53 research outputs found
Outcome and renal function following salvage surgery for bilateral Wilms tumor: a single-institution experience
Objective: The aim of this study was to determine the surgical outcomes and renal function following salvage surgery for bilateral Wilms tumor (BWT). Summary background data The challenge for the surgeon treating BWT lies in striking a fine balance between renal preservation and oncological clearance.Methods: This is a retrospective review of medical records in a tertiary care hospital in India. Nine children with BWT who presented between 2005 and 2015 were reviewed and followed up through telephone. Survival rates were calculated using the Kaplan–Meier method. A P value of less than 0.05 was considered statistically significant.Results: Seven (78%) of nine children were boys and two (22%) were syndromic. Six (67%) children presented at less than 1 year of age. Eight (89%) children presented with an abdominal mass. There were no metastases at presentation. All children underwent trucut biopsy and neoadjuvant chemotherapy. Six children underwent surgery: four underwent bilateral nephron sparing surgery (NSS) and two underwent unilateral nephrectomy with contralateral NSS. Tumor recurred in two children. The mean follow-up was 38 months (range: 5–108 months). Creatinine clearance (CrCl) improved postoperatively in all children. Postoperative hypertension was transient and resolved with improvement in CrCl.Conclusion: Children with BWT in the Indian subcontinent may be younger than those in the rest of the world. NSS yields good outcomes even for recurrences. Postoperative hypertension is transient in the majority of patients and correlated with improvement in CrCl. Prognosis is related to operability and syndromic association.Keywords: bilateral Wilms, creatinine clearance, hypertension, nephron sparing surger
Vitamin C activates young LINE-1 elements in mouse embryonic stem cells via H3K9me3 demethylation
Background
Vitamin C (vitC) enhances the activity of 2-oxoglutarate-dependent dioxygenases, including TET enzymes, which catalyse DNA demethylation, and Jumonji-domain histone demethylases. The epigenetic remodelling promoted by vitC improves the efficiency of induced pluripotent stem cell derivation, and is required to attain a ground-state of pluripotency in embryonic stem cells (ESCs) that closely mimics the inner cell mass of the early blastocyst. However, genome-wide DNA and histone demethylation can lead to upregulation of transposable elements (TEs), and it is not known how vitC addition in culture media affects TE expression in pluripotent stem cells.
Results
Here we show that vitC increases the expression of several TE families, including evolutionarily young LINE-1 (L1) elements, in mouse ESCs. We find that TET activity is dispensable for L1 upregulation, and that instead it occurs largely as a result of H3K9me3 loss mediated by KDM4A/C histone demethylases. Despite increased L1 levels, we did not detect increased somatic insertion rates in vitC-treated cells. Notably, treatment of human ESCs with vitC also increases L1 protein levels, albeit through a distinct, post-transcriptional mechanism.
Conclusion
VitC directly modulates the expression of mouse L1s and other TEs through epigenetic mechanisms, with potential for downstream effects related to the multiple emerging roles of L1s in cellular function.This work was supported by grants from the Wellcome Trust/Royal Society (101225/Z/13/Z) and MRC (MR/X008487/1) to M.R.B.; and BBSRC (BB/T000031/1) to M.R.B. and J.M.F.Peer reviewe
Rapid and High-Throughput pan-Orthopoxvirus Detection and Identification using PCR and Mass Spectrometry
The genus Orthopoxvirus contains several species of related viruses, including the causative agent of smallpox (Variola virus). In addition to smallpox, several other members of the genus are capable of causing human infection, including monkeypox, cowpox, and other zoonotic rodent-borne poxviruses. Therefore, a single assay that can accurately identify all orthopoxviruses could provide a valuable tool for rapid broad orthopovirus identification. We have developed a pan-Orthopoxvirus assay for identification of all members of the genus based on four PCR reactions targeting Orthopoxvirus DNA and RNA helicase and polymerase genes. The amplicons are detected using electrospray ionization-mass spectrometry (PCR/ESI-MS) on the Ibis T5000 system. We demonstrate that the assay can detect and identify a diverse collection of orthopoxviruses, provide sub-species information and characterize viruses from the blood of rabbitpox infected rabbits. The assay is sensitive at the stochastic limit of PCR and detected virus in blood containing approximately six plaque-forming units per milliliter from a rabbitpox virus-infected rabbit
Mitochondrial 2,4-dienoyl-CoA Reductase Deficiency in Mice Results in Severe Hypoglycemia with Stress Intolerance and Unimpaired Ketogenesis
The mitochondrial β-oxidation system is one of the central metabolic pathways of energy metabolism in mammals. Enzyme defects in this pathway cause fatty acid oxidation disorders. To elucidate the role of 2,4-dienoyl-CoA reductase (DECR) as an auxiliary enzyme in the mitochondrial β-oxidation of unsaturated fatty acids, we created a DECR–deficient mouse line. In Decr−/− mice, the mitochondrial β-oxidation of unsaturated fatty acids with double bonds is expected to halt at the level of trans-2, cis/trans-4-dienoyl-CoA intermediates. In line with this expectation, fasted Decr−/− mice displayed increased serum acylcarnitines, especially decadienoylcarnitine, a product of the incomplete oxidation of linoleic acid (C18:2), urinary excretion of unsaturated dicarboxylic acids, and hepatic steatosis, wherein unsaturated fatty acids accumulate in liver triacylglycerols. Metabolically challenged Decr−/− mice turned on ketogenesis, but unexpectedly developed hypoglycemia. Induced expression of peroxisomal β-oxidation and microsomal ω-oxidation enzymes reflect the increased lipid load, whereas reduced mRNA levels of PGC-1α and CREB, as well as enzymes in the gluconeogenetic pathway, can contribute to stress-induced hypoglycemia. Furthermore, the thermogenic response was perturbed, as demonstrated by intolerance to acute cold exposure. This study highlights the necessity of DECR and the breakdown of unsaturated fatty acids in the transition of intermediary metabolism from the fed to the fasted state
Global Surveillance of Emerging Influenza Virus Genotypes by Mass Spectrometry
Effective influenza surveillance requires new methods capable of rapid and inexpensive genomic analysis of evolving viral species for pandemic preparedness, to understand the evolution of circulating viral species, and for vaccine strain selection. We have developed one such approach based on previously described broad-range reverse transcription PCR/electrospray ionization mass spectrometry (RT-PCR/ESI-MS) technology.Analysis of base compositions of RT-PCR amplicons from influenza core gene segments (PB1, PB2, PA, M, NS, NP) are used to provide sub-species identification and infer influenza virus H and N subtypes. Using this approach, we detected and correctly identified 92 mammalian and avian influenza isolates, representing 30 different H and N types, including 29 avian H5N1 isolates. Further, direct analysis of 656 human clinical respiratory specimens collected over a seven-year period (1999-2006) showed correct identification of the viral species and subtypes with >97% sensitivity and specificity. Base composition derived clusters inferred from this analysis showed 100% concordance to previously established clades. Ongoing surveillance of samples from the recent influenza virus seasons (2005-2006) showed evidence for emergence and establishment of new genotypes of circulating H3N2 strains worldwide. Mixed viral quasispecies were found in approximately 1% of these recent samples providing a view into viral evolution.Thus, rapid RT-PCR/ESI-MS analysis can be used to simultaneously identify all species of influenza viruses with clade-level resolution, identify mixed viral populations and monitor global spread and emergence of novel viral genotypes. This high-throughput method promises to become an integral component of influenza surveillance
Actin Dynamics Regulate Multiple Endosomal Steps during Kaposi's Sarcoma-Associated Herpesvirus Entry and Trafficking in Endothelial Cells
The role of actin dynamics in clathrin-mediated endocytosis in mammalian cells is unclear. In this study, we define the role of actin cytoskeleton in Kaposi's sarcoma-associated herpesvirus (KSHV) entry and trafficking in endothelial cells using an immunofluorescence-based assay to visualize viral capsids and the associated cellular components. In contrast to infectivity or reporter assays, this method does not rely on the expression of any viral and reporter genes, but instead directly tracks the accumulation of individual viral particles at the nuclear membrane as an indicator of successful viral entry and trafficking in cells. Inhibitors of endosomal acidification reduced both the percentage of nuclei with viral particles and the total number of viral particles docking at the perinuclear region, indicating endocytosis, rather than plasma membrane fusion, as the primary route for KSHV entry into endothelial cells. Accordingly, a viral envelope protein was only detected on internalized KSHV particles at the early but not late stage of infection. Inhibitors of clathrin- but not caveolae/lipid raft-mediated endocytosis blocked KSHV entry, indicating that clathrin-mediated endocytosis is the major route of KSHV entry into endothelial cells. KSHV particles were colocalized not only with markers of early and recycling endosomes, and lysosomes, but also with actin filaments at the early time points of infection. Consistent with these observations, transferrin, which enters cells by clathrin-mediated endocytosis, was found to be associated with actin filaments together with early and recycling endosomes, and to a lesser degree, with late endosomes and lysosomes. KSHV infection induced dynamic actin cytoskeleton rearrangements. Disruption of the actin cytoskeleton and inhibition of regulators of actin nucleation such as Rho GTPases and Arp2/3 complex profoundly blocked KSHV entry and trafficking. Together, these results indicate an important role for actin dynamics in the internalization and endosomal sorting/trafficking of KSHV and clathrin-mediated endocytosis in endothelial cells
Preperitoneal bladder augmentation: Feasibility and results
Introduction: Bladder augmentation is an important part of pediatric reconstructive urology. This study was conducted to assess the feasibility and results of our technique of preperitoneal bladder augmentation.
Materials and Methods: Thirty-three children underwent preperitoneal bladder augmentation for small inelastic bladders who had failed medical management or needed undiversion. The underlying diagnosis included neurogenic bladder, valve bladder, bladder exstrophy, non-neurogenic neurogenic, ectopic ureters, and urogenital sinus. The operative procedure involved placing the entire augmentation in the preperitoneal or subcutaneous space after bivalving the native bladder. The augment segment of the bowel with its pedicle was brought into the preperitoneal space through a small opening in the parietal peritoneum. A Mitrofanoff port was also provided where needed.
Results: Preperitoneal augmentation provided an adequately compliant, good volume bladder except in children with bladder exstrophy or previous abdominal surgery. There was a good cystometric recovery, with resolution of hydronephrosis and incontinence. Vesicoureteral reflux resolved in 24 of 26 units. In the 13 children who were uremic preoperatively, there was a significant decrease in serum creatinine levels, although 9 children continued to have supra-normal serum creatinine. Surgical complications seen were within expectations. There was no incidence of intraperitoneal leak, which is the main projected benefit of this procedure over the traditional “intraperitoneal” method of augmentation.
Conclusions: The preperitoneal augmentation provides an adequate, safe, and low-pressure reservoir of urine except in cases of bladder exstrophy and previous abdominal surgery
Metanephric stromal tumour: A rare pediatric benign stromal specific renal neoplasm
A case of incidentally detected Metanephric Stromal Tumour (MST) is reported here. This is a rare, recently recognized pediatric benign stromal specific renal neoplasm. A review of the English literature revealed only five cases after its original description by Argani et al. Recognition of this entity can spare a child from potentially toxic adjuvant chemotherapy that might be used to treat malignant lesions which are part of the differential diagnosis, particularly clear cell sarcoma of kidney (CCSK)
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