RESUMEN
Reduction-oxidation (redox) chemistry plays a vital role in human homeostasis. These reactions play critical roles in energy generation, as part of innate immunity, and in the generation of secondary messengers with various functions such as cell cycle progression or the release of neurotransmitters. Despite this cornerstone role, if left unchecked, the body can overproduce reactive oxygen species (ROS) or reactive nitrogen species (RNS). When these overwhelm endogenous antioxidant systems, oxidative stress (OS) occurs. In neonates, OS has been associated with retinopathy of prematurity (ROP), leukomalacia, and bronchopulmonary dysplasia (BPD). Given its broad spectrum of effects, research has started to examine whether OS plays a role in necrotizing enterocolitis (NEC). In this paper, we will discuss the basics of redox chemistry and how the human body keeps these in check. We will then discuss what happens when these go awry, focusing mostly on NEC in neonates.
Asunto(s)
Enterocolitis Necrotizante , Oxidación-Reducción , Estrés Oxidativo , Especies de Nitrógeno Reactivo , Especies Reactivas de Oxígeno , Humanos , Enterocolitis Necrotizante/metabolismo , Enterocolitis Necrotizante/patología , Especies Reactivas de Oxígeno/metabolismo , Recién Nacido , Especies de Nitrógeno Reactivo/metabolismo , Antioxidantes/metabolismo , AnimalesRESUMEN
BACKGROUND: The notable decline in the number of Tregs within Necrotizing enterocolitis (NEC) intestinal tissuesï¼contribute to excessive inflammation and necrosis, yet the precise underlying factors remain enigmatic. Ferroptosis, a novel cell death stemming from a disrupted lipid redox metabolism, is the focus of this investigation. Specifically, this study delves into the ferroptosis of Treg cells in the context of NEC and observes the protective effects exerted by vitamin E intervention, which aims to mitigate ferroptosis of Treg cells. METHODS: To investigate the reduction of Treg cells in NEC intestine, we analyzed its association with ferroptosis from multiple angles. We constructed a mouse with a specific knockout of Gpx4 in Treg cells, aiming to examine the impact of Treg cell ferroptosis on NEC intestinal injury and localized inflammation. Ultimately, we employed vitamin E treatment to mitigate ferroptosis in NEC intestine's Treg cells, monitoring the subsequent amelioration in intestinal inflammatory damage. RESULTS: The diminution of Treg cells in NEC is attributed to ferroptosis stemming from diminished GPX4 expression. Gpx4-deficient Treg cells exhibit impaired immunosuppressive function and are susceptible to ferroptosis. This ferroptosis of Treg cells exacerbates intestinal damage and inflammatory response in NEC. Notably, Vitamin E can inhibit the ferroptosis of Treg cells, subsequently alleviating intestinal damage and inflammation in NEC. Additionally, Vitamin E bolsters the anti-lipid peroxidation capability of Treg cells by upregulating the expression of GPX4. CONCLUSION: In the context of NEC, the ferroptosis of Treg cells represents a significant factor contributing to intestinal tissue damage and an exaggerated inflammatory response. GPX4 is pivotal for the viability and functionality of Treg cells. Vitamin E exhibits the capability to mitigate the ferroptosis of Treg cells, thereby enhancing their number and function, which plays a crucial role in mitigating intestinal tissue damage and inflammatory response in NEC.
Asunto(s)
Enterocolitis Necrotizante , Ferroptosis , Fosfolípido Hidroperóxido Glutatión Peroxidasa , Linfocitos T Reguladores , Vitamina E , Animales , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Vitamina E/farmacología , Fosfolípido Hidroperóxido Glutatión Peroxidasa/metabolismo , Fosfolípido Hidroperóxido Glutatión Peroxidasa/genética , Ratones , Enterocolitis Necrotizante/metabolismo , Enterocolitis Necrotizante/patología , Enterocolitis Necrotizante/tratamiento farmacológico , Modelos Animales de Enfermedad , Inflamación/metabolismo , Inflamación/patología , Humanos , Ratones Noqueados , Intestinos/patologíaRESUMEN
BACKGROUND: Neonatal necrotizing enterocolitis (NEC) is one of the most prevalent and severe intestinal emergencies in newborns. The inflammatory activation of macrophages is associated with the intestinal injury of NEC. The neuroimmune regulation mediated by α7 nicotinic acetylcholine receptor (α7nAChR) plays an important role in regulating macrophage activation and inflammation progression, but in NEC remains unclear. This study aims to explore the effect of macrophage α7nAChR on NEC. METHODS: Mice NEC model were conducted with high-osmolarity formula feeding, hypoxia, and cold stimulation. The α7nAChR agonist PNU-282987 and mTOR inhibitor rapamycin were treated by intraperitoneal injections in mice. The expression and distribution of macrophages, α7nAChR, and phospho-mammalian target of rapamycin (p-mTOR) in the intestines of NEC patients and mice was assessed using immunohistochemistry, immunofluorescence, and flow cytometry. The expression of NLRP3, activated caspase-1 and IL-1ß in mice intestines was detected by flow cytometry, western blot or ELISA. In vitro, the mouse RAW264.7 macrophage cell line was also cultured followed by various treatments. Expression of p-mTOR, NLRP3, activated caspase-1, and IL-1ß in macrophages was determined. RESULTS: Macrophages accumulated in the intestines and the expression of α7nAChR in the mucosal and submucosal layers of the intestines was increased in both the NEC patients and mice. The p-mTOR and CD68 were increased and co-localized in intestines of NEC patients. In vitro, α7nAChR agonist PNU-282987 significantly reduced the increase of NLRP3, activated caspase-1, and IL-1ß in macrophages. PNU-282987 also significantly reduced the increase of p-mTOR. The effect was blocked by AMPK inhibitor compound C. The expression of NLRP3, activated caspase-1, and IL-1ß was inhibited after mTOR inhibitor rapamycin treatment. In NEC model mice, PNU-282987 reduced the expression of p-mTOR, NLRP3, activated caspase-1, and IL-1ß in the intestine. Meanwhile, rapamycin significantly attenuated NLRP3 activation and the release of IL-1ß. Moreover, the proportion of intestinal macrophages and intestinal injury decreased after PNU-282987 treatment. CONCLUSION: Macrophage α7nAChR activation mitigates NLRP3 inflammasome activation by modulating mTOR phosphorylation, and subsequently alleviates intestinal inflammation and injury in NEC.
Asunto(s)
Benzamidas , Compuestos Bicíclicos con Puentes , Enterocolitis Necrotizante , Interleucina-1beta , Macrófagos , Proteína con Dominio Pirina 3 de la Familia NLR , Transducción de Señal , Serina-Treonina Quinasas TOR , Receptor Nicotínico de Acetilcolina alfa 7 , Animales , Receptor Nicotínico de Acetilcolina alfa 7/metabolismo , Receptor Nicotínico de Acetilcolina alfa 7/agonistas , Enterocolitis Necrotizante/tratamiento farmacológico , Enterocolitis Necrotizante/metabolismo , Enterocolitis Necrotizante/patología , Enterocolitis Necrotizante/inmunología , Ratones , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Humanos , Compuestos Bicíclicos con Puentes/farmacología , Compuestos Bicíclicos con Puentes/uso terapéutico , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Macrófagos/inmunología , Interleucina-1beta/metabolismo , Benzamidas/farmacología , Benzamidas/uso terapéutico , Recién Nacido , Transducción de Señal/efectos de los fármacos , Células RAW 264.7 , Modelos Animales de Enfermedad , Masculino , Ratones Endogámicos C57BL , Animales Recién Nacidos , FemeninoRESUMEN
Necrotizing enterocolitis (NEC) is a complex, multifactorial gastrointestinal disorder predominantly affecting preterm infants. The pathogenesis of this condition involves a complex interplay between intestinal barrier dysfunction, microbial dysbiosis, and an altered immune response. This study investigates the potential role of endogenous hyaluronan (HA) in both the early phases of intestinal development and in the context of NEC-like intestinal injury. We treated neonatal CD-1 mouse pups with PEP1, a peptide inhibiting HA receptor interactions, from postnatal days 8 to 12. We evaluated postnatal intestinal developmental indicators, such as villi length, crypt depth, epithelial cell proliferation, crypt fission, and differentiation of goblet and Paneth cells, in PEP1-treated animals compared with those treated with scrambled peptide. PEP1 treatment significantly impaired intestinal development, as evidenced by reductions in villi length, crypt depth, and epithelial cell proliferation, along with a decrease in crypt fission activity. These deficits in PEP1-treated animals correlated with increased susceptibility to NEC-like injuries, including higher mortality rates, and worsened histological intestinal injury. These findings highlight the role of endogenous HA in supporting intestinal development and protecting against NEC.
Asunto(s)
Enterocolitis Necrotizante , Homeostasis , Ácido Hialurónico , Intestinos , Animales , Ácido Hialurónico/farmacología , Ácido Hialurónico/metabolismo , Enterocolitis Necrotizante/patología , Enterocolitis Necrotizante/metabolismo , Enterocolitis Necrotizante/tratamiento farmacológico , Ratones , Homeostasis/efectos de los fármacos , Intestinos/patología , Intestinos/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Animales Recién Nacidos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patología , Mucosa Intestinal/efectos de los fármacos , Modelos Animales de EnfermedadRESUMEN
Intestinal dysbiosis is believed to play a role in the development of necrotizing enterocolitis (NEC). The efficacy of JNK-inhibitory peptide (CPJIP) in treating NEC was assessed. Treatment with CPJIP led to a notable reduction in p-JNK expression in IEC-6 cells and NEC mice. Following LPS stimulation, the expression of RNA and protein of claudin-1, claudin-3, claudin-4 and occludin was significantly decreased, with this decrease being reversed by CPJIP administration, except for claudin-3, which remained consistent in NEC mice. Moreover, the expression levels of the inflammatory factors TNF-α, IL-1ß and IL-6 were markedly elevated, a phenomenon that was effectively mitigated by the addition of CPJIP in both IEC-6 cells and NEC mice. CPJIP administration resulted in improved survival rates, ameliorated microscopic intestinal mucosal injury, and increased the total length of the intestines and colon in NEC mice. Additionally, CPJIP treatment led to a reduction in serum concentrations of FD-4, D-lactate and DAO. Furthermore, our results revealed that CPJIP effectively inhibited intestinal cell apoptosis and promoted cell proliferation in the intestine. This study represents the first documentation of CPJIP's ability to enhance the expression of tight junction components, suppress inflammatory responses, and rescue intestinal cell fate by inhibiting JNK activation, ultimately mitigating intestinal severity. These findings suggest that CPJIP has the potential to serve as a promising candidate for the treatment of NEC.
Asunto(s)
Apoptosis , Enterocolitis Necrotizante , Inflamación , Mucosa Intestinal , Enterocolitis Necrotizante/tratamiento farmacológico , Enterocolitis Necrotizante/metabolismo , Enterocolitis Necrotizante/patología , Animales , Ratones , Mucosa Intestinal/metabolismo , Mucosa Intestinal/efectos de los fármacos , Mucosa Intestinal/patología , Inflamación/metabolismo , Inflamación/tratamiento farmacológico , Inflamación/patología , Apoptosis/efectos de los fármacos , Péptidos/farmacología , Modelos Animales de Enfermedad , Proliferación Celular/efectos de los fármacos , Ratones Endogámicos C57BL , Línea Celular , Ratas , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Lipopolisacáridos , Funcion de la Barrera IntestinalRESUMEN
AIMS: Neonatal necrotizing enterocolitis (NEC) is a leading cause of intestine inflammatory disease, and macrophage is significantly activated during NEC development. Posttranslational modifications (PTMs) of proteins, particularly ubiquitination, play critical roles in immune response. This study aimed to investigate the effects of ubiquitin-modified proteins on macrophage activation and NEC, and discover novel NEC-related inflammatory proteins. MATERIALS AND METHODS: Proteomic and ubiquitin proteomic analyses of intestinal macrophages in NEC/healthy mouse pups were carried out. In vitro macrophage inflammation model and in vivo NEC mouse model, as well as clinical human samples were used for further verification the inhibitor of nuclear factor-κB kinase α (IKKα) ubiquitination on NEC development through Western blot, immunofluorescence, quantitative real-time polymerase chain reaction (qRT-PCR) and flow cytometry. KEY FINDINGS: We report here that IKKα was a new ubiquitin-modified protein during NEC through ubiquitin proteomics, and RING finger protein 31 (RNF31) acted as an E3 ligase to be involved in IKKα degradation. Inhibition of IKKα ubiquitination and degradation with siRNF31 or proteasome inhibitor decreased nuclear factor-κB (NF-κB) activation, thereby decreasing the expression of pro-inflammatory factors and M1 macrophage polarization, resulting in reliving the severity of NEC. SIGNIFICANCE: Our study suggests the activation of RNF31-IKKα-NF-κB axis triggering NEC development and suppressing RNF31-mediated IKKα degradation may be therapeutic strategies to be developed for NEC treatment.
Asunto(s)
Enterocolitis Necrotizante , Quinasa I-kappa B , Inflamación , FN-kappa B , Ubiquitina-Proteína Ligasas , Ubiquitinación , Animales , Femenino , Humanos , Recién Nacido , Masculino , Ratones , Animales Recién Nacidos , Modelos Animales de Enfermedad , Enterocolitis Necrotizante/metabolismo , Enterocolitis Necrotizante/patología , Quinasa I-kappa B/metabolismo , Inflamación/metabolismo , Inflamación/patología , Intestinos/patología , Macrófagos/metabolismo , Ratones Endogámicos C57BL , FN-kappa B/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitina-Proteína Ligasas/genéticaRESUMEN
BACKGROUND: Imbalanced intestinal microbiota and damage to the intestinal barrier contribute to the development of necrotizing enterocolitis (NEC). Autoinducer-2 (AI-2) plays a crucial role in repairing intestinal damage and reducing inflammation. OBJECTIVE: This study aimed to investigate the impact of AI-2 on the expression of intestinal zonula occludens-1 (ZO-1) and occludin proteins in NEC. We evaluated its effects in vivo using NEC mice and in vitro using lipopolysaccharide (LPS)-stimulated intestinal cells. METHODS: Pathological changes in the intestines of neonatal mice were assessed using histological staining and scoring. Cell proliferation was measured using the cell counting kit-8 (CCK-8) assay to determine the optimal conditions for LPS and AI-2 interventions. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to analyze the mRNA levels of matrix metalloproteinase-3 (MMP3), protease activated receptor-2 (PAR2), interleukin-1ß (IL-1ß), and IL-6. Protein levels of MMP3, PAR2, ZO-1, and occludin were evaluated using western blot, immunohistochemistry, or immunofluorescence. RESULTS: AI-2 alleviated NEC-induced intestinal damage (P < 0.05) and enhanced the proliferation of damaged IEC-6 cells (P < 0.05). AI-2 intervention reduced the mRNA and protein expressions of MMP3 and PAR2 in intestinal tissue and cells (P < 0.05). Additionally, it increased the protein levels of ZO-1 and occludin (P < 0.05), while reducing IL-1ß and IL-6 mRNA expression (P < 0.05). CONCLUSION: AI-2 intervention enhances the expression of tight junction proteins (ZO-1 and occludin), mitigates intestinal damage in NEC neonatal mice and IEC-6 cells, potentially by modulating PAR2 and MMP3 signaling. AI-2 holds promise as a protective intervention for NEC. AI-2 plays a crucial role in repairing intestinal damage and reducing inflammation.
Asunto(s)
Enterocolitis Necrotizante , Metaloproteinasa 3 de la Matriz , Receptor PAR-2 , Transducción de Señal , Animales , Humanos , Ratones , Animales Recién Nacidos , Línea Celular , Proliferación Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Enterocolitis Necrotizante/patología , Enterocolitis Necrotizante/tratamiento farmacológico , Enterocolitis Necrotizante/metabolismo , Homoserina/análogos & derivados , Homoserina/farmacología , Mucosa Intestinal/patología , Mucosa Intestinal/efectos de los fármacos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/inmunología , Intestinos/patología , Intestinos/efectos de los fármacos , Lactonas/farmacología , Lipopolisacáridos , Metaloproteinasa 3 de la Matriz/metabolismo , Metaloproteinasa 3 de la Matriz/genética , Ratones Endogámicos C57BL , Ocludina/metabolismo , Ocludina/genética , Receptor PAR-2/metabolismo , Receptor PAR-2/genética , Transducción de Señal/efectos de los fármacos , Proteína de la Zonula Occludens-1/metabolismo , Proteína de la Zonula Occludens-1/genéticaRESUMEN
Necrotizing enterocolitis (NEC) is one of the most common and serious intestinal illnesses in newborns and seriously affects their long-term prognosis and survival. Butyrate is a short-chain fatty acid that can relieve intestinal inflammation, but its mechanism of action is unclear. Results from an in vivo neonatal rat model has shown that butyrate caused an improved recovery from NEC. These protective effects were associated with the metabolite of hesperetin, as determined by metabolomics and molecular biological analysis. Furthermore, transcriptomics combined with inhibitor assays were used to investigate the mechanism of action of hesperetin in an in vitro NEC model (IEC-6 cells exposed to LPS) to further investigate the mechanism by which butyrate attenuates NEC. The transcriptomics analysis showed that the PI3K-Akt signaling pathway was involved in the anti-NEC effect of hesperitin. Subsequently, the results using an inhibitor of PI3K (LY294002) indicated that the suppression could be explained by the hesperetin-induced expression of tight junction (TJ) proteins by potentially blocking the PI3K-Akt signaling pathway. In summary, the present study demonstrated that butyrate could improve recovery from NEC with a hesperetin metabolite, causing potential inhibition of the phosphorylation of the PI3K-Akt signaling pathway, resulting in the increased expression of TJ proteins. These findings reveal a potential new therapeutic pathway for the treatment of NEC.
Asunto(s)
Enterocolitis Necrotizante , Hesperidina , Fosfatidilinositol 3-Quinasas , Proteínas Proto-Oncogénicas c-akt , Ratas Sprague-Dawley , Transducción de Señal , Enterocolitis Necrotizante/tratamiento farmacológico , Enterocolitis Necrotizante/metabolismo , Enterocolitis Necrotizante/patología , Hesperidina/farmacología , Animales , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal/efectos de los fármacos , Fosfatidilinositol 3-Quinasas/metabolismo , Ratas , Animales Recién Nacidos , Modelos Animales de Enfermedad , Butiratos/farmacología , Línea CelularRESUMEN
OBJECTIVE: Neonatal necrotizing enterocolitis (NEC) is a serious intestinal inflammatory disease. We investigated intestinal fatty acid binding protein (I-FABP), I-FABP mRNA, and interleukin-6 (IL-6) as potential diagnostic biomarkers in NEC. METHODS: Forty mice were subjected to hypoxic-ischemic intestinal injury, and then serum I-FABP protein and mRNA levels were quantified. Ileal tissue pathological scores were determined by hematoxylin and eosin staining. I-FABP expression levels and translocation in these tissues were detected using western blotting and immunofluorescence, respectively. Samples from 30 human neonates with NEC and 30 healthy neonates had serum I-FABP protein/mRNA and IL-6 levels measured. RESULTS: The mouse ileal tissue pathological score and I-FABP levels, as well as serum I-FABP and I-FABP mRNA levels, were significantly higher in the model group than in the control group. Serum I-FABP, I-FABP mRNA, and IL-6 levels were significantly higher in human neonates with NEC than in the healthy group. Logistic regression and receiver operating curve analyses revealed that I-FABP protein/mRNA and IL-6 levels could be diagnostic biomarkers for NEC. CONCLUSIONS: I-FABP protein/mRNA and IL-6 levels are useful biomarkers of intestinal ischemic injury in neonates with NEC. The combined detection of I-FABP protein/mRNA and IL-6 is recommended rather than using a single biomarker.
Asunto(s)
Biomarcadores , Modelos Animales de Enfermedad , Enterocolitis Necrotizante , Proteínas de Unión a Ácidos Grasos , Interleucina-6 , Ratones Endogámicos BALB C , ARN Mensajero , Enterocolitis Necrotizante/metabolismo , Enterocolitis Necrotizante/sangre , Enterocolitis Necrotizante/patología , Enterocolitis Necrotizante/genética , Enterocolitis Necrotizante/diagnóstico , Animales , Proteínas de Unión a Ácidos Grasos/sangre , Proteínas de Unión a Ácidos Grasos/genética , Proteínas de Unión a Ácidos Grasos/metabolismo , Interleucina-6/sangre , Interleucina-6/genética , Recién Nacido , Humanos , Biomarcadores/sangre , Biomarcadores/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Mensajero/sangre , Ratones , Masculino , Femenino , Animales Recién Nacidos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patología , Íleon/metabolismo , Íleon/patología , Estudios de Casos y Controles , Curva ROCRESUMEN
Necrotising enterocolitis (NEC) has a complex pathophysiology but the common end-point is ischaemia reperfusion injury (IRI) and intestinal necrosis. We have previously reported that RIC significantly reduces the intestinal injury in a rat model of NEC. Here we describe the changes in intestinal mRNA occurring in the intestine of animals exposed to IRI, both with and without RIC. Related rat-pups were randomly assigned to four groups: SHAM, IRI only, RIC only and RIC + IRI. IRI animals, underwent 40 min of intestinal ischaemia, and 90 min of reperfusion. Animals that underwent RIC had three cycles of 5 min of alternating ischaemia/reperfusion by means of a ligature applied to the hind limb. Samples from the terminal ileum were immediately stored in RNA-preserving media for later next generation sequencing and transciptome analysis using R v 3.6.1. Differential expression testing showed that 868 genes differentially expressed in animals exposed to RIC alone compared to SHAM and 135 in the IRI and RIC group compared to IRI alone. Comparison between these two sets showed that 25 genes were differentially expressed in both groups. Pro-inflammatory molecules: NF-ĸß2, Cxcl1, SOD2 and Map3k8 all show reduced expression in response to RIC. Targeted gene analysis revealed increased expression in PI3K which is part of the so-called RISK-pathway which is a key part of the protective mechanisms of RIC in the heart. Overall, this transcriptomic analysis shows that RIC provides a protective effect to the intestine via anti-inflammatory pathways. This could be particularly relevant to treating and preventing NEC.
Asunto(s)
Modelos Animales de Enfermedad , Enterocolitis Necrotizante , Perfilación de la Expresión Génica , Daño por Reperfusión , Animales , Enterocolitis Necrotizante/genética , Enterocolitis Necrotizante/patología , Enterocolitis Necrotizante/metabolismo , Ratas , Daño por Reperfusión/genética , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Precondicionamiento Isquémico/métodos , TranscriptomaRESUMEN
BACKGROUND & AIMS: Necrotizing enterocolitis (NEC) is a life-threatening disease affecting mostly the ileum of preemies. Intestinal epithelial cell (IEC) apoptosis contributes to NEC pathogenesis. However, how scattered crypt IEC apoptosis leads to NEC with excessive villus epithelial necrosis remains unclear. METHODS: A novel triple-transgenic mouse model, namely, 3xTg-iAPcIEC (inducible apoptosis phenotype in crypt-IEC), was developed to induce IEC-specific overexpression of Fasl transgene using doxycycline (Dox)-inducible tetO-rtTA system and villin-cre technology. The 3-days-old neonatal 3xTg-iAPcIEC mice and their littermate controls were subcutaneously (s.c.) challenged with a single dose of Dox. Intestinal tissues were processed at different time points to examine scattered crypt IEC apoptosis-mediated NEC development. Gene knockout technology, antibody-mediated cell depletion, and antibiotic-facilitated Gram-positive bacteria depletion were used to study mechanisms. RESULTS: Treatment of 3xTg-iAPcIEC mouse pups with Dox induces scattered crypt IEC apoptosis followed by crypt inflammation and excessive villous necrosis resembling NEC. This progression correlated with elevated Ifng, Rip3, CD8+ T cells, and Gram-positive bacteria in the ileum. Mechanistically, IFN-γ and RIP3-activated signals mediate the effect of scattered crypt IEC apoptosis on the induction of intestinal crypt inflammation and villous necrosis. Meanwhile, pathophysiological events of CD8+ T cell infiltration and dysbiosis with Gram-positive bacteria primarily contribute to excessive villous inflammation and necrosis. Notably, blocking any of these events protects against NEC development in 3xTg-iAPcIEC mouse pups, underlining their central roles in NEC pathogenesis. CONCLUSIONS: Scattered crypt IEC apoptosis induces NEC in mouse pups via IFN-γ, RIP3, CD8+ T cells, and Gram-positive bacteria-mediated comprehensive pathophysiological events. Our findings may advance knowledge in the prevention and treatment of NEC.
Asunto(s)
Apoptosis , Modelos Animales de Enfermedad , Enterocolitis Necrotizante , Proteína Ligando Fas , Interferón gamma , Mucosa Intestinal , Ratones Transgénicos , Animales , Enterocolitis Necrotizante/patología , Enterocolitis Necrotizante/metabolismo , Ratones , Mucosa Intestinal/patología , Mucosa Intestinal/microbiología , Mucosa Intestinal/inmunología , Interferón gamma/metabolismo , Proteína Ligando Fas/metabolismo , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Proteína Serina-Treonina Quinasas de Interacción con Receptores/genética , Células Epiteliales/patología , Células Epiteliales/metabolismo , Linfocitos T CD8-positivos/inmunología , Necrosis , Animales Recién Nacidos , Doxiciclina/farmacología , Humanos , Íleon/patología , Íleon/inmunologíaRESUMEN
BACKGROUND Necrotizing enterocolitis (NEC) is a potentially life-threatening disease that affects the intestine of the neonate, causing necrosis and general inflammation. Treatment consists of feeding cessation and antibiotic therapy. In more severe cases, surgical intervention is necessary. Recently, different NEC models have been used to study the development of novel diagnostic and therapeutic methods. This work modified an experimental NEC model in rat pups by a single exposure of animals to NEC-causing factors and testing the impact of mother's milk on prevalence of the disease. MATERIAL AND METHODS Fifty rat pups were subjected to the NEC protocol, in which they were exposed to 100% nitrogen atmosphere and cold stress for set periods of time and formula feeding with exposure to mother's milk and artificial milk. Twenty-nine pups were used for control. After a set time of 72 h, bowel fragments were obtained and examined histologically by hematoxylin-eosin staining with a modified 3-grade scale. RESULTS Histological features of NEC were present in most of the samples (10/14) in the group exposed to 1 min of hypoxia (P=0.016), 10 min of cold stress (P=0.4) and formula feeding every 3 h with no mother's milk (P=0.001). In the group of 11 animals with the same stress conditions but fed mother's milk right after birth, only 1 sample of NEC was present. CONCLUSIONS The modified experimental NEC model based on formula feeding and single exposure to hypothermia and hypoxia was assessed statistically and histologically. In this model, mother's milk had a protective effect against necrotizing enterocolitis.
Asunto(s)
Animales Recién Nacidos , Modelos Animales de Enfermedad , Enterocolitis Necrotizante , Hipoxia , Leche , Animales , Enterocolitis Necrotizante/patología , Ratas , Hipoxia/complicaciones , Leche/metabolismo , Hipotermia , Femenino , Incidencia , Ratas Sprague-DawleyRESUMEN
The near-infrared spectroscopy is often used to distinguish small bowel necrosis due to necrotizing enterocolitis (NEC). The characteristic bands of small bowel necrosis, as an important basis for evaluating the confidence of the differentiation results, are challenging to identify quickly. In this study, we proposed to identify characteristic bands of lesion samples based on hyperspectral imaging (HSI) and cellwise outlier detection. Rabbits were used as an animal model to simulate the clinical symptoms of NEC. The rabbits were detected at intervals of 10, 30, 60, and 90 min. The characteristic bands were identified within the same rabbit, between different rabbits and at different times. The result showed the bands near 763 nm, corresponding to the absorption peak of deoxyhemoglobin, were the characteristic bands separating samples with NEC. The identification result was plausible because hypoxia was the main cause of NEC. The method was easy to perform.
Asunto(s)
Algoritmos , Enterocolitis Necrotizante , Intestino Delgado , Necrosis , Espectroscopía Infrarroja Corta , Animales , Conejos , Intestino Delgado/patología , Intestino Delgado/diagnóstico por imagen , Enterocolitis Necrotizante/patología , Enterocolitis Necrotizante/diagnóstico por imagen , Imágenes HiperespectralesRESUMEN
BACKGROUND AND AIM: Necrotizing Enterocolitis (NEC) is an inflammation-associated ischemic necrosis of the intestine. To investigate the effects of extra virgin olive oil (EVOO) on inflammation, oxidative stress, apoptosis, and histological changes in NEC-induced newborn rats. MATERIALS AND METHODS: 24 rats were randomly divided into three groups: control, NEC and NEC + EVOO. NEC induction was performed using hypoxia-hyperoxia, formula feeding, and cold stress. The NEC + EVOO group received 2 ml/kg EVOO with high phenolic content by gavage twice a day for 3 days. 3 cm of bowel including terminal ileum, cecum, and proximal colon was excised. RESULTS: Weight gain and clinical disease scores were significantly higher in the NEC + EVOO group than in the NEC group (p < 0.001). EVOO treatment caused significant decreases in IL1ß, IL6 levels (p = 0.016, p = 0.029 respectively) and EGF, MDA levels (p = 0.032, p = 0.013 respectively) compared to NEC group. Significant decreases were observed in IL6 gene expression in the NEC + EVOO group compared to the NEC group (p = 0.002). In the group NEC + EVOO, the number of Caspase-3 positive cells was found to be significantly reduced (p < 0.001) and histopathological examination revealed minimal changes and significantly lower histopathological scores (p < 0.001). CONCLUSION: Phenol-rich EVOO prevents intestinal damage caused by NEC by inhibiting inflammation, oxidative stress, apoptosis.
Asunto(s)
Enterocolitis Necrotizante , Interleucina-6 , Ratas , Animales , Aceite de Oliva/uso terapéutico , Aceite de Oliva/farmacología , Interleucina-6/metabolismo , Enterocolitis Necrotizante/patología , Estrés Oxidativo , Apoptosis , Inflamación , Fenoles/farmacología , Fenoles/uso terapéutico , Modelos Teóricos , Animales Recién NacidosRESUMEN
BACKGROUND: Recombinant human IGF-1/binding protein-3 (rhIGF-1/BP3) is currently being tested in phase II clinical trials in premature infants to prevent bronchopulmonary dysplasia, but its impact on the neonatal intestine remains unclear. The aim of this study was to determine whether rhIGF-1/BP3 protects against necrotizing enterocolitis (NEC) in mice and to investigate the mechanisms involved. METHODS: Neonatal mice were dam fed or injected intraperitoneally with rhIGF-1/BP3 (or vehicle) and submitted to an experimental NEC model. Serum IGF-1 was assessed by ELISA and intestinal vascular endothelial growth factor (VEGF) and VEGF receptor 2 (VEGFR2) expression by Western blot. Intestinal endothelial cell proliferation, and enterocyte proliferation and migration were examined by immunofluorescence. Pup survival and histological intestinal injury were determined. RESULTS: In pups exposed to experimental NEC, serum IBP3-bound IGF-1 level was decreased. Exogenous rhIGF-1/BP3 preserved VEGF and VEGFR2 protein expression, decreased vascular permeability, and preserved endothelial cell proliferation in the small intestine. Furthermore, rhIGF-1/BP3 promoted enterocyte proliferation and migration, which effects were attenuated by inhibiting VEGFR2 signaling, decreased enterocyte apoptosis and decreased systemic and intestinal inflammation. rhIGF-1/BP3 improved survival and reduced the incidence of severe intestinal injury in experimental NEC. CONCLUSIONS: Exogenous rhIGF-1/BP3 protects neonatal mice against experimental NEC via multiple mechanisms. IMPACT: Exogenous rhIGF-1/BP3 preserves intestinal microvascular development and integrity, promotes enterocyte proliferation and migration, decreases local and systemic inflammation, and protects neonatal mice against NEC. The article adds pre-clinical evidence of a protective role for rhIGF-1/BP3 on the premature gut. It provides evidence supporting the use of rhIGF1/BP3 in premature neonates to protect against NEC.
Asunto(s)
Animales Recién Nacidos , Proliferación Celular , Modelos Animales de Enfermedad , Enterocolitis Necrotizante , Factor I del Crecimiento Similar a la Insulina , Proteínas Recombinantes , Factor A de Crecimiento Endotelial Vascular , Receptor 2 de Factores de Crecimiento Endotelial Vascular , Animales , Enterocolitis Necrotizante/prevención & control , Enterocolitis Necrotizante/metabolismo , Enterocolitis Necrotizante/patología , Ratones , Factor I del Crecimiento Similar a la Insulina/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Proteínas Recombinantes/farmacología , Factor A de Crecimiento Endotelial Vascular/metabolismo , Enterocitos/metabolismo , Humanos , Intestinos/patología , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/metabolismo , Ratones Endogámicos C57BL , Movimiento Celular , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patología , Células Endoteliales/metabolismo , FemeninoRESUMEN
OBJECTIVE: To investigate the accuracy of preoperative and intraoperative diagnosis via comparison to pathologic diagnosis in spontaneous intestinal perforation (SIP) vs. necrotizing enterocolitis (NEC). STUDY DESIGN: A retrospective review of neonates <1500 g treated for pneumoperitoneum between 07/2004-09/2022 was conducted. Patients treated for NEC medically prior to diagnosis and those treated with drain only were excluded. Fleiss' Kappa analysis assessed agreement between all three diagnoses: preoperative, intraoperative, and pathologic. RESULT: Overall, 125 patients were included with mean birthweight 834.2 g (SD:259.2) and mean gestational age 25.8 weeks (SD:2.2). Preoperative and intraoperative diagnoses agreed in 90.3%, intraoperative and pathologic agreed in 71.1%, and preoperative and pathologic agreed in 75.2% of patients. Fleiss' Kappa was 0.55 (95% CI:0.43,0.68), indicating moderate agreement between the three diagnoses. CONCLUSION: Our study shows moderate agreement between preoperative, intraoperative, and pathologic diagnoses. Further studies investigating the clinical characteristics of SIP and NEC are needed to improve diagnostic accuracy and management.
Asunto(s)
Enterocolitis Necrotizante , Enfermedades Fetales , Enfermedades del Recién Nacido , Perforación Intestinal , Cirujanos , Femenino , Recién Nacido , Humanos , Lactante , Enterocolitis Necrotizante/diagnóstico , Enterocolitis Necrotizante/cirugía , Enterocolitis Necrotizante/patología , Perforación Intestinal/diagnóstico , Perforación Intestinal/cirugía , Estudios RetrospectivosRESUMEN
Necrotizing enterocolitis (NEC) is one of the most common acute gastrointestinal diseases in preterm infants. Recent studies have found that NEC is not only caused by changes in the intestinal environment but also by the failure of multiple systems and organs, including the liver. The accumulation of bile acids (BAs) in the ileum and the disorder of ileal BA transporters are related to the ileum injury of NEC. Inflammatory factors such as tumor necrosis factor (TNF)-α and interleukin (IL)-18 secreted by NEC also play an important role in regulating intrahepatic BA transporters. As an important link connecting the liver and intestinal circulation, the bile acid metabolic pathway plays an important role in the regulation of intestinal microbiota, cell proliferation, and barrier protection. In this review, we focus on how bile acids explore the dynamic changes of bile acid metabolism in necrotizing enterocolitis and the potential therapeutic value of targeting the bile acid signaling pathways.
Asunto(s)
Enterocolitis Necrotizante , Recien Nacido Prematuro , Recién Nacido , Humanos , Ácidos y Sales Biliares/metabolismo , Enterocolitis Necrotizante/etiología , Enterocolitis Necrotizante/metabolismo , Enterocolitis Necrotizante/patología , Íleon/metabolismo , Intestinos/patología , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
The effect of MDANP effects on ER stress signalling not well known or elucidated. Endoplasmic reticulum (ER) stress plays a critical role in necrotizing enterocolitis (NEC) pathogenesis through the PERK-eIF2É-QRICH1 axis. The present study aimed to explore the protective effects of MDANP in NEC development. Firstly, a function screening was designed to identify the candidate peptides in human milk, and then the identified peptides were validated in NEC patients. In vivo, NEC was induced in mice pups and divided into four groups: (1) control group, (2) NEC group, (3) MDANP + NEC group, and (4) NS + NEC group. In vitro, lentivirus-mediated QRICH1 silencing, was used to transfect NCM460 cell lines, then stimulated with LPS. After LPS stimulation, cells were treated with chemically synthesized MDANP, and the essential proteins in the QRICH1 signalling pathway in cells were tested and compared. After the small-scale screening, a peptide (SKSKKFRRPDIQYPDATDED) named MDANP was determined as the principal peptide. Its protective effect against NEC through inhibiting the expression of ERS key proteins and impeding the intestinal cells' apoptosis was observed in the animal models. Furthermore, the inhibitive effect of MDANP on apoptosis of intestinal epithelial cells through modulating the PERK-eIF2É-QRICH1 ERS pathway was also confirmed in vitro. Taken together, our data suggest that MDANP effectively ameliorates apoptosis in NEC through attenuating PERK-eIF2É-QRICH1.
Asunto(s)
Enterocolitis Necrotizante , Recién Nacido , Ratones , Animales , Humanos , Enterocolitis Necrotizante/patología , Lipopolisacáridos/farmacología , Intestinos/patología , Línea Celular , Péptidos/farmacología , Péptidos/metabolismo , Mucosa Intestinal/metabolismo , Modelos Animales de EnfermedadRESUMEN
Introdução:A enterocolite necrosante é uma doença que pode afetar o trato gastrointestinal de recém-nascidos,cujas manifestações clínicas podem ser caracterizadas por vômitos biliosos, sangue nas fezes, distensão abdominal, além de alterações nos parâmetros hemodinâmicos e instabilidade térmica. As populações mais vulneráveis a essa enfermidade são recém-nascidos de baixo peso,expostos ao ambiente de terapia intensiva neonatal. Objetivos: Identificar o perfil de recém-nascidos e os fatores maternos e neonatais associados à ocorrência de óbitos por enterocolite necrosante, em maternidade de referência do Ceará-Brasil. Metodologia: Trata-se de coorte retrospectiva, estudo que objetiva a descrição da incidência de determinado evento ao longo do tempo, além do estabelecimento de relações causais entre os fatores associados ao acontecimento. Incluíram-serecém-nascidos que tiveram óbitos por enterocolite necrosante entre 2019 e 2021, comficha de investigação de óbitos neonatais preenchida corretamente, não sendo excluído nenhum recém-nascido, totalizando amostra de 29 óbitos.Resultados: Identificou-se que o perfil dos recém-nascidos foi,em maioria, deprematuros e com baixo peso e fatores de risco para outras doenças associadas,como a sepse, o que acarretourealização de procedimentos invasivos e internação em ambiente de terapia intensiva neonatal.Conclusões: A prematuridade e o baixo peso ao nascer foram as variáveis relevantes no estudo e podem estar associadas à piora das condições clínicas do recém-nascido e ao desenvolvimento de enterocolite necrosante (AU).
Introduction: Necrotizing Enterocolitis is a disease that can affect the gastrointestinal tract of newborns, whose clinical manifestations can be characterized by bilious vomiting, blood in stool, abdominal distension, in addition to changes in hemodynamic parameters and thermal instability. The populations most vulnerable to this disease are low birth weight newborns exposed to the neonatal intensive care environment. Objectives: This study aimed to identify the profile of newborns and maternal and neonatal factors associated with the occurrence of deaths from necrotizing enterocolitis in a reference maternity hospital in Ceará, Brazil. Methodology: This is a retrospective cohort study seeking to describe the incidence ofa particular event over time, as well as establish causal relationships between the factors associated with the event. The study population comprised newborns who died from necrotizing enterocolitis between 2019 and 2021, who had neonatal death investigation forms filled out correctly, with no newborns being excluded, totaling a sample of 29 deaths. Results: It was identified that the profile of newborns was mostly premature, of low birth weight and with risk factors for other associated diseases such as sepsis, leading to invasive procedures and hospitalization in a neonatal intensive care environment. Conclusions: Prematurity and low birth weight were relevant variables in the study and may be associated with worsening of the newborn's clinical conditionsand development of necrotizing enterocolitis (AU).
ntroducción:La Enterocolitis Necrotizante es enfermedad que puede afectar el tracto gastrointestinal del recién nacido, cuyas manifestaciones clínicas pueden caracterizarse por vómitos biliosos, sangre en las heces, distensión abdominal, además de cambios en los parámetros hemodinámicos e inestabilidad térmica.Las poblaciones más vulnerables a esta enfermedad son recién nacidos con bajo peso expuestos al entorno de cuidados intensivos neonatales.Objetivos: Identificar el perfil de recién nacidos y los factores maternos y neonatales asociados a la ocurrencia de muertes por enterocolitis necrotizante, en maternidad de referencia en el Ceará-Brasil.Metodología: Estudio de cohorte retrospectivo, para describir la incidencia de determinado evento a lo largo del tiempo, además de establecer relaciones causales entre los factores asociados al evento.Se incluyeron recién nacidos fallecidos por enterocolitis necrotizante entre 2019 y 2021, quienes tuvieron formulario de investigación de muerte neonatal correctamente diligenciado, no excluyéndose ningún recién nacido, totalizando muestra de 29 defunciones.Resultados:El perfil de los recién nacidos fue mayoritariamente prematuro y de bajo peso al nacer y con factores de riesgo para otras enfermedades asociadas, como sepsis, con procedimientos invasivos y hospitalización en ambiente de cuidados intensivosneonatales.Conclusiones:La prematuridad y el bajo peso al nacer fueron variables relevantes en el estudio y pueden estar asociados con empeoramiento de las condiciones clínicas de recién nacidos y desarrollo de enterocolitis necrotizante (AU).