Rebecca M. Rentea

Intestinal alkaline phosphatase administration in newborns is protective of gut barrier function in a neonatal necrotizing enterocolitis rat model

BACKGROUND Previously, we have shown that supplementation of intestinal alkaline phosphatase (IAP) decreased severity of necrotizing enterocolitis (NEC)-associated intestinal injury. We hypothesized that IAP administration is protective of intestinal epithelial barrier function in a dose-dependent manner. METHODS Control rat pups were vaginally delivered and breast-fed. Premature rats were divided into 4 groups: formula fed with lipopolysaccharide and hypoxia (NEC) or additional daily bovine IAP 40, 4, or 0.4 U/kg (NEC + IAP 40 U, IAP 4 U, or IAP 0.4 U). RESULTS Necrotizing enterocolitis is associated with decreased IAP protein expression and activity. Supplemental IAP increases IAP activity in intestinal homogenates and decreased NEC injury score in a dose-dependent manner. Intestinal injury as measured by fluorescein isothiocyanate-dextran flux from ileal loops showed increased permeability vs control, but supplemental IAP reversed this. Tight junction proteins claudin-1, claudin-3, occludin, and zonula occludin 1 were elevated in the NEC and IAP-treated groups with differences in expression patterns. No differences in messenger RNA levels were observed on postinjury day 3. Intestinal alkaline phosphatase administration decreases intestinal NEC injury in a dose-dependent manner. CONCLUSION Early enteral supplemental IAP may reduce NEC-related injury and may be useful for preserving the intestinal epithelial barrier function.

Intestinal alkaline phosphatase prevents the systemic inflammatory response associated with necrotizing enterocolitis

BACKGROUND Necrotizing enterocolitis (NEC) is the most common surgical emergency in neonates, with an incidence of 0.5-2.4 cases per 1000 live births and a mortality rate between 10% and 50%. Neonates affected by NEC develop a septic injury that is associated with increased risk of neurological impairment due to intraventricular bleeding and chronic lung disease. Intestinal alkaline phosphatase (IAP) is an endogenous protein that has been shown to inactivate the endotoxin lipopolysaccharide (LPS), and has recently been used successfully as an adjunct to treat sepsis in adult patients. We tested the hypothesis that systemic, exogenous IAP will mitigate the inflammatory response as measured by serum levels of proinflammatory cytokines in a rat model of NEC. METHODS Newborn Sprague-Dawley rats were divided into groups. Control pups were dam fed. NEC was induced by feeding formula containing LPS and exposure to intermittent hypoxia. NEC pups were given intraperitoneal injections of 4 or 40 glycine units (U) of IAP or placebo twice daily. Intestine and serum was collected for cytokine analysis as well as measurement of alkaline phosphatase activity. RESULTS Systemic IAP administration significantly increased serum alkaline phosphatase activity in a dose- and time-dependent fashion. The proinflammatory cytokines tumor necrosis factor α, interleukin 6, and interleukin 1β were significantly increased in NEC rats versus controls on days 2 and 3. Importantly, treatment with 40 U systemic IAP decreased these proinflammatory cytokines back to near-control levels. CONCLUSIONS Systemic IAP administration appears effective in mitigating the systemic inflammatory response associated with NEC, and may prove to be a valuable adjunctive treatment for NEC.

Pediatric appendicitis: state of the art review

Appendicitis is a common cause of abdominal pain in children. The diagnosis and treatment of the disease have undergone major changes in the past two decades, primarily as a result of the application of an evidence-based approach. Data from several randomized controlled trials, large database studies, and meta-analyses have fundamentally affected patient care. The best diagnostic approach is a standardized clinical pathway with a scoring system and selective imaging. Non-operative management of simple appendicitis is a reasonable option in selected cases, with the caveat that data in children remain limited. A minimally invasive (laparoscopic) appendectomy is the current standard in US and European children’s hospitals. This article reviews the current ‘state of the art’ in the evaluation and management of pediatric appendicitis.

Intestinal alkaline phosphatase administration in newborns decreases systemic inflammatory cytokine expression in a neonatal necrotizing enterocolitis rat model

BACKGROUND Supplementation of intestinal alkaline phosphatase (IAP), an endogenous protein expressed in the intestines, decreases the severity of necrotizing enterocolitis (NEC)-associated intestinal injury and permeability. We hypothesized that IAP administration is protective in a dose-dependent manner of the inflammatory response in a neonatal rat model. MATERIALS AND METHODS Pre- and full-term newborn Sprague-Dawley rat pups were sacrificed on day of life 3. Control pups were vaginally delivered and dam fed. Preterm pups were delivered via cesarean section and exposed to intermittent hypoxia and formula feeds containing lipopolysaccharide (NEC) with and without IAP. Three different standardized doses were administered to a group of pups treated with 40, 4, and 0.4U/kg of bovine IAP (NEC+IAP40, IAP4, or IAP0.4U). Reverse transcription-real-time polymerase chain reaction (RT-PCR) for inducible nitric oxide synthase (iNOS) and tumor necrosis factor (TNF)-α on liver and lung tissues and serum cytokine analysis for interleukin (IL)-1β, IL-6, IL-10, and TNF-α were performed. Data were analyzed by Kruskal-Wallis and Mann-Whitney tests, expressed as mean±standard error of the mean and P≤0.05 considered significant. RESULTS Levels of cytokines IL-1β, IL-6, and TNF-α increased significantly in NEC versus control, returning to control levels with increasing doses of supplemental enteral IAP. Hepatic and pulmonary TNF-α and iNOS messenger ribonucleic acid expressions increased in NEC, and the remaining elevated despite IAP supplementation. CONCLUSIONS Proinflammatory cytokine expression is increased systemically with intestinal NEC injury. Administration of IAP significantly reduces systemic proinflammatory cytokine expression in a dose-dependent manner. Early supplemental enteral IAP may reduce NEC-related injury and be useful for reducing effects caused by a proinflammatory cascade.

Enteral intestinal alkaline phosphatase administration in newborns decreases iNOS expression in a neonatal necrotizing enterocolitis rat model.

PURPOSE To determine if intestinal alkaline phosphatase (IAP) decreases intestinal injury resulting from experimentally induced necrotizing enterocolitis (NEC). We hypothesized that IAP administration prevents the initial development of NEC related intestinal inflammation. METHODS Pre- and full-term newborn Sprague-Dawley rat pups were sacrificed on day 1 of life. Pre-term pups were exposed to intermittent hypoxia and formula containing LPS to induce NEC. Select NEC pups were given 40, 4 or 0.4 units/kg of bovine IAP (NEC+IAP40u, IAP4u or IAP0.4u) enterally, once daily. Ileal sections were evaluated by real-time PCR (qRT-PCR) for IAP, iNOS, IL-1β, IL-6, and TNF-α mRNA and immunofluorescence for 3-nitrotyrosine (3-NT). RESULTS Experimentally induced NEC decreased IAP mRNA expression by 66% (p ≤ 0.001). IAP supplementation increased IAP mRNA expression to control. Supplemental enteral IAP decreased nitrosative stress as measured by iNOS mRNA expression and 3-NT staining in the NEC stressed pups (p ≤ 0.01), as well as decreased intestinal TNF-α mRNA expression. In addition, IAP decreased LSP translocation into the serum in the treated pups. CONCLUSIONS We conclude that enterally administered IAP prevents NEC-related intestinal injury and inflammation. Enteral IAP may prove a useful strategy in the prevention of NEC in preterm neonates.

Early Enteral Stressors in Newborns Increase Inflammatory Cytokine Expression in a Neonatal Necrotizing Enterocolitis Rat Model

INTRODUCTION Inflammation in the premature intestine is a key factor that leads to the development of necrotizing enterocolitis (NEC). Activation of nuclear factor kappa B (NF-κB) and subsequent inflammation increases the severity of NEC. The aim of this study was to investigate the early temporal expression of inflammatory markers and activation of NF-κB in a neonatal rat model of NEC. METHODS Pre- and full-term newborn Sprague-Dawley rat pups were sacrificed at birth, 1.5, 4, 8, and 24 hours after receiving their first feed. Control pups were vaginally delivered and mother fed; NEC was induced by a combination of gavage feeding formula, hypoxia, and enteral lipopolysaccharide (LPS); and formula fed pups were fed every 4 hours with infant formula. Ileal tissue was collected for immunohistochemistry, real-time polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay. Serum was collected for cytokine content. Fold change of expression of inducible nitric oxide synthase (iNOS), interleukin (IL)-1β, IL-6, tumor necrosis factor-α (TNF-α), IL-10, NF-κB p65, and IκBα used RT-PCR. Data were analyzed by paired two-tailed t test, expressed as mean ± standard error of the mean, and p ≤ 0.05 considered significant. RESULTS No histologic injury was evident in ileal sections. At 1.5 h, iNOS expression increased twofold over control in NEC pups (2.1 vs. 1.0, p ≤ 0.05) and remained elevated at 24 h (0.7 vs. 9.4, p ≤ 0.05). IL-1β and IL-6 reached a peak at 24 h in NEC tissue compared with control. IL-10 expression rose in NEC pups after 4 h of insult and remained elevated in formula and NEC stressed pups. Coincident with an increase in p65 translocation into the nucleus and a reduction of IκBα detected in the cytoplasm, increased transcription of IκBα occurs. CONCLUSION These findings suggest that NF-κB activation initiates inflammation early in the course of NEC resulting in increased proinflammatory protein expression, underscoring the importance of the inflammatory response in this NEC model, which precedes evidence of histological injury.

Intestinal NADPH Oxidase 2 Activity Increases in a Neonatal Rat Model of Necrotizing Enterocolitis

Necrotizing enterocolitis (NEC) is a complication of prematurity. The etiology is unknown, but is related to enteral feeding, ischemia, infection, and inflammation. Reactive oxygen species production, most notably superoxide, increases in NEC. NADPH oxidase (NOX) generates superoxide, but its activity in NEC remains unknown. We hypothesize that NOX-derived superoxide production increases in NEC. Newborn Sprague-Dawley rats were divided into control, formula-fed, formula/LPS, formula/hypoxia, and NEC (formula, hypoxia, and LPS). Intestinal homogenates were analyzed for NADPH-dependent superoxide production. Changes in superoxide levels on days 0-4 were measured. Inhibitors for nitric oxide synthase (L-NAME) and NOX2 (GP91-ds-tat) were utilized. RT-PCR for eNOS, NOX1, GP91phox expression was performed. Immunofluorescence studies estimated the co-localization of p47phox and GP91phox in control and NEC animals on D1, D2, and D4. NEC pups generated more superoxide than controls on D4, while all other groups were unchanged. NADPH-dependent superoxide production was greater in NEC on days 0, 3, and 4. GP91-ds-tat decreased superoxide production in both groups, with greater inhibition in NEC. L-NAME did not alter superoxide production. Temporally, superoxide production varied minimally in controls. In NEC, superoxide generation was decreased on day 1, but increased on days 3-4. GP91phox expression was higher in NEC on days 2 and 4. NOX1 and eNOS expression were unchanged from controls. GP91phox and p47phox had minimal co-localization in all control samples and NEC samples on D1 and D2, but had increased co-localization on D4. In conclusion, this study proves that experimentally-induced NEC increases small intestinal NOX activity. All components of NEC model are necessary for increased NOX activity. NOX2 is the major source, especially as the disease progresses.

Contemporary Management of Appendicitis in Children.

Appendicitis presents less classically than commonly thought. Use of algorithms, imaging, and laboratory studies in combination should guide the workup for diagnosis. Standardized definitions and quality improvement initiatives are important in dealing with children with appendicitis to improve outcomes. Perforated appendicitis should be managed laparoscopic if possible. Appendicitis can be managed nonoperatively in selected children, but the risk of recurrent appendicitis is unknown.

The effectiveness of costal cartilage excision in children for slipping rib syndrome

PURPOSE Slipping rib syndrome (SRS) is an elusive diagnosis. Previous reports have been single cases or small series. We previously reported a small multicenter review with encouraging early results. We now describe our matured experience. METHODS This is a follow-up study of patients with SRS from 2006 to 2015. Included are 5 previously analyzed patients and 25 new patients. Patients were called to review current symptoms, course, and satisfaction. RESULTS From 2006 to 2015, 30 patients underwent 38 operations. Eight underwent re-operation. All had reproducible pain localized to the costal margin, 60% had a popping sensation, and 23% were bilateral. 86% were female. Median age of symptom onset was 14 (IQR 13.75-15) years, while median age at diagnosis was 16 (IQR 15-17). Contact was possible with 18/30 patients, and mean follow up time was 1.3years. 72% of those felt they were cured, and 44% rated satisfaction a 10/10 (mean 7.84). Of those not cured, all reported significant improvement. CONCLUSIONS Costal cartilage excision is an effective treatment for SRS and should be considered early in the workup of costal margin pain in a normally active population. Case Series with no Comparison Group - Level IV.

Utility of pediatric female fertility preservation discussions following pelvic radiation

IntroductionWhile many childhood cancers are curable with therapy, adverse consequences in fertility exist. We sought to assess the number of female patients with pelvic tumors receiving radiation therapy, and the proportion that undergo measures for fertility preservation (FP).MethodsA total of 53 female patients treated with pelvic tumors from 2000 to 2016 were retrospectively identified.Results19 (34%) of these patients underwent pelvic radiation therapy (pXRT). Three of the patients received pXRT for palliative treatment. Of the 19 female patients receiving pXRT, six (31%) were prepubertal and 13 (68%) were postpubertal. Three patients (16%) had documentation of a discussion of FP measures prior to pXRT. One was prepubertal and the others were post-pubertal. Six patients (32%) were evaluated by endocrinology after radiation therapy, diagnosed with ovarian failure, and placed on hormone therapy. Current guidelines recommend discussion of FP in pre-and postpubertal patients with cancer. This 16-year retrospective review of female patients that underwent pXRT for pelvic tumors demonstrated < 17% of patients have documentation of a discussion of FP measures.ConclusionFemale pediatric patients who underwent chemotherapy and pXRT suffer a high rate of premature ovarian failure, high morbidity and mortality as well as low rates of documented FP discussions. Based on these findings we have established a multi-disciplinary fertility preservation team available for consultation and a protocol for discussing and documenting the impact of pXRT, along with other treatments, on fertility.Level of evidenceIII.