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The mammalian Fem1b gene encodes a homolog of FEM-1, a protein in the sex-determination pathway of the nematode Caenorhabditis elegans. Fem1b and FEM-1 proteins each contain a VHL-box motif that mediates their interaction with certain E3 ubiquitin ligase complexes. In C. elegans, FEM-1 negatively regulates the transcription factor TRA-1, and functions as an E3 ubiquitin ligase substrate recognition subunit to target TRA-1 for ubiquitylation. TRA-1 is homologous to the mammalian Gli1 protein, a transcription factor that mediates Hedgehog signaling as well as having Hedgehog-independent functions. Whether the interaction between nematode FEM-1 and TRA-1 proteins is conserved, between corresponding mammalian homologs, has not been reported. Herein, we show that Fem1b interacts with Gli1 within cells, and directly binds Gli1. Fem1b also promotes ubiquitylation of Gli1, suppresses transcriptional activation by Gli1, and attenuates an oncogenic Gli1 autoregulatory loop in cancer cells, all dependent on the VHL-box of Fem1b. These findings have implications for understanding the cellular functions of Fem1b, and the regulation of Gli1 oncoprotein activity., (Copyright © 2013 Elsevier Inc. All rights reserved.)

We have shown that mucosal HIV-1 recombinant pox viral vaccination can induce high, avidity HIV-specific CD8(+) T cells with reduced interleukin (IL)-4 and IL-13 expression compared to, systemic vaccine delivery. In the current study how these cytokines act to regulate anti-viral CD8(+) T, cell avidity following HIV-1 recombinant pox viral prime-boost vaccination was investigated. Out of a panel of T cell avidity markers tested, only CD8 expression levels were found to be enhanced on, KdGag197-205 (HIV)-specific CD8(+) T cells obtained from IL-13(-/-), IL-4(-/-) and signal transducer and, activator of transcription of 6 (STAT6)(-/-) mice compared to wild-type (WT) controls following, vaccination. Elevated CD8 expression levels in this instance also correlated with polyfunctionality, (interferon (IFN)-γ, tumour necorsis factor (TNF)-α and IL-2 production) and the avidity of HIVspecific CD8(+) T cells. Furthermore, mucosal vaccination and vaccination with the novel adjuvanted IL-13 inhibitor (i.e. IL-13Rα2) vaccines significantly enhanced CD8 expression levels on HIV-specific CD8(+), T cells, which correlated with avidity. Using anti-CD8 antibodies that blocked CD8 availability on CD8(+), T cells, it was established that CD8 played an important role in increasing HIV-specific CD8(+) T cell avidity and polyfunctionality in IL-4(-/-), IL-13(-/-) and STAT6(-/-) mice compared to WT controls, following vaccination. Collectively, our data demonstrate that IL-4 and IL-13 dampen CD8 expression levels on anti-viral CD8(+) T cells, which can down-regulate anti-viral CD8(+) T cell avidity and, polyfunctionality following HIV-1 recombinant pox viral vaccination. These findings can be exploited to, design more efficacious vaccines not only against HIV-1, but many chronic infections where high, avidity CD8(+) T cells help protection., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

The study had 2 major purposes. First, it showed that the same task (Brown-Peterson task) can be used to test the memory abilities of both young children and adults, given an appropriate distracter task. Second, it illustrated that children can perform as accurately as adults on working memory tasks when prompted to use memory techniques such as rehearsal. Specifically, a modified version of the Brown-Peterson task (typically used with adults), tested working memory of adults and children aged 5-6 years. The modification was reciting the pledge of allegiance as the distracter task, as the pledge is at a level where young children are just learning it, and thus they know it, but not exceptionally well. Adults would have previously overlearned it as children but may not have recited it recently. This allows for the use of the same distracter task, thus allowing for a direct comparison of children and adults. Under experimental conditions, both groups showed a typical Brown-Peterson decay curve, with children showing a steeper decay than adults. With no distracter, adults performed at ceiling level, but only when rehearsal was encouraged did the accuracy of recalling the trigrams in the Brown-Peterson task improve for children, resulting in similar performance as adults.

Internal ribosome entry sites/segments (IRESs) were first discovered over 20 years ago in picornaviruses, followed by the discovery of two other types of IRES in hepatitis C virus (HCV), and the dicistroviruses, which infect invertebrates. In the meantime, reports of IRESs in eukaryotic cellular mRNAs started to appear, and the list of such putative IRESs continues to grow to the point in which it now stands at ~100, 80% of them in vertebrate mRNAs. Despite initial skepticism from some quarters, there now seems universal agreement that there is genuine internal ribosome entry on the viral IRESs. However, the same cannot be said for cellular mRNA IRESs, which continue to be shrouded in controversy. The aim of this article is to explain why vertebrate mRNA IRESs remain controversial, and to discuss ways in which these controversies might be resolved.

With the hope of understanding how interleukin (IL)-4 and IL-13 modulated quality of anti-viral CD8(+) T cells, we evaluated the expression of receptors for these cytokines following a range of viral infections (e.g. pox viruses and influenza virus). Results clearly indicated that unlike other IL-4/IL-13 receptor subunits, IL-4 receptor α (IL-4Rα) was significantly down-regulated on anti-viral CD8(+) T cells in a cognate antigen dependent manner. The infection of gene knockout mice and wild-type (WT) mice with vaccinia virus (VV) or VV expressing IL-4 confirmed that IL-4, IL-13 and signal transducer and activator of transcription 6 (STAT6) were required to increase IL-4Rα expression on CD8(+) T cells, but not interferon (IFN)-γ. STAT6 dependent elevation of IL-4Rα expression on CD8(+) T cells was a feature of poor quality anti-viral CD8(+) T cell immunity as measured by the production of IFN-γ and tumor necrosis factor α (TNF-α) in response to VV antigen stimulation in vitro. We propose that down-regulation of IL-4Rα, but not the other IL-4/IL-13 receptor subunits, is a mechanism by which CD8(+) T cells reduce responsiveness to IL-4 and IL-13. This can improve the quality of anti-viral CD8(+) T cell immunity. Our findings have important implications in understanding anti-viral CD8(+) T cell immunity and designing effective vaccines against chronic viral infections.

Intranasal infection with vaccinia virus co-expressing interferon epsilon (VV-HIV-IFN-ε) was used to evaluate the role of IFN-ε in mucosal immunity. VV-HIV- IFN-ε infection induced a rapid VV clearance in lung that correlated with (i) an elevated lung VV-specific CD8(+)CD107a(+)IFN-γ(+) population expressing activation markers CD69/CD103, (ii) enhanced lymphocyte recruitment to lung alveoli with reduced inflammation, and (iii) an heightened functional/cytotoxic CD8(+)CD4(+) T-cell subset (CD3(hi)CCR7(hi)CD62L(lo)) in lung lymph nodes. These responses were different to that observed with intranasal VV-HA-IFN-α(4) or VV-HA-IFN-β infections. When IFN-ε was used in an intranasal/intramuscular heterologous HIV prime-boost immunization, elevated HIV-specific effector, but not memory CD8(+)T cells responses, were observed in spleen, genito-rectal nodes, and Peyer's patch. Homing marker α4β7 and CCR9 analysis indicated that unlike other type I IFNs, IFN-ε could promote migration of antigen-specific CD8(+)T cells to the gut. Our results indicate that IFN-ε has a unique role in the mucosae and most likely can be used to control local lung and/or gut infections (i.e., microbicide) such as tuberculosis, HIV-1, or sexually transmitted diseases.

PTB (polypyrimidine tract-binding protein) is an abundant and widely expressed RNA-binding protein with four RRM (RNA recognition motif) domains. PTB is involved in numerous post-transcriptional steps in gene expression in both the nucleus and cytoplasm, but has been best characterized as a regulatory repressor of some ASEs (alternative splicing events), and as an activator of translation driven by IRESs (internal ribosome entry segments). We have used a variety of approaches to characterize the activities of PTB and its molecular interactions with RNA substrates and protein partners. Using splice-sensitive microarrays we found that PTB acts not only as a splicing repressor but also as an activator, and that these two activities are determined by the location at which PTB binds relative to target exons. We have identified minimal splicing repressor and activator domains, and have determined high resolution structures of the second RRM domain of PTB binding to peptide motifs from the co-repressor protein Raver1. Using single-molecule techniques we have determined the stoichiometry of PTB binding to a regulated splicing substrate in whole nuclear extracts. Finally, we have used tethered hydroxyl radical probing to determine the locations on viral IRESs at which each of the four RRM domains bind. We are now combining tethered probing with single molecule analyses to gain a detailed understanding of how PTB interacts with pre-mRNA substrates to effect either repression or activation of splicing.

Optimal mobility, defined as relative ease and freedom of movement in all of its forms, is central to healthy aging. Mobility is a significant consideration for research, practice, and policy in aging and public health. We examined the public health burdens of mobility disability, with a particular focus on leading public health interventions to enhance walking and driving, and the challenges and opportunities for public health action. We propose an integrated mobility agenda, which draws on the lived experience of older adults. New strategies for research, practice, and policy are needed to move beyond categorical promotion programs in walking and driving to establish a comprehensive program to enhance safe mobility in all its forms.

Translation termination in eukaryotes occurs in response to a stop codon in the ribosomal A-site and requires two release factors (RFs), eRF1 and eRF3, which bind to the A-site as an eRF1/eRF3/GTP complex with eRF1 responsible for codon recognition. After GTP hydrolysis by eRF3, eRF1 triggers hydrolysis of the polypeptidyl-tRNA, releasing the completed protein product. This leaves an 80S ribosome still bound to the mRNA, with deacylated tRNA in its P-site and at least eRF1 in its A-site, which needs to be disassembled and released from the mRNA to allow further rounds of translation. The first step in recycling is dissociation of the 60S ribosomal subunit, leaving a 40S/deacylated tRNA complex bound to the mRNA. This is mediated by ABCE1, which is a somewhat unusual member of the ATP-binding cassette family of proteins with no membrane-spanning domain but two essential iron-sulfur clusters. Two distinct pathways have been identified for subsequent ejection of the deacylated tRNA followed by dissociation of the 40S subunit from the mRNA, one executed by a subset of the canonical initiation factors (which therefore starts the process of preparing the 40S subunit for the next round of translation) and the other by Ligatin or homologous proteins. However, although this is the normal sequence of events, there are exceptions where the termination reaction is followed by reinitiation on the same mRNA (usually) at a site downstream of the stop codon. The overwhelming majority of such reinitiation events occur when the 5'-proximal open reading frame (ORF) is short and can result in significant regulation of translation of the protein-coding ORF, but there are also rare examples, mainly bicistronic viral RNAs, of reinitiation after a long ORF. Here, we review our current understanding of the mechanisms of termination, ribosome recycling, and reinitiation after translation of short and long ORFs., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Translation initiation dependent on the foot-and-mouth disease virus (FMDV) internal ribosome entry site (IRES) occurs at two sites (Lab and Lb), 84 nucleotides (nt) apart. In vitro translation of an mRNA comprising the IRES and Lab-Lb intervening segment fused to a chloramphenicol acetyltransferase (CAT) reporter has been used to study the parameters influencing the ratio of the two products and the combined product yield as measures of relative initiation site usage and productive ribosome recruitment, respectively. With wild-type mRNA, ∼40% of initiation occurred at the Lab site, which was increased to 90% by optimization of its context, but decreased to 20% by mutations that reduced downstream secondary structure, with no change in recruitment in either case. Inserting 5 nt into the pyrimidine-rich tract located just upstream of the Lab site increased initiation at this site by 75% and ribosome recruitment by 50%. Mutating the Lab site to RCG or RUN codons decreased recruitment by 20 to 30% but stimulated Lb initiation by 20 to 40%. An antisense oligodeoxynucleotide annealing across the Lab site inhibited initiation at both sites. These and related results lead to the following conclusions. Recruitment by the wild-type IRES is limited by its short oligopyrimidine tract. At least 90% of internal ribosome entry occurs at the Lab AUG, but initiation at this site is restricted by its poor context, despite a counteracting effect of downstream secondary structure. Initiation at the Lb site is by ribosomes that access it by linear scanning from the original entry site, and not by an independent entry process.

Picornavirus mRNAs contain IRESs that sustain their translation during infection, when host protein synthesis is shut off. The major classes of picornavirus IRESs (Types 1 and 2) have distinct structures and sequences, but initiation on both is determined by their specific interaction with eIF4G. We report here that Aichivirus (AV), a member of the Kobuvirus genus of Picornaviridae, contains an IRES that differs structurally from Type 1 and Type 2 IRESs. Its function similarly involves interaction with eIF4G, but its eIF4G-interacting domain is structurally distinct, although it contains an apical eIF4G-interacting motif similar to that in Type 2 IRESs. Like Type 1 and Type 2 IRESs, AV IRES function is enhanced by pyrimidine tract-binding protein (PTB), but the pattern of PTB's interaction with each of these IRESs is distinct. Unlike all known IRESs, the AV IRES is absolutely dependent on DHX29, a requirement imposed by sequestration of its initiation codon in a stable hairpin.

Here, we report that the untreated rabbit reticulocyte lysate contains over 300 different endogenous microRNAs together with the major components of the RNA-induced silencing complex and thus can be used as a model in vitro system to study the effects of microRNAs on gene expression. By using this system, we were able to show that microRNA hybridization to its target resulted in a very rapid and strong inhibition of expression that was exerted exclusively at the level of translation initiation with no involvement of transcript degradation or deadenylation. Moreover, we demonstrate that the magnitude of microRNA-induced repression can only be recapitulated in the context of a competitive translating environment. By using a wide spectrum of competitor cellular and viral RNAs, we could further show that competition was not exerted at the level of general components of the translational machinery, but relied exclusively on the presence of the poly(A) tail with virtually no involvement of the cap structure.

Polypyrimidine tract binding protein (PTB) is an RNA-binding protein with four RNA-binding domains (RBDs). It is a major regulator of alternative splicing and also stimulates translation initiation at picornavirus IRESs (internal ribosome entry sites). The sites of interaction of each RBD with two picornaviral IRESs have previously been mapped. To establish which RBD-IRES interactions are essential for IRES activation, point mutations were introduced into the RNA-binding surface of each RBD. Three such mutations were sufficient to inactivate RNA-binding by any one RBD, but the sites of the other three RBD-IRES interactions remained unperturbed. Poliovirus IRES activation was abrogated by inactivation of RBD1, 2, or 4, but the RBD3-IRES interaction was superfluous. Stimulation of the encephalomyocarditis virus IRES was reduced by inactivation of RBD1, 3, or 4, and abrogated by mutation of RBD2, or both RBDs 3 and 4. Surprisingly, therefore, the binding of PTB in its normal orientation does not guarantee IRES activation; three native RBDs are sufficient for correct binding but not for activation if the missing RBD-IRES interaction is critical.

Numerous human immunodeficiency virus (HIV)-1 vaccines have been developed over the last three decades, but to date an effective HIV-1 vaccine that can be used for prophylactic or therapeutic purposes in humans has not been identified. The failures and limited successes of HIV-1 vaccines have highlighted the gaps in our knowledge with regard to fundamental immunity against HIV-1 and have provided insights for vaccine strategies that may be implemented for designing more effective HIV-1 vaccines in the future. Recent studies have shown that robust mucosal immunity, high avidity and polyfunctional T cells, and broadly neutralizing antibodies are important factors governing the induction of protective immunity against HIV-1. Furthermore, optimization of vaccine delivery methods for DNA or live viral vector-based vaccines, elucidating the immune responses of individuals who remain resistant to HIV-1 infections and also understanding the core immune responses mediating protection against simian immunodeficiency viruses (SIV) and HIV-1 in animal models following vaccination, are key aspects to be regarded for designing more effective HIV-1 vaccines in the future.

Termination-dependent reinitiation is used to co-ordinately regulate expression of the M1 and BM2 open-reading frames (ORFs) of the dicistronic influenza B segment 7 RNA. The start codon of the BM2 ORF overlaps the stop codon of the M1 ORF in the pentanucleotide UAAUG and ∼10% of ribosomes terminating at the M1 stop codon reinitiate translation at the overlapping AUG. BM2 synthesis requires the presence of, and translation through, 45 nt of RNA immediately upstream of the UAAUG, known as the 'termination upstream ribosome binding site' (TURBS). This region may tether ribosomal 40S subunits to the mRNA following termination and a short region of the TURBS, motif 1, with complementarity to helix 26 of 18S rRNA has been implicated in this process. Here, we provide further evidence for a direct interaction between mRNA and rRNA using antisense oligonucleotide targeting and functional analysis in yeast cells. The TURBS also binds initiation factor eIF3 and we show here that this protein stimulates reinitiation from both wild-type and defective TURBS when added exogenously, perhaps by stabilising ribosome-mRNA interactions. Further, we show that the position of the TURBS with respect to the UAAUG overlap is crucial, and that termination too far downstream of the 18S complementary sequence inhibits the process, probably due to reduced 40S tethering. However, in reporter mRNAs where the restart codon alone is moved downstream, termination-reinitiation is inhibited but not abolished, thus the site of reinitiation is somewhat flexible. Reinitiation on distant AUGs is not inhibited in eIF4G-depleted RRL, suggesting that the tethered 40S subunit can move some distance without a requirement for linear scanning.

Evidence for the impact of prehospital, physician-delivered advanced cardiac life support (ACLS) on survival from out-of-hospital cardiac arrest is conflicting. The prospective observational study by Yasunaga and co-workers demonstrates an improved survival at 1 month associated with prehospital physician-delivered ACLS over emergency life-saving technician-delivered ACLS. These effects are additive to the survival benefit seen with bystander-initiated cardiopulmonary resuscitation (BCPR) compared with no BCPR. The present commentary places these findings in the context of the existing literature and discusses some of the unresolved controversies., (© 2011 BioMed Central Ltd)

As childhood obesity has reached epidemic proportions, it is critical to devise interventions that target the root causes of obesity and its risk factors. The two main components of childhood obesity are physical inactivity and improper nutrition, and it is becoming increasingly evident that the built environment can determine the level of exposure to these risk factors. Through a multidisciplinary literature review, we investigated the association between various built environment attributes and childhood obesity. We found that neighborhood features such as walkability/bikeability, mixed land use, accessible destinations, and transit increase resident physical activity; also that access to high-caloric foods and convenience stores increases risk of overweight and obesity, whereas the presence of neighborhood supermarkets and farmers' markets is associated with lower childhood body mass index and overweight status. It is evident that a child's built environment impacts his access to nutritious foods and physical activity. In order for children, as well as adults, to prevent onset of overweight or obesity, they need safe places to be active and local markets that offer affordable, healthy food options. Interventions that are designed to provide safe, walkable neighborhoods with access to necessary destinations will be effective in combating the epidemic of obesity., (© 2011 Mount Sinai School of Medicine.)

Objective: To systematically review evidence relating to the clinical efficacy of oseltamivir, zanamivir and amantadine in the prevention of influenza., Methods: RCTs evaluating these interventions in seasonal prophylaxis and post-exposure prophylaxis were identified using electronic bibliographic databases and handsearching of retrieved articles., Results: Oseltamivir was effective in preventing symptomatic laboratory-confirmed influenza (SLCI) in seasonal prophylaxis in healthy adults and at-risk elderly subjects and in post-exposure prophylaxis within households of mixed composition. Post-exposure prophylaxis using oseltamivir for paediatric contacts was observed to prevent SLCI. Zanamivir prevented SLCI in seasonal prophylaxis in healthy adults, at-risk adults and adolescents and in post-exposure prophylaxis within mixed households, with a trend for seasonal and post-exposure preventative effects in elderly subjects. Evidence for amantadine prophylaxis across subgroups was very limited. However, amantadine prevented SLCI in seasonal prophylaxis in healthy adults and in outbreak control amongst adolescent subjects. Interventions were reported to be well tolerated by subjects, with a relatively low proportion of subjects experiencing drug-related adverse events and drug-related withdrawals., Conclusions: Evidence was identified for the efficacy of oseltamivir and zanamivir in preventing influenza in a range of population subgroups. The evidence base for amantadine was considerably more limited., (Crown Copyright © 2010. Published by Elsevier Ltd. All rights reserved.)

Tethered hydroxyl-radical probing has been used to determine the orientation of binding of polypyrimidine tract-binding protein (PTB) to the poliovirus type 1 (Mahoney) (PV-1(M)) internal ribosome entry site/segment (IRES)-the question of which RNA-binding domain (RBD) binds to which sites on the IRES. The results show that under conditions in which PTB strongly stimulates IRES activity, a single PTB is binding to the IRES, a finding which was confirmed by mass spectrometry of PTB/IRES complexes. RBDs1 and 2 interact with the basal part of the Domain V irregular stem loop, very close to the binding site of eIF4G, and RBDs3 and 4 interact with the single-stranded regions flanking Domain V. The binding of PTB is subtly altered in the presence of the central domain (p50) of eIF4G, and p50 binding is likewise modified if PTB is present. This suggests that PTB stimulates PV-1(M) IRES activity by inducing eIF4G to bind in the optimal position and orientation to promote internal ribosome entry, which, in PV-1(M), is at an AUG triplet 30 nt downstream of the base of Domain V.

Translation initiation site usage on the human rhinovirus 2 internal ribosome entry site (IRES) has been examined in a mixed reticulocyte lysate/HeLa cell extract system. There are two relevant AUG triplets, both in a base-paired hairpin structure (domain VI), with one on the 5' side at nucleotide (nt) 576, base paired with the other at nt 611, which is the initiation site for polyprotein synthesis. A single residue was inserted in the apical loop to put AUG-576 in frame with AUG-611, and in addition another in-frame AUG was introduced at nt 593. When most of the IRES was deleted to generate a monocistronic mRNA, the use of these AUGs conformed to the scanning ribosome model: improving the AUG-576 context increased initiation at this site and decreased initiation at downstream sites, whereas the converse was seen when AUG-576 was mutated to GUA; and AUG-593, when present, took complete precedence over AUG-611. Under IRES-dependent conditions, by contrast, much less initiation occurred at AUG-576 than in a monocistronic mRNA with the same AUG-576 context, mutation of AUG-576 decreased initiation at downstream sites by approximately 70%, and introduction of AUG-593 did not completely abrogate initiation at AUG-611, unless the apical base pairing in domain VI was destroyed by point mutations. These results indicate that ribosomes first bind at the AUG-576 site, but instead of initiating there, most of them are transferred to AUG-611, the majority by strictly linear scanning and a substantial minority by direct transfer, which is possibly facilitated by the occasional persistence of base pairing in the apical part of the domain VI stem.

Codon-anticodon interactions are central to both the initiation and elongation phases of eukaryotic mRNA translation. The obvious difference is that the interaction takes place in the ribosomal A-site during elongation, whereas the 40S ribosomal subunit and associated initiation factors scan the mRNA sequence in search of an initiation codon with Met-tRNA(i) bound in the P-site, ceasing once codon-anticodon interaction is established at the AUG. As an indirect test of whether the two mechanisms of mRNA sequence inspection are basically similar or not, the effects of six different uridine analog substitutions in the mRNA were examined in reticulocyte lysate translation assays and 80S initiation complex formation assays. Four constructs, each with the same reporter coding sequence, were used, differing in whether the initiation codon was AUG or ACG, and in whether the 5'-UTR had U residues or not. Three analogs (5-bromoU, 5-aminoallylU, and pseudoU) inhibited both elongation and initiation, but the other three had striking differential effects. Ribothymidine had a negligible effect on elongation but caused a approximately 50% inhibition of initiation, with little effect on actual AUG recognition, which implies that inhibition must have occurred at some earlier step in initiation. In complete contrast, 2' deoxyU was prohibitive to elongation but had no effect on initiation, and 4-thioU actually stimulated initiation but quite strongly inhibited elongation processivity. These results show that the detailed mechanisms of inspection of the mRNA sequence during scanning-dependent initiation and elongation must be considerably different.

The present work describes a methodology for patterning biomolecules on silicon-based analytical devices that reconciles 3-D biological functionalization with standard resist lift-off techniques. Unlike classic sol-gel approaches in which the biomolecule of interest is introduced within the sol mixture, a two-stage scenario has been developed. It consists first of patterning micrometer/submicrometer polycondensate scaffold structures, using classic microfabrication tools, that are then loaded with native biomolecules via a second simple incubation step under biologically friendly environmental conditions. The common compatibility issue between the biological and microfabrication worlds has been circumvented because native recognition biomolecules can be introduced into the host scaffold downstream from all compatibility issues. The scaffold can be generated on any silicon substrate via the polycondensation of aminosilane, namely, aminopropyltriethoxy silane (APTES), under conditions that are fully compatible with resist mask lithography. The scaffold porosity and high primary amine content allow proteins and nucleic acid sequences to penetrate the polycondensate and to interact strongly, thus giving rise to micrometer/submicrometer 3-D structures exhibiting high biological activity. The integration of such a biopatterning approach in the microfabrication process of silicon analytical devices has been demonstrated via the successful completion of immunoassays and nucleic acid assays.

Background: Timing of repair of congenital diaphragmatic hernia (CDH) in babies that require stabilization on extracorporeal membrane oxygenation (ECMO) remains controversial. Although many centers delay operation until physiologic stabilization has occurred or ECMO is no longer needed, we repair soon after ECMO has been initiated. The purpose of this study is to determine if our approach has achieved acceptable morbidity and mortality., Methods: Charts of live-born babies with CDH treated at our institution between 1993 and 2007 were retrospectively reviewed. Data were then compared with The Congenital Diaphragmatic Hernia Study Group and Extracorporeal Life Support Organization registries., Results: Forty-eight (39%) patients required ECMO Thirty-four of these 48 neonates were cannulated before operative repair. Venoarterial ECMO was used exclusively. The mean (SD) time of repair from cannulation was 55 (21) hours. Survival for this subset of patients was 71%. Three patients (8.8%) who underwent repair on ECMO experienced surgical site hemorrhage that required intervention., Conclusion: Early repair of CDH in neonates on ECMO can be accomplished with acceptable rates of morbidity and mortality., (Copyright 2010 Elsevier Inc. All rights reserved.)

Protein synthesis is principally regulated at the initiation stage (rather than during elongation or termination), allowing rapid, reversible and spatial control of gene expression. Progress over recent years in determining the structures and activities of initiation factors, and in mapping their interactions in ribosomal initiation complexes, have advanced our understanding of the complex translation initiation process. These developments have provided a solid foundation for studying the regulation of translation initiation by mechanisms that include the modulation of initiation factor activity (which affects almost all scanning-dependent initiation) and through sequence-specific RNA-binding proteins and microRNAs (which affect individual mRNAs).

Background: Expression of the minor virion structural protein VP2 of the calicivirus murine norovirus (MNV) is believed to occur by the unusual mechanism of termination codon-dependent reinitiation of translation. In this process, following translation of an upstream open reading frame (ORF) and termination at the stop codon, a proportion of 40S subunits remain associated with the mRNA and reinitiate at the AUG of a downstream ORF, which is typically in close proximity. Consistent with this, the VP2 start codon (AUG) of MNV overlaps the stop codon of the upstream VP1 ORF (UAA) in the pentanucleotide UAAUG., Principal Findings: Here, we confirm that MNV VP2 expression is regulated by termination-reinitiation and define the mRNA sequence requirements. Efficient reintiation is dependent upon 43 nt of RNA immediately upstream of the UAAUG site. Chemical and enzymatic probing revealed that the RNA in this region is not highly structured and includes an essential stretch of bases complementary to 18S rRNA helix 26 (Motif 1). The relative position of Motif 1 with respect to the UAAUG site impacts upon the efficiency of the process. Termination-reinitiation in MNV was also found to be relatively insensitive to the initiation inhibitor edeine., Conclusions: The termination-reinitiation signal of MNV most closely resembles that of influenza BM2. Similar to other viruses that use this strategy, base-pairing between mRNA and rRNA is likely to play a role in tethering the 40S subunit to the mRNA following termination at the VP1 stop codon. Our data also indicate that accurate recognition of the VP2 ORF AUG is not a pre-requisite for efficient reinitiation of translation in this system.

Bacterial colonies are spatially complex structures whose physiology is profoundly dependent on interactions between cells and with the underlying semi-solid substratum. Here, we use bacterial colonies as a model of a microbial community to evaluate the potential of laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) to delineate elemental distributions within colonies with minimal pre-treatment. To reduce water content of the colony and limit undesirable absorption of laser energy, we compared methods of preparing 24h-old colonies of Escherichia coli TG1 on agar for laser ablation. Colonies on excised agar segments dried on chromatography paper were superior to colonies dried in a dessicator or by prolonged incubation, with respect to signal magnitude, signal:noise ratio and background signal. Having optimised laser scan speed (10 microm s(-1)) and laser beam diameter (100 microm), further improvements were achieved by growing colonies on nylon membranes over agar, which were then transferred to the ablation chamber without further treatment. Repeated line rasters across individual membrane-supported colonies yielded three-dimensional elemental maps of colonies, revealing a convex morphology consistent with visual inspection. By normalising isotope counts for P, Mn, Zn, Fe and Ca against Mg, the most abundant cellular divalent cation, we sought elemental heterogeneity within the colony. The normalised concentration of Mn in the perimeter was higher than in the colony interior, whereas the converse was true for Ca. LA-ICP-MS is a novel and powerful method for probing elemental composition and organisation within microbial communities and should find numerous applications in, for example, biofilm studies.

Purpose: Contrast enema is the initial study of choice for simple meconium ileus to confirm diagnosis and to relieve obstruction. Despite favorable historically published results, our clinical impression suggests decreased effectiveness of the contrast enema resulting in more surgical interventions in contemporary practice., Methods: A retrospective multiinstitutional review for a 12-year period was conducted for neonates diagnosed with meconium ileus by contrast enema. The neonates were divided into 2 groups-historic group (HG = before 2002) and contemporary group (CG = after 2002). T test was used for comparison of continuous variables and chi(2) for categorical data., Results: Thirty-seven total patients were identified (21 females and 16 males). Obstruction was relieved in 8 neonates (22% overall success rate). Average enema attempt per patient was decreased in the CG group compared to HG (1.4 vs 1.9). The success rate in the CG group was 5.5% (1/18) compared to 39% (7/18) in HG., Conclusions: In this review, success of contrast enema for relief of meconium ileus has significantly decreased over time. These findings may be because of reluctance to repeat enemas, change in radiologist experience, or use of contrast agent. As a result, higher rates of operative intervention are now observed. In stable patients, surgeons should recommend repeat enemas before exploration.

A strong cell-mediated immune response is critical for controlling viral infections and is regulated by a number of cytokines, including IL-12 and IL-18. Indeed, some viruses have evolved to specifically target these pathways to counter the host immune response. Orthopoxviruses, including ectromelia virus, encode immune evasion molecules that specifically target IL-18 and IFN-gamma. We hypothesized that IL-12 and IL-18 are pivotal for induction of IFN-gamma production and subsequent generation of an effective host response to ectromelia virus infection. In this study, we demonstrate that absence of both IL-12p40 and IL-18 resulted in increased susceptibility to infection that was associated with skewing of the cytokine response to Th2 and a reduction in NK and CTL responses. The decrease in CTL response correlated with a defect in CD8(+) T cell proliferation and lower numbers of virus-specific CD8(+) T cells. Lack of either IL-12p40 and/or IL-18 was also associated with reduced numbers of CD8(+) T cells at sites of infection and with an increase in the numbers of splenic T regulatory cells. Taken together, our data indicate that IL-12p40 and IL-18 act in concert and play an important antiviral role through the up-regulation of IFN-gamma production and cell-mediated immune responses.

The Farm Bill is meant to supplement and secure farm incomes, ensure a stable food supply, and support the American farm economy. Over time, however, it has evolved into a system that creates substantial health impacts, both directly and indirectly. By generating more profit for food producers and less for family farmers; by effectively subsidizing the production of lower-cost fats, sugars, and oils that intensify the health-destroying obesity epidemic; by amplifying environmentally destructive agricultural practices that impact air, water, and other resources, the Farm Bill influences the health of Americans more than is immediately apparent. In this article, we outline three major public health issues influenced by American farm policy. These are (1) rising obesity; (2) food safety; and (3) environmental health impacts, especially exposure to toxic substances and pesticides.

Polypyrimidine tract binding (PTB) protein is a regulator of alternative pre-mRNA splicing, and also stimulates the initiation of translation dependent on many viral internal ribosome entry segments/sites (IRESs). It has four RNA-binding domains (RBDs), but although the contacts with many IRESs have been mapped, the orientation of binding (i.e., which RBD binds to which site in the IRES) is unknown. To answer this question, 16 derivatives of PTB1, each with a single cysteine flanking the RNA-binding surface in an RBD, were constructed and used in directed hydroxyl radical probing with the encephalomyocarditis virus IRES. The results, together with mass spectrometry data on the stoichiometry of PTB binding to different IRES derivatives, show that the minimal IRES binds a single PTB in a unique orientation, with RBD1 and RBD2 binding near the 3' end, and RBD3 contacting the 5' end, thereby constraining and stabilizing the three-dimensional structural fold of the IRES.

Background: The diagnosis of pyloric stenosis (PS) by physical examination is a lost art that has been replaced by radiology-performed ultrasound (US). The purpose of this study is to demonstrate that the diagnosis of PS can be made solely upon the surgeons US evaluation., Methods: Surgical ultrasonographers included 2 senior general surgery residents and 2 pediatric surgery residents without prior formal US experience. These surgeons underwent proctored training in the use of US for PS. Measurements including channel length and muscle thickness were recorded at bedside. A positive examination included muscle thickness more than 4 mm and channel length more than 16 mm. Patients with positive results underwent pyloromyotomy. Negative results were confirmed with a repeat US through the radiology department, and infants without PS were subsequently referred for appropriate medical management., Results: Thirty-two consecutive patients with suspected PS were evaluated using surgeon-performed ultrasonography. All examinations were diagnostically accurate. There were no false-positive or false-negative result. Seven patients (22%) were correctly determined to be negative for PS. The remaining 25 infants underwent successful pyloromyotomy with resolution of symptoms., Conclusion: Surgeons who have undergone focused training to perform US for PS can diagnose the condition without confirmatory testing by a radiologist.

Background: Probiotic fortified diet reduces bacterial colonization and translocation in a short-term neonatal rabbit model when continuously challenged with pathogen. The purpose of this study was to determine if live probiotic diet could remain effective at decreasing colonization/translocation of pathogens in a long-term neonatal rabbit model without ill effects of the probiotic outside the gastrointestinal (GI) tract., Methods: Rabbit pups were born via cesarean delivery 1 day preterm and assigned to 2 diets: a newly formulated diet (controls) vs the same diet fortified with the live probiotic Lactoccocus lactis. Enterobacter cloacae was added to both preparations before each feed. Pups were gavage fed twice daily, and weights were recorded. Rabbits were sacrificed on day 7, and organs were harvested and cultured for target organism growth., Results: The probiotic fortified diet resulted in a significant decrease in Enterobacter translocation to the liver and decreased colonization in the stomach and lungs. There was no evidence of Lactococccus translocation or colonization outside of the GI tract., Conclusion: This probiotic fortified diet was effective at decreasing pathogenic bacteria colonization and translocation in a long-term neonatal model. The addition of L lactis to the diet resulted in appropriate growth without any colonization or translocation of the probiotic outside of the GI tract.

Background/purpose: We previously reported that a diet acidified with citric acid effectively reinforces gastric acid protection against bacterial colonization and translocation. In this study, our objective was to examine a biologically acidified formula hypothesized to be more physiologic than formula acidified with free acid. This study was Institutional Animal Care and Use Committee (IACUC) approved and designed to determine whether this diet is better tolerated and equally effective to acidification with citric acid against gut colonization and subsequent bacterial translocation in a premature infant rabbit model., Methods: A total of 89 rabbit pups born via cesarean delivery 1 day preterm were randomly assigned to 3 feeding groups: Pelargon Nestle at pH 4.55; NAN Nestle, a control diet at pH 7.0 with similar composition; and NAN Nestle acidified in the laboratory with citric acid at pH 4.55. Pups were gavage fed every 12 hours with Enterobacter cloacae challenges of 10 colony-forming units per milliliter of diet per feed and killed on day 3 of life. Lungs, liver, spleen, mesenteric lymph nodes, stomach, and cecum were cultured and quantitatively analyzed for target organism growth and statistically analyzed using chi(2) and Kruskal-Wallis tests., Results: Pelargon, compared to acidified NAN and NAN, significantly reduced the incidence of gastric colonization (15/33 [45%], 21/27 [78%], and 25/29 [86%], respectively; P < .01) and pulmonary colonization (10/33 [30%], 19/27 [70%], 21/29 [72%]; P < .01). Comparing the bacterial logs of colonized groups, the same benefit is observed in the lungs (0.77 +/- 1.22, 1.89 +/- 1.41, 2.12 +/- 1.47; P < .01). Gut colonization and bacterial translocation were equivalent between treatment groups (mesenteric lymph nodes: 10/33 [30%], 11/27 [40%], 8/29 [27%]; spleen: 10/33 [30%], 7/27 [26%], 8/29 [27%]; liver: 10/33 [30%], 6/27 [22%], 9/29 [31%]; cecum: 33/33 [100%], 27/27 [100%], 29/29 [100%])., Conclusion: Biologically acidified formula demonstrated superior protection against pulmonary and gastric colonization compared to normal pH and diets acidified with free acid. Its effects may potentially reduce clinical pulmonary infection.

Despite growing evidence of the direct and indirect effects of the built environment on public health, planners, who shape the built environment, and public health professionals, who protect the public's health, rarely interact. Most public health professionals have little experience with urban planners, zoning boards, city councils, and others who make decisions about the built environment. Likewise, few planners understand the health implications of design, land use, or transportation decisions. One strategy for bridging this divide is the development of interdisciplinary courses in planning and public health that address the health implications of the built environment. Professional networking and Internet-based searches in 2007 led to the identification of six primarily graduate-level courses in the U.S. that address the links between the built environment and public health. Common content areas in most of the identified courses included planning and public health histories, health disparities, interdisciplinary approaches, air and water quality, physical activity, social capital, and mental health. Instructors of these courses collaborated on course content, assignments, and evaluations to develop a model curriculum that follows an active learning-centered approach to course design. The proposed model curriculum is adaptable by both planning and public health departments to promote interdisciplinary learning. Results show that students gain planning and public health perspectives through this instruction, benefiting from active-learning opportunities. Faculty implementation of the proposed interdisciplinary model curriculum will help bridge the divide between the built environment and public health and enable both planners and public health professionals to value, create, and promote healthy environments.

Purpose: We will demonstrate that a surgical resident with proven accuracy in the diagnosis of hypertrophic pyloric stenosis (HPS) can teach other surgeons to diagnose HPS with reproducible accuracy., Methods: A surgical resident with proven sonographic accuracy in diagnosing HPS instructed 5 other surgical residents in the technique. Consecutive patients referred to pediatric surgery with a presumed clinical diagnosis of HPS were examined, and measurements of residents were compared with formal radiology studies. Each surgeon was proctored for 5 examinations before independent evaluation and was blinded to results from both radiologists and other residents. Results were evaluated using Student's t test; P less than .05 was considered significant., Results: Seventy-one patients were evaluated by 5 surgical residents. Residents were diagnostically accurate in all cases. There was no statistically significant difference between pyloric muscle thickness or channel length measurements obtained by radiology and any of the residents., Conclusion: Surgeon-performed ultrasound examination for the diagnosis of HPS is accurate and reproducible through surgeon-to-surgeon instruction on appropriate technique. This skill is a valuable asset in the initial surgical evaluation of any patient with suspected HPS, expediting appropriate management.

Viruses do not carry their own protein biosynthesis machinery and the translation of viral proteins therefore requires that the virus usurps the machinery of the host cell. To allow optimal translation of viral proteins at the expense of cellular proteins, virus families have evolved a variety of methods to repress the host translation machinery, while allowing effective viral protein synthesis. Many viruses use noncanonical mechanisms that permit translation of their own RNAs under these conditions. Viruses have also developed mechanisms to evade host innate immune responses that would repress translation under conditions of viral infection, in particular PKR activation in response to double-stranded RNA (dsRNA). Importantly, the study of viral translation mechanisms has enormously enhanced our understanding of many aspects of the cellular protein biosynthesis pathway and its components. A number of unusual mechanisms of translation initiation that were first discovered in viruses have since been observed in cellular mRNAs, and it has become apparent that a diverse range of translation mechanisms operates in eukaryotes, allowing subtle regulation of this essential process., (Copyright © 2009 Elsevier Inc. All rights reserved.)

Escherichia coli possesses two major systems for inorganic phosphate (P(i)) uptake. The Pst system (pstSCAB) is inducible by low phosphate concentrations whereas the low-affinity transporter (pitA) has been described as constitutively expressed. PitA catalyses transport of metal [Mg(II), Ca(II)]-phosphate complexes, and mutations in pitA confer Zn(II) resistance. Here we report that pitA transcription is not constitutive; activity of a single-copy pitA-lacZ transcriptional fusion (monolysogen) was maximal at high extracellular Zn(II) (150 microM), in the absence of added P(i), and in a well-defined pitA mutant strain. Intracellular zinc levels were unaffected by adding Zn(II) to the medium for both the wild-type and mutant strains. However, in the wild-type strain, Mg levels (per gram of dry biomass) fell by eightfold in cells grown with added Zn(II) and by 20-fold when Zn(II) and P(i) were added to cultures. Mutation of pitA reduced the effects of external Zn(II) and phosphate levels on Mg pools, consistent with competition or inhibition by Zn(II) of PitA. The mechanism of pitA regulation by extracellular Zn(II) and P(i) is unknown but appears not to involve Fur or other well-characterized regulators.

Background/purpose: Secure placement of peritoneal dialysis (PD) catheters in the pelvis has been described by various techniques. We describe minimally invasive placement using an Endo Close device, securing the catheter in the pelvis, and compare this method with standard open technique in children., Methods: A retrospective institutional review was conducted for children requiring PD access from 2001 to 2007. Patients were grouped into laparoscopic with secure placement (SP) and open placement (OP) groups. Groups were cohort-matched based on age, paying particular attention to the number of catheter migrations., Results: Twenty-seven patients underwent 36 procedures in SP, whereas 23 patients in OP had 32 catheter-related procedures. Exit site infections were decreased in SP (0.57 vs 1.33 episodes per patient-year). There was no difference in the number of catheter migrations (3 vs 5); however, time to migration was statistically longer in the SP group (9 vs. 2.4 months, P < .05)., Conclusions: Laparoscopic placement of PD catheters using a securing suture in the pelvis is a more durable technique when compared to open placement. Extending the catheter migration time is important in children when PD is used as a bridge to renal transplantation.

Coupled expression of the M1 and BM2 open-reading frames (ORFs) of influenza B from the dicistronic segment 7 mRNA occurs by a process of termination-dependent reinitiation. The AUG start codon of the BM2 ORF overlaps the stop codon of the upstream M1 ORF in the pentanucleotide UAAUG, and BM2 synthesis is dependent upon translation of the M1 ORF and termination at the stop codon. Here, we have investigated the mRNA sequence requirements for BM2 expression. Termination-reinitiation is dependent upon 45 nucleotide (nt) of RNA immediately upstream of the UAAUG pentanucleotide, which includes an essential stretch complementary to 18S rRNA helix 26. Thus, similar to the caliciviruses, base-pairing between mRNA and rRNA is likely to play a role in tethering the 40S subunit to the mRNA following termination at the M1 stop codon. Consistent with this, repositioning of the M1 stop codon more than 24 nt downstream from the BM2 start codon inhibited BM2 expression. RNA structure probing revealed that the RNA upstream of the UAAUG overlap is not highly structured, but upon encountering the M1 stop codon by the ribosome, a stem-loop may form immediately 5' of the ribosome, with the 18S rRNA complementary region in the apical loop and in close proximity to helix 26. Mutational analysis reveals that the normal requirements for start site selection in BM2 expression are suspended, with little effect of initiation codon context and efficient use of noncanonical initiation codons. This suggests that the full complement of initiation factors is not required for the reinitiation process.

Purpose: Evidence-based guidelines for the treatment of pediatric snakebite injuries are lacking because they occur infrequently in most centers., Methods: We reviewed our experience treating snakebites from January 1995 through December 2005. Demographic (eg, age, sex, geographic location) and clinical information (eg, location of bite, species of snake, vital signs, laboratories, treatment, hospital length of stay) were obtained., Results: Over the last decade, we have treated 114 children with confirmed snakebites. Mean age was 7.3 +/- 4.2 years (range, 1-17 years), and snakebites were more common in males (n = 68, 60%). All bites occurred on the extremities, and lower extremity bites were more common (n = 71, 62%). Copperheads inflicted the most bite injuries (n = 65, 57%), followed by rattlesnakes (n = 9, 8%) and cottonmouths (n = 7, 6%). The snake was not identified in 33 (29%) cases. Seven (6%) children were treated with Crotalidae antivenin. Of the children treated with antivenin, only 4 met criteria for treatment, and 1 had an anaphylactic reaction. If compartment syndrome was suspected based on neurovascular examination, compartment pressures were measured. Only 2 (1.8%) patients required fasciotomies. Over the last 2 years, we have stopped empiric treatment with antibiotics and have not observed any infectious complications. Average hospital length of stay was 30 +/- 25 hours., Conclusions: Most children bitten by pit vipers can be managed conservatively with analgesics and elevation of the affected extremity. Treatment with Crotalidae antivenin, antibiotics, and fasciotomy is rarely indicated.

Background: Robot-assisted surgery must be evaluated before its acceptance as an option for standard therapy in the pediatric population. Our objective is a comparison of results using the robot system with results for the laparoscopic and open approaches., Methods: Following IRB approval, robot-assisted procedures were case-matched with controls, selected from 1994 to 2005. Data for 150 Nissen cases were divided equally into 3 groups [robot (R), laparoscopic (L), and open (O)], comparing surgical times, length of hospitalization, and outcomes., Results: The average age (R = 117+/-64 months, L = 107+/-71 months, O = 85+/-55 months, P<0.05) and weight (R = 37+/-23 kg, L = 33+/-24 kg, O = 24+/-17 kg, P<0.05) of the open group were lower comparatively. Robot operative times proved significantly longer compared with laparoscopic and open time (R = 160+/-61 min, L = 107+31 min, O = 73+/-27 min, P<0.05). The robot had 2 conversions (2/50, 4%), comparable to the laparoscopic conversion rate (1/50, 2%). Open cases resulted in longer hospitalization [R = 2.94+/-4.5 days, L = 3.54+/-7.8 days, O = 3.5+/-2.8, P<0.05]. Complication rates were equivalent between groups. The most common complication with the da Vinci and laparoscopic approaches was tight wrap requiring dilation [R = 4/50 (8%) and L = 3/50 (6%)]., Conclusion: Robot-assisted surgery is equivalent to standard laparoscopic surgery in terms of complications and length of stay, with both having significantly increased operation times but reduced length of stay compared with open surgery. Further experience with this technology is needed to overcome the learning curve and reduce operative times.

Purpose: Current organizational guidelines for the management of isolated spleen and liver injuries are based on injury grade. We propose that management based on hemodynamic status is safe in children and results in decreased length of stay (LOS) and resource use compared to current grade-based guidelines., Methods: Patients with spleen or liver injuries for a 5-year period were identified using our institutional trauma registry. All patients were managed using a pathway based on hemodynamic status. Charts were reviewed for demographics, mechanism, hematrocrit values, transfusion requirement, imaging, injury grade, LOS, and outcome. Exclusion criteria included penetrating mechanism, associated injuries altering LOS or ambulation status, combined spleen/liver injury, initial operative management or death. Statistical comparison was performed using Student's t test; P < .05 is significant., Results: One hundred one patients (50 spleen, 51 liver) meeting inclusion criteria were identified. Average actual LOS for all patients was 1.9 days vs 3.2 projected days based on American Pediatric Surgical Association guidelines (P < .0001). Actual vs projected LOS for grades III to V was 2.5 vs 4.3 days (P < .0001). All patients returned to full activity without complication., Conclusions: Isolated blunt spleen and liver injuries, regardless of grade, can be safely managed using a pathway based on hemodynamic status, resulting in decreased LOS and resource use compared to current guidelines.

Background: The frequency of synonymous codon usage varies widely between organisms. Suboptimal codon content limits expression of viral, experimental or therapeutic heterologous proteins due to limiting cognate tRNAs. Codon content is therefore often adjusted to match codon bias of the host organism. Codon content also varies between genes within individual mammalian species. However, little attention has been paid to the consequences of codon content upon translation of host proteins., Methodology/principal Findings: In comparing the splicing repressor activities of transfected human PTB and its two tissue-restricted paralogs-nPTB and ROD1-we found that the three proteins were expressed at widely varying levels. nPTB was expressed at 1-3% the level of PTB despite similar levels of mRNA expression and 74% amino acid identity. The low nPTB expression was due to the high proportion of codons with A or U at the third codon position, which are suboptimal in human mRNAs. Optimization of the nPTB codon content, akin to the "humanization" of foreign ORFs, allowed efficient translation in vivo and in vitro to levels comparable with PTB. We were then able to demonstrate that all three proteins act as splicing repressors., Conclusions/significance: Our results provide a striking illustration of the importance of mRNA codon content in determining levels of protein expression, even within cells of the natural host species.

Purpose: The purpose of the study was to identify the spectrum of disease characteristics of pheochromocytoma in children., Methods: Records of 21 consecutive children diagnosed with pheochromocytoma were reviewed. Patients' age, sex, presentation, associated conditions, diagnostic and imaging modalities used, preoperative preparation, operative details, outcome, and follow-up were recorded., Results: The study included 21 children. Patients were diagnosed clinically and confirmed by biochemical tests. Tumors were localized by imaging studies and all were confirmed pathologically. Patients included 17 with adrenal and 4 with extra-adrenal tumors including 1 in the urinary bladder. Seventeen had sporadic and 4 had familial pheochromocytoma. Associated manifestations were the predominant features in 1 of the 4 patients with familial pheochromocytoma. The patient with bladder pheochromocytoma presented with gross hematuria. Hypertension and visual disturbances were prominent findings in the other patients with sporadic form. Two patients (1 sporadic and 1 familial) had malignant pheochromocytoma. One patient with benign pheochromocytoma had multiple recurrences in chromaffin-containing sites. All patients were treated surgically. Seventeen patients were treated preoperatively with alpha-adrenergic blockade. Two patients continued to have significant visual disturbances. One patient with malignant pheochromocytoma died of the disease, and 1 with recurrent pheochromocytoma had neurologic consequences., Conclusions: Pheochromocytoma in children has unique characteristics. To our knowledge, this series is one of the largest reports of adrenal pheochromocytoma in children. It also reflects the spectrum of pheochromocytoma in this age group.

Background/purpose: Acidified diets are protective against intestinal bacterial colonization and translocation. Probiotic diets are designed to modulate the intestinal flora to enhance mucosal immunity. This study was designed to determine if formula acidified with live probiotic decreases bacterial gut colonization and translocation, and is equally tolerated as other acidified diets., Methods: One hundred twenty-eight rabbit pups delivered via cesarean section [cesarean delivery, cesarean birth, abdominal delivery] were randomly assigned to 4 feeding groups: NAN Nestle (control, pH 7.0), NAN acidified with citric acid (pH 4.55), biologically acidified Pelargon (pH 4.55), and NAN with live Lactococcus lactis culture (pH 4.2). Pups were gavage fed every 12 hours with Enterobacter cloacae challenges of 10 colony-forming units/mL per feed and killed on day of life 3. Lungs, liver, spleen, mesenteric lymph nodes (MLNs), stomach, and cecum were cultured and quantitatively analyzed for target organism growth. Results were analyzed using chi(2) tests., Results: NAN with live probiotic culture, when compared with Pelargon, acidified NAN, and NAN, significantly reduced the incidence of Enterobacter pulmonary colonization (P < .01), bacterial translocation (liver, P < .025; spleen and MLN, P < .05), and gastric and intestinal colonization (P < .001 for both)., Conclusion: Probiotic-fortified formula provides superior protection against pulmonary and gastrointestinal bacterial colonization and translocation compared with neutral and acidified formulas, and is equally tolerated.

Background/purpose: At our institution, patients undergoing selected operative procedures are referred to a protocol for telephone follow-up by surgical specialty nurses. Our objective was to review our experience with this protocol to determine if telephone follow-up is a safe and preferred alternative to the traditional postoperative clinic visit., Methods: Records of patients followed up by telephone over 6 months were evaluated for information regarding each patient's postoperative course. Records included telephone follow-up forms, clinic notes, and emergency department records., Results: A total of 563 patients underwent a total of 601 procedures. Seventy-six percent (n = 427) were successfully contacted postoperatively; 24% (n = 136) did not respond to multiple contact attempts. Forty-five requests for clinic follow-up resulted in 27 actual visits, 10 from families not satisfied with telephone contact alone. Most families contacted (382/427, 90%) were satisfied and did not request an appointment. A total of 43 postoperative clinic or emergency department evaluations resulted in 9 interventions (1.6% complication rate)., Conclusion: Postoperative follow-up by telephone using a structured protocol is a safe alternative to routine clinic follow-up for patients undergoing selected procedures and is preferred by patients' families.