Necrotizing Enterocolitis Group Research Articles

Profile analysis reveals endogenous RNAs regulate necrotizing enterocolitis progression

Necrotizing enterocolitis (NEC) is one of the most common and devastating gastrointestinal diseases in preterm newborns, and its underlying mechanisms remain unclear. Non-coding RNAs (ncRNAs) play critical roles in intestinal diseases; however, little is known about their roles in the development of NEC. To gain a deeper understanding of the pathophysiological mechanism of NEC, long non-coding RNAs (lncRNAs), microRNAs (miRNAs) and mRNAs were detected in an NEC rat model. In total, 1820 lncRNAs, 118 miRNAs and 929 mRNAs were differentially expressed in NEC group. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated that these molecules were enriched in apoptosis, autophagic cell death, TLR4 signaling pathway, Notch signaling pathway, and mTOR signaling pathway. These pathways are thought to be closely associated with NEC. Furthermore, a lncRNA-miRNA interaction network was constructed, and four of the novel, differentially expressed lncRNAs with large changes were randomly verified using quantitative polymerase chain reaction (qPCR). The GO and KEGG pathway analysis of these four lncRNAs showed that they were associated with the negative regulation of TLR4 signaling pathway and Notch signaling pathway. In conclusion, our study revealed that these differentially expressed lncRNAs may participate in the development of NEC via interactions with miRNAs and may serve as possible biomarkers and target genes in NEC.

Changes of long non-coding RNA expression profiles in intestinal tissues from neonatal necrotizing enterocolitis.

Long non-coding RNAs (lncRNAs) were involved in the development and regulation of necrotizing enterocolitis (NEC) in premature infants. To investigate the changes of lncRNA expression profile in intestinal tissues of NEC for its possible mechanisms. Intestinal samples were collected from 11 patients with NEC who needed surgery(the NEC group), and 7 from neonatal non-NEC patients with surgery (the Control group).LncRNA's changes in intestinal samples (3 in the Control group and 3 in the NEC group) were analyzed with high-throughput sequencing.Part of the remaining samples were detected by real-time polymerase chain reaction (real-time PCR), and the results were used to validate the results of high-throughput sequencing. Gene Ontology (GO) analysis and KEGG signaling pathway analysis were performed on differentially expressed genes. There were 5 257 different lncRNAs between the control group and the NEC group. The results of up-regulated lncRNAs (NONHSAG008675.3, NONHSAG020715.2, NONHSAG038187.2) and down-regulated lncRNA (NONHSAG028744.3) were confirmed to be consistent with the results of high-throughput sequencing. Expressions of DUOX2, IL-6, TNF, and SAA1 were up-regulated in intestinal tissues of NEC. GO analysis showed that the different lncRNAs were involved in regulation of stimulation, molecular junction and function, and signal transduction and transcription. KEGG analysis identified mainly biological pathways involved in inflammatory bowel disease, PI3K-Akt, NF-κB, etc. LncRNAs might be involved in the pathogenesis of NEC and the inflammation-related lncRNAs may be one of the key factors.

Risk factors of necrotizing enterocolitis in neonates with sepsis: A retrospective case-control study

Sepsis, a severe infectious disease in the neonatal period, is considered a risk factor for necrotizing enterocolitis (NEC). To investigate the specific risk factors for NEC in septic infants, septic infants admitted to our center from January 2010 to April 2018 were included. Septic neonates with proven NEC (Bell’s stage ⩾II) were enrolled in the NEC group, and those without NEC were enrolled in the control group. Demographics, clinical characteristics, and risk factors were compared between the two groups. Univariate and logistic regression analyses were used to evaluate the potential risk factors for NEC. A total of 610 septic neonates were included, of whom 78 (12.8%) had complicated NEC. The univariate analysis indicated that infants with NEC had a lower birth weight, a lower gestational age, and older age on admission than those without NEC (P < 0.05). Higher rates of anemia, prolonged rupture of membranes (PROM) (⩾18 h), pregnancy-induced hypertension, late-onset sepsis (LOS), red blood cell transfusion and hypoalbuminemia were observed in the NEC group than in the non-NEC group (P<0.05). Logistic regression analysis revealed LOS (P = 0.000), red blood cell transfusion (P = 0.001) and hypoalbuminemia (P = 0.001) were associated with the development of NEC. Among NEC infants, those who needed red blood cell transfusion had a longer hospitalization duration than those who did not need transfusion (P < 0.05). LOS, red blood cell transfusion and hypoalbuminemia were independent risk factors for the development of NEC in infants with sepsis. Taking measures to reduce the occurrence of hypoproteinemia and severe anemia may help to reduce the occurrence of NEC in septic neonates.

Early Biomarkers in Neonatal Necrotizing Enterocolitis: A Pilot Study.

Necrotizing enterocolitis (NEC) is a frequent serious disease of the digestive system in neonates. It is considered as an important cause of serious neonatal complication and death. Therefore, its early suspicion and proper management are important. Early and sensitive detection of neonatal NEC through determination of levels of fecal calprotectin (FCP), serum levels of procalcitonin (PCT), high-sensitivity C-reactive protein (hs-CRP), epithelial neutrophil activating peptide-78 (ENA-78), and interleukin 18 (IL-18). This prospective case control study was conducted in Tanta University Hospital from June 2016 to March 2018. The study included 20 healthy neonates (control group) and 20 NEC newborn patients. They were all subjected to the measurement of levels of FCP and serum levels of hs-CRP, PCT, ENA-78, IL-18, Malondialdehyde (MDA), and total antioxidant capacity (TAC). Receiver operating characteristic (ROC) curve analysis was conducted for FCP, ENA-78, PCT, hs-CRP, and IL-18. The study found a detectable increase in FCP level and serum levels of hs-CRP, PCT, ENA-78, IL-18, and MDA in NEC group in comparison to their levels in the control group. Also, it found a detectable decline in the levels of TAC in comparison to its level in the control group. FCP, ENA-78, and PCT can be considered as early markers for diagnosis of NEC.

Clinical characteristics of neonatal fulminant necrotizing enterocolitis in a tertiary Children's hospital in the last 10 years.

The aim of this retrospective study was to explore the risk factors and clinical characteristics related to neonatal fulminant necrotizing enterocolitis (NEC). From 1 November 2007 to 31 October 2017, 352 neonates who were diagnosed with NEC (Bell stage ≥ΠB) and admitted to the Children’s Hospital of Chongqing Medical University were enrolled. Among these patients, 112 (31.82%) cases fulfilled the definition of fulminant NEC, and 62.5% (70/112) of fulminant cases presented a poor prognosis. All the survivors in the fulminant NEC group underwent surgery. Those in the fulminant NEC group were more likely to have the following clinical features: sepsis preceding NEC (P<0.001), abdominal distention (P<0.001), bowel sound disappearance (P = 0.001), leukopenia or neutropenia (P<0.001), C-reactive protein (CRP) <10 mg/L (P = 0.003), procalcitonin (PCT) < 2 μg/L (P<0.001), pH ≤7.2 (P<0.001), and radiographic evidence of pneumoperitoneum (P<0.001) or seroperitoneum on ultrasonography (P = 0.017). In conclusion, fulminant NEC is characterized by urgent onset and prompt deterioration, potentially resulting in death. The lack of unique characteristics makes it difficult to recognize by medical caregivers. Close observation, early detection and timely surgical intervention may improve the prognosis.

Protective effect of Lactobacillus reuteri against oxidative stress in neonatal mice with necrotizing enterocolitis

To investigate the protective effect of L. reuteri DSM17938 strain against oxidative stress in a neonatal mouse model of necrotizing enterocolitis (NEC) and explore the possible mechanism. Ninety-six 10-day-old neonatal C57BL/6J mice were equally randomized into control group, NEC group, and NEC+ L. reuteri group. The pathological changes of the ileocecal intestinal tissue were evaluated with HE staining and double-blind pathological scoring. The mRNA and protein expressions of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in the intestinal tissues were detected using quantitative real-time PCR and ELISA, respectively. Colorimetric assays were used to determine the activity of superoxide dismutase (SOD) and its inhibition rate, malondialdehyde (MDA), glutathione (GSH), oxidized glutathione (GSSG), and GSSG/ GSH ratio. Compared with those in the control group, the neonatal mice in NEC group showed significant weight loss (P < 0.05), obvious intestinal injury, increased pathological scores (P < 0.05), increased expressions of TNF-α and IL-1β mRNA and proteins (P < 0.05), decreased SOD activity and inhibition rate, decreased GSH, and significantly increased MDA, GSSG, and GSSG/GSH ratios (P < 0.05). Treatment with L. reuteri obviously decreased the pathological scores, expressions of TNF-α and IL-1β (P < 0.05), MDA, GSSG, and GSSG/GSH ratio (P < 0.05), and significantly increased SOD activity, its inhibition rate, and GSH level in the mice with NEC, but the survival rate was not significantly different between NEC and L. reuteri-treated groups (P > 0.05). L. reuteri DSM17938 can offer protection against NEC in mice by reducing oxidative stress and increasing antioxidant capacity of the intestinal tissue to suppress intestinal inflammations.

The MicroRNA Landscapes Profiling Reveals Potential Signatures of Necrotizing Enterocolitis in Infants.

The purpose of this study is to extract potential signatures involved in necrotizing enterocolitis (NEC). The data set GSE68054 was acquired from Gene Expression Omnibus repertory, and differentially expressed genes were screened between NEC and control group. After that, the target prediction analysis and functional enrichment analysis were carried out. Moreover, protein-protein interaction (PPI) and functional analyses of top 20 key genes in PPI network were undertaken. In addition, drug-gene interaction prediction analysis was also carried out to screen the key genes for NEC treatment. Totally, there were 16 differentially expressed microRNAs (DEmiRNAs; 2 upregulated and 14 downregulated miRNAs) between NEC and controls. Furthermore, 354 DEmiRNAs-targets interactions were predicted, including KDR-miR-200c-3p and YWHAG-miR-200a-3p. Several genes such as BDNF, KDR, YWHAG, YWHAE, and YWHAB acted as hub genes in PPI network and functional analysis implied that multiple genes, including YWHAG, YWHAE, and YWHAB, were strongly linked with positive regulation of protein insertion into mitochondrial membrane involved in apoptotic signaling pathway, whereas BDNF and KDR were markedly enriched in PI3K-Akt signaling pathway and neurotrophin signaling pathway, respectively. Interestingly, we found that KDR showed strong correlations with 16 drugs based on drug-gene interaction predictive analysis. YWHAG, YWHAE, YWHAB, BDNF, and KDR might serve as promising therapeutic targets for NEC management. In addition, miR-200a-3p was also possibly involved in the molecular mechanism of CNEC, presenting deeper insights into NEC diagnosis and treatment.

Lactoferrin Reduces Necrotizing Enterocolitis Severity by Upregulating Intestinal Epithelial Proliferation.

Necrotizing enterocolitis (NEC) is a devastating intestinal illness in premature infants characterized by severe intestinal inflammation. Despite medical interventions, NEC mortality remains alarmingly high, which necessitates improved therapies. Lactoferrin is among the most abundant proteins in human milk and has important immunomodulatory functions. While previous studies have indicated protective effects of lactoferrin against neonatal sepsis and NEC, the underlying mechanism remains unclear. We hypothesize that lactoferrin downregulates inflammation and upregulates proliferation in intestinal epithelium during NEC injury. NEC was induced by hypoxia, gavage feeding of hyperosmolar formula and lipopolysaccharide between postnatal day P5 and P9 (n = 8). Breastfed mice were used as control (n = 7). Lactoferrin (0.3 g/kg/day) was administered once daily by gavage from P6 to P8 in both NEC (NEC + Lac; n = 9) and control mice (Cont + Lac; n = 5). Distal ileum was harvested on P9 and analyzed for disease severity, inflammation, and proliferation. Groups were compared using one-way ANOVA and t-test appropriately; p < 0.05 was considered significant. Compared to NEC group, lactoferrin-treated NEC mice had reduced disease severity, reduced inflammation markers IL-6 and TNF-α expression and increased intestinal stem cell marker Lgr5 + expression. Lactoferrin-treated NEC mice exhibited increased nuclear β-catenin, indicating upregulated Wnt pathway, and increased Ki67 positivity, suggesting enhanced proliferation. Furthermore, lactoferrin administration to control mice did not affect intestinal inflammation as well as Lgr5 + stem cell expression and epithelial proliferation. This supports the safety of lactoferrin administration and indicates that the beneficial effects of lactoferrin are present when intestinal injury such as NEC is present. Lactoferrin administration reduces the intestinal injury in experimental NEC by downregulating inflammation and upregulating cell proliferation. This beneficial effect of lactoferrin in stimulating cell proliferation is mediated by the Wnt pathway. This experimental study provides insights on the mechanism of action of lactoferrin in NEC and the role of lactoferrin in enteral feeding.

Role of lactadherin in intestinal barrier integrity in experimental neonatal necrotizing enterocolitis

Necrotizing enterocolitis (NEC) is one of the most widespread and devastating gastrointestinal diseases in neonates. Destruction of the intestinal barrier is the main underlying cause of NEC. The aim of this study was to determine the role of lactadherin in preventing NEC in a neonatal rat model and investigate the molecular mechanism of lactadherin-mediated protection of the intestinal barrier. Neonatal rats were divided into three groups: dam feeding (DF), NEC (NEC), and NEC supplemented with 10 μg/(g·day) recombinant human lactadherin (NEC+L). Intestinal permeability, tissue damage, and cell junction protein expression and localization were evaluated. We found that lactadherin reduced weight loss caused by NEC, reduced the incidence of NEC from 100% to 46.7%, and reduced the mean histological score for tissue damage to 1.40 compared with 2.53 in the NEC group. Intestinal permeability of lactadherin-treated rats was significantly reduced when compared with that of the NEC group. In addition, the expression levels of JAM-A, claudin 3, and E-calcium in the ileum of NEC group animals increased compared with those in the ileum of DF group animals, and these levels decreased in the NEC+L group. Lactadherin changed the localization of claudin 3, occludin, and E-cadherin in epithelial cells. The mechanism underlying lactadherin-mediated protection of the intestinal barrier might be restoring the correct expression levels and localization of tight junction and adherent junction proteins. These findings suggest a new candidate agent for the prevention of NEC in newborns.

Application of abdominal sonography in diagnosis of infants with necrotizing enterocolitis.

The purpose of this study was to explore the diagnostic significance of abdominal sonography (AUS) in infants with Necrotizing enterocolitis (NEC) admitted to a neonatal intensive care unit to better evaluate the ability of AUS to differentiate necrotizing enterocolitis from other intestinal diseases.All patients diagnosed with NEC at the Department of General Surgery and Neonatal Surgery, Qilu Children‘s Hospital between 1st, Jun, 2010 and 30th, Dec, 2015. The logistic regression analysis and the area under receiver operating characteristic (ROC) curve (AUCs) were also used to identify the sonographic factors for diagnosing NEC.For the entire cohort of 91 patients, we divided these patients into suspected NEC (n = 35) group and definite NEC (n = 56) group. After adjusting for competing sonographic factors, we identified that thick bowel wall (more than 2.5 mm) (P = .013, OR: 1.246), intramural gas (pneumatosis intestinalis) (P = .002, OR:1.983), portal venous gas (P = .022, OR:1.655) and reduced peristalsis (P = .011, OR:1.667) were independent diagnostic factors associated with NEC. We built a logistic model to diagnose NEC according to the results of multivariable logistic regression analysis. We found the AUROC for thick bowel wall (more than 2.5 mm), intramural gas (pneumatosis intestinalis), portal venous gas and reduced peristalsis were significantly lower than the AUROC for the logistic model was 0.841 (95% CI: 0.669 to 0.946).We found that thick bowel wall (more than 2.5 mm), intramural gas (pneumatosis intestinalis), portal venous gas and reduced peristalsis were independent diagnostic factors associated with NEC. The logistic model was significantly superior to the single sonographic parameter for diagnosing NEC.

Recombinant human soluble thrombomodulin reduces the severity and incidence of necrotizing enterocolitis in a newborn rat model.

Necrotizing enterocolitis (NEC) remains the leading cause of death in preterm infants. Recombinant human soluble thrombomodulin (rTM) has been reported to have anti-inflammatory effects as well as antithrombogenic effects. The aim of this study was to evaluate the effect of rTM in a rat NEC model. NEC was induced by enteral feeding with hyperosmolar formula, gavage administration of lipopolysaccharide and asphyxia stress. Controls were fed by their mother ad libitum. In the treatment group, rTM was administered subcutaneously twice (once each on the first and second day). All animals surviving beyond 96h or that developed signs of distress were euthanized. The ileum was harvested for a histological evaluation and the measurement of the mRNA and protein expression. The rate of NEC-like intestinal injury in the treatment group (9/25, 36%) was significantly lower than in the NEC group (25/34, 73.5%). Tissue levels of TNF-α, IL-6 and HMGB1 were significantly elevated in the NEC group, whereas those in the treatment group were decreased to similar values as in the control group. Our experimental study showed that rTM is able to reduce the severity and incidence of NEC. It may be an alternative option for the treatment of NEC.

Gastrointestinal sequelae and growth impairment at school age following necrotising enterocolitis in the newborn period.

To evaluate gastrointestinal sequelae and growth impairment at school age in children who suffered from necrotising enterocolitis (NEC). This historic cohort study compared all surviving children born in Denmark between 1 January 2002 and 31 December 2011 with NEC in the newborn period, to surviving children without NEC, but same gestational age, birthweight and year of birth. Outcomes were investigated through a parental questionnaire, including gastrointestinal and growth-related outcomes. We performed exploratory ad hoc analysis, by adjusting for possible confounding and by dividing NEC children into surgical and medical. In total, 163 children with NEC (50%) and 237 (36%) without NEC completed the parental questionnaire. Episodes of diarrhoea were more often reported in the NEC group (p=0.0002). The increased risk seemed to be limited to those who underwent surgery for NEC. The absence from school (1.67 versus 1.31days), rate of low height for age (17.9 versus 12.1%) and weight (29.9 versus 31.6kg) did not differ significantly between children with NEC and children without NEC. Our findings suggest that long-term gastrointestinal complications following NEC appeared to be of little clinical importance at the population level and therefore do not encourage specific routine follow-up.

Calprotectin and Intestinal Fatty Acid Binding Protein (I-FABP) Level in Preterm Neonates with Necrotizing Enterocolitis

Necrotizing enterocolitis (NEC) is inflammatory state of intestinal tissue which mostly occurred in preterm neonates and associated with ischemia and inflammation. This study was aimed to investigate calprotectin level (inflammatory marker) and intestinal fatty acid binding protein (I-FABP) (ischemia marker) in preterm neonates with NEC. This research was designed as cross sectional which involve 32 preterm neonates divided into 2 groups as follows: NEC group (n=16 subjects) and control group (n=16 subjects). Diagnosis of NEC was established by clinical and radiological signs (abdominal distension and pneumatosis). Fecal calprotectin and urinary I-FABP were measured using ELISA method. Results showed that fecal calprotectin and urinary I-FABP was significantly higher in NEC group as compared to control group ( Mann-Whitney test , p<0.05). Both calprotectin and I-FABP was positively correlated with NEC state (Spearman correlation test, p=0.000, r=0.866). Moreover, calprotectin and I-FABP level was positively correlated with grade (Bell’s classification) and type (Gordon’s classification) of NEC (Spearman correlation test, p<0.05). We concluded that calprotectin and I-FABP level was higher in NEC group. Moreover, I-FABP but not calprotectin, were correlated with grade and type of NEC.

The relation between serum levels of epidermal growth factor and necrotizing enterocolitis in preterm neonates.

PurposeNecrotizing enterocolitis (NEC) is one of the most serious complications of prematurity. Many risk factors can contribute to the development of NEC. The epidermal growth factor (EGF) plays a major role in intestinal barrier function, increases intestinal enzyme activity, and improves nutrient transport. The aim of this study was to assess the role of epidermal growth factor in the development of NEC in preterm neonates.MethodsIn this study, 130 preterm neonates were included and divided into 3 groups, as follows: group 1, 40 preterm neonates with NEC; group 2, 50 preterm neonates with sepsis; and group 3, 40 healthy preterm neonates as controls. The NEC group was then subdivided into medical and surgical NEC subgroups. The serum EGF level was measured using enzyme-linked immunosorbent assay.ResultsSerum EGF levels (pg/dL) were significantly lower in the NEC group (median [interquartile range, IQR], 9.6 [2–14]) than in the sepsis (10.1 [8–14]) and control groups (11.2 [8–14], P<0.001), with no significant difference between the sepsis and control groups, and were positively correlated with gestational age (r=0.7, P<0.001). A binary logistic regression test revealed that low EGF levels and gestational ages could significantly predict the development of NEC. The receiver-operating characteristic curve for EGF showed an optimal cutoff value of 8 pg/mL, with 73.3% sensitivity, 98% specificity, and an area under the curve of 0.92.ConclusionThe patients with NEC in this study had significantly lower serum EGF levels (P<0.001), which indicated that EGF could be a reliable marker of NEC in preterm neonates.

Early diagnosis of necrotizing enterocolitis by plasma RELMβ and thrombocytopenia in preterm infants: A pilot study

As the inflammatory regulators, Resistin-like molecule β (RELMβ) and Resistin might be potential biomarkers of necrotizing enterocolitis (NEC), while thrombocytopenia is often related to the severity of NEC, clinical observation suggests that thrombocytopenia might be an early biomarker of NEC. The aim of this study was to evaluate whether RELMβ, Resistin and thrombocytopenia could be biomarkers for early diagnosis of NEC in preterm infants. From January 2016 to March 2018, twenty-nine NEC preterm infants who were diagnosed with NEC (Bell's stage ≥Ⅱ) by two independent neonatologists and twenty-nine non NEC preterm infants at neonatal intensive care unit in our hospital were enrolled in this case-control study. Preterm infants with a history of serious infections (sepsis, pneumonia), asphyxia, and congenital malformations were excluded from the study. The plasma RELMβ and Resistin were evaluated by enzyme linked immunosorbent assay (ELISA) and serum platelet levels were measured directly by ordinary light microscope at the diagnosis of NEC (Bell's stage ≥Ⅱ). Plasma RELMβ levels in NEC group were significantly higher than control group (P<0.05). The optimal cut-off value of RELMβ determined by receiver operating characteristic curve (ROC) was 378.3ng/L. The overall estimates for sensitivity and specificity of high RELMβ concentrations in the detection of neonatal NEC were 71.4% and 91.7%, respectively. No significant difference was found in plasma Resistin levels between two groups (P>0.05). If platelet level was less than 157×109/L, the sensitivity and specificity were 69.34% and 82.87%, respectively. Interestingly, the combination of RELMβ and thrombocytopenia increased sensitivity and specificity to 82.89% and 93.21%, respectively. The combination of RELMβ and thrombocytopenia was a reliable biomarker for the early diagnosis of NEC in this study with 82.89% sensitivity and 93.21% specificity, respectively.

Associations between Red Blood Cell Transfusions and Necrotizing Enterocolitis in Very Low Birth Weight Infants: Ten-Year Data of a Tertiary Neonatal Unit.

Background and Objective: Necrotizing enterocolitis (NEC) remains an important cause of mortality in preterm neonates. There are many risk factors for NEC; however, probably the most controversial one is red blood cell transfusions (RBCT). The data concerning the link between NEC and RBCT has been conflicting. Therefore, we aimed to analyze the association between NEC and RBCT in Neonatal Intensive Care Unit (NICU) at the Hospital of Lithuanian University of Health Sciences. Materials and Methods: We used the Very Low Birth Weight (VLBW) Infants database to match all infants with ≥2a Bell’s stage NEC admitted between 1 January 2005 and 31 December 2014 (n = 54) with a control group (n = 54) of similar gestational age and birth weight and without NEC. We analyzed the charts of these infants and performed statistical analysis on 20 clinical variables including RBCT. Results: The main clinical and demographic characteristics did not differ between the two groups. All variables associated with RBCT (receipt of any RBCT, the number of transfusions and the volume transfused in total) were significantly higher in the NEC group both before the onset of NEC and throughout the hospitalization. RBCT increased the odds of NEC even after adjustment for confounding factors. In addition, we found that congenital infection was more abundant in the NEC group and increased the odds of NEC 2.7 times (95% confidence interval CI (1.1, 6.3), p = 0.024). Conclusions: A higher number and the total volume of RBCT are associated with an increased risk of NEC in VLBW infants. The presence of congenital infection might identify the infants at risk.

Inhibition of HMGB1 improves necrotizing enterocolitis by inhibiting NLRP3 via TLR4 and NF-κB signaling pathways.

To explore the relationship between high-mobility group box 1 (HMGB1) and NLR pyrin domain containing 3 (NLRP3) in the development of necrotizing enterocolitis (NEC). NEC rat models were constructed and treated with HMGB1 inhibitor glycyrrhizin (GL) with different concentration. An inflammatory condition of intestinal tissue in newborn NEC rats was observed by hematoxylin and eosin staining. The messenger RNA (mRNA) and protein expression of HMGB1, NLRP3, toll-like receptor 4 (TLR4), nuclear factor-κB (NF-κB), and caspase 1 were determined by real-time polymerase chain reaction and western blot analysis, respectively. The content of interleukin (IL)-1β and tumor necrosis factor-α (TNF-α) was determined by enzyme-linked immunosorbent assay. Human intestinal epithelial cell lines were induced to NEC by lipopolysaccharides (LPSs). LPS-induced cells were transfected with small interfering RNA-HMGB1 and NLRP3 plasmid vector. The mRNA and protein expression of HMGB1, NLRP3, TLR4, NF-κB, caspase 1, IL-1β, and TNF-α were determined by real-time PCR and western blot analysis, respectively. The mRNA and protein expression of HMGB1 and NLRP3 in the NEC group was significantly higher than the control group. Inhibition of HMGB1 expression improved intestinal inflammation in newborn NEC rats. The expression of HMGB1, NLRP3, TLR4, NF-κB, and caspase 1 was upregulated in NEC and was weakened after treating with GL. LPS induction to intestinal epithelial cells markedly increased the expression of HMGB1, NLRP3, TLR4, NF-κB, caspase 1, IL-1β, and TNF-α. The knockdown of HMGB1 abolished the increase of expression, whereas further transfection with NLRP3 plasmid vector recovered the increase. HMGB1 and NLRP3 were all upregulated in the development of NEC. Inhibition on HMGB1 could improve the intestinal inflammation in NEC by inhibiting NLRP3 via TLR4 and NF-κB signaling pathways.

The contribution of morphine sulfate to the development of necrotizing enterocolitis in preterm infants: a matched casecontrol study.

Zvizdic Z, Milisic E, Jonuzi A, Terzic S, Zvizdic D. The contribution of morphine sulfate to the development of necrotizing enterocolitis in preterm infants: a matched case-control study. Turk J Pediatr 2019; 61: 513-519. The aim of the study was to determine whether morphine sulfate administration is associated with an increased risk of necrotizing enterocolitis (NEC) in preterm infants supported by mechanical ventilation due to respiratory failure. The matched case-control retrospective study was carried out at the Neonatal Intensive Care Unit (NICU) of the University Clinical Center Sarajevo, on 122 preterm infants classified into total NEC group and control group. The total NEC group was further divided into medical NEC and surgical NEC subgroups. The association between the use and duration of morphine sulfate infusion and the development of NEC was evaluated in both unadjusted and adjusted analysis. Preterm infants who developed NEC were on mechanical ventilation more frequently compared to premature infants without signs of NEC (Mann- Whitney U test; p=0.0031). A positive correlation between the frequency of receiving morphine sulfate and the development of NEC was observed (Chi square test of independence; p=0.0001). The risk of NEC in preterm infants was increased by the use of morphine sulfate. Validation of this observation in other populations is warranted.

Complications of newborn enterostomies.

AIMTo evaluate the occurrence and severity of enterostomy complications in newborns suffering from different intestinal disorders.METHODSA 10-year retrospective cohort study (2008-2017) investigated newborns that underwent enterostomy formation and reversal for different intestinal disorders. Only infants less than 28 d old at the time of enterostomy creation were included in the study (corrected age was applied in the cases of preterm neonates). The patients were divided into two groups according to their underlying diseases. Group 1 included infants suffering from necrotizing enterocolitis (NEC), whereas Group 2 included newborns diagnosed with intestinal disorders other than NEC, such as meconium obstruction, anorectal malformation, focal intestinal perforation, ileus, intestinal atresia and volvulus. The primary outcome measure was enterostomy-related morbidity. The data were analyzed statistically using Pearson’s χ2 test or Fisher’s exact test for categorical variables and the Wilcoxon-Mann-Whitney U-Test for continuous variables.RESULTSIn total, 76 infants met the inclusion criteria and were evaluated for enterostomy-related complications. Neither group showed significant differences regarding gender, gestational age, weight at birth or weight at enterostomy formation. Infants suffering from NEC (Group 1) were significantly older at enterostomy formation than the neonates of Group 2 [median (range), 11 (2-75) d vs 4 (1-101) d, P = 0.004)]. Significantly more ileostomies were created in Group 1 [47 (92.2%) vs 16 (64.0%), P = 0.007], whereas colostomies were performed significantly more often in Group 2 [2 (3.9%) vs 8 (32.0%), P = 0.002]. The initiation of enteral nutrition after enterostomy was significantly later in Group 1 infants than in Group 2 infants [median (range), 5 (3-13) vs 3 (1-9), P < 0.001]. The overall rate of one or more complications in patients of both groups after enterostomy formation was 80.3%, with rates of 86.3% in Group 1 and 68.0% in Group 2 (P = 0.073). Most patients suffered from two complications (23.7%). Four or more complications occurred in 21.6% of the infants in Group 1 and in 12.0% of the infants in Group 2 (P = 0.365). Following enterostomy closure, at least one complication was observed in 26.0% of the patients (30.6% in Group 1 and 16.7% in Group 2, P = 0.321). The occurrence of complications was not significantly different between neonates with NEC and infants with other intestinal disorders. 48 (65.8%) patients required no treatment or only pharmacological treatment for the complications that occurred [Clavien-Dindo-Classification (CDC) < III], while 25 (34.2%) required surgery to address the complications (CDC ≥ III). Early reversal of the enterostomy was performed significantly more often (P = 0.003) and the time to full enteral nutrition after closure was significantly longer [median (range), 7 (3-87) d vs 12 (5-93) d, P = 0.006] in infants with a CDC grading ≥ III than in infants with a CDC grading < III.CONCLUSIONComplications occur in almost all infants with enterostomies. The majority of these complications are minor and do not require surgical treatment. There is a clear trend that neonates with NEC have a higher risk for developing complications than those without NEC.

Obestatin Reduces Intestinal Damage in Experimental Necrotizing Enterocolitis in Newborn Rats.

To investigate the effect of pretreatment with obestatin (OB), an endogenous hormone also found in mother's milk, in experimental necrotizing enterocolitis (NEC). Pups were randomized into four groups: control, OB-control, NEC, and OB-NEC. NEC was induced by asphyxia and hypothermia in the NEC and OB-NEC groups. OB was administered to the OB-control and OB-NEC groups. Macroscopic scoring of the intestinal tract was evaluated and tissue samples were obtained for histopathological and biochemical examination on the fourth day. OB improved the macroscopic appearance of the gut and the clinical score during the experiment (p < 0.05). The rate of occurrence of NEC in the OB-NEC group was lower than the NEC group (p = 0.001). OB prevented necrosis and reduced the number of apoptotic cells in the OB-NEC group compared with the NEC group (p = 0.006). Furthermore, interleukin-6 and malondialdehyde levels in the OB-NEC group were lower than the NEC group (p < 0.05). OB reduced intestinal damage and prevented necrosis through anti-inflammatory and antiapoptotic effects in experimental NEC. This effect of OB should be confirmed in clinical studies. Furthermore, future research should investigate whether OB plays a role in NEC pathogenesis or NEC is associated with OB levels in the serum and in breast milk.