Karl G. Sylvester

Doxycycline sclerotherapy as primary treatment of head and neck lymphatic malformations in children

PURPOSE The authors report their experience with doxycycline sclerotherapy as primary treatment of head and neck lymphatic malformations (LMs) in children. METHODS A retrospective chart review was used to collect data on 11 patients treated with doxycycline sclerotherapy for LMs of the head and neck at our institution since 2003. Radiographic imaging allowed classification of patient LM as macrocystic, microcystic, or mixed according to previously published guidelines. Only patients with macrocystic or mixed lesions were offered doxycycline sclerotherapy. Radiographic imaging and physical examination were used to determine efficacy of treatment. After each treatment, the clinical and radiographic response was characterized as excellent (> or = 95% decrease in lesion size), satisfactory (> or = 50% decrease in volume and asymptomatic), or poor (< 50% decrease in volume or symptomatic). RESULTS Eleven patients underwent a total of 23 sclerotherapies with an average of 2 treatments per patient (range, 1-4). All 7 patients with macrocystic lesions achieved complete clinical resolution with an average radiographic resolution of 93%. The 4 patients with mixed lesions achieved only partial clinical resolution and an average of 73% radiographic resolution. No patient experienced any adverse effects related to the treatment. At a median follow-up of 8 months, 2 patients (18%) experienced lesion recurrence in the setting of concomitant infection. CONCLUSION Doxycycline sclerotherapy is safe and effective as a primary treatment modality for macrocystic and mixed LMs of the head and neck in the pediatric population.

SIRT1 and c-Myc promote liver tumor cell survival and predict poor survival of human hepatocellular carcinomas.

The increased expression of SIRT1 has recently been identified in numerous human tumors and a possible correlation with c-Myc oncogene has been proposed. However, it remains unclear whether SIRT1 functions as an oncogene or tumor suppressor. We sought to elucidate the role of SIRT1 in liver cancer under the influence of c-Myc and to determine the prognostic significance of SIRT1 and c-Myc expression in human hepatocellular carcinoma. The effect of either over-expression or knock down of SIRT1 on cell proliferation and survival was evaluated in both mouse and human liver cancer cells. Nicotinamide, an inhibitor of SIRT1, was also evaluated for its effects on liver tumorigenesis. The prognostic significance of the immunohistochemical detection of SIRT1 and c-Myc was evaluated in 154 hepatocellular carcinoma patients. SIRT1 and c-Myc regulate each other via a positive feedback loop and act synergistically to promote hepatocellular proliferation in both mice and human liver tumor cells. Tumor growth was significantly inhibited by nicotinamide in vivo and in vitro. In human hepatocellular carcinoma, SIRT1 expression positively correlated with c-Myc, Ki67 and p53 expression, as well as high á-fetoprotein level. Moreover, the expression of SIRT1, c-Myc and p53 were independent prognostic indicators of hepatocellular carcinoma. In conclusion, this study demonstrates that SIRT1 expression supports liver tumorigenesis and is closely correlated with oncogenic c-MYC expression. In addition, both SIRT1 and c-Myc may be useful prognostic indicators of hepatocellular carcinoma and SIRT1 targeted therapy may be beneficial in the treatment of hepatocellular carcinoma.

Wnt/β-Catenin Signaling Protects Mouse Liver against Oxidative Stress-induced Apoptosis through the Inhibition of Forkhead Transcription Factor FoxO3

Background: Wnt/β-catenin signaling regulates various hepatocellular processes; however, it remains unexplored whether β-catenin provides hepatocyte protection against oxidative stress-induced apoptosis. Results: Mice with β-catenin-deficient hepatocytes demonstrate significantly increased hepatotoxin-induced liver injury. Conclusion: Hepatic β-catenin signaling confers hepatocyte protection against oxidative stress-induced apoptosis. Significance: Our findings have relevance for potential future therapies directed at hepatocyte protection, regeneration, and anti-cancer treatment. Numerous liver diseases are associated with extensive oxidative tissue damage. It is well established that Wnt/β-catenin signaling directs multiple hepatocellular processes, including development, proliferation, regeneration, nutrient homeostasis, and carcinogenesis. It remains unexplored whether Wnt/β-catenin signaling provides hepatocyte protection against hepatotoxin-induced apoptosis. Conditional, liver-specific β-catenin knockdown (KD) mice and their wild-type littermates were challenged by feeding with a hepatotoxin 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet to induce chronic oxidative liver injury. Following the DDC diet, mice with β-catenin-deficient hepatocytes demonstrate increased liver injury, indicating an important role of β-catenin signaling for liver protection against oxidative stress. This finding was further confirmed in AML12 hepatocytes with β-catenin signaling manipulation in vitro using paraquat, a known oxidative stress inducer. Immunofluorescence staining revealed an intense nuclear FoxO3 staining in β-catenin-deficient livers, suggesting active FoxO3 signaling in response to DDC-induced liver injury when compared with wild-type controls. Consistently, FoxO3 target genes p27 and Bim were significantly induced in β-catenin KD livers. Conversely, SGK1, a β-catenin target gene, was significantly impaired in β-catenin KD hepatocytes that failed to inactivate FoxO3. Furthermore, shRNA-mediated deletion of FoxO3 increased hepatocyte resistance to oxidative stress-induced apoptosis, confirming a proapoptotic role of FoxO3 in the stressed liver. Our findings suggest that Wnt/β-catenin signaling is required for hepatocyte protection against oxidative stress-induced apoptosis. The inhibition of FoxO through its phosphorylation by β-catenin-induced SGK1 expression reduces the apoptotic function of FoxO3, resulting in increased hepatocyte survival. These findings have relevance for future therapies directed at hepatocyte protection, regeneration, and anti-cancer treatment.

Wnt–β-catenin Signaling Protects against Hepatic Ischemia and Reperfusion Injury in Mice

BACKGROUND & AIMS Ischemia and reperfusion injury are common causes of oxidative tissue damage associated with many liver diseases and hepatic surgery. The Wnt-β-catenin signaling pathway is an important regulator of hepatic development, regeneration, and carcinogenesis. However, the role of Wnt signaling in the hepatocellular response to ischemia-reperfusion (I/R) injury has not been determined. METHODS Hepatic injury following ischemia or I/R was investigated in hepatocyte-specific, β-catenin-deficient mice, as well as Wnt1-overexpressing and wild-type (control) mice. RESULTS Wnt-β-catenin signaling was affected by the cellular redox balance in hepatocytes. Following ischemia or I/R, mice with β-catenin-deficient hepatocytes were significantly more susceptible to liver injury. Conversely, mice that overexpressed Wnt1 in hepatocytes were resistant to hepatic I/R injury. Hypoxia inducible factor (HIF)-1α signaling was reduced in β-catenin-deficient liver but increased in hepatocytes that overexpressed Wnt1 under hypoxia and following I/R, indicating an interaction between β-catenin and HIF-1α signaling in the liver. The mechanism by which Wnt signaling protects against liver injury involves the role of β-catenin as a transcriptional coactivator of HIF-1α signaling, which promotes hepatocyte survival under hypoxic conditions. CONCLUSIONS Cellular redox balance affects Wnt-β-catenin signaling, which protects against hypoxia and I/R injury. These findings might be used to develop strategies for protection of hepatocytes, regeneration of liver, and inhibition of carcinogenesis.

Urine Protein Biomarkers for the Diagnosis and Prognosis of Necrotizing Enterocolitis in Infants

OBJECTIVES To test the hypothesis that an exploratory proteomics analysis of urine proteins with subsequent development of validated urine biomarker panels would produce molecular classifiers for both the diagnosis and prognosis of infants with necrotizing enterocolitis (NEC). STUDY DESIGN Urine samples were collected from 119 premature infants (85 NEC, 17 sepsis, 17 control) at the time of initial clinical concern for disease. The urine from 59 infants was used for candidate biomarker discovery by liquid chromatography/mass spectrometry. The remaining 60 samples were subject to enzyme-linked immunosorbent assay for quantitative biomarker validation. RESULTS A panel of 7 biomarkers (alpha-2-macroglobulin-like protein 1, cluster of differentiation protein 14, cystatin 3, fibrinogen alpha chain, pigment epithelium-derived factor, retinol binding protein 4, and vasolin) was identified by liquid chromatography/mass spectrometry and subsequently validated by enzyme-linked immunosorbent assay. These proteins were consistently found to be either up- or down-regulated depending on the presence, absence, or severity of disease. Biomarker panel validation resulted in a receiver-operator characteristic area under the curve of 98.2% for NEC vs sepsis and an area under the curve of 98.4% for medical NEC vs surgical NEC. CONCLUSIONS We identified 7 urine proteins capable of providing highly accurate diagnostic and prognostic information for infants with suspected NEC. This work represents a novel approach to improving the efficiency with which we diagnose early NEC and identify those at risk for developing severe, or surgical, disease.

β-Catenin regulates hepatic mitochondrial function and energy balance in mice.

BACKGROUND & AIMS Wnt signaling regulates hepatic function and nutrient homeostasis. However, little is known about the roles of β-catenin in cellular respiration or mitochondria of hepatocytes. METHODS We investigated β-catenins role in the metabolic function of hepatocytes under homeostatic conditions and in response to metabolic stress using mice with hepatocyte-specific deletion of β-catenin and their wild-type littermates, given either saline (sham) or ethanol (as a model of binge drinking and acute ethanol intoxication). RESULTS Under homeostatic conditions, β-catenin-deficient hepatocytes demonstrated mitochondrial dysfunctions that included impairments to the tricarboxylic acid cycle and oxidative phosphorylation (OXPHOS) and decreased production of adenosine triphosphate (ATP). There was no evidence for redox imbalance or oxidative cellular injury in the absence of metabolic stress. In mice with β-catenin-deficient hepatocytes, ethanol intoxication led to significant redox imbalance in the hepatocytes and further deterioration in mitochondrial function that included reduced OXPHOS, fatty acid oxidation (FAO), and ATP production. Ethanol feeding significantly increased liver steatosis and oxidative damage, compared with wild-type mice, and disrupted the ratio of nicotinamide adenine dinucleotide. β-catenin-deficient hepatocytes also had showed disrupted signaling of Sirt1/peroxisome proliferator-activated receptor-α signaling. CONCLUSIONS β-catenin has an important role in the maintenance of mitochondrial homeostasis, regulating ATP production via the tricarboxylic acid cycle, OXPHOS, and fatty acid oxidation; β-catenin function in these systems is compromised under conditions of nutrient oxidative stress. Reagents that alter Wnt-β-catenin signaling might be developed as a useful new therapeutic strategy for treatment of liver disease.

Risk Factors for Parenteral Nutrition–associated Liver Disease Following Surgical Therapy for Necrotizing Enterocolitis

Objective:The aim of the study was to prospectively determine risk factors for the development of parenteral nutrition–associated liver disease (PNALD) in infants who underwent surgery for necrotizing enterocolitis (NEC), the most common cause of intestinal failure in children. Patients and Methods:From February 2004 to February 2007, we diagnosed 464 infants with NEC, of whom 180 had surgery. One hundred twenty-seven patients were available for full analysis. PNALD was defined as serum direct bilirubin ≥2 mg/dL or ALT ≥2 × the upper limit of normal in the absence of sepsis after ≥14 days of exposure to PN. Results:Median gestational age was 26 weeks and 68% were boys. Seventy percent of the cohort developed PNALD and the incidence of PNALD varied significantly across the 6 study sites, ranging from 56% to 85% (P = 0.05). Multivariable logistic regression analysis identified small-bowel resection or creation of jejunostomy (odds ratio [OR] 4.96, 95% confidence interval [CI] 1.97–12.51, P = 0.0007) and duration of PN in weeks (OR 2.37, 95% CI 1.56–3.60, P < 0.0001) as independent risk factors for PNALD. Preoperative exposure to PN was also associated with the development of PNALD; the risk of PNALD was 2.6 (95% CI 1.5–4.7; P = 0.001) times greater in patients with ≥4 weeks of preoperative PN compared with those with less preoperative PN use. Breast milk feedings, episodes of infection, and gestational age were not related to the development of PNALD. Conclusions:The incidence of PNALD is high in infants with NEC undergoing surgical treatment. Risk factors for PNALD are related to signs of NEC severity, including the need for small-bowel resection or proximal jejunostomy, as well as longer exposure to PN. Identification of these and other risk factors can help in the design of clinical trials for the prevention and treatment of PNALD and for clinical assessment of patients with NEC and prolonged PN dependence.

The Surgical Management of Necrotizing Enterocolitis

Necrotizing enterocolitis (NEC), a common cause of neonatal morbidity and mortality, is strongly associated with prematurity and typically occurs following initiation of enteral feeds. Mild NEC is adequately treated by cessation of enteral feeding, empiric antibiotics, and supportive care. Approximately 50% of affected infants will develop progressive intestinal necrosis requiring urgent operation. Several surgical techniques have been described, but there is no clear survival benefit for any single operative approach. While debate continues regarding the optimal surgical management for infants with severe NEC, future progress will likely depend on the development of improved diagnostic tools and preventive therapies.

Risk Factors for Intestinal Failure in Infants with Necrotizing Enterocolitis: A Glaser Pediatric Research Network Study

OBJECTIVE To determine risk factors for intestinal failure (IF) in infants undergoing surgery for necrotizing enterocolitis (NEC). STUDY DESIGN Infants were enrolled in a multicenter prospective cohort study. IF was defined as the requirement for parenteral nutrition for >or= 90 days. Logistic regression was used to identify predictors of IF. RESULTS Among 473 patients enrolled, 129 had surgery and had adequate follow-up data, and of these patients, 54 (42%) developed IF. Of the 265 patients who did not require surgery, 6 (2%) developed IF (OR 31.1, 95% CI, 12.9 - 75.1, P < .001). Multivariate analysis identified the following risk factors for IF: use of parenteral antibiotics on the day of NEC diagnosis (OR = 16.61, P = .022); birth weight < 750 grams, (OR = 9.09, P < .001); requirement for mechanical ventilation on the day of NEC diagnosis (OR = 6.16, P = .009); exposure to enteral feeding before NEC diagnosis (OR=4.05, P = .048); and percentage of small bowel resected (OR = 1.85 per 10 percentage point greater resection, P = .031). CONCLUSION The incidence of IF among infants undergoing surgical treatment for NEC is high. Variables characteristic of severe NEC (low birth weight, antibiotic use, ventilator use, and greater extent of bowel resection) were associated with the development of IF.

Outcomes of sutureless gastroschisis closure

INTRODUCTION A new technique of gastroschisis closure in which the defect is covered with sterile dressings and allowed to granulate without suture repair was first described in 2004. Little is known about the outcomes of this technique. This study evaluated short-term outcomes from the largest series of sutureless gastroschisis closures. METHODS AND PATIENTS A retrospective case control study of 26 patients undergoing sutureless closure between 2006 and 2008 was compared to a historical control group of 20 patients with suture closure of the abdominal fascia between 2004 and 2006. Four major outcomes were assessed: (1) time spent on ventilator, (2) time to initiating enteral feeds, (3) time to discharge from the neonatal intensive care unit, and (4) rate of complications. RESULTS In multivariate analysis, sutureless closure of gastroschisis defects independently reduced the time to extubation as compared to traditional closure (5.0 vs 12.1 days, P = .025). There was no difference in time to full enteral feeds (16.8 vs 21.4 days, P = .15) or time to discharge (34.8 vs 49.7 days, P = .22) with sutureless closure. The need for silo reduction independently increased the time to extubation (odds ratio, 4.2; P = .002) and time to enteral feeds (odds ratio, 5.2; P < .001). Small umbilical hernias were seen in all patients. CONCLUSION Sutureless closure of uncomplicated gastroschisis is a safe technique that reduces length of intubation and does not significantly alter the time required to reach full enteral feeds or hospital discharge.