BACKGROUND AND AIMS:Necrotizing enterocolitis (NEC) is a life-threatening condition in premature infants, marked by acute intestinal necrosis. NEC develops in part after activation of the lipopolysaccharide receptor toll-like receptor 4 (TLR4) by intestinal microbes in the intestinal epithelium. Previous authors have shown an increased risk of NEC in human infants after cytomegalovirus (CMV) infection, which can affect mitochondrial function. We now seek to explore the impact and the mechanisms of CMV infection on NEC severity and its relationship with TLR4 signaling and mitochondria function. METHODS:NEC was induced in newborn mice with and without CMV infection. RNA sequencing and gene set enrichment analysis were performed to identify effects on inflammatory and metabolic pathways. The role of TLR4 signaling and mitochondrial function were investigated in wild-type and Tlr4-deficient mice. The adenosine receptor agonist 5'-N-ethylcarboxamido adenosine was tested for its ability to reduce CMV-induced effects on NEC severity. RESULTS:CMV infection significantly increased NEC severity in wild-type mice. Mechanistically, CMV infection triggered proinflammatory pathways, disrupted cellular metabolism, and upregulated Tlr4 expression, leading to mitochondrial dysfunction and nuclear factor-kB translocation. These effects were notably absent in Tlr4-deficient mice. 5'-N-ethylcarboxamido adenosine treatment reversed CMV-induced NEC severity by reducing mitochondrial dysfunction and TLR4-driven nuclear factor-kB activation. CONCLUSIONS:CMV infection worsens NEC severity in mice by amplifying TLR4 signaling, inflammation, and mitochondrial dysfunction. Targeting CMV and its influence on TLR4 may offer novel therapeutic approaches for NEC.

INTRODUCTION:Traumatic brain injury (TBI) is a leading cause of death and disability worldwide. Although TBI pathophysiology has been thoroughly investigated, the effectivity of therapeutic approaches for TBI is still lacking. Our group has developed a novel approach of noninvasive transcutaneous auricular vagus nerve stimulation (taVNS) in a mouse model of TBI to investigate its impact on neuroinflammation. METHODS:A murine-controlled cortical impact model was used, and results were analyzed on postinjury days (PIDs) 3 and 7. The experimental groups included (1) sham C57BL/6 wild type (WT), (2) TBI wild type, (3) sham-taVNS, and (4) TBI-taVNS. The animals underwent anesthesia, off-site stimulation, or taVNS for 30 minutes. The short- and long-term groups received two sessions of taVNS treatment and were tested on PIDs 3 and 7, respectively. A combination of real-time polymerase chain reaction and immunohistochemistry was used to validate the success of the model and to quantify gene expression associated with microglial and astrocyte activation. Student's t test and one-way analysis of variance were used for statistical analysis, with significance achieved when p  < 0.05. RESULTS:Transcutaneous auricular vagus nerve stimulation (VNS) activated the solitary tract nucleus and the dorsal motor nucleus of the vagus nerve as evidenced by a significant upregulation of the neuronal activation marker c-Fos, indicating vagus nerve activation. Transcutaneous auricular VNS treatment reduced the expression of pro-inflammatory microglial markers Tnf (1.69 ± 0.17 vs. 3.615 ± 0.86, p  < 0.05) and Lcn2 (64.15 ± 14 vs. 337.7 ± 104.8, p  < 0.01) in the ipsilateral injured cortex on PIDs 3 and 7, respectively. Transcutaneous auricular VNS also increased the expression of anti-inflammatory microglial marker Arg1 (55 ± 6.47 vs. 30.49 ± 3.94, p  < 0.01) and astrocyte Gfap reactivity (8,582 ± 826 vs. 4,569 ± 554.3, p  < 0.01) on PID 3. ( J Trauma Acute Care Surg . 2026;100: 707-713. Copyright © 2025 Wolters Kluwer Health, Inc. All rights reserved.).

Electronic cigarettes, also known as e-cigarettes or vapes, are battery-operated devices that heat and produce aerosols for inhalation. Maternal nicotine e-cigarette use during pregnancy can have detrimental effects on fetal growth and development, which may be in part due to deleterious effects of nicotine vapor exposure on innate immunity of the developing fetus tethered to the harmful chemicals and toxins present in the vaping device. Understanding these effects is crucial for identifying potential risks to fetal health and developing interventions that may mitigate the impact of maternal stress on the developing immune system. We hypothesized that murine maternal exposure to nicotine e-cigarette vapor would lead to neonatal innate immune activation. Our preliminary studies showed that pregnant mice chronically (five times per week, from gestational day 14 through 21, or birth) exposed to e-cigarette vapor in-utero exhibited alterations in inflammatory response, as demonstrated through the increased production of pro-inflammatory cytokines in the gut, including TLR4, IL-6, TNF, and IL-1b, by RT-PCR. We also show the activation of neonatal immune cells, including microglia in the brain and neutrophils in the lung via immunohistochemistry. Consistent with our hypothesis, these findings reveal maternal vaping induces immunological signaling and leads to innate immune activation in neonatal mouse lung, gut, and brain, suggesting a role in adverse immunological outcomes. Supported by NIH/NIGMS R35GM141956; Vivien Thomas Scholars Initiative. Innate Immune Responses and Host Defense: Molecular Mechanisms (INM)

Short bowel syndrome (SBS) leads to severe morbidity and mortality. Intestinal adaptation is crucial in improving outcomes. To understand the human gene pathways associated with adaptation, we perform single-cell transcriptomic analysis of human small intestinal organoids explanted from mice with experimental SBS. We show that transmembrane ion pathways, specifically the transepithelial zinc transport pathway genes SLC39A4 and SLC39A5, are upregulated in SBS. This discovery is corroborated by an external dataset, bulk RT-qPCR, and Western blots. Oral zinc supplementation is shown to improve survival and weight gain of SBS mice and increase the proliferation of intestinal crypt cells in vitro. Finally, we identify the upregulation of SLC39A5 and associated transcription factor KLF5 in biopsied intestinal tissue specimens from patients with SBS. Thus, we identify zinc supplementation as a potential therapy for SBS and describe a xenotransplantation model that provides a platform for discovery in other intestinal diseases. SBS leads to severe morbidity and mortality. Here, using a xenotransplantation model, the authors uncover the role of zinc transport pathways in intestinal adaptation in SBS, highlighting the potential of zinc supplementation as a therapeutic approach.

BACKGROUND Traumatic brain injury (TBI) leads to acute gastrointestinal dysfunction and mucosal damage, resulting in feeding intolerance. C-C motif chemokine receptor 2 (Ccr2+) monocytes are crucial immune cells that regulate the gut's inflammatory response via the brain-gut axis. Using Ccr2ko mice, we investigated the intricate interplay between these cells to better elucidate the role of systemic inflammation after TBI. METHODS A murine-controlled cortical impact model was used, and results were analyzed on postinjury days 1 and 3. The experimental groups included (1) sham C57Bl/6 wild type (WT), (2) TBI WT, (3) sham Ccr2ko, and (4) TBI Ccr2ko. Mice were euthanized on postinjury days 1 and 3 to harvest the ileum and study intestinal dysfunction and serotonergic signaling using a combination of quantitative real-time polymerase chain reaction, immunohistochemistry, fluorescein isothiocyanate-dextran motility assays, and flow cytometry. Student's t test and one-way analysis of variance were used for statistical analysis, with significance achieved when p < 0.05. RESULTS Traumatic brain injury resulted in severe dysfunction and dysmotility of the small intestine in WT mice as established by significant upregulation of inflammatory cytokines iNOS, Lcn2, TNFα, and IL1β and the innate immunity receptor toll-like receptor 4 (Tlr4). This was accompanied by disruption of genes related to serotonin synthesis and degradation. Notably, Ccr2ko mice subjected to TBI showed substantial improvements in intestinal pathology. Traumatic brain injury Ccr2ko groups demonstrated reduced expression of inflammatory mediators (iNOS, Lcn2, IL1β, and Tlr4) and improvement in serotonin synthesis genes, including tryptophan hydroxylase 1 (Tph1) and dopa decarboxylase (Ddc). CONCLUSION Our study reveals a critical role for Ccr2+ monocytes in modulating intestinal homeostasis after TBI. Ccr2+ monocytes aggravate intestinal inflammation and alter gut-derived serotonergic signaling. Therefore, targeting Ccr2+ monocyte-dependent responses could provide a better understanding of TBI-induced gut inflammation. Further studies are required to elucidate the impact of these changes on brain neuroinflammation and cognitive outcomes.

Background: Radiographic diagnosis of necrotizing enterocolitis (NEC) is challenging. Deep learning models may improve accuracy by recognizing subtle imaging patterns. We hypothesized it would perform with comparable accuracy to that of senior surgical residents. Methods: This cohort study compiled 494 anteroposterior neonatal abdominal radiographs (214 images NEC, 280 other) and randomly divided them into training, validation, and test sets. Transfer learning was utilized to fine-tune a ResNet-50 deep convolutional neural network (DCNN) pre-trained on ImageNet. Gradient-weighted Class Activation Mapping (Grad-CAM) heatmaps visualized image regions of greatest relevance to the pretrained neural network. Senior surgery residents at a single institution examined the test set. Resident and DCNN ability to identify pneumatosis on radiographic images were measured via area under the receiver operating curves (AUROC) and compared using DeLong's method. Results: The pretrained neural network achieved AUROC of 0.918 (95% CI, 0.837-0.978) with an accuracy of 87.8% with five false negative and one false positive prediction. Heatmaps confirmed appropriate image region emphasis by the pretrained neural network. Senior surgical residents had a median area under the receiver operating curve of 0.896, ranging from 0.778 (95% CI 0.615-0.941) to 0.991 (95% CI 0.971-0.999) with zero to five false negatives and one to eleven false positive predictions. The deep convolutional neural network performed comparably to each surgical resident's performance (p > 0.05 for all comparisons). Conclusions: A deep convolutional neural network trained to recognize pneumatosis can quickly and accurately assist clinicians in promptly identifying NEC in clinical practice. Level of Evidence: III (study type: Study of Diagnostic Test, study of nonconsecutive patients without a universally applied "gold standard") (c) 2024 Elsevier Inc. All rights are reserved, including those for text and data mining, AI training, and similar technologies.

BACKGROUND & AIMS: The abdominal discomfort experienced by patients with colitis may be attributable in part to the presence of small intestinal dysmotility, yet mechanisms linking colonic inflammation with small-bowel motility remain largely unexplored. We hypothesize that colitis results in small intestinal hypomotility owing to a loss of enteroendocrine cells (EECs) within the small intestine that can be rescued using serotonergic-modulating agents. METHODS: Male C57BL/6J mice, as well as mice that over- express (EECOVER) or lack (EECDEL) NeuroD1 & thorn; enteroendocrine cells, were exposed to dextran sulfate sodium (DSS) colitis (2.5% or 5% for 7 days) and small intestinal motility was assessed by 70-kilodalton fluorescein isothiocyanate-dextran fluorescence transit. EEC number and differentiation were evaluated by immunohistochemistry, terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling staining, and quantitative reversetranscriptase polymerase chain reaction. Mice were treated with the 5-hydroxytryptamine receptor 4 agonist prucalopride (5 mg/kg orally, daily) to restore serotonin signaling. RESULTS: DSS-induced colitis was associated with a significant small-bowel hypomotility that developed in the absence of significant inflammation in the small intestine and was associated with a significant reduction in EEC density. EEC loss occurred in conjunction with alterations in the expression of key serotonin synthesis and transporter genes, including Tph1, Ddc, and Slc6a4. Importantly, mice overexpressing EECs revealed improved small intestinal motility, whereas mice lacking EECs had worse intestinal motility when exposed to DSS. Finally, treatment of DSSexposed mice with the 5-hydroxytryptamine receptor 4 agonist prucalopride restored small intestinal motility and attenuated colitis. CONCLUSIONS: Experimental DSS colitis induces significant small-bowel dysmotility in mice owing to enteroendocrine loss that can be reversed by genetic modulation of EEC or administering serotonin analogs, suggesting novel therapeutic approaches for patients with symptomatic colitis.

Efforts to develop an artificial intestine for treatment of short bowel syndrome have thus far failed to achieve coordinated peristalsis. To address this, we introduce an implantable, biomimetic, magnetically-actuated peristaltic pumping apparatus. The system consists of a magnetic pump surrounding an intestinal graft and a rotating external magnetic field generator that produces peristaltic motion in the graft as the field direction alternates. We performed computational finite element modeling to simulate deformation and stress concentration on the pump. We then tested the ability of the system to produce fluid flow through porcine subintestinal submucosa at actuation frequencies from 0 Hz to 4 Hz. The system achieved a maximal flow rate of 6.03 mL/min at 3 Hz actuation frequency, which is in the physiological range of human intestinal flow rates. This device represents a novel proof-of-concept for achieving untethered peristalsis in an implantable intestinal graft that is primed for preclinical testing in animal models of short bowel syndrome.

BackgroundProbiotic administration may decrease the incidence of necrotizing enterocolitis (NEC) through mechanisms that are largely unknown. We investigated the effects of probiotics on intestinal epigenetics and assessed their effects on intestinal inflammation and motility using both ileum-predominant and combined ileo-colitis mouse NEC models.MethodsC57BL/6 J mice were gavage-fed a multi-strain probiotic from postnatal days 3-11, consisting of B. infantis, B. lactis, and S. thermophilus. From p8, mice were exposed to ileo-colitis NEC involving formula containing NEC bacteria and 0.5% DSS. DNA methylation was measured using the Infinium Methylation Assay. Gastrointestinal motility was assessed by 70 Kd FITC-dextran transit time. Probiotic colonization was measured in probiotic-fed mice by qPCR.ResultsProbiotic administration caused significant changes in the small intestine's epigenetic signature, a reduction in NEC severity, and improved intestinal motility. The effects of probiotics were more pronounced in the ileo-colitis NEC model.ConclusionsThese findings shed light on the role of probiotics in two clinically relevant models of NEC, add additional insights into their underlying mechanism of action, and reveal unanticipated epigenetic modifications to the intestinal mucosa after their use.ImpactThese findings shed light on the role of multi-strain probiotics in two clinically relevant animal models of NEC, and add additional insights into their underlying mechanism of actionThis study provides a new, clinically relevant model for the study of NEC including administration of 0.5% DSS, to include ileal dominant and ileo-colonic dominant phenotypes of the disease.These results reveal that clinically relevant strains of probiotic bacteria can exert epigenetic effects on the small intestine in mice, and can attenuate the epigenetic changes induced by NEC.

Necrotizing enterocolitis (NEC) is a devastating gastrointestinal disorder in premature infants that causes significant morbidity and mortality. Research efforts into the pathogenesis of NEC have discovered a pivotal role for the gram-negative bacterial receptor, Toll-like receptor 4 (TLR4), in its development. TLR4 is activated by dysbiotic microbes within the intestinal lumen, which leads to an exaggerated inflammatory response within the developing intestine, resulting in mucosal injury. More recently, studies have identified that the impaired intestinal motility that occurs early in NEC has a causative role in disease development, as strategies to enhance intestinal motility can reverse NEC in preclinical models. There has also been broad appreciation that NEC also contributes to significant neuroinflammation, which we have linked to the effects of gut-derived pro-inflammatory molecules and immune cells which activate microglia in the developing brain, resulting in white matter injury. These findings suggest that the management of the intestinal inflammation may secondarily be neuroprotective. Importantly, despite the significant burden of NEC on premature infants, these and other studies have provided a strong rationale for the development of small molecules with the capability of reducing NEC severity in pre-clinical models, thus guiding the development of specific anti-NEC therapies. This review summarizes the roles of TLR4 signaling in the premature gut in the pathogenesis of NEC, and provides insights into optimal clinical management strategies based upon findings from laboratory studies.

Necrotizing enterocolitis (NEC) is a devastating disease in premature infants and the leading cause of death and disability from gastrointestinal disease in this vulnerable population. Although the pathophysiology of NEC remains incompletely understood, current thinking indicates that the disease develops in response to dietary and bacterial factors in the setting of a vulnerable host. As NEC progresses, intestinal perforation can result in serious infection with the development of overwhelming sepsis. In seeking to understand the mechanisms by which bacterial signaling on the intestinal epithelium can lead to NEC, we have shown that the gram-negative bacterial receptor toll-like receptor 4 is a critical regulator of NEC development, a finding that has been confirmed by many other groups. This review article provides recent findings on the interaction of microbial signaling, the immature immune system, intestinal ischemia, and systemic inflammation in the pathogenesis of NEC and the development of sepsis. We will also review promising therapeutic approaches that show efficacy in pre-clinical studies.

Necrotizing enterocolitis (NEC) is the leading cause of morbidity and mortality in premature infants. One of the most devastating complications of NEC is the development of NEC-induced brain injury, which manifests as impaired cognition that persists beyond infancy and which represents a proinflammatory activation of the gut-brain axis. Given that oral administration of the human milk oligosaccharides (HMOs) 2 '-fucosyllactose (2 '-FL) and 6 '-sialyslactose (6 '-SL) significantly reduced intestinal inflammation in mice, we hypothesized that oral administration of these HMOs would reduce NEC-induced brain injury and sought to determine the mechanisms involved. We now show that the administration of either 2 '-FL or 6 '-SL significantly attenuated NEC-induced brain injury, reversed myelin loss in the corpus callosum and midbrain of newborn mice, and prevented the impaired cognition observed in mice with NEC-induced brain injury. In seeking to define the mechanisms involved, 2 '-FL or 6 '-SL administration resulted in a restoration of the blood-brain barrier in newborn mice and also had a direct anti-inflammatory effect on the brain as revealed through the study of brain organoids. Metabolites of 2 '-FL were detected in the infant mouse brain by nuclear magnetic resonance (NMR), whereas intact 2 '-FL was not. Strikingly, the beneficial effects of 2 '-FL or 6 '-SL against NEC-induced brain injury required the release of the neurotrophic factor brain-derived neurotrophic factor (BDNF), as mice lacking BDNF were not protected by these HMOs from the development of NEC-induced brain injury. Taken in aggregate, these findings reveal that the HMOs 2 '-FL and 6 '-SL interrupt the gut-brain inflammatory axis and reduce the risk of NEC-induced brain injury. NEW & NOTEWORTHY This study reveals that the administration of human milk oligosaccharides, which are present in human breast milk, can interfere with the proinflammatory gut-brain axis and prevent neuroinflammation in the setting of necrotizing enterocolitis, a major intestinal disorder seen in premature infants.

BACKGROUND:The use of synthetic mesh is considered too high risk, and therefore, not an option when closing a contaminated abdominal fascial defect. This study evaluated the clinical outcomes when using synthetic mesh combined with vacuum-assisted closure (VAC) dressing to close these facial defects. MATERIALS AND METHODS:From 2010 to 2016, a retrospective review was performed, including 34 patients in a single rural trauma center who underwent a damage control laparotomy in the presence of a contaminated or infected field. Definitive abdominal closure with a bridging polypropylene mesh along with the application of a VAC dressing was done in all cases. Data collection included baseline demographics, operative indication, postoperative complications, mortality and length of follow up. RESULTS:Median age of the patients was 67 y (IQR 40-87 y), with 22 (65%) being male at the time of operation. The median duration of clinical follow-up was 15.15 mo. The observed complications included three fistulas, two hernias, nine draining sinus tracts, and three mesh explanations with an overall complication rate of 41.1%. Although the absolute observed fistula rate was 8.8% (3 cases), the adjusted mesh-related fistulas formation rate after chart review was 0.0%. No mortalities were attributed directly to mesh-related complication. CONCLUSIONS:This study found no mesh-related fistulas when using a synthetic mesh along with a VAC dressing for abdominal closure in a contaminated field. These results may provide a platform for further study regarding the safety of this technique.