RESUMEN
INTRODUCTION: Massive intestinal loss resulting in short bowel syndrome has been linked to intestinal failure associated liver disease. Efforts to elucidate the driving force behind the observed hepatic injury have identified inflammatory mediators, alterations in the microbiome, extent of structural and functional intestinal adaptation, and toxic shifts in the bile acid pool. In the present study, we posit that ileocecal resection interrupts the delivery of these hepatotoxic substances to the liver by physically disrupting the enterohepatic circulation, thereby shielding the liver from exposure to the aforementioned noxious stimuli. METHODS: Mice underwent sham, 50% proximal, or 50% distal small bowel resection (SBR), with or without tauroursodeoxycolic acid supplementation. Enterohepatic signaling and nonsense-mediated ribonucleic acid (RNA) decay were evaluated and correlated with hepatic injury. RESULTS: When compared to 50% proximal SBR, mice that underwent ileocecal resection exhibited reduced hepatic oxidative stress and exhibited a more physiological bile acid profile with increased de novo bile acid synthesis, enhanced colonic bile acid signaling, and reduced hepatic proliferation. Distal intestinal resection promoted an adaptive response including via the nonsense-mediated RNA decay pathway to satisfactorily process injurious messenger RNA and successfully maintain homeostasis. By contrast, this adaptive response was not observed in the proximal SBR group and hepatic injury persisted. CONCLUSIONS: In summary, interruption of enterohepatic circulation via ileocecal resection abrogates the liver's exposure to toxic and inflammatory mediators while promoting physiological adaptations in bile acid metabolism and maintaining existing homeostatic pathways.
Asunto(s)
Hepatopatías , ARN , Ratones , Animales , ARN/metabolismo , Hígado/cirugía , Hígado/metabolismo , Hepatopatías/metabolismo , Ácidos y Sales Biliares/metabolismo , Mediadores de Inflamación/metabolismoRESUMEN
Massive intestinal resection is a regrettably necessary but life-saving intervention for progressive or fulminant necrotizing enterocolitis (NEC). However, the resultant short bowel syndrome (SBS) poses its own array of challenges and complications. Within hours of such an abrupt loss of intestinal length, the intestine begins to adapt. Our ability to understand this process of intestinal adaptation has proven critical in our ability to clinically treat the challenging problem of short bowel syndrome. This review first highlights key data relating to intestinal adaptation including structural and functional changes, biochemical regulation, and other factors affecting the magnitude of intestinal adaptation responses. We then focus on intestinal rehabilitation as it relates to strategies to enhance intestinal adaptation while meeting nutritional needs and preventing complications of parenteral nutrition.
Asunto(s)
Enterocolitis Necrotizante , Síndrome del Intestino Corto , Recién Nacido , Humanos , Síndrome del Intestino Corto/cirugía , Intestinos , Nutrición Parenteral , Adaptación Fisiológica , Enterocolitis Necrotizante/cirugíaRESUMEN
BACKGROUND: Massive small bowel resection (SBR) is associated with liver injury and fibrosis. Efforts to elucidate the driving force behind hepatic injury have identified multiple factors, including the generation of toxic bile acid metabolites. METHODS: Sham, 50% proximal, and 50% distal SBR were carried out in C57BL/6 mice to determine the effect of jejunal (proximal SBR) versus ileocecal resection (distal SBR) on bile acid metabolism and liver injury. Tissues were harvested at 2 and 10-week postoperative timepoints. RESULTS: When compared with 50% proximal SBR, mice that underwent distal SBR exhibited less hepatic oxidative stress as verified by decreased mRNA expression of tumor necrosis factor-α (TNFα, p ≤ 0.0001), nicotinamide adenine dinucleotide phosphate oxidase (NOX, p ≤ 0.0001), and glutathione synthetase (GSS, p ≤ 0.05). Distal SBR mice also exhibited a more hydrophilic bile acid profile with reduced abundance of insoluble bile acids (cholic acid (CA), taurodeoxycholic acid (TCA), and taurolithocholic acid (TLCA)), and increased abundance of soluble bile acids (tauroursodeoxycholic acid (TUDCA)). In contrast with proximal SBR, ileocecal resection alters enterohepatic circulation leading to reduced oxidative stress and promotes physiological bile acid metabolism. CONCLUSION: These findings challenge the notion that preservation of the ileocecal region is beneficial in patients with short bowel syndrome. Administration of selected bile acids may present potential therapy to mitigate resection-associated liver injury. LEVEL OF EVIDENCE: III-Case-Control Study.
Asunto(s)
Ácidos y Sales Biliares , Hígado , Ratones , Animales , Estudios de Casos y Controles , Ratones Endogámicos C57BL , Hígado/cirugía , Hígado/metabolismo , Circulación EnterohepáticaRESUMEN
BACKGROUND: Resection-associated liver steatosis, injury, and fibrosis is a devastating complication associated with massive small bowel resection (SBR). Peroxisome proliferator-activated receptor-alpha (PPARα) is a key regulator of intestinal lipid transport and metabolism whose expression is selectively increased after SBR. Here we asked if attenuating intestinal PPARα signaling would prevent steatosis and liver injury after SBR. METHODS: Pparα was deleted selectively in adult mouse intestine using a tamoxifen-inducible Cre-LoxP breeding schema. Mice underwent 50% SBR. At 10 weeks post-operatively, metabolic phenotyping, body composition analysis, in vivo assessment of lipid absorption and intestinal permeability, and assessment of adaptation and liver injury was completed. RESULTS: Pparα intestinal knockout and littermate control mice were phenotypically similar in terms of weight trends and body composition after SBR. All mice demonstrated intestinal adaptation with increased villus height and crypt depth; however, Pparα intestinal knockout mice exhibited decreased villus growth at 10 weeks compared to littermate controls. Liver injury and fibrosis were similar between groups as assessed by serum AST and ALT levels, Sirius Red staining, and hepatic expression of Col1a1 and Acta2. CONCLUSIONS: Inducible intestinal deletion of Pparα influences structural adaptation but does not mitigate liver injury after SBR. These findings suggest that enterocyte PPARα signaling in adult mice is dispensable for resection-induced liver injury. The results are critical for understanding the contribution of intestinal lipid metabolic signaling pathways to the pathogenesis of hepatic injury associated with short bowel syndrome.
Asunto(s)
Intestino Delgado , PPAR alfa , Animales , Ratones , Adaptación Fisiológica , Intestino Delgado/cirugía , Lípidos , Ratones Endogámicos C57BL , Ratones Noqueados , PPAR alfa/genética , PPAR alfa/metabolismoRESUMEN
Short bowel syndrome (SBS) is associated with diminished levels of serum fats caused by unknown mechanisms. We have shown that mesenteric lymphatics remodel to a more primitive state one week after small bowel resection (SBR); therefore, this study focuses on the effect of chronic lymphatic remodeling and magnitude of resection on intestinal lipid uptake and transport. C57BL6 and Prox1 creER-Rosa26LSLTdTomato (lymphatic reporter) mice underwent 50% or 75% proximal SBR or sham operations. Functional transport of lipids and fecal fat content was measured and lymphatic vasculature was compared via imaging. There was a significant reduction in functional transport of cholesterol and triglyceride after SBR with increasing loss of bowel, mirrored by a progressive increase in fecal fat content. We also describe significant morphological changes in the lymphatic vasculature in both the lamina propria and mesentery. Intestinal lymphatic drainage assay in vivo demonstrated a marked reduction of systemic absorption after resection. Intestinal lymphatic vessels significantly remodel in the setting of chronic SBS. This remodeling may account at least in part for impaired intestinal uptake and transport of fat via the compromised lymphatic architecture. We believe that these changes may contribute to the development of intestinal failure associated liver disease (IFALD), a major morbidity in patients with SBS.
Asunto(s)
Enfermedades Intestinales , Vasos Linfáticos , Síndrome del Intestino Corto , Animales , Absorción Intestinal , Intestinos , Lípidos , Vasos Linfáticos/diagnóstico por imagen , Ratones , Ratones Endogámicos C57BLRESUMEN
The unfolded protein response (UPR) is a complex adaptive signaling pathway activated by the accumulation of misfolded proteins in the endoplasmic reticulum (ER). ER stress (ERS) triggers a cascade of responses that converge upon C/EBP homologous protein (CHOP) to drive inflammation and apoptosis. Herein, we sought to determine whether liver injury and fibrosis after small bowel resection (SBR) were mediated by a maladaptive hepatic ERS/UPR. C57BL/6 mice underwent 50% proximal SBR or sham operation. Markers of liver injury and UPR/ERS pathways were analyzed. These were compared with experimental groups including dietary fat manipulation, tauroursodeoxycholic acid (TUDCA) treatment, distal SBR, and global CHOP knockout (KO). At 10 wk, proximal SBR had elevated alanine aminotransferase/aspartate aminotransferase (ALT/AST) (P < 0.005) and greater hepatic tumor necrosis factor-α (TNFα) (P = 0.001) and collagen type 1 α1 (COL1A1) (P = 0.02) than shams. SBR livers had increased CHOP and p-eIF2α, but were absent in activating transcription factor 4 (ATF4) protein expression. Low-fat diet (LFD), TUDCA, and distal SBR groups had decreased liver enzymes, inflammation, and fibrosis (P < 0.05). Importantly, they demonstrated reversal of hepatic UPR with diminished CHOP and robust ATF4 signal. CHOP KO-SBR had decreased ALT but not AST compared with wild-type (WT)-SBR (P = 0.01, P = 0.12). There were no differences in TNFα and COL1A1 (P = 0.09, P = 0.50). SBR-induced liver injury, fibrosis is associated with a novel hepatic UPR/ERS response characterized by increased CHOP and decreased ATF4. LFD, TUDCA, and ileocecal resection rescued the hepatic phenotype and reversed the UPR pattern. Global CHOP KO only partially attenuated liver injury. This underscores the significance of disruptions to the gut/liver axis after SBR and potentiates targets to mitigate the progression of intestinal failure-associated liver disease.NEW & NOTEWORTHY The unfolded protein response (UPR) is a complex signaling cascade that converges upon C/EBP-homologous protein (CHOP). Under conditions of chronic cellular stress, the UPR shifts from homeostatic to proapoptotic leading to inflammation and cell death. Here, we provide evidence that small bowel resection-induced liver injury and fibrosis are mediated by a maladaptive hepatic UPR. Low-fat diet, TUDCA treatment, and ileocecal resection rescued the hepatic phenotype and reversed the UPR pattern.
Asunto(s)
Enfermedad Hepática Crónica Inducida por Sustancias y Drogas , Factor de Necrosis Tumoral alfa , Animales , Apoptosis/genética , Estrés del Retículo Endoplásmico , Fibrosis , Inflamación/metabolismo , Cirrosis Hepática , Ratones , Ratones Endogámicos C57BL , Factor de Transcripción CHOP/genética , Factor de Transcripción CHOP/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Respuesta de Proteína DesplegadaRESUMEN
BACKGROUND: Murine ileocecal resection (ICR) has been used to investigate intestinal adaptation. The established model often includes the sacrifice of significant length of the proximal colon. Here, we optimized a highly selective vascular approach to the ICR, with primary jejunal-colic anastomosis yielding maximal colonic preservation. MATERIALS AND METHODS: Forty C57BL/6 mice underwent a highly vascularly selective ICR. The terminal branches of the ileocecal artery are isolated apart from the mesenteric branches supplying the small bowel to be resected. The distal 50% of small bowel and cecum are resected; a primary jejuno-colonic anastomosis is performed. Animals were sacrificed at postoperative weeks 2 (n = 10) and 10 (n = 29). Proximal 50% small bowel resection (SBR) with jejuno-ileal anastomosis was also performed for comparison. RESULTS: The entire colon (with exception of the cecum) was preserved in 100% of animals. Ninety-seven percent of animals survived to postoperative week 10, and all exhibited structural adaptation in the remnant small intestine epithelium. Crypts deepened by 175%, and villi lengthened by 106%, versus 39% and 29% in the proximal SBR cohort, respectively. Colonic proliferation, structural adaptation, and functional adaptation (measured by p-histone 3, luminal-facing apical crypt border size, and sucrase isomaltase, respectively) were increased in ICR compared with proximal SBR. CONCLUSIONS: Highly selective isolation of the cecal vasculature allows for greater colon preservation and yields enhanced remnant intestine epithelial adaptation. ICR is also associated with greater colonic adaptation and unique plasticity toward an intestinal phenotype. These findings underscore major differences between resection sites and offer insights into the critical adaptive mechanisms in response to massive intestinal loss.
Asunto(s)
Intestino Delgado , Síndrome del Intestino Corto , Adaptación Fisiológica/fisiología , Animales , Colon/cirugía , Humanos , Mucosa Intestinal , Intestino Delgado/fisiología , Intestino Delgado/cirugía , Yeyuno/cirugía , Ratones , Ratones Endogámicos C57BL , Síndrome del Intestino Corto/cirugíaRESUMEN
Adult stem cells maintain regenerative tissue structure and function by producing tissue-specific progeny, but the factors that preserve their tissue identities are not well understood. The small and large intestines differ markedly in cell composition and function, reflecting their distinct stem cell populations. Here we show that SATB2, a colon-restricted chromatin factor, singularly preserves LGR5+ adult colonic stem cell and epithelial identity in mice and humans. Satb2 loss in adult mice leads to stable conversion of colonic stem cells into small intestine ileal-like stem cells and replacement of the colonic mucosa with one that resembles the ileum. Conversely, SATB2 confers colonic properties on the mouse ileum. Human colonic organoids also adopt ileal characteristics upon SATB2 loss. SATB2 regulates colonic identity in part by modulating enhancer binding of the intestinal transcription factors CDX2 and HNF4A. Our study uncovers a conserved core regulator of colonic stem cells able to mediate cross-tissue plasticity in mature intestines.
Asunto(s)
Colon , Íleon , Animales , Mucosa Intestinal , Ratones , Organoides , Células MadreRESUMEN
Loss of functional small bowel surface area causes short bowel syndrome (SBS), intestinal failure, and parenteral nutrition (PN) dependence. The gut adaptive response following resection may be difficult to predict, and it may take up to 2 yr to determine which patients will wean from PN. Here, we examined features of gut microbiota and bile acid (BA) metabolism in determining adaptation and ability to wean from PN. Stool and sera were collected from healthy controls and from patients with SBS (n = 52) with ileostomy, jejunostomy, ileocolonic, and jejunocolonic anastomoses fed with PN plus enteral nutrition or who were exclusively enterally fed. We undertook 16S rRNA gene sequencing, BA profiling, and 7α-hydroxy-4-cholesten-3-one (C4) quantitation with LC-MS/MS and serum amino acid analyses. Patients with SBS exhibited altered gut microbiota with reduced gut microbial diversity compared with healthy controls. We observed differences in the microbiomes of patients with SBS with ileostomy versus jejunostomy, jejunocolonic versus ileocolonic anastomoses, and PN dependence compared with those who weaned from PN. Stool and serum BA composition and C4 concentrations were also altered in patients with SBS, reflecting adaptive changes in enterohepatic BA cycling. Stools from patients who were weaned from PN were enriched in secondary BAs including deoxycholic acid and lithocholic aicd. Shifts in gut microbiota and BA metabolites may generate a favorable luminal environment in select patients with SBS, promoting the ability to wean from PN. Proadaptive microbial species and select BA may provide novel targets for patient-specific therapies for SBS.NEW & NOTEWORTHY Loss of intestinal surface area causes short bowel syndrome, intestinal failure, and parenteral nutrition dependence. We analyzed the gut microbiota and bile acid metabolome of a large cohort of short bowel syndrome adult patients with different postsurgical anatomies. We report a novel analysis of the microbiome of patients with ileostomy and jejunostomy. Enrichment of specific microbial and bile acid species may be associated with the ability to wean from parenteral nutrition.
Asunto(s)
Ácidos y Sales Biliares/metabolismo , Heces/microbiología , ARN Ribosómico 16S/metabolismo , Síndrome del Intestino Corto/metabolismo , Adaptación Fisiológica/fisiología , Cromatografía Liquida , Microbioma Gastrointestinal/fisiología , Humanos , Intestino Delgado/metabolismo , Metaboloma/fisiología , Microbiota/fisiologíaRESUMEN
Congenital diaphragmatic hernia (CDH) is a severe congenital anomaly that is often accompanied by other anomalies. Although the role of genetics in the pathogenesis of CDH has been established, only a small number of disease-associated genes have been identified. To further investigate the genetics of CDH, we analyzed de novo coding variants in 827 proband-parent trios and confirmed an overall significant enrichment of damaging de novo variants, especially in constrained genes. We identified LONP1 (lon peptidase 1, mitochondrial) and ALYREF (Aly/REF export factor) as candidate CDH-associated genes on the basis of de novo variants at a false discovery rate below 0.05. We also performed ultra-rare variant association analyses in 748 affected individuals and 11,220 ancestry-matched population control individuals and identified LONP1 as a risk gene contributing to CDH through both de novo and ultra-rare inherited largely heterozygous variants clustered in the core of the domains and segregating with CDH in affected familial individuals. Approximately 3% of our CDH cohort who are heterozygous with ultra-rare predicted damaging variants in LONP1 have a range of clinical phenotypes, including other anomalies in some individuals and higher mortality and requirement for extracorporeal membrane oxygenation. Mice with lung epithelium-specific deletion of Lonp1 die immediately after birth, most likely because of the observed severe reduction of lung growth, a known contributor to the high mortality in humans. Our findings of both de novo and inherited rare variants in the same gene may have implications in the design and analysis for other genetic studies of congenital anomalies.
Asunto(s)
Proteasas ATP-Dependientes/genética , Proteasas ATP-Dependientes/fisiología , Anomalías Craneofaciales/genética , Variaciones en el Número de Copia de ADN , Anomalías del Ojo/genética , Trastornos del Crecimiento/genética , Hernias Diafragmáticas Congénitas/genética , Luxación Congénita de la Cadera/genética , Proteínas Mitocondriales/genética , Proteínas Mitocondriales/fisiología , Mutación Missense , Osteocondrodisplasias/genética , Anomalías Dentarias/genética , Animales , Estudios de Casos y Controles , Estudios de Cohortes , Anomalías Craneofaciales/patología , Anomalías del Ojo/patología , Femenino , Trastornos del Crecimiento/patología , Hernias Diafragmáticas Congénitas/patología , Luxación Congénita de la Cadera/patología , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Osteocondrodisplasias/patología , Linaje , Anomalías Dentarias/patologíaRESUMEN
The biogenesis of high-density lipoprotein (HDL) requires apoA1 and the cholesterol transporter ABCA1. Although the liver generates most of the HDL in the blood, HDL synthesis also occurs in the small intestine. Here, we show that intestine-derived HDL traverses the portal vein in the HDL3 subspecies form, in complex with lipopolysaccharide (LPS)-binding protein (LBP). HDL3, but not HDL2 or low-density lipoprotein, prevented LPS binding to and inflammatory activation of liver macrophages and instead supported extracellular inactivation of LPS. In mouse models involving surgical, dietary, or alcoholic intestinal insult, loss of intestine-derived HDL worsened liver injury, whereas outcomes were improved by therapeutics that elevated and depended upon raising intestinal HDL. Thus, protection of the liver from injury in response to gut-derived LPS is a major function of intestinally synthesized HDL.
Asunto(s)
Intestino Delgado/metabolismo , Lipoproteínas HDL3/metabolismo , Hepatopatías/prevención & control , Hígado/metabolismo , Vena Porta/metabolismo , Proteínas de Fase Aguda/metabolismo , Adulto , Animales , Proteínas Portadoras/metabolismo , HDL-Colesterol/sangre , HDL-Colesterol/metabolismo , Enterocitos/metabolismo , Humanos , Intestino Delgado/cirugía , Macrófagos del Hígado/inmunología , Macrófagos del Hígado/metabolismo , Receptores de Lipopolisacáridos/metabolismo , Lipopolisacáridos/metabolismo , Lipoproteínas HDL3/sangre , Hígado/patología , Cirrosis Hepática/patología , Cirrosis Hepática/prevención & control , Hepatopatías/patología , Receptores X del Hígado/metabolismo , Glicoproteínas de Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Unión Proteica , Transducción de Señal , Receptor Toll-Like 4/metabolismoRESUMEN
Surgical removal of the intestine, lifesaving in catastrophic gastrointestinal disorders of infancy, can result in a form of intestinal failure known as short bowel syndrome (SBS). Bloodstream infections (BSIs) are a major challenge in pediatric SBS management. BSIs require frequent antibiotic therapy, with ill-defined consequences for the gut microbiome and childhood health. Here, we combine serial stool collection, shotgun metagenomic sequencing, multivariate statistics and genome-resolved strain-tracking in a cohort of 19 patients with surgically-induced SBS to show that antibiotic-driven intestinal dysbiosis in SBS enriches for persistent intestinal colonization with BSI causative pathogens in SBS. Comparing the gut microbiome composition of SBS patients over the first 4 years of life to 19 age-matched term and 18 preterm controls, we find that SBS gut microbiota diversity and composition was persistently altered compared to controls. Commensals including Ruminococcus, Bifidobacterium, Eubacterium, and Clostridium species were depleted in SBS, while pathobionts (Enterococcus) were enriched. Integrating clinical covariates with gut microbiome composition in pediatric SBS, we identified dietary and antibiotic exposures as the main drivers of these alterations. Moreover, antibiotic resistance genes, specifically broad-spectrum efflux pumps, were at a higher abundance in SBS, while putatively beneficial microbiota functions, including amino acid and vitamin biosynthesis, were depleted. Moreover, using strain-tracking we found that the SBS gut microbiome harbors BSI causing pathogens, which can persist intestinally throughout the first years of life. The association between antibiotic-driven gut dysbiosis and enrichment of intestinal pathobionts isolated from BSI suggests that antibiotic treatment may predispose SBS patients to infection. Persistence of pathobionts and depletion of beneficial microbiota and functionalities in SBS highlights the need for microbiota-targeted interventions to prevent infection and facilitate intestinal adaptation.
Asunto(s)
Antibacterianos/uso terapéutico , Disbiosis/tratamiento farmacológico , Disbiosis/etiología , Microbioma Gastrointestinal/efectos de los fármacos , Sepsis/tratamiento farmacológico , Sepsis/etiología , Síndrome del Intestino Corto/complicaciones , Adolescente , Niño , Preescolar , Estudios de Cohortes , Disbiosis/microbiología , Femenino , Humanos , Masculino , Missouri , Síndrome del Intestino Corto/microbiologíaRESUMEN
INTRODUCTION: Prior data suggest that infants with gastroschisis are at high risk for hypothermia and infectious complications (ICs). This study evaluated the associations between perioperative hypothermia (PH) and ICs in gastroschisis using a multi-institutional cohort. METHODS: Retrospective review of infants with gastroschisis who underwent abdominal closure from 2013-2017 was performed at 7 children's hospitals. Any-IC and surgical site infection (SSI) were stratified against the presence or absence of PH, and perioperative characteristics associated with PH and SSI were determined using multivariable logistic regression. RESULTS: Of 256 gastroschisis neonates, 42% developed PH, with 18% classified as mild hypothermia (35.5-35.9⯰C), 10.5% as moderate (35.0-35.4⯰C), and 13% severe (<35⯰C). There were 82 (32%) ICs with 50 (19.5%) being SSIs. No associations between PH and any-IC (p = 0.7) or SSI (p = 0.98) were found. Pulmonary comorbidities (odds ratio (OR)=3.76, 95%CI:1.42-10, p = 0.008) and primary closure (OR=0.21, 95%CI:0.12-0.39, p<0.001) were associated with PH, while silo placement (OR=2.62, 95%CI:1.1-6.3, p = 0.03) and prosthetic patch (OR=3.42, 95%CI:1.4-8.3, p = 0.007) were associated with SSI on multivariable logistic regression. CONCLUSIONS: Primary abdominal closure and pulmonary comorbidities are associated with PH in gastroschisis, however PH was not associated with increased risk of ICs. Independent risk factors for SSI include silo placement and prosthetic patch closure.
Asunto(s)
Gastrosquisis , Hipotermia , Niño , Gastrosquisis/complicaciones , Gastrosquisis/epidemiología , Gastrosquisis/cirugía , Humanos , Hipotermia/epidemiología , Hipotermia/etiología , Lactante , Recién Nacido , Estudios Retrospectivos , Factores de Riesgo , Infección de la Herida Quirúrgica/epidemiología , Infección de la Herida Quirúrgica/etiología , Resultado del TratamientoRESUMEN
Intestinal failure-associated liver disease is a major morbidity associated with short bowel syndrome. We sought to determine if the obesity-resistant mouse strain (129S1/SvImJ) conferred protection from liver injury after small bowel resection (SBR). Using a parenteral nutrition-independent model of resection-associated liver injury, C57BL/6J and 129S1/SvImJ mice underwent a 50% proximal SBR or sham operation. At postoperative week 10, hepatic steatosis, fibrosis, and cholestasis were assessed. Hepatic and systemic inflammatory pathways were evaluated using oxidative markers and abundance of tissue macrophages. Potential mechanisms of endotoxin resistance were also explored. Serum lipid levels were elevated in all mouse lines. Hepatic triglyceride levels were no different between mouse strains, but there was an increased accumulation of free fatty acids in the C57BL/6J mice. Histological and serum markers of hepatic fibrosis, steatosis, and cholestasis were significantly elevated in resected C57BL/6J SBR mice as well as oxidative stress markers and macrophage recruitment in both the liver and visceral white fat in C57BL/6J mice compared with sham controls and the 129S1/SvImJ mouse line. Serum endotoxin levels were significantly elevated in C57BL/6J mice with significant elevation of hepatic TLR4 and reduction in PPARα expression levels. Despite high levels of serum lipids, 129S1/SvImJ mice did not develop liver inflammation, fibrosis, or cholestasis after SBR, unlike C57BL/6J mice. These data suggest that the accumulation of hepatic free fatty acids as well as increased endotoxin-driven inflammatory pathways through PPARα and TLR4 contribute to the liver injury seen in C57BL/6J mice with short bowel syndrome.NEW & NOTEWORTHY Unlike C57BL/6 mice, the 129S1/SvImJ strain is resistant to liver inflammation and injury after small bowel resection. These disparate outcomes are likely due to the accumulation of hepatic free fatty acids as well as increased endotoxin-driven inflammatory pathways through PPARα and TLR4 in C57BL/6 mice with short bowel syndrome.
Asunto(s)
Hepatopatías/etiología , Hígado/metabolismo , Síndrome del Intestino Corto/metabolismo , Tejido Adiposo Blanco/metabolismo , Animales , Biomarcadores/sangre , Procedimientos Quirúrgicos del Sistema Digestivo , Modelos Animales de Enfermedad , Endotoxinas/sangre , Ácidos Grasos no Esterificados/metabolismo , Intestino Delgado/cirugía , Lípidos/sangre , Cirrosis Hepática/metabolismo , Hepatopatías/metabolismo , Ratones , Ratones Endogámicos C57BL , Obesidad/metabolismo , Triglicéridos/metabolismoRESUMEN
BACKGROUND: Short bowel syndrome resulting from small bowel resection (SBR) is associated with significant morbidity and mortality. Many adverse sequelae including steatohepatitis and bacterial overgrowth are thought to be related to increased bacterial translocation, suggesting alterations in gut permeability. We hypothesized that after intestinal resection, the intestinal barrier is altered via toll-like receptor 4 (TLR4) signaling at the intestinal level. METHODS: B6 and intestinal-specific TLR4 knockout (iTLR4 KO) mice underwent 50% SBR or sham operation. Transcellular permeability was evaluated by measuring goblet cell associated antigen passages via two-photon microscopy. Fluorimetry and electron microscopy evaluation of tight junctions (TJ) were used to assess paracellular permeability. In parallel experiments, single-cell RNA sequencing measured expression of intestinal integral TJ proteins. Western blot and immunohistochemistry confirmed the results of the single-cell RNA sequencing. RESULTS: There were similar number of goblet cell associated antigen passages after both SBR and sham operation (4.5 versus 5.0, P > 0.05). Fluorescein isothiocyanate-dextran uptake into the serum after massive SBR was significantly increased compared with sham mice (2.13 ± 0.39 ng/µL versus 1.62 ± 0.23 ng/µL, P < 0.001). SBR mice demonstrated obscured TJ complexes on electron microscopy. Single-cell RNA sequencing revealed a decrease in TJ protein occludin (21%) after SBR (P < 0.05), confirmed with immunostaining and western blot analysis. The KO of iTLR4 mitigated the alterations in permeability after SBR. CONCLUSIONS: Permeability after SBR is increased via changes at the paracellular level. However, these alterations were prevented in iTLR4 mice. These findings suggest potential protein targets for restoring the intestinal barrier and obviating the adverse sequelae of short bowel syndrome.
Asunto(s)
Mucosa Intestinal/metabolismo , Síndrome del Intestino Corto/etiología , Uniones Estrechas/metabolismo , Receptor Toll-Like 4/metabolismo , Animales , Ratones Endogámicos C57BL , Ratones Noqueados , Permeabilidad , Síndrome del Intestino Corto/metabolismo , Uniones Estrechas/ultraestructura , Receptor Toll-Like 4/genéticaRESUMEN
BACKGROUND: Short bowel syndrome (SBS) results from significant loss of small intestinal length. In response to this loss, adaptation occurs, with Epidermal Growth Factor Receptor (EGFR) being a key driver. Besides enhanced enterocyte proliferation, we have revealed that adaptation is associated with angiogenesis. Further, we have found that small bowel resection (SBR) is associated with diminished oxygen delivery and elevated levels of hypoxia-inducible factor 1-alpha (HIF1α). METHODS: We ablated EGFR in the epithelium and endothelium as well as HIF1α in the epithelium, ostensibly the most hypoxic element. Using these mice, we determined the effects of these genetic manipulations on intestinal blood flow after SBR using photoacoustic microscopy (PAM), intestinal adaptation and angiogenic responses. Then, given that endothelial cells require a stromal support cell for efficient vascularization, we ablated EGFR expression in intestinal subepithelial myofibroblasts (ISEMFs) to determine its effects on angiogenesis in a microfluidic model of human small intestine. RESULTS: Despite immediate increased demand in oxygen extraction fraction measured by PAM in all mouse lines, were no differences in enterocyte and endothelial cell EGFR knockouts or enterocyte HIF1α knockouts by POD3. Submucosal capillary density was also unchanged by POD7 in all mouse lines. Additionally, EGFR silencing in ISEMFs did not impact vascular network development in a microfluidic device of human small intestine. CONCLUSIONS: Overall, despite the importance of EGFR in facilitating intestinal adaptation after SBR, it had no impact on angiogenesis in three cell types-enterocytes, endothelial cells, and ISEMFs. Epithelial ablation of HIF1α also had no impact on angiogenesis in the setting of SBS.
Asunto(s)
Subunidad alfa del Factor 1 Inducible por Hipoxia/fisiología , Intestino Delgado/irrigación sanguínea , Neovascularización Fisiológica , Síndrome del Intestino Corto/cirugía , Animales , Receptores ErbB/genética , Receptores ErbB/fisiología , Femenino , Células Endoteliales de la Vena Umbilical Humana , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Intestino Delgado/metabolismo , Masculino , Ratones , Técnicas Analíticas Microfluídicas , Miofibroblastos , Síndrome del Intestino Corto/metabolismoRESUMEN
PURPOSE: Congenital diaphragmatic hernia (CDH) is associated with significant mortality and long-term morbidity in some but not all individuals. We hypothesize monogenic factors that cause CDH are likely to have pleiotropic effects and be associated with worse clinical outcomes. METHODS: We enrolled and prospectively followed 647 newborns with CDH and performed genomic sequencing on 462 trios to identify de novo variants. We grouped cases into those with and without likely damaging (LD) variants and systematically assessed CDH clinical outcomes between the genetic groups. RESULTS: Complex cases with additional congenital anomalies had higher mortality than isolated cases (P = 8 × 10-6). Isolated cases with LD variants had similar mortality to complex cases and much higher mortality than isolated cases without LD (P = 3 × 10-3). The trend was similar with pulmonary hypertension at 1 month. Cases with LD variants had an estimated 12-17 points lower scores on neurodevelopmental assessments at 2 years compared with cases without LD variants, and this difference is similar in isolated and complex cases. CONCLUSION: We found that the LD genetic variants are associated with higher mortality, worse pulmonary hypertension, and worse neurodevelopment outcomes compared with non-LD variants. Our results have important implications for prognosis, potential intervention and long-term follow up for children with CDH.
Asunto(s)
Hernias Diafragmáticas Congénitas , Niño , Hernias Diafragmáticas Congénitas/genética , Humanos , Recién Nacido , Estudios RetrospectivosRESUMEN
OBJECTIVE: To describe current hospital guidelines and the opinions of extracorporeal membrane oxygenation leaders at U.S. children's hospitals concerning the use of extracorporeal membrane oxygenation for coronavirus disease 2019-positive pediatric patients. DESIGN: Confidential, self-administered questionnaire. SETTING: One hundred twenty-seven U.S. pediatric extracorporeal membrane oxygenation centers. SUBJECTS: Extracorporeal membrane oxygenation center program directors and coordinators. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: In March 2020, a survey was sent to 127 pediatric extracorporeal membrane oxygenation centers asking them to report their current hospital extracorporeal membrane oxygenation guidelines for coronavirus disease 2019-positive patients. Respondents were also asked their opinion on three ethical dilemmas including: prioritization of children over adults for extracorporeal membrane oxygenation use, institution of do-not-resuscitate orders, and the use of extracorporeal cardiopulmonary resuscitation for coronavirus disease 2019-positive patients. Forty-seven extracorporeal membrane oxygenation centers had enacted guidelines including 46 (100%) that offer venovenous-extracorporeal membrane oxygenation and 42 (89%) that offer venoarterial-extracorporeal membrane oxygenation for coronavirus disease 2019-positive pediatric patients. Forty-four centers (94%) stated that the indications for extracorporeal membrane oxygenation candidacy in coronavirus disease 2019 disease were similar to those used in other viral illnesses, such as respiratory syncytial virus or influenza. Most program directors (98%) did not endorse that children hospitalized with coronavirus disease 2019 should be made do-not-resuscitate and had variable opinions on whether children should be given higher priority over adults when rationing extracorporeal membrane oxygenation. Over half of program directors (60%) did not support the use of extracorporeal cardiopulmonary resuscitation for coronavirus disease 2019. CONCLUSIONS: The majority of pediatric extracorporeal membrane oxygenation centers have proactively established guidelines for the use of extracorporeal membrane oxygenation for coronavirus disease 2019-related illnesses. Further work is needed to help guide the fair allocation of extracorporeal membrane oxygenation resources and to determine the appropriateness of extracorporeal cardiopulmonary resuscitation.
Asunto(s)
Infecciones por Coronavirus/terapia , Oxigenación por Membrana Extracorpórea/métodos , Hospitales Pediátricos , Neumonía Viral/terapia , Guías de Práctica Clínica como Asunto , Adulto , Betacoronavirus , COVID-19 , Reanimación Cardiopulmonar , Niño , Oxigenación por Membrana Extracorpórea/legislación & jurisprudencia , Femenino , Humanos , Lactante , Masculino , Persona de Mediana Edad , Pandemias , Pediatría , Órdenes de Resucitación , SARS-CoV-2 , Encuestas y Cuestionarios , Estados UnidosRESUMEN
After 50% proximal small bowel resection (SBR) in mice, we have demonstrated hepatic steatosis, impaired glucose metabolism without insulin resistance, and increased pancreatic islet area. We sought to determine the consequences of SBR on pancreatic ß-cell morphology, proliferation, and expression of a key regulatory hormone, glucagon-like peptide-1 (GLP-1). C57BL/6 mice underwent 50% SBR or sham operation. At 10 wk, pancreatic insulin content and secretion was measured by ELISA. Immunohistochemistry was performed to determine structural alterations in pancreatic α-and ß-cells. Western blot analysis was used to measure GLP-1R expression, and immunoassay was used to measure plasma insulin and GLP-1. Experiments were repeated by administering a GLP-1 agonist (exendin-4) to a cohort of mice following SBR. After SBR, there was pancreatic islet hypertrophy and impaired glucose tolerance. The proportion of α and ß cells was not grossly altered. Whole pancreas and pancreatic islet insulin content was not significantly different; however, SBR mice demonstrated decreased insulin secretion in both static incubation and islet perfusion experiments. The expression of pancreatic GLP-1R was decreased approximately twofold after SBR, compared with sham and serum GLP-1, was decreased. These metabolic derangements were mitigated after administration of the GLP-1 agonist. Following massive SBR, there is significant hypertrophy of pancreatic islet cells with morphologically intact α- and ß-cells. Significantly reduced pancreatic insulin release in both static and dynamic conditions demonstrate a perturbed second phase of insulin secretion. GLP-1 is a key mediator of this amplification pathway. Decreased expression of serum GLP-1 and pancreatic GLP-1R in face of no change in insulin content presents a novel pathway for enteropancreatic glucose regulation following SBR.NEW & NOTEWORTHY Metabolic changes occur following intestinal resection; however, the effects on pancreatic function are unknown. Prior studies have demonstrated that glucagon-like protein-1 (GLP-1) signaling is a crucial player in the improved insulin sensitivity after bariatric surgery. In this study, we explore the effect of massive small bowel resection on gut hormone physiology and provide novel insights into the enteropancreatic axis.