Protocolized approach to the management of congenital diaphragmatic hernia: benefits of reducing variability in care.

PURPOSE: Variable approaches to the care of infants with congenital diaphragmatic hernia (CDH) by multiple providers may contribute to inconsistent care. Our institution developed a comprehensive evidence-based protocol to standardize the management of CDH infants. This report reviews patient outcomes before and after the implementation of the protocol. METHODS: Retrospective chart review of CDH infants managed with individualized care (preprotocol group, January 1997-December 2001, n = 22) or on the protocol (Protocol group, January 2002-July 2009, n = 47). Survival and other categorical variables were compared by chi(2) analysis, and continuous variables were compared using 1-sided analysis of variance analysis, with significance defined as P < .05. RESULTS: Survival to discharge was significantly greater in the Protocol group (40/47; 85%) than the preprotocol group (12/22; 52%; P = .006), although mean gestational age, mean birth weight, and expected survival were not statistically different between the 2 groups. The use of supportive therapies, including high-frequency jet ventilation, inhaled nitric oxide, and extracorporeal life support, was similar between groups as well. CONCLUSIONS: Since the implementation of a management protocol for infants with CDH, survival has improved significantly compared with expected survival and preprotocol controls. Reduction in the variability of care through use of an evidence-based protocol may improve the survival of CDH infants.

Neuroblastoma survival and death: an in vitro model of hypoxia and metabolic stress.

Heterogeneous oxygen tension and access to metabolites in solid tumors may produce variability in response to adjuvant therapy. To better understand these microenvironmental features, we examined survival and proliferation of neuroblastoma (NB) cells in an in vitro model of hypoxia and metabolite deprivation. Human NB cells (SH-SY5Y) were subjected to a "self-generated" diffusion gradient of nutrient and oxygen deprivation in a modified in vitro "sandwich model." In this model, the extent of both hypoxia and metabolite deprivation were individually altered, and the effects of each were studied. Cellular proliferation was confirmed by proliferating cell nuclear antigen (PCNA) immunocytochemistry and morphology and hypoxia by vascular endothelial growth factor (VEGF) and pimonidazole immunocytochemistry. We examined apoptotic cell death using TUNEL analysis, assaying for plasma membrane transfer of phosphotidylserine and the presence of the anti-apoptotic protein Bcl-2 using immunocytochemistry. As predicted, cellular survival diminished with increasing duration and severity of hypoxia and metabolite deprivation; oxygen deprivation was determined to be the more important contributory factor to early survival and proliferation. PCNA immunocytochemistry confirmed decreasing fractions of proliferating cells as a function of distance from oxygen and metabolites. VEGF and Bcl-2 immunoreactivity increased with prolonged exposure and increased extent of oxygen/metabolite deprivation. TUNEL analysis and phosphotidylserine transfer demonstrated cellular death of hypoxic and metabolite-deprived NB cells in a manner consistent with a mitochondrial apoptotic pathway. This in vitro model demonstrates that increasing the severity of hypoxia and metabolite deprivation results in diminished proliferation and greater apoptotic death, observations analogous to that of clinical NB tumors.

Induced differentiation affords neuroblastoma cells protection from hypoxic injury.

BACKGROUND/PURPOSE: Neuroblastoma (NB) tumors uniquely exhibit heterogeneous cellular differentiation. Advanced histologic differentiation portends favorable clinical outcome, and differentiation therapy with retinoic acid has shown promise clinically. Although cellular hypoxia is a common feature of solid tumors, how this influences differentiation in NB is not known. The authors' hypothesis contends that induced differentiation may alter the susceptibility of NB cells to hypoxic injury. METHODS: Human NB cells (SH-SY5Y) were subjected to a self-generated diffusion gradient of oxygen and nutrient deprivation in an in vitro "sandwich" model as previously established. Differentiation was induced by the addition of either all-trans-retinoic acid (RA; 5 micromol/L) or the phorbol ester 12-0-tetradeconoylphorbol-13-acetate (TPA; 16 nmol/L). Cellular survival is quantified as the viable cell border width, the viability of which is confirmed by substrate adherence, MTT assay, and trypan blue exclusion. Differentiation and proliferation were evaluated by tyrosine hydroxylase and proliferating cell nuclear antigen (PCNA) immunocytochemistry, respectively. RESULTS: Addition of RA or TPA improved NB cell survival under conditions of hypoxia/metabolite deprivation (8.4 +/- 0.3 mm v 4.8 +/- 0.2 mm; RA v control at 24 hours, P <.05). This improvement was evident at all time-points examined (2 hours to 4 days) for both NB cell lines tested. Proliferation was diminished with addition of either TPA or RA. Treated cells displayed advanced differentiation by morphologic and immunocytochemical criteria. Nontreated NB cells did not differentiate under hypoxic conditions alone. Neither RA nor TPA treatment affected survival rate in non-NB cell lines (ie, hepatocyte HepG2 cells). CONCLUSIONS: Pharmacologic differentiation protects NB cells from hypoxia and metabolite deprivation in vitro. Hypoxic conditions do not induce differentiation. Although clinical differentiation therapy may well limit cell proliferation, it also may afford resistance against hypoxia-induced injury and death, antiangiogenesis therapy notwithstanding.

Expression of trkB in human neuroblastoma in relation to MYCN expression and retinoic acid treatment.

Expression of full-length trkB can be found in some highly malignant neuroblastoma tumors with an amplified MYCN gene. This contrasts sympathetic neuroblasts, from which neuroblastomas are thought to arise, which neither express trkB nor are dependent on the p145(trkB) ligands, brain-derived neurotrophic factor (BDNF) or neurotrophin-4/5, for their normal development. In this study we show that trkB was expressed in two out of five neuroblastoma tumors with amplified MYCN, while no trkB expression was observed when the MYCN gene was overexpressed in a non-MYCN-amplified neuroblastoma cell line. This shows that MYCN overexpression per se is not sufficient to induce trkB expression. trkB expression and BDNF responsiveness in neuroblastoma cells can be induced by all-trans-retinoic acid (RA). When SH-SY5Y cells were stimulated with a combination of RA and BDNF, norepinephrine and tyrosine hydroxylase levels were unaltered, showing that the cells did not change toward a more catecholaminergic sympathetic phenotype. However, expression of growth-associated protein 43, indicative of a neuronal phenotype, was elevated. Vesicular acetylcholine transporter, choline acetyl transferase, and neuropeptide tyrosine mRNA levels also increased in RA-BDNF-treated cells, which could suggest that these cells develop into a sympathetic cholinergic phenotype. In addition, treatment with RA-induced expression of the platelet-derived growth factor receptor-alpha. As previously shown for BDNF, platelet-derived growth factor stimulated growth of the RA-treated cells, findings that could have clinical relevance. If these receptors mediate a mitogenic signal in vivo also, this might limit the effect of RA treatment on neuroblastoma patients.

Fatal complication from a balloon-expandable tracheal stent in a child: a case report.

OBJECTIVE: The use of airway stents in the pediatric population is uncommon, reflected in the few patient series reported in the literature. We describe a fatal complication of tracheal stent placement in an 18-month-old child with spondylothoracic dysplasia. DESIGN: Case report. SETTING: Intensive care unit of a tertiary academic pediatric center. PATIENT: An 18-month-old child with spondylothoracic dysplasia who underwent tracheal stent placement for tracheomalacia. INTERVENTIONS: Management of an acute upper-airway hemorrhage. MAIN RESULTS: The patient died, despite aggressive interventions. CONCLUSION: Use of tracheal stents in pediatric patients with tracheomalacia is not without risks; tracheal erosion with severe hemoptysis is an infrequent but devastating complication of this intervention.

Colonic perforation complicating posteroanterior fusion using fibular strut autograft for a high-grade spondylolisthesis.

Posteroanterior fusion using a fibular strut autograft has been advocated for the surgical treatment of high-grade lumbosacral spondylolisthesis. We report here the treatment of a 14-year-old girl using an S2-L5 fibular autograft, which resulted in the postoperative complication of a sigmoid colon perforation. Techniques for recognizing, treating, and avoiding this complication are presented.