Do hematological biomarkers predict surgical necrotizing enterocolitis?

Necrotizing enterocolitis (NEC) is a devastating gastrointestinal emergency in preterm infants and the clinical presentation of NEC may vary with gestational age. We lack reliable biomarkers for early diagnosis of NEC limiting timely intervention. Hematological changes in NEC are actively researched for their potential role as biomarkers. The pattern and severity of hematological abnormalities have been correlated with rapid progression, the need for surgery, increased risk of mortality, and morbidity. In this issue of Pediatric Research, Chong et al. report GA-specific hematological biomarkers in preterm infants with NEC that could predict the need for surgery. Thrombocytopenia at NEC onset was an independent predictor of surgical intervention in extremely preterm infants. Persistent thrombocytopenia and lymphopenia at 72 h and elevated C-reactive protein at 48 h after NEC onset, predicted surgery in infants of 28 to <32 weeks GA. Persistent thrombocytopenia at 24 h after the onset of NEC was predictive of mortality in infants who underwent surgery. Well-designed, prospective, multi-center studies are needed to confirm the role of hematological biomarkers in early diagnosis and prognostication in NEC.

Outcomes by disease onset, sex, and intervention in neonates with SIP and surgical NEC.

BACKGROUND: Outcomes of infants following surgical necrotizing enterocolitis (NEC) and spontaneous intestinal perforation (SIP) categorized by the age of onset, interventions, and sex are not well defined. METHODS: Retrospective comparison of infants categorized by age of onset (NEC at <10, 10-20, and >20 days) and SIP at <7 versus ≥7 days), sex, and intervention [Penrose Drain (PD) vs. laparotomy]. RESULTS: A total of 114 infants had NEC and 37 had SIP. On multinomial logistic regression, infants with NEC/SIP onset >20 days had significantly lower odds of small bowel involvement (aOR = 0.07, 95% CI: 0.01-0.33, p = 0.001), higher necrosis (aOR = 3.59, 95% CI: 1.34-9.65, p = 0.012) and higher CRP (p = 0.004) than onset <10 days. Initial laparotomy was associated with more bowel loss (24.1 cm [12.3; 40.6] vs.12.1 [8.00; 23.2]; p = 0.001), small and large intestine involvement (47.1% vs 17.2%; p = 0.01), and ileocecal valve resection (42% vs. 19.4%; p = 0.036) than initial PD therapy. Females underwent fewer small bowel resections (52.3% vs 73.6%; p = 0.025) but had higher surgical morbidity (53.7% vs. 24.7%.; p = 0.001) than males. CONCLUSION: Clinical, radiological, and histopathological presentation and outcomes in preterm infants with surgical NEC/SIP are associated with age of disease onset, sex, and initial intervention. IMPACT: Neonates with surgical NEC onset >20 days had more severe necrosis, inflammation, kidney injury, and bowel loss than those with <10 days. Initial laparotomy was associated with later age onset, more bowel loss, and ileocecal valve resection compared to initial PD treatment, but not with differences in mortality or length of stay. Female sex was associated with lower maturity, more placental malperfusion, less often small bowel involvement, lower pre-NEC hematocrit as well as higher surgical morbidity than males. Whether the management of surgical NEC and SIP should differ by the age of onset requires further investigation.

Necrotizing Enterocolitis-Associated Acute Kidney Injury-Transforming the Paradigm.

Necrotizing enterocolitis (NEC) is one of the most common conditions requiring emergency surgery in the neonatal intensive care unit and is associated with a septic shock-like state contributing to multiorgan dysfunction. NEC affects 6 to 10% of very low-birth-weight infants and remains a leading cause of death. The occurrence of severe acute kidney injury (AKI) following surgical NEC is a harbinger of multiple morbidities. This review presents current evidence about the clinical impact of NEC-associated AKI on the clinical outcomes. Studies evaluating nephroprotective strategies to prevent AKI and its consequences are greatly needed to improve the postoperative recovery and clinical outcomes in neonates with NEC. Future observational studies and clinical trials in preterm infants with NEC prioritize measuring short-term (AKI) and longer term (chronic kidney disease) kidney outcomes. KEY POINTS: · Severe AKI is common following surgical NEC.. · Severe AKI following NEC is associated with poor clinical outcomes.. · Studies evaluating nephroprotective strategies to prevent AKI and its consequences are needed.. IMPACT: · Severe AKI (stage 2 and 3) occurs in 32.6% of neonates after NEC diagnosis and in 58.7% following surgical NEC diagnosis.. · NEC-associated AKI is associated with severe postoperative course, moderate-to-severe bronchopulmonary dysplasia, surgical complications, brain injury, and longer hospital stay in preterm infants.. · Severity of NEC-associated AKI can be utilized by bedside providers for the prognostication of clinical outcomes in preterm infants..

Placenta and Intestinal Injury in Preterm Infants.

Necrotizing enterocolitis (NEC) is one of the most common gastrointestinal conditions affecting 6 to 10% of low-birth-weight infants and remains a leading cause of death. The risk factors associated with NEC are complex and multifactorial, including preterm birth and intrauterine exposure to inflammation and hypoxia. Chorioamnionitis has been associated with intestinal injury in animal and human clinical studies. This review presents current evidence about the clinical impact of the intrauterine environment on intestinal injury during pregnancy and postpregnancy. We present information from our own clinical and laboratory research in conjunction with information collected from an extensive search in the databases PubMed, EMBASE, and Scopus. Prospective multicenter studies, including accurate and precise clinical, maternal, and laboratory predictors (e.g., inflammatory biomarkers), will help identify the mechanisms associated with the placental pathology, the development of NEC, and the impact of in utero-triggered inflammation on the clinical outcomes. Filling the knowledge gap to link the inflammatory surge to postnatal life will aid in identifying at-risk infants for NEC in a timely manner and facilitate the development of novel immunomodulatory treatments or interventions to improve the outcomes of these vulnerable infants. KEY POINTS: · Placental inflammatory and vascular lesions are associated with NEC severity.. · Higher grade chorioamnionitis with a fetal response is associated with an increased risk of surgical NEC.. · There is a need for routine bedside utilization of placenta pathology in clinical decision-making..

Clinical and Growth Correlates of Retinopathy of Prematurity in Preterm Infants with Surgical Necrotizing Enterocolitis and Intestinal Perforation.

OBJECTIVE: This study aimed to identify the clinical and growth parameters associated with retinopathy of prematurity (ROP) in infants with necrotizing enterocolitis (NEC) and spontaneous ileal perforation (SIP). STUDY DESIGN: We conducted a retrospective cohort study that compared clinical data before and after NEC/SIP onset in neonates, categorizing by any ROP and severe ROP (type 1/2) status. RESULTS: The analysis included 109 infants with surgical NEC/SIP. Sixty infants (60/109, 55%) were diagnosed with any ROP, 32/109 (29.3%) infants (22% type 1 and 7.3% type 2) with severe ROP. On univariate analysis, those with severe ROP (32/109, 39.5%) were of lower median gestational age (GA, 23.8 weeks [23.4, 24.6] vs. 27.3 [26.3, 29.0], p < 0.001), lower median birth weight (625 g [512, 710] vs. 935 [700, 1,180], p < 0.001) and experienced higher exposure to clinical chorioamnionitis (22.6 vs. 2.13%, p < 0.006), and later median onset of ROP diagnosis (63.0 days [47.0, 77.2] vs. 29.0 [19.0, 41.0], p < 0.001), received Penrose drain placement more commonly (19 [59.4%] vs. 16 [34.0%], p = 0.04), retained less residual small bowel (70.0 cm [63.1, 90.8] vs. 90.8 [72.0, 101], p = 0.007) following surgery, were exposed to higher FiO2 7 days after birth (p = 0.001), received ventilation longer and exposed to higher FiO2 at 2 weeks (p < 0.05) following NEC and developed acute kidney injury (AKI) more often (25 [86.2%] vs. 20 [46.5%], p = 0.002) than those without ROP. Those with severe ROP had lower length, weight for length, and head circumference z scores. In an adjusted Firth's logistic regression, GA (adjusted odds ratio [aOR] = 0.51, 95% confidence interval [CI]: [0.35, 0.76]) and diagnosis at later age (aOR = 1.08, 95% CI: [1.03, 1.13]) was shown to be significantly associated with any ROP. CONCLUSION: Infants who develop severe ROP following surgical NEC/SIP are likely to be younger, smaller, have been exposed to more O2, develop AKI, and grow poorly compared with those did not develop severe ROP. KEY POINTS: · Thirty percent of infants with NEC/SIP had severe ROP.. · Those with severe ROP had poor growth parameters before and after NEC/SIP.. · Risk factors based ROP prevention strategies are needed to have improved ophthalmic outcomes..

Gestational age-specific clinical correlates of acute kidney injury in preterm infants with necrotizing enterocolitis.

BACKGROUND: To study the gestational age-specific risk factors and outcomes of severe acute kidney injury (AKI) in neonates with necrotizing enterocolitis (NEC). METHODS: Retrospective cohort study comparing gestational age (GA)-specific clinical data between infants without severe AKI (stage 0/1 AKI) and those with severe AKI (stages 2 and 3 AKI) stratified by GA ≤27 and >27 weeks. RESULTS: Infants with GA ≤27 weeks had double the rate of severe AKI (46.3% vs. 20%). In infants with GA >27 weeks, male sex, outborn, and nephrotoxic medication exposure were associated with severe AKI. On multivariable logistic regression, in infants with GA ≤27 weeks, surgical NEC (OR 35.08 (CI 5.05, 243.73), p < 0.001) and ostomy (OR 6.2(CI 1.29, 29.73), p = 0.027) were associated with significantly higher odds of severe AKI. Surgical NEC infants with GA >27 weeks and severe AKI were significantly more likely to be outborn, have later NEC onset, need dopamine, and have longer hospitalization (158 days [110; 220] vs.75.5 days [38.8; 105]; p = 0.007 than those with non-severe AKI. CONCLUSION: In neonates with NEC, surgical intervention was associated with moderate-to-severe AKI in infants with GA ≤27 weeks and with longer hospitalization in infants with GA >27 weeks. IMPACT: In both cohorts need for surgery, stoma, cholestasis, and mechanical ventilation were associated with severe AKI; however, the infants with GA <27 weeks had twice the risk of severe AKI than GA >27 weeks group. The longer exposure to nephrotoxic medication and referral need were significant risk factors for AKI in GA >27 weeks group. GA-specific kidney protective and monitoring strategies to prevent AKI and its consequences are needed to improve the clinical outcomes in neonates with NEC. Understanding the risk factors and short- and long-term outcomes unique to different GA groups will help inform those strategies.

Current Patterns of Probiotic Use in U.S. Neonatal Intensive Care Units: A Multi-Institution Survey.

OBJECTIVE: Probiotic supplementation is associated with health benefits in preterm infants. The 2021 American Academy of Pediatrics (AAP) statement on probiotic use advised caution, citing heterogeneity and absence of federal regulation. We assessed the impact of the AAP statement and current institution-wide patterns of probiotic use across neonatal intensive care units (NICU) across the United States. STUDY DESIGN: A cross-sectional web-based institutional survey using REDCap was emailed to 430 Children's Hospital Neonatal Consortium (CHNC) and Pediatrix Medical Group institutions. The survey captured data on probiotic formulations, supplementation, initiation and cessation criteria, reasons for discontinuation, interest in initiating, and AAP statement's impact. RESULTS: Ninety-five (22.1%) hospitals, including 42/46 (91%) CHNC and 53/384 (14%) Pediatrix institutions, completed the survey. Thirty-seven (39%) currently use probiotics. Fourteen different probiotic formulations were reported. The common criteria for initiation were birth weight <1,500 g and gestational age <32 weeks. Parental consent or assent was obtained at only 30% of institutions. Five hospitals (11%) with prior probiotic use discontinued solely due to the AAP statement. Overall, 23 (24%) of hospitals indicated that the AAP statement significantly influenced their decision regarding probiotic use. Nineteen of 51 nonusers (37%) are considering initiation. CONCLUSION: Probiotic use in preterm infants is likely increasing in NICUs across the United States, but significant variability exists. The 2021 AAP statement had variable impact on NICUs' decision regarding probiotic use. The growing interest in adopting probiotics and the significant interhospital variability highlight the need for better regulation and consensus guidelines to ensure standardized use. KEY POINTS: · Probiotic use in preterm infants is likely increasing in U.S. NICUs, but clinical variability exists.. · The AAP statement on probiotic use in preterm infants had a modest impact on current practices.. · There's a need for better product regulation and consensus guidelines to ensure standardized use..

Association of Placental Pathologic Findings with the Severity of Necrotizing Enterocolitis in Preterm infants - A Matched Case-Control Study.

OBJECTIVE: To determine the association of placental pathology with the severity of necrotizing enterocolitis (NEC) in preterm infants. METHODS: This single-center matched case-control study included infants with NEC (n = 107) and gestational age and birth weight-matched controls (n = 130), born between 2013 and 2020. Placentas were evaluated according to the Amsterdam Placental Workshop Group Consensus Statement. RESULTS: Acute histologic chorioamnionitis with the fetal response was significantly more common in infants with surgical NEC vs. medical NEC (35.4% vs. 15.3%; p = 0.02). On regression model, infants with multiple placental pathologies (OR 2.16; 95% CI 1.01 - 4.73; p = 0.04) and maternal vascular malperfusion (OR 2.2; 95% CI 1.12 - 4.51; p = 0.02) had higher odds of either medical or surgical NEC than controls. CONCLUSION: Infants with multiple placental lesions, including placental inflammatory and vascular lesions, were at higher risk of medical or surgical NEC in the postnatal period.

Clinical impact of NEC-associated sepsis on outcomes in preterm infants.

OBJECTIVE: To determine risk factors and outcomes of necrotizing enterocolitis (NEC)-associated sepsis in infants with NEC. METHODS: A retrospective review comparing demographic and clinical information in infants with and without NEC-associated sepsis (defined as positive blood culture at the time of NEC onset). RESULTS: A total of 209 infants with medical (n = 98) and surgical NEC (n = 111) had a median gestational age of 27 weeks (IQR 25; 30.5) and a median birth weight of 910 g [IQR 655; 1138]. Fifty of 209 (23.9%) infants had NEC-associated sepsis. Infants with NEC-associated sepsis had lower median GA (26.4 vs. 27.4 weeks; p = 0.01), lower birth weight (745 vs. 930 g; p = 0.009), were more likely mechanically ventilated [p < 0.001], received dopamine [p < 0.001], had more evidence of acute kidney injury [60% vs. 38.4%, p = 0.01], longer postoperative ileus (16 [13.0; 22.0] vs. 12 [8; 16] days; p = 0.006), higher levels of C-reactive protein, lower platelet counts, longer hospitalization compared to infants without NEC-associated sepsis. On multivariate regression, cholestasis was an independent risk factor for NEC-associated sepsis (OR 2.94; 95% CI 1.1-8.8, p = 0.038). CONCLUSION: NEC-associated sepsis was associated with greater hemodynamic support, acute kidney injury, longer postoperative ileus, and hospitalization on bivariate analysis, and cholestasis was associated with higher odds of sepsis on multi regression analysis. IMPACT: NEC-associated sepsis was present in 24% of infants with NEC. Gram-positive bacteria, Gram-negative bacteria, and Candida were found in 15.3%, 10.5%, and 2.8% of cases, respectively. Infants with NEC-associated sepsis had a greater inflammatory response (CRP levels), received more blood transfusion before NEC onset, frequently needed assisted ventilation ionotropic support, and had acute kidney injury after NEC onset. NEC infants with Gram-negative sepsis had higher portal venous gas, received more platelet transfusions before NEC onset, and had higher CRP levels and lower median lymphocyte counts at 24 h after NEC onset than those with Gram-positive sepsis.

Brain injury in preterm infants with surgical necrotizing enterocolitis: clinical and bowel pathological correlates.

BACKGROUND: The objective of this study was to determine the risk factors and outcomes of white matter brain injury (WMBI) on magnetic resonance imaging (MRI) at term-equivalent age in infants with surgical necrotizing enterocolitis (NEC). METHODS: This retrospective study compared clinical/pathological information between infants with and those without WMBI. RESULTS: Out of 69 infants with surgical NEC, 17 (24.6%) had mild WMBI, 13 (18.8%) had moderate WMBI, and six (8.7%) had severe WMBI on the brain MRI. Several clinical factors (gestational age, more red blood cell (RBC) transfusions before NEC onset, pneumoperitoneum, earlier NEC onset age, postoperative ileus, acute kidney injury (AKI) by serum creatinine, postnatal steroids, hospital stay) and histopathological findings (necrosis, hemorrhage) had univariate associations with WMBI. Associations with RBC transfusion (odds ratio (OR) 23.6 [95% confidence interval (CI): 4.73-117.97]; p = 0.0001), age at NEC onset (OR 0.30 [95%CI: 0.11-0.84]; p = 0.021), necrosis (OR 0.10 [95%CI: 0.01-0.90]; p = 0.040), and bowel hemorrhage (OR 7.79 [95%CI: 2.19-27.72]; p = 0.002) persisted in multivariable association with grade 3-4 WMBI. The infants with WMBI had lower mean motor, cognitive, language scores, and higher ophthalmic morbidity at 2 years of age. CONCLUSIONS: The WMBI was most likely associated with earlier NEC onset, higher RBC transfusions, and less necrosis and greater hemorrhage lesions on intestinal pathology in preterm infants with surgical NEC. IMPACT: In preterm infants with surgical NEC, brain MRI showed injury in the white matter in 52%, gray matter in 10%, and cerebellar region in 30%. Preterm infants with severe WMBI (grade 3-4) had less necrosis and greater hemorrhagic lesions on histopathology of the bowel. Preterm infants with WMBI were more likely to have a more severe postoperative course, AKI, and longer length of hospitalization. Neuroprotective strategies to prevent brain injury in preterm infants with surgical NEC are needed with the goal of improving the neurodevelopmental outcomes.

Clinical Correlates of Moderate-to-Severe Bronchopulmonary Dysplasia in Preterm Infants following Surgical Necrotizing Enterocolitis.

OBJECTIVE: The aim of the study is to determine clinical correlates of moderate to severe bronchopulmonary dysplasia (BPD) in preterm infants following surgical necrotizing enterocolitis (NEC). STUDY DESIGN: This is a retrospective, single-center cohort study comparing patients with moderate to severe BPD to patients with non/mild BPD among surgical NEC infants. BPD was defined by NIH 2001 consensus definition. RESULTS: Of 92 consecutive neonates with surgical NEC, 77% (71/92) had moderate/severe BPD and 22% (21/92) had non/mild BPD. The patent ductus arteriosus (PDA) was significantly higher in those developing moderate/severe BPD (67.6% [48/71]) than non/mild BPD (28.6% [6/21]; p = 0.001). Postoperatively, infants with moderate/severe BPD had more severe acute kidney injury (AKI; 67.6 [48/71] vs. 28.6% [6/21]; p = 0.001), were intubated longer (40.5 [interquartile (IQR): 12, 59] vs. 6 days [IQR: 2, 13]; p <0.001), received more parenteral nutrition (109 [IQR: 77, 147] vs. 55 days [IQR: 19, 70]; p <0.001), developed higher surgical morbidity (46.5 [33/71] vs. 14.3% [3/21]; p = 0.008), had more intestinal failure (62.5 vs. 13.3%; p <0.001), required a longer hospital stay (161 [IQR: 112, 186] vs. 64 days [IQR: 20, 91]; p <0.001), and were more likely to need home oxygen. In a multivariable analysis, lower birth weight (OR = 0.3, [95% confidence interval (CI): 0.1-0.5]; p = 0.001), PDA (OR = 10.3, [95% CI: 1.6-65.4]; p = 0.014), and longer parenteral nutritional days (OR = 8.8; [95% CI: 2.0-43.0]; p = 0.005) were significantly and independently associated with higher odds of moderate/severe versus non-/mild BPD. CONCLUSION: Development of moderate/severe BPD occurred in the majority of preterm infants with surgical NEC in this consecutive series. Preterm infants with moderate/severe BPD were more likely to have a PDA before NEC. Development of moderate/severe BPD was associated with significantly greater burden and duration of postoperative morbidity following surgical NEC. Identifying surgical NEC infants at increased risk of moderate/severe BPD and developing lung protection strategies may improve surgical NEC outcomes. KEY POINTS: · Three-fourths of preterm infants experienced severe lung injury following surgical NEC.. · The infants with severe moderate/severe BPD were most likely associated with greater duration of postoperative morbidity.. · There is need to understand and develop lung protective strategies in infants with surgical NEC..

Incomplete resection of necrotic bowel may increase mortality in infants with necrotizing enterocolitis.

BACKGROUND: Infants with advanced necrotizing enterocolitis (NEC) often need surgical resection of necrotic bowel. We hypothesized that incomplete resection of NEC lesions, signified by the detection of necrotic patches in margins of resected bowel loops, results in inferior clinical outcomes. METHODS: We reviewed the medical records of infants with surgical NEC in the past 15 years for demographic, clinical, and histopathological data. We also developed statistical models to predict mortality and hospital stay. RESULTS: Ninety infants with surgical NEC had a mean (±standard error) gestational age of 27.3 ± 0.4 weeks, birth weight 1008 ± 48 g, NEC onset at 25.2 ± 2.4 days, and resected bowel length of 29.2 ± 3.2 cm. Seventeen (18.9%) infants who had complete resection of the necrosed bowel had fewer (4; 23.5%) deaths and shorter lengths of hospital stay. In contrast, a group of 73 infants with some necrosis within the margins of resected bowel had significantly more (34; 46.6%) deaths and longer hospital stay. The combination of clinical and histopathological data gave better regression models for mortality and hospital stay. CONCLUSION: In surgical NEC, incomplete resection of necrotic bowel increased mortality and the duration of hospitalization. Regression models combining clinical and histopathological data were more accurate for mortality and the length of hospital stay. IMPACT: In infants with surgical NEC, complete resection of necrotic bowel reduced mortality and hospital stay. Regression models combining clinical and histopathological information were superior at predicting mortality and hospital stay than simpler models focusing on either of these two sets of data alone. Prediction of mortality improved with the combination of antenatal steroids, chorioamnionitis, and duration of post-operative ileus, with severity of inflammation and hemorrhages in resected intestine. Length of hospital stay was shorter in infants with higher gestational ages, but longer in those with greater depth of necrosis or needing prolonged parenteral nutrition or supervised feedings.

Severe acute kidney injury in neonates with necrotizing enterocolitis: risk factors and outcomes.

BACKGROUND: To study the risk factors and outcomes of severe acute kidney injury (AKI) in neonates with necrotizing enterocolitis. METHODS: Retrospective chart review of 202 neonates with necrotizing enterocolitis (NEC) (Bell stage >IIa) from 2013 to 2018. AKI was defined as per-modified neonatal Kidney Disease: Improving Global Outcomes criteria. Demographic, clinical, and outcome data were compared between neonates without severe AKI (stage 0 and 1 AKI) and those with severe AKI (stage 2 and 3 AKI). RESULTS: Severe AKI occurred in 66/202 (32.6%) of neonates after NEC diagnosis and after 61/104 (58.7%) of surgical NEC diagnoses. On adjusted model, surgical NEC [adjusted odds ratio (aOR) = 30.6; 95% confidence interval (CI) = 8.9, 130.6], outborn [aOR = 3.9; 95% CI = 1.54, 11.0], exposure to antenatal steroids [aOR = 3.0; 95% CI = 1.1, 8.9], and positive blood culture sepsis [aOR = 3.5; 95% CI = 1.3, 10.0] had increased odds for severe AKI. Those with severe AKI required longer hospitalization [124 days (interquartile range (IQR) 88-187) vs. 82 days (IQR 42-126), p < 0.001]. CONCLUSIONS: Severe AKI is common in neonates with NEC who require surgical intervention, are outborn, have positive blood culture sepsis, and receive antenatal steroids. Severe AKI is associated with a significantly longer length of hospitalization. IMPACT: Neonates with NEC, who are transferred from outside hospitals, require surgical NEC management, and/or have a positive blood culture at NEC onset are at the highest odds for severe (stages 2 and 3) AKI. Assessment of urine output is important for patients with NEC. Without it, 11% of those with severe AKI would have been misdiagnosed using serum creatinine alone. Kidney-protective strategies in the pre-, peri-, and postoperative period may improve the morbidity and mortality associated with severe AKI in neonates with NEC.

Necrotizing enterocolitis leads to disruption of tight junctions and increase in gut permeability in a mouse model.

BACKGROUND: Necrotizing enterocolitis (NEC) is a leading cause of death in preterm infants. Neonates weighing <1500 grams are at the highest risk for acquiring NEC, with a prevalence of nearly 7-10%, mortality up to 30%, and several long-term complications among survivors. Despite advancements in neonatal medicine, this disease remains a challenge to treat. The aim of this study is to investigate the effect of NEC on gut epithelial tight junctions and its barrier function using a NEC mouse model. METHODS: Three-day old C57BL/6 mouse pups were fed with Esbilac formula every 3 hours and then subjected to hypoxia twice daily followed by cold stress. Dam fed pups from the same litters served as controls. Pups were observed and sacrificed 96 hours after the treatments and intestines were removed for experiments. The successful induction of NEC was confirmed by histopathology. Changes in tight junction proteins in NEC intestines were studied by western blotting and immunofluorescent microscopy using specific protein markers. The gut leakage in NEC was visualized using biotin tracer molecules. RESULTS: Our study results demonstrate that we induced NEC in >50% of experimental pups, pups lost nearly 40% of weight and their intestines showed gross changes and microscopic changes associated with NEC. There were inflammatory changes with loss of tight junction barrier function and disruption of tight junction claudin proteins in the intestines of NEC mouse model. We have demonstrated for the first time that NEC intestines develop increased leakiness as visualized by biotin tracer leakage. CONCLUSIONS: NEC leads to breakdown of epithelial barrier due to changes in tight junction proteins with increased leakiness which may explain the transmigration of microbes and microbial products from the gut lumen into the blood stream leading to sepsis like signs clinically witnessed.

Transfusion-associated necrotizing enterocolitis in preterm infants: an updated meta-analysis of observational data.

BACKGROUND: The number of observational studies that report an association between packed red blood cell (PRBC) transfusions and necrotizing enterocolitis (NEC) has increased. The primary objective of this study was to evaluate the association between PRBC transfusions and NEC in observational studies. METHODS: Medline, Embase and Cochrane Library databases as well as the Pediatrics Academic Societies abstract archives were systematically searched to identify observational studies that investigated the association between PRBC transfusions and NEC. Key search terms included premature infant, blood transfusion and necrotizing enterocolitis. The generic inverse variance method with a random-effects model was used to meta-analyze selected studies. Odds ratios (ORs) and confidence intervals (CIs) were calculated. RESULTS: A meta-analysis of 17 observational studies that reported the association between PRBC transfusions and NEC was performed. The meta-analysis revealed no evidence of an association between PRBC transfusions and a higher risk of NEC (OR: 0.96; 95% CI: 0.53-1.71; P=0.88). The effect estimates that suggested an association between PRBC transfusion and NEC in matched case-control studies (OR: 1.20; 95% CI: 0.58-2.47; P=0.63) differed from those reported in cohort studies (OR: 0.51; 95% CI: 0.34-0.75; P=<0.01). CONCLUSIONS: This updated meta-analysis of predominantly low-to-moderate quality observational studies suggests that there is no significant association between PRBC transfusions and NEC. A higher quality of evidence on this topic is needed.

Necrotizing enterocolitis in a mouse model leads to widespread renal inflammation, acute kidney injury, and disruption of renal tight junction proteins.

BACKGROUND: Necrotizing enterocolitis (NEC) is a devastating condition affecting premature infants and leads to high mortality and chronic morbidity. Severe form of NEC is associated with acute renal failure, fluid imbalance, hyponatremia, and acidosis. We investigated the effect of NEC on tight junction (TJ) proteins in kidneys using a NEC mouse model to investigate the basis for the observed renal dysfunction. METHODS: NEC was induced in C57BL/6 mice by formula feeding and subjecting them to periods of hypoxia and cold stress. NEC was confirmed by gross and histological examination. We studied various markers of inflammation in kidneys and investigated changes in expression of several TJ proteins and AQP2 using immunofluorecent staining and western blotting. RESULTS: We found markedly increased expression of NFκB, TGFß, and ERK1/2 along with claudin-1, -2, -3, -4, -8, and AQP-2 in NEC kidneys. The membrane localization of claudin-2 was altered in the NEC kidneys and its immunostaining signal at TJ was disrupted. CONCLUSION: NEC led to a severe inflammatory response not only in the gut but also in the kidneys. NEC increased expression of several TJ proteins and caused disruption of claudin-2 in renal tubules. These observed changes can help explain some of the clinical findings observed in NEC.

The VPS33B-binding protein VPS16B is required in megakaryocyte and platelet α-granule biogenesis.

Patients with platelet α or dense δ-granule defects have bleeding problems. Although several proteins are known to be required for δ-granule development, less is known about α-granule biogenesis. Our previous work showed that the BEACH protein NBEAL2 and the Sec1/Munc18 protein VPS33B are required for α-granule biogenesis. Using a yeast two-hybrid screen, mass spectrometry, coimmunoprecipitation, and bioinformatics studies, we identified VPS16B as a VPS33B-binding protein. Immunoblotting confirmed VPS16B expression in various human tissues and cells including megakaryocytes and platelets, and also in megakaryocytic Dami cells. Characterization of platelets from a patient with arthrogryposis, renal dysfunction, and cholestasis (ARC) syndrome containing mutations in C14orf133 encoding VPS16B revealed pale-appearing platelets in blood films and electron microscopy revealed a complete absence of α-granules, whereas δ-granules were observed. Soluble and membrane-bound α-granule proteins were reduced or undetectable, suggesting that both releasable and membrane-bound α-granule constituents were absent. Immunofluorescence microscopy of Dami cells stably expressing GFP-VPS16B revealed that similar to VPS33B, GFP-VPS16B colocalized with markers of the trans-Golgi network, late endosomes and α-granules. We conclude that VPS16B, similar to its binding partner VPS33B, is essential for megakaryocyte and platelet α-granule biogenesis.

Is Necrotizing Enterocolitis and Spontaneous Intestinal Perforation Part of Same Disease Spectrum - New Insights?

The diagnosis of NEC is based on the presence of pneumatosis, dilated bowel loops, portal venous gas, or pneumoperitoneum on the abdominal x-ray. Published studies suggest that the appearance of pneumatosis most likely depends on the gestational age, with a shift occurring between 27-28 weeks. For infants of gestational age under 27 weeks, pneumoperitoneum is the most likely presentation of bowel injury due to the thin bowel wall and the colonization of the gut with the non-gas-producing bacteria. Assessment of postoperative morbidity and white matter injury on the brain MRI at term equivalent age in a cohort of preterm infants failed to identify differences between SIP and NEC groups when confirmed by histology. These findings illustrate the difficulty in conclusively identifying cases as SIP or NEC, particularly when gestational age is considered and raise speculation that both conditions lie on the same spectrum of intestinal injury.