Rapunzel Syndrome: a rare cause of acute small bowel obstruction

The Rapunzel syndrome is a very rare condition where trichobezoar hasextended up to the small bowel. Here we are reporting a rare case of Rapunzel syndrome in an adolescent girl with history of trichophagia who presented with small bowel obstruction. Patient underwent exploratory laparotomy and bezoar was removed through gastrotomy. Post-operative course was unremarkable.KEY WORDS: Rapunzel syndrome; Trichobezoars; Psychotherapy; Gastrotom

Macrophage migration inhibitory factor downregulation: a novel mechanism of resistance to anti-angiogenic therapy.

Anti-angiogenic therapies for cancer such as VEGF neutralizing antibody bevacizumab have limited durability. While mechanisms of resistance remain undefined, it is likely that acquired resistance to anti-angiogenic therapy will involve alterations of the tumor microenvironment. We confirmed increased tumor-associated macrophages in bevacizumab-resistant glioblastoma patient specimens and two novel glioblastoma xenograft models of bevacizumab resistance. Microarray analysis suggested downregulated macrophage migration inhibitory factor (MIF) to be the most pertinent mediator of increased macrophages. Bevacizumab-resistant patient glioblastomas and both novel xenograft models of resistance had less MIF than bevacizumab-naive tumors, and harbored more M2/protumoral macrophages that specifically localized to the tumor edge. Xenografts expressing MIF-shRNA grew more rapidly with greater angiogenesis and had macrophages localizing to the tumor edge which were more prevalent and proliferative, and displayed M2 polarization, whereas bevacizumab-resistant xenografts transduced to upregulate MIF exhibited the opposite changes. Bone marrow-derived macrophage were polarized to an M2 phenotype in the presence of condition-media derived from bevacizumab-resistant xenograft-derived cells, while recombinant MIF drove M1 polarization. Media from macrophages exposed to bevacizumab-resistant tumor cell conditioned media increased glioma cell proliferation compared with media from macrophages exposed to bevacizumab-responsive tumor cell media, suggesting that macrophage polarization in bevacizumab-resistant xenografts is the source of their aggressive biology and results from a secreted factor. Two mechanisms of bevacizumab-induced MIF reduction were identified: (1) bevacizumab bound MIF and blocked MIF-induced M1 polarization of macrophages; and (2) VEGF increased glioma MIF production in a VEGFR2-dependent manner, suggesting that bevacizumab-induced VEGF depletion would downregulate MIF. Site-directed biopsies revealed enriched MIF and VEGF at the enhancing edge in bevacizumab-naive patients. This MIF enrichment was lost in bevacizumab-resistant glioblastomas, driving a tumor edge M1-to-M2 transition. Thus, bevacizumab resistance is driven by reduced MIF at the tumor edge causing proliferative expansion of M2 macrophages, which in turn promotes tumor growth

Rectovaginal Fistulas Not Involving the Rectovaginal Septum Should Be Treated Like Anal Fistulas: A New Concept and Proposal for a Reclassification of Rectovaginal Fistulas

Pankaj Garg,1 Laxmikant Ladukar,2 Vipul D Yagnik,3 Kaushik Bhattacharya,4 Gurleen Kaur5 1Department of Colorectal Surgery, Garg Fistula Research Institute (GFRI), Panchkula, Haryana, India; 2Department of Surgery, Ladukar Surgical Hospital, Bramhapuri, Maharashtra, India; 3Department of Surgery, Banas Medical College and Research Institute, Palanpur, Gujarat, India; 4Department of Surgery, MGM Medical College and LSK Hospital, Kishanganj, Bihar, India; 5Department of Pharmacology, Adesh Medical College and Hospital, Shahbad, Haryana, IndiaCorrespondence: Pankaj Garg, Chief Colorectal Surgeon, Colorectal Surgery, Garg Fistula Research Institute (GFRI), 1042, Sector-15, Panchkula, Haryana, 134113, India, Email [email protected]: Many rectovaginal fistulas(RVF), especially low RVF, do not involve/penetrate the RV-septum, but due to lack of proper nomenclature, such fistulas are also managed like RVF (undertaking repair of RV-septum) and inadvertently lead to the formation of a high RVF (involving RV-septum) in many cases. Therefore, REctovaginal Fistulas, Not Involving the Rectovaginal Septum, should be Treated like Anal fistulas(RENISTA) to prevent any risk of injury to the RV septum. This concept(RENISTA) was tested in this study.Methods: RVFs not involving RV-septum were managed like anal fistulas, and the RV-septum was not cut/incised. MRI, objective incontinence scoring, and anal manometry were done preoperatively and postoperatively. High RVF (involving RV-septum) were excluded.Results: Twenty-seven patients with low RVF (not involving RV-septum) were operated like anal fistula[age:35.2± 9.2 years, median follow-up-15 months (3– 36 months)]. 19/27 were low fistula[ 1/3 EAS involved) and underwent a sphincter-sparing procedure. Three patients were excluded. The fistula healed well in 22/24 (91.7%) patients and did not heal in 2/24 (8.3%). The healing was confirmed on MRI, and there was no significant change in mean incontinence scores and anal pressures on tonometry. RV-septum injury did not occur in any patient.Conclusions: RVF not involving RV-septum were managed like anal fistulas with a high cure rate and no significant change in continence. RV-septum injury or formation of RVF with septum involvement did not occur in any patient. The RENISTA concept was validated in the present study. A new classification was developed to prevent any inadvertent injury to the RV-septum.Keywords: rectovaginal, fistula, anal, incontinence, recurrence, scoring system, fistulotomy, classificatio

Genome-wide expression profiling of urinary bladder implicates desmosomal and cytoskeletal dysregulation in the bladder exstrophy-epispadias complex

The bladder exstrophy-epispadias complex (BEEC) represents a spectrum of urological abnormalities where part, or all, of the distal urinary tract fails to close during development, becoming exposed on the outer abdominal wall. While the etiology of BEEC remains unknown, strong evidence exists that genetic factors are implicated. To understand the pathways regulating embryonic bladder development and to identify high-probability BEEC candidate genes, we conducted a genome-wide expression profiling (GWEP) study using normal and exstrophic human urinary bladders and human and mouse embryologic bladder-precursor tissues. We identified 162 genes differentially expressed in both embryonic and postnatal human samples. Pathway analysis of these genes revealed 11 biological networks with top functions related to skeletal and muscular system development, cellular assembly and development, organ morphology, or connective tissue disorders. The two most down-regulated genes desmin (DES, fold-change, -74.7) and desmuslin (DMN, fold-change, -53.0) are involved in desmosome mediated cell-cell adhesion and cytoskeletal architecture. Intriguingly, the sixth most overexpressed gene was desmoplakin (DSP, fold-change, +48.8), the most abundant desmosomal protein. We found 30% of the candidate genes to be directly associated with desmosome structure/function or cytoskeletal assembly, pointing to desmosomal and/or cytoskeletal deregulation as an etiologic factor for BEEC. Further findings indicate that p63, PERP, SYNPO2 and the Wnt pathway may also contribute to BEEC etiology. This study provides the first expression profile of urogenital genes during bladder development and points to the high-probability candidate genes for BEEC

A Comparison of Four Treatments for Generalized Convulsive Status Epilepticus

ABSTRACT Background and Methods Although generalized convulsive status epilepticus is a life-threatening emergency, the best initial drug treatment is uncertain. We conducted a five-year randomized, doubleblind, multicenter trial of four intravenous regimens: diazepam (0.15 mg per kilogram of body weight) followed by phenytoin (18 mg per kilogram), lorazepam (0.1 mg per kilogram), phenobarbital (15 mg per kilogram), and phenytoin (18 mg per kilogram). Patients were classified as having either overt generalized status epilepticus (defined as easily visible generalized convulsions) or subtle status epilepticus (indicated by coma and ictal discharges on the electroencephalogram, with or without subtle convulsive movements such as rhythmic muscle twitches or tonic eye deviation). Treatment was considered successful when all motor and electroencephalographic seizure activity ceased within 20 minutes after the beginning of the drug infusion and there was no return of seizure activity during the next 40 minutes. Analyses were performed with data on only the 518 patients with verified generalized convulsive status epilepticus as well as with data on all 570 patients who were enrolled. Results Three hundred eighty-four patients had a verified diagnosis of overt generalized convulsive status epilepticus. In this group, lorazepam was successful in 64.9 percent of those assigned to receive it, phenobarbital in 58.2 percent, diazepam and phenytoin in 55.8 percent, and phenytoin in 43.6 percent (P=0.02 for the overall comparison among the four groups). Lorazepam was significantly superior to phenytoin in a pairwise comparison (P=0.002). Among the 134 patients with a verified diagnosis of subtle generalized convulsive status epilepticus, no significant differences among the treatments were detected (range of success rates, 7.7 to 24.2 percent). In an intention-to-treat analysis, the differences among treatment groups were not significant, either among the patients with overt status epilepticus (P=0.12) or among those with subtle status epilepticus (P=0.91). There were no differences among the treatments with respect to recurrence during the 12- hour study period, the incidence of adverse reactions, or the outcome at 30 days. Conclusions As initial intravenous treatment for overt generalized convulsive status epilepticus, lorazepam is more effective than phenytoin. Although lorazepam is no more efficacious than phenobarbital or diazepam and phenytoin, it is easier to use. (N Engl J Med 1998;339:792-8.

Cell Culture Replication of a Genotype 1b Hepatitis C Virus Isolate Cloned from a Patient Who Underwent Liver Transplantation

The introduction of the genotype 2a isolate JFH1 was a major breakthrough in the field of hepatitis C virus (HCV), allowing researchers to study the complete life cycle of the virus in cell culture. However, fully competent culture systems encompassing the most therapeutically relevant HCV genotypes are still lacking, especially for the highly drug-resistant genotype 1b. For most isolated HCV clones, efficient replication in cultured hepatoma cells requires the introduction of replication-enhancing mutations. However, such mutations may interfere with viral assembly, as occurs in the case of the genotype 1b isolate Con1. In this study, we show that a clinical serum carrying a genotype 1b virus with an exceptionally high viral load was able to infect Huh7.5 cells. Similar to previous reports, inoculation of Huh7.5 cells by natural virus is very inefficient compared to infection by cell culture HCV. A consensus sequence of a new genotype 1b HCV isolate was cloned from the clinical serum (designated Barcelona HCV1), and then subjected to replication studies. This virus replicated poorly in a transient fashion in Huh7.5 cells after electroporation with in vitro transcribed RNA. Nonetheless, approximately 3 weeks post electroporation and thereafter, core protein-positive cells were detected by immunofluorescence. Surprisingly, small amounts of core protein were also measurable in the supernatant of electroporated cells, suggesting that HCV particles might be assembled and released. Our findings not only enhance the current method of cloning in vitro HCV replication-competent isolates, but also offer valuable insights for the realization of fully competent culture systems for HCV

Structural Basis for Broad Neutralization of Hepatitis C Virus Quasispecies

Monoclonal antibodies directed against hepatitis C virus (HCV) E2 protein can neutralize cell-cultured HCV and pseudoparticles expressing envelopes derived from multiple HCV subtypes. For example, based on antibody blocking experiments and alanine scanning mutagenesis, it was proposed that the AR3B monoclonal antibody recognized a discontinuous conformational epitope comprised of amino acid residues 396–424, 436–447, and 523–540 of HCV E2 envelope protein. Intriguingly, one of these segments (436–447) overlapped with hypervariable region 3 (HVR3), a domain that exhibited significant intrahost and interhost genetic diversity. To reconcile these observations, amino-acid sequence variability was examined and homology-based structural modelling of E2 based on tick-borne encephalitis virus (TBEV) E protein was performed based on 413 HCV sequences derived from 18 subjects with chronic hepatitis C. Here we report that despite a high degree of amino-acid sequence variability, the three-dimensional structure of E2 is remarkably conserved, suggesting broad recognition of structural determinants rather than specific residues. Regions 396–424 and 523–540 were largely exposed and in close spatial proximity at the surface of E2. In contrast, region 436–447, which overlaps with HVR3, was >35 Å away, and estimates of buried surface were inconsistent with HVR3 being part of the AR3B binding interface. High-throughput structural analysis of HCV quasispecies could facilitate the development of novel vaccines that target conserved structural features of HCV envelope and elicit neutralizing antibody responses that are less vulnerable to viral escape

Genetic variability of hepatitis C virus before and after combined therapy of interferon plus ribavirin

We present an analysis of the selective forces acting on two hepatitis C virus genome regions previously postulated to be involved in the viral response to combined antiviral therapy. One includes the three hypervariable regions in the envelope E2 glycoprotein, and the other encompasses the PKR binding domain and the V3 domain in the NS5A region. We used a cohort of 22 non-responder patients to combined therapy (interferon alpha-2a plus ribavirin) for which samples were obtained before initiation of therapy and after 6 or/and 12 months of treatment. A range of 25-100 clones per patient, genome region and time sample were sequenced. These were used to detect general patterns of adaptation, to identify particular adaptation mechanisms and to analyze the patterns of evolutionary change in both genome regions. These analyses failed to detect a common adaptive mechanism for the lack of response to antiviral treatment in these patients. On the contrary, a wide range of situations were observed, from patients showing no positively selected sites to others with many, and with completely different topologies in the reconstructed phylogenetic trees. Altogether, these results suggest that viral strategies to evade selection pressure from the immune system and antiviral therapies do not result from a single mechanism and they are likely based on a range of different alternatives, in which several different changes, or their combination, along the HCV genome confer viruses the ability to overcome strong selective [email protected]; [email protected]; [email protected]; [email protected]; [email protected]; [email protected]; [email protected]; [email protected]

The Disulfide Bonds in Glycoprotein E2 of Hepatitis C Virus Reveal the Tertiary Organization of the Molecule

Hepatitis C virus (HCV), a major cause of chronic liver disease in humans, is the focus of intense research efforts worldwide. Yet structural data on the viral envelope glycoproteins E1 and E2 are scarce, in spite of their essential role in the viral life cycle. To obtain more information, we developed an efficient production system of recombinant E2 ectodomain (E2e), truncated immediately upstream its trans-membrane (TM) region, using Drosophila melanogaster cells. This system yields a majority of monomeric protein, which can be readily separated chromatographically from contaminating disulfide-linked aggregates. The isolated monomeric E2e reacts with a number of conformation-sensitive monoclonal antibodies, binds the soluble CD81 large external loop and efficiently inhibits infection of Huh7.5 cells by infectious HCV particles (HCVcc) in a dose-dependent manner, suggesting that it adopts a native conformation. These properties of E2e led us to experimentally determine the connectivity of its 9 disulfide bonds, which are strictly conserved across HCV genotypes. Furthermore, circular dichroism combined with infrared spectroscopy analyses revealed the secondary structure contents of E2e, indicating in particular about 28% β-sheet, in agreement with the consensus secondary structure predictions. The disulfide connectivity pattern, together with data on the CD81 binding site and reported E2 deletion mutants, enabled the threading of the E2e polypeptide chain onto the structural template of class II fusion proteins of related flavi- and alphaviruses. The resulting model of the tertiary organization of E2 gives key information on the antigenicity determinants of the virus, maps the receptor binding site to the interface of domains I and III, and provides insight into the nature of a putative fusogenic conformational change