Colonization of fecal microbiota from patients with neonatal necrotizing enterocolitis exacerbates intestinal injury in germfree mice subjected to necrotizing enterocolitis-induction protocol via alterations in butyrate and regulatory T cells.

BACKGROUND: Necrotizing enterocolitis (NEC) remains a life-threatening disease in neonates. Numerous studies have shown a correlation between the intestinal microbiota and NEC, but the causal link remains unclear. This study aimed to demonstrate the causal role of gut microbiota in NEC and explore potential mechanisms involved. METHODS: Eighty-one fecal samples from patients with NEC and eighty-one matched controls (matched to the NEC infants by gestational age, birth weight, date of birth, mode of delivery and feeding patterns) were collected. To explore if altered gut microbiota contributes to the pathogenesis of NEC, fecal microbiota transplantation (FMT) was carried out in germ-free (GF) mice prior to a NEC-induction protocol that included exposure to hypoxia and cold stress. Butyric acid was also administered to demonstrate its role in NEC. The fecal microbiota from patients and mice were analyzed by 16S rRNA gene sequencing analysis. Short chain fatty acid (SCFA) levels were measured by gas chromatography-mass spectrometry (GC-MS). The ontogeny of T cells and regulatory T cells (Tregs) in lamina propria mononuclear cells (LPMC) from the ileum of patients and mice were isolated and analyzed by flow cytometry.The transcription of inflammatory cytokines was quantified by qRT-PCR. RESULTS: NEC patients had increased Proteobacteria and decreased Firmicutes and Bacteroidetes compared to fecal control samples, and the level of butyric acid in the NEC group was lower than the control group. FMT in GF mice with samples from NEC patients achieved a higher histological injury scores when compared to mice that received FMT with control samples. Alterations in microbiota and butyrate levels were maintained in mice following FMT. The ratio of Treg/CD4+T (Thelper) cells was reduced in both NEC patients and mice modeling NEC following FMT. CONCLUSIONS: The microbiota was found to have NEC and the microbial butyrate-Treg axis was identified as a potential mechanism for the observed effects.

Sodium Butyrate Alleviates Intestinal Inflammation in Mice with Necrotizing Enterocolitis.

OBJECTIVE: To determine the role of sodium butyrate in intestinal inflammation via regulation of high-mobility group box-1 (HMGB1), we analyzed the potential mechanism in necrotizing enterocolitis (NEC) in a neonatal mouse model. METHODS: A NEC model was created with hypoxia and cold exposure and artificial overfeeding. C57BL/6 neonatal mice were randomized into three groups: the control, untreated NEC, and sodium butyrate (150 mM)-pretreated NEC groups. Pathological variations in ileocecal intestinal tissue were observed by HE staining and scored in a double-blind manner. The mRNA expression levels of HMGB1, Toll-like receptor 4 (TLR4), nuclear factor-κB (NF-κB), and inflammatory cytokines in intestinal tissues were determined by quantitative real-time PCR. The protein levels of HMGB1 and associated cytokines in intestinal tissues were evaluated using ELISA. The relative protein expression levels of TLR4 and NF-κB in intestinal tissues were quantified by western blot. RESULTS: Sodium butyrate administration improved the body weight and survival rate of NEC mice; relieved intestinal pathological injury; reduced the intestinal expression of HMGB1, TLR4, NF-κB, interleukin- (IL-) 1ß, IL-6, IL-8, and TNF-α; and increased the intestinal expression of IL-10 (P < 0.05). Treatment with butyrate decreased the proportion of opportunistic Clostridium_sensu_stricto_1 and Enterococcus and increased the proportion of beneficial Firmicutes and Lactobacillus in the NEC model. CONCLUSIONS: Sodium butyrate intervention relieves intestinal inflammation and partially corrects the disrupted intestinal flora in mice with NEC.

The emerging role of the gut virome in necrotizing enterocolitis.

Necrotizing enterocolitis (NEC) is the most common gastrointestinal emergency in neonates, particularly preterm infants. Many factors can lead to NEC, but microbial dysbiosis is one of the most important risk factors that can induce this disease. Given the major role of the gut virome in shaping bacterial homeostasis, virome research is a fledgling but rapidly evolving area in the field of microbiome that is increasingly connected to human diseases, including NEC. This review provides an overview of the development of the gut virome in newborns, discusses its emerging role in NEC, and explores promising therapeutic applications, including phage therapy and fecal virome transplantation.

Novel predictors of intravenous immunoglobulin resistance in patients with Kawasaki disease: a retrospective study.

Objective: The aim of this study was to investigate the predictive value of systemic immune inflammation index (SII), systemic inflammatory response index (SIRI), and pan-immune inflammation value (PIV) in predicting intravenous immunoglobulin (IVIG) resistance in children diagnosed with Kawasaki disease (KD). Methods: The clinical data of pediatric patients diagnosed with Kawasaki disease and admitted to our hospital between January 2006 and December 2022 were retrospectively analyzed. Results: In total, 771 children diagnosed with KD were included in this study, 86 (11.2%) of whom were diagnosed with IVIG resistance. The correlation between SII, SIRI, PIV and IVIG resistance was evaluated using univariate testing, binary logistic regression analysis, and receiver operating characteristic (ROC) curve analysis. Our study found that the SII, SIRI, and PIV were independent risk factors (p=0.001, p<0.001, and p=0.02, respectively). The area under the ROC curve (AUC) values of the SII, SIRI, and PIV were 0.626 (95% confidence interval (CI): 0.553-0.698, p<0.001), 0.571 (95% CI: 0.500-0.642, p=0.032), and 0.568 (95% CI: 0.495-0.641, p=0.040), respectively, and the cutoff values were 2209.66, 3.77, and 1387.825, respectively. Conclusion: The SII, SIRI, and PIV have potential value in predicting IVIG resistance in patients with KD.

Exogenous autoinducer-2 alleviates intestinal damage in necrotizing enterocolitis via PAR2/MMP3 signaling pathway.

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.

Kawasaki disease complicated with shock syndrome, macrophage activation syndrome, and acute abdomen in children: Two case reports.

Background: Kawasaki disease (KD) is an acute systemic vasculitis that can involve multiple organs. Few reports have been published about KD patients presenting with multiple complications such as acute abdomen, KD shock syndrome (KDSS), and macrophage activation syndrome (MAS). Case Description: We present the cases of two males (9 and 12 years old) diagnosed with KD accompanied by rare manifestations. Case 1 is a 9-year-old male treated for acute appendicitis, KDSS, and MAS. Case 2 is a 12-year-old male who presented with KDSS, MAS, and an ileal perforation. They were treated with intravenous immunoglobulin, aspirin, high-dose corticosteroids, vasoactive drugs, and symptomatic treatment, with good outcomes. Conclusions: Clinicians should be aware of the possibility of KD in the presence of fever and unusual manifestations, such as severe inflammatory indicators and acute abdomen that is nonresponsive to antibiotic therapy. Meanwhile, KD-related unusual complications should be recognized, such as KDSS and MAS.

Kawasaki Disease Associated Acute Abdomen: Most Require No Surgery.

Objective: To summarize the clinical features and treatment experiences of patients with Kawasaki disease (KD)-associated acute abdomen (KD-AA). Methods: We conducted a retrospective case-control study of patients with KD-AA treated at our hospital between January 2006 and November 2022. Results: Of the 917 children with KD, 43 (4.7%) presented with AA. Of these, 33 with complete information were included in the KD-AA group. Patients with KD-AA were significantly older, with higher neutrophil rate, C-reactive protein, procalcitonin, and alanine transaminase levels and lower hemoglobin, albumin, and serum sodium levels. Additionally, more patients with KD-AA presented with aseptic meningitis and KD shock syndrome than those with KD alone (all p<0.05). The two groups did not differ in the incidence of intravenous immunoglobulin (IVIG) resistance, incomplete KD, or coronary artery abnormalities. All patients received aspirin and IVIG therapy, with nine receiving a second dose of IVIG and 11 receiving corticosteroids. Only two patients with KD-AA underwent surgery, and the prognosis of all patients with KD-AA was good. Conclusion: KD-AA should be suspected in febrile children with abdominal symptoms. Prompt diagnosis of KD-AA is important for early effective treatment to avoid unnecessary surgical harm. KD, complicated by acute abdomen, has a good prognosis.

Clinical Analysis of Acute Suppurative Thyroiditis in 18 Children.

Objective: To summarize our clinical experience with the diagnosis and treatment of children with acute suppurative thyroiditis (AST). Methods: We retrospectively reviewed the clinical data of 18 children with AST treated at our hospital between January 2009 and May 2022. Results: There were 8 boys and 10 girls, aged 7.8 ± 3.8 years at admission. The main clinical manifestations were fever (88.9%), neck pain (100%), and neck mass (100%). Blood and pus cultures were performed in 9 and 15 patients, respectively. All blood cultures were negative, while positive pus cultures were noted in eight cases (six Streptococcus spp., one Staphylococcus spp., and one Streptococcus spp. and Staphylococcus spp. mixed infection). Additionally, all patients received antibiotic treatment: three received antibiotics alone, seven received antibiotics and ultrasound (US)-guided needle aspiration, seven received antibiotics as well as surgical incision and drainage, and one received antibiotics, US-guided needle aspiration in addition to surgical incision and drainage. Consequently, the average length of hospital stay in patients who received antibiotics and US-guided needle aspiration was 9.1±2.9 days compared to 14.0±2.0 days in patients in the antibiotics alone group and 13.0 ± 2.2 days in patients in the antibiotics and surgical incision and drainage group. Follow-up was conducted in 15 of the 18 patients. Three patients relapsed, and the prognosis of the other patients was good. Conclusion: AST has atypical clinical symptoms at the early stage. Regular monitoring of the thyroid gland using ultrasonography is strongly advised in unsure cases. Antibiotics combined with US-guided aspiration is a safe, effective, and minimally invasive treatment for AST in children and can reduce hospital stay. However, surgery may be necessary, particularly in the presence of complications. It is strongly recommended that patients with recurrence be examined for anatomical abnormalities and undergo radical treatment.

The decrease of fucosylation in intestinal epithelium is related to the development of necrotizing enterocolitis.

Necrotizing enterocolitis (NEC) is a devastating neonatal gastrointestinal emergency. Fucosylated glycans on intestinal epithelial cells (IECs) play a central role in the maintenance of intestinal homeostasis. Nevertheless, its association with necrotizing enterocolitis is not clear. We examined paraffin-embedded intestinal specimens from participants and found that the NEC patients showed lower intestinal epithelial fucosylation levels than the control patients. In the mouse model of NEC, the percentage of fucosylated epithelial cells (F-ECs) and ILC3s was decreased. Also, the expression levels of IL-22 and Fut2 were reduced. Moreover, the critical role of epithelial fucosylation in NEC was further confirmed by administering the anti-IL-22 antibody, which caused an increase in histological damage, body weight loss, intestinal permeability and proinflammatory cytokine release correlated with a reduction of F-ECs. Overall, intestinal fucosylation deficiency led to increased susceptibility and severity of NEC. Further studies are needed to determine whether modification of intestinal fucosylation affects the development of NEC.

Exogenous Autoinducer-2 Rescues Intestinal Dysbiosis and Intestinal Inflammation in a Neonatal Mouse Necrotizing Enterocolitis Model.

Autoinducer-2 (AI-2) is believed to be a bacterial interspecies signaling molecule that plays an important role in the regulation of the physiological behaviors of bacteria. The effect of AI-2 on the process of necrotizing enterocolitis (NEC) is unknown, and the aim of this study was to study the effect of AI-2 in a mouse NEC model. C57BL/6 mouse pups were randomly divided into three groups: the control group, the NEC group, and the NEC+AI-2 (NA) group. Exogenous AI-2 (500 nM) was added to the formula milk of the NA group. The concentrations of fecal AI-2 and flora were tested. The expression of cytokines, TLR4 and NF-κB in intestinal tissue was detected. The AI-2 level was significantly decreased in the NEC group (P<0.05). Compared with the NEC group, the intestinal injury scores, expression of TLR4, NF-kB, and proinflammatory factors (IL-1ß, IL-6, IL-8 and TNF-α) were reduced, and expression of anti-inflammatory factor (IL-10) was increased in the NA group mice (P<0.05). At the phylum level, the Proteobacteria abundance in the NA group was significantly increased, while the Bacteroidota abundance in the control group was significantly increased (P<0.05). At the genus level, Helicobacter and Clostridium_sensu_stricto_1 exhibited significantly greater abundance in the NEC group than in the other two groups, while Lactobacillus had the opposite trend (P<0.05). In addition, the abundances of Klebsiella, Rodentibacter and Enterococcus were significantly higher in the NA group than in the NEC and control groups (P < 0.05). Exogenous AI-2 partially reverses flora disorder and decreases inflammation in an NEC mouse model.

Autoinducer-2 May Be a New Biomarker for Monitoring Neonatal Necrotizing Enterocolitis.

Autoinducer-2 (AI-2) has a widely accepted role in bacterial intra- and interspecies communication. Little is known about the relationships between AI-2 and NEC. This study found that AI-2 levels in patients and in a NEC mouse model were detected using the Vibrio harveyi BB170 assay system. Bacterial communities of the newborns' stool microbiota (NEC acute group, NEC recovery group, control group, and antibiotics-free group) and of the NEC mouse model (NEC group and control group) were detected by high-throughput sequencing. Intestinal histopathological changes were observed after HE staining. The AI-2 level in the NEC acute group (44.75 [40.17~65.52]) was significantly lower than that in the control group, NEC recovery group and antibiotics-free group. The overall microbiota compositions of each group at the phylum level were not significantly different. The proportions of Enterococcus, Clostridium_sensu_stricto_1, Peptoclostridium, and Veillonella had significant differences among the 4 groups at the genus level. In animal experiments, the AI-2 level in feces of NEC mice (56.89 ± 11.87) was significantly lower than that in the feces of control group mice (102.70 ± 22.97). The microbiota compositions of NEC and control group mice at the phylum level were not significantly different. At the genus level, Klebsiella, Clostridium_sensu_stricto_1, and Peptoclostridium abundances in the NEC group increased significantly compared with those in the control group (P < 0.05). In addition, Lactobacillus, Pasteurella, and Parabacteroides abundances in the NEC group decreased significantly compared with those in the normal control group (P < 0.05), while Lactobacillus, Pasteurella, and Parabacteroides abundances had the opposite trend. The AI-2 concentration decreased significantly in the acute phase of NEC and increased gradually in the convalescent phase. We conclude that the concentration of AI-2 was correlated with intestinal flora disorder and different stages of disease. AI-2 may be a new biomarker for the diagnosis and monitoring of NEC. Trial Registry: ClinicalTrials.gov; ChiCTR-ROC-17013746; URL: www.clinicaltrials.gov.

[Study on esophageal motor function against reflux after esophagogastric anastomosis with mucosal valve].

OBJECTIVE: To find an effective operative procedure against reflux after esophagogastric anastomosis with mucosal valve. METHODS: Four hundred and sixty-four patients with esophageal or cardiac cancer were randomly divided into three groups according to anastomosis modes. Group A underwent esophagogastric anastomosis with mucosal valve (175 cases), group B with mechanical stapler (151 cases) and group C one layer anastomosis with handcraft suture (138 cases). The gastroesophageal reflux index (GERI) was examined by isotope,and 24 h esophageal pH was also monitored. The esophageal motor function was compared among three groups. RESULTS: The reflux rates were 0, 33.3%, and 6.7% in group A, B, C respectively. The esophageal motor function and the 24 h esophageal pH monitoring indicated that the various indexes were approaching to the normal level in group A, but the various indexes in group B and C were significantly different from the normal values (P< 0.05). CONCLUSION: The esophagogastric anastomosis with mucosal valve has better antireflux effect and can prevent the reflux esophagitis after esophageal or cardiac cancer eradication.