoptotic cleaved caspase 3 and 9 within the colons of aged, burn-injured mice. Finally, there was a decrease in the expression of antimicrobial peptide ang4, and not camp in the colons of aged, burn-injured mice. Taken together, these data suggest that in the colon, disruption of proliferation and apoptosis in aged burn-injured mice occurs primarily in the nonepithelial compartment.NEW & NOTEWORTHY Aged mice have more senescent cells in their colons and burn injury in aged mice leads to suppression of proliferation markers in the colon, but not in epithelial cells or cultured organoids. Colonic expression of stem cell marker lgr5 is reduced in colon from aged, burn-injured mice, and a proapoptotic caspase cascade was seen in this cohort. Finally, antimicrobial peptide ang4 expression is decreased in colons from both aged and aged, burn-injured mice.

The gastrointestinal manifestations of cystic fibrosis (CF) are a continued source of morbidity and mechanistic uncertainty despite recent advances in CF care. We sought to characterize intestinal glucose demand in a mouse model of CF to better understand CF intestinal disease. We assessed in vivo systemic glucose uptake from circulation, including intestinal glucose demand, using ¹⁸F-fluorodeoxyglucose positron emission tomography (PET) imaging studies in wild-type (WT) and CF mice. RNA-sequencing studies with complementary assessments of protein expression and functional metabolism were performed to identify the responsible glucose transporter and relevant metabolic pathways. Finally, morphologic and histologic differences between the CF and WT small intestine were investigated. Increased glucose uptake from circulation to CF intestine was detected with the most prominent increases seen in CF jejunum and ileum. Increased mRNA and protein expression of GLUT1 was evident in whole intestinal tissue and isolated crypts, suggesting that GLUT1 is responsible for mediating the increased glucose uptake from the blood supply. We found transcriptional and functional enrichment of glycolysis in the CF jejunum and ileum. Proliferative intestinal adaptations, including increased intestinal length and weight, in addition to increased villi length and crypt depth, were observed in CF mice. The increased intestinal glucose uptake from circulation and increased glycolysis, in combination with the morphologic and histologic changes in the CF intestine, are suggestive of a proliferative adaptive response and increased intestinal glucose demand in CF. This work may yield novel markers of CF disease status and new therapeutic approaches. NEW & NOTEWORTHY We found transcriptional, protein level, and functional evidence of increased intestinal glucose uptake from circulation and increased glycolysis in a cystic fibrosis mouse model. These findings in the context of hyperplastic morphologic and histologic changes in the cystic fibrosis intestine are indicative of an adaptive response. Our work elucidates new mechanisms of intestinal disease in cystic fibrosis and identifies altered intestinal glucose uptake and glycolysis as potential markers of disease status and gastrointestinal cancer risk.

Fibroblast-like cells (FLCs) exist in the smooth muscle layers of visceral organs, yet in many instances their functional role(s) have not been identified. FLCs express platelet-derived growth factor receptor (PDGFR) α and are a novel class of excitable cells recently described in visceral organs. Crenolanib is a benzamidine quinolone derivative originally developed as an inhibitor of PDGFR to treat certain solid tumors with PDGFRα over-expression mutations. In the present study we used crenolanib to disrupt PDGFRα expression and signaling in the GI tracts of BALB/c mice. Intra-peritoneal injections of crenolanib (100 μg/g body weight) or DMSO control vehicle were given to littermates from postpartum P1 through P15. Crenolanib injected mice were smaller in size and weight. The gastrointestinal tracts were also shorter and appeared partially distended. qPCR revealed down-regulation of key gene transcripts involved in PDGFRα cell signaling including Pdgfrα, Kcnn3 and P2ry1. Confocal immunofluorescence demonstrated significant decreases in PDGFRα and SK3 protein expression. c-Kit expression was slightly inhibited but gastric, intestinal and colonic pacemaker activity was not affected by crenolanib. Purinergic inhibitory post-junctional motor responses were greatly attenuated in the GI tracts of crenolanib treated animals compared to vehicle treated controls in response to electric field evoked nerve stimulation. These data provide evidence for a functional role of PDGFRα⁺ cells in inhibitory neuroeffector motor responses throughout the gastrointestinal tract.

Radiation exposure impairs rapidly renewing tissues like the intestinal epithelium, yet translational insights from murine models have been limited by species-specific responses. Here, we use human intestinal organoids (HIOs) derived from jejunal epithelium to evaluate human epithelial responses to low-dose proton and photon (γ) radiation. γ irradiation induces a unique developmental and metabolic shift in crypt-like organoids, including enrichment of amino acid metabolism pathways and activation of fetal-associated transcription factors and morphology. Integrated multiomic profiling reveals serotonin biosynthesis as a central regenerative node. HIOs can complement animal models and are emerging as a powerful tool in modeling human radiation responses and identifying candidate biomarkers for intestinal injury.NEW & NOTEWORTHY Radiation damages the intestinal epithelium, but murine models often fail to capture human-specific responses. Using human intestinal organoids, we show that γ irradiation triggers a distinct developmental and metabolic reprogramming, including enrichment of amino acid metabolism and induction of fetal-associated transcription factors and morphology. These findings highlight human organoids as a translational platform to model radiation injury and uncover candidate biomarkers for intestinal damage.

Some forms of gastroesophageal reflux disease (GERD) are associated with crural diaphragm (CD) dysfunction, suggesting that GERD may be influenced by skeletal muscle deficiencies. Skeletal muscle atrophy has been strongly linked to alterations in the ubiquitin-proteasome system, the primary pathway for protein degradation. This study aimed to assess the expression of muscle atrophy-related proteins in the CD of patients with reflux esophagitis compared with those without esophagitis. In addition, we examined the correlation between these proteins, esophagitis severity, and esophageal acid exposure. CD biopsies were obtained from 15 volunteers (8 males, 7 females; mean age 43 yr) during antireflux laparoscopic Nissen fundoplication (GERD group) or gallbladder surgery (control group). The GERD group was further classified based on the Los Angeles classification into grades A (n = 5), B (n = 7), and C (n = 3). We analyzed key signaling pathways involved in muscle atrophy, including AKT, phosphorylated AKT (pAKT), muscle-specific RING finger 1 protein (MuRF-1), and muscle atrophy F-box (MAFbx/atrogin-1), normalized to glyceraldehyde 3-phosphate dehydrogenase (GAPDH). No significant differences were observed in MuRF-1, pAKT/AKT ratio, or MAFbx/atrogin-1 expression between the control and GERD groups. However, MuRF-1 expression was significantly elevated in the GERD C group compared with GERD B group. The control group showed no differences from GERD A or B. Notably, MuRF-1 expression correlated with esophageal total reflux time in the supine position. These findings suggest that increased MuRF-1 expression may contribute to CD fiber atrophy and weakness in patients with GERD, potentially impairing gastroesophageal junction function and influencing disease progression.NEW & NOTEWORTHY This study demonstrated, for the first time, an increased activation of the ubiquitin-proteasome pathway and elevated MuRF-1 expression in the crural diaphragm of humans with moderate reflux esophagitis. It showed a positive correlation between the supine reflux time and MuRF-1 expression, suggesting a molecular mechanism associated with diaphragm fiber atrophy and weakness. These findings highlight a potential link between diaphragm degradation and reflux esophagitis, which may modulate gastroesophageal reflux and symptoms.

Fructose consumption contributes to metabolic dysfunction-associated steatohepatitis (MASH). Retatrutide is a novel triple receptor agonist that improves obesity and hepatic steatosis in humans. The aims of this study were to develop a shortened and clinically relevant dietary mouse model of diet-induced steatohepatitis, and to evaluate the effects of a retatrutide intervention in this model. C57BL/6N mice were subjected to a single fructose binge (10 mg/g body wt), or a new 31-day mouse model of diet-induced steatohepatitis using a Western diet, fructose, and sucrose in the drinking water, and a final fructose binge with or without retatrutide. A single fructose binge resulted in significantly elevated alanine aminotransferase (ALT) and hepatic triglyceride levels in female mice after 6 h; male mice showed less hepatotoxicity. The novel 31-day feeding model significantly increased body weight, ALT levels, hepatic triglycerides and cholesterol, and hepatic inflammatory markers in female and male mice compared with their chow-fed controls. The overall hepatic gene expression profile per RNA sequencing of treated mice correlated with that of human MASH in children and adults. Retatrutide intervention over the final 2 weeks of the 31-day mouse model significantly reduced body weight, ALT levels, hepatic triglycerides and cholesterol, and hepatic inflammatory markers in female mice compared with their vehicle-treated counterparts. Our findings indicate that female mice develop more severe liver injury due to a single fructose binge than males. The novel 31-day mouse model induces robust steatohepatitis and correlates with human disease. An intervention with retatrutide improves steatohepatitis in this shortened mouse model.NEW & NOTEWORTHY Female mice are more prone to liver injury due to a single fructose binge compared with male mice. The new 31-day mouse model induces robust steatohepatitis in mice and correlates with MASLD in children and adults. An intervention with retatrutide improves steatohepatitis in this novel mouse model, indicating despite its short duration, the model can be used to trial pharmacological interventions.

Lithogenic diet exposure disrupts biliary lipid homeostasis to promote precipitation of excess biliary cholesterol; however, the underlying pathogenic signaling mechanism remains unclear. Protein kinase Cbeta (PKCβ) is involved in regulating hepatic cholesterol and bile acid metabolism. In this study, we aimed to identify the initiating signaling and biological changes in the liver upon loss of hepatic PKCβ function under lithogenic stress. Transcriptome analysis of the liver revealed that hepatic deletion of PKCβ altered the expression of 183 liver genes, 118 of which were upregulated and 65 were downregulated. We identified marked increases in the expression of genes involved in bile acid biosynthesis (Cyp7a1 and Cyp8b1) and a decrease in retinol metabolism (Cyp26b1) as the most relevant changes, with blunted expression of genes involved in bile acid and phosphatidylcholine transporters. Mechanistic studies revealed that the hepatic PKCβ deficiency was associated with reduced ERK1/2 phosphorylation in concert with increased p38^MAPK phosphorylation in the liver. Overexpression of PKCβ in the liver blocked p38^MAPK activation as well as resulted in increased ERK1/2 phosphorylation and was accompanied by suppression of both Cyp7a1 and Cyp8b1 expression, demonstrating that hepatic PKCβ functions as a positive regulator of ERK1/2 to suppress the expression of both genes by antagonizing p38^MAPK. Furthermore, depletion of liver p38^MAPK in PKCβ^Liv-/- mice resulted in enhanced ERK1/2 phosphorylation and suppression of Cyp7a1 and Cyp8b1 expression. The findings yielded by this study support our understanding of the intricate interplay among PKCβ, p38^MAPK, and ERK1/2 signaling in vivo and provide valuable insights into potential therapeutic targets for the development of novel strategies to combat cholelithiasis.NEW & NOTEWORTHY This study underscores the pivotal role of hepatic PKCβ in controlling biliary lipid composition under lithogenic stress. Our findings on the distinct and combined effects of downstream p38^MAPK and ERK1/2 offer key insights into the mechanisms driving lithogenic diet-induced dysregulation of biliary lipid composition. This research reveals that the potential of PKCβ/p38^MAPK/ERK1/2 signaling axis offers the possibility for the integration of different inputs to modulate the signaling output balancing in a way most appropriate for context.

Mucosal homeostasis requires coordinated immune regulation and epithelial repair. Inflammatory bowel disease (IBD) arises from disrupted coordination between the immune system and intestinal epithelium, where resolution and repair must occur in parallel. Interleukin-6 (IL-6) plays a dual role: it promotes epithelial regeneration but destabilizes regulatory T cells (Tregs). We aimed to determine the contribution of Treg IL-6 receptor (IL-6R) signaling to intestinal inflammation and epithelial integrity. We developed a conditional knockout mouse model in which IL-6R was deleted from Tregs (Treg IL-6R knockout). These mice were subjected to dextran sodium sulfate (DSS)-induced colitis and a T cell transfer model of colitis. Soluble IL-6R production by Tregs was assessed in vitro, and transcriptional changes in epithelial cells were analyzed by RNA-seq. Human colonic organoids from patients with IBD were treated with IL-6 or hyper-IL-6 (IL-6/sIL-6R fusion protein) to test downstream signaling effects. Tregs lacking IL-6R improved colitis to a similar extent as control Tregs in the adoptive transfer model, indicating intact suppressive function. However, Treg IL-6R knockout mice were more susceptible to DSS colitis than controls, suggesting a physiologic role for Treg IL-6R signaling in epithelial protection. In vitro, Tregs shed soluble IL-6R, enabling IL-6 trans-signaling to epithelial cells. Intestinal epithelial cells from Treg IL-6R knockout mice compared with WTcre controls revealed widespread transcriptional downregulation of genes related to survival and repair pathways at baseline, and impaired transcriptional responses following DSS treatment. In human organoids, IL-6 trans-signaling elicited stronger STAT3 activation than IL-6 alone. These findings reveal a previously unrecognized role for Treg-derived IL-6R in promoting epithelial resilience and maintaining mucosal homeostasis.NEW & NOTEWORTHY This study reveals a novel role for Treg-derived IL-6R in supporting epithelial repair. Despite preserved immune-suppressive capacity, deletion of IL-6R from Tregs impairs epithelial transcription and worsens injury in colitis. We demonstrate that human intestinal organoids preferentially respond to trans- over classic IL-6 signaling. These findings introduce a Treg-specific role in immune-epithelial cross talk relevant to mucosal healing and inflammatory bowel disease.

Excessive intake of fructose and fats disrupts hepatocyte function by overwhelming endoplasmic reticulum (ER) capacity, leading to unresolved protein stress and progression to metabolic dysfunction-associated steatohepatitis (Shepherd EL, Saborano R, Northall E, Matsuda K, Ogino H, Yashiro H, Pickens J, Feaver RE, Cole BK, Hoang SA, Lawson MJ, Olson M, Figler RA, Reardon JE, Nishigaki N, Wamhoff BR, Günther UL, Hirschfield G, Erion DM, Lalor PF. JHEP Rep 3: 100217, 2021). Ketohexokinase (KHK), the primary enzyme for fructose metabolism, is increasingly recognized for nonmetabolic roles (Peng C, Yang P, Zhang D, Jin C, Peng W, Wang T, Sun Q, Chen Z, Feng Y, Sun Y. Acta Pharm Sin B 14: 2959-2976, 2024; Li X, Qian X, Peng LX, Jiang Y, Hawke DH, Zheng Y, Xia Y, Lee JH, Cote G, Wang H, Wang L, Qian CN, Lu Z. Nat Cell Biol 18: 561-571, 2016), but its function in regulating ER proteostasis under nutrient stress remains unclear. We show that steatogenic conditions synergistically induce lipid accumulation and robust KHK expression, accompanied by activation of the IRE1α-XBP1 branch of the unfolded protein response. This adaptive axis was observed in HepG2 cells, primary hepatocytes from Gubra Amylin NASH, (GAN) diet-fed mice, and liver biopsies from MASLD patients, establishing a conserved KHK-IRE1α axis across species. Khk knockdown disrupted this balance, causing accumulation of misfolded and ubiquitinated proteins, proteotoxic stress, and a shift toward PERK-CHOP-driven apoptosis. Similar signatures in Khk-deficient mouse livers further underscore KHK's role in sustaining ER homeostasis. Our findings identify KHK as a dual-function enzyme: a metabolic gatekeeper of fructose flux and a proteostatic regulator that safeguards hepatocyte survival. Although KHK contributes to steatosis, its complete loss destabilizes ER proteostasis, suggesting that selective inhibition of KHK enzymatic activity may offer therapeutic benefit without compromising ER function.NEW & NOTEWORTHY This study uncovers a noncanonical role for ketohexokinase (KHK) in maintaining ER proteostasis during nutrient overload. In hepatocytes exposed to fructose and saturated fat, KHK promotes adaptive IRE1α-XBP1 signaling and prevents proteotoxic stress and apoptosis. These findings position KHK as a metabolic checkpoint linking fructose metabolism to ER stress resolution and offer new insight into liver survival pathways relevant to MASLD and MASH.