MUSHROOM BODY INFLUENCE ON LOCOMOTOR ACTIVITY AND CIRCADIAN RHYTHMS IN DROSOPHILA MELANOGASTER

Whether or not mechanisms underlying circadian locomotor rhythms and learning are related anatomically through the mushroom bodies (MBs) was investigated by monitoring behavioral rhythmicity in flies with MB lesions induced by chemical ablation and by mutations in five different genes. All flies tested were later examined histologically to assess (1) MB neuroanatomy, and (2) the condition of the putative pacemaker cells -- the ventral Lateral Neurons (LN v s) and their terminals that project to the vicinity of the MB calyces. All groups of flies had normal rhythms except for mushroom body miniature ( mbm ; only in a wild-type Berlin genetic background) and mushroom body defect ( mud ). MB ablation had no effect on the gender-specific differences in the rhythmic activity profile that are typical of wild-type flies. However, ablated males had a slightly longer period than untreated males and were more active under constant dark conditions. LN v s and their arborization patterns appeared normal in MB-ablated and in most mutant flies. Activity defects of mbm flies were attributed to genetic background rather than to the mutation alone. Misrouted LN v projections and ～14% arrhythmia of mud flies were uncorrelated and attributed to pleiotropy rather than to specific effects of MB lesions. Our results imply that MBs are not involved in circadian activity rhythms but that they do have an inhibitory effect on activity levels of male flies.     

Clinical features and outcome of T-cell acute lymphoblastic leukemia in childhood with respect to alterations at the TAL1 locus: a Pediatric Oncology Group study

Alteration of the TAL1 locus is the most common nonrandom genetic defect in childhood T-cell acute lymphoblastic leukemia (T-ALL). To determine if rearrangements of the TAL1 proto-oncogene confer a distinct leukemic phenotype, we studied leukemic peripheral blood or bone marrow samples from 182 children with newly diagnosed T-ALL enrolled on Pediatric Oncology Group treatment protocols. Forty-eight (26%) of the samples had a local rearrangement of the TAL1 locus. Demographic and clinical features were compared for patient subgroups with and without TAL1 rearrangements. The only clinical correlates that were significantly associated with TAL1 gene rearrangements were higher white blood cell count (P = .017) and higher hemoglobin (P = .007) at diagnosis. Immunophenotypically, samples with altered TAL1 were more likely to be CD2+ (P = .001) and lack CD10 (cALLa) expression (P = .007) than those without the rearrangement. There was a trend toward improved event-free survival (EFS) in patients with TAL1 rearrangements (4-year EFS was 44% +/- 7% for patients without the rearrangements v 59% +/- 11% for those with rearrangements), but the difference was not significant (P = .34). The role of TAL1 in leukemogenesis has yet to be clearly defined, and the prognostic significance of TAL1 gene rearrangements in T-ALL deserves further study.     

Phenotype diversity in type 1 Gaucher disease: discovering the genetic basis of Gaucher disease/hematologic malignancy phenotype by individual genome analysis

Gaucher disease (GD), an inherited macrophage glycosphingolipidosis, manifests with an extraordinary variety of phenotypes that show imperfect correlation with mutations in the GBA gene. In addition to the classic manifestations, patients suffer from increased susceptibility to hematologic and nonhematologic malignancies. The mechanism(s) underlying malignancy in GD is not known, but is postulated to be secondary to macrophage dysfunction and immune dysregulation arising from lysosomal accumulation of glucocerebroside. However, there is weak correlation between GD/cancer phenotype and the systemic burden of glucocerebroside-laden macrophages. Therefore, we hypothesized that genetic modifier(s) may underlie the GD/cancer phenotype. In the present study, the genetic basis of GD/T-cell acute lymphoblastic lymphoma in 2 affected siblings was deciphered through genomic analysis. GBA gene sequencing revealed homozygosity for a novel mutation, D137N. Whole-exome capture and massively parallel sequencing combined with homozygosity mapping identified a homozygous novel mutation in the MSH6 gene that leads to constitutional mismatch repair deficiency syndrome and increased cancer risk. Enzyme studies demonstrated that the D137N mutation in GBA is a pathogenic mutation, and immunohistochemistry confirmed the absence of the MSH6 protein. Therefore, precise phenotype annotation followed by individual genome analysis has the potential to identify genetic modifiers of GD, facilitate personalized management, and provide novel insights into disease pathophysiology.     

Comparison of the hepatitis B virus core, surface and polymerase gene substitution rates in chronically infected patients

For phylogenetic comparison of hepatitis B virus (HBV) isolates, often a region of the HBV surface gene is analysed. Because the surface gene completely overlaps the polymerase gene, its evolution is constrained, and it may not be the best choice for genetic comparison of HBV isolates. Analysing serial sample pairs of 33 chronically HBV-infected, untreated patients, with a cumulative follow-up of 184 years, the synonymous and nonsynonymous substitution rates of a part of the overlapping HBV surface and polymerase genes were compared to those of a nonoverlapping part of the HBV core gene. The substitution rate of the HBV core gene was higher (8.15 × 10(-4) vs 4.57 × 10(-4) substitutions/site/year) than that of the surface gene. The difference was mainly due to a significantly lower synonymous substitution rate in the surface gene, with dN/dS ratios of 0.412 in the core gene and 0.986 in the surface gene. Contrary to the core gene, the number of substitutions in the surface gene was higher in low viraemic hosts, who control HBV infection by suppressing replication. The number of substitutions in the core gene correlated more strongly with the duration of follow-up. The overlapping HBV surface and polymerase genes experience strong negative selection, which limits the number of substitutions. Because the HBV core gene reflects the duration of infection more accurately, it is more suitable for the analysis of short-term viral evolution and of hepatitis B transmission chains.     

Nephrotoxicity and Kidney Transport Assessment on 3D Perfused Proximal Tubules

Proximal tubules in the kidney play a crucial role in reabsorbing and eliminating substrates from the body into the urine, leading to high local concentrations of xenobiotics. This makes the proximal tubule a major target for drug toxicity that needs to be evaluated during the drug development process. Here, we describe an advanced in vitro model consisting of fully polarized renal proximal tubular epithelial cells cultured in a microfluidic system. Up to 40 leak-tight tubules were cultured on this platform that provides access to the basolateral as well as the apical side of the epithelial cells. Exposure to the nephrotoxicant cisplatin caused a dose-dependent disruption of the epithelial barrier, a decrease in viability, an increase in effluent LDH activity, and changes in expression of tight-junction marker zona-occludence 1, actin, and DNA-damage marker H2A.X, as detected by immunostaining. Activity and inhibition of the efflux pumps P-glycoprotein (P-gp) and multidrug resistance protein (MRP) were demonstrated using fluorescence-based transporter assays. In addition, the transepithelial transport function from the basolateral to the apical side of the proximal tubule was studied. The apparent permeability of the fluorescent P-gp substrate rhodamine 123 was decreased by 35% by co-incubation with cyclosporin A. Furthermore, the activity of the glucose transporter SGLT2 was demonstrated using the fluorescent glucose analog 6-NBDG which was sensitive to inhibition by phlorizin. Our results demonstrate that we developed a functional 3D perfused proximal tubule model with advanced renal epithelial characteristics that can be used for drug screening studies.     

A double mobility acetabular implant for primary hip arthroplasty in patients at high risk of dislocation

IntroductionDislocation following total hip replacement continues to be a problem for which no completely satisfactory solution has been found. Several methods have been proposed to reduce the incidence of hip dislocations with varying degrees of success, including elevated rim liners, constrained liners and large diameter bearings. We present our experience with the double mobility acetabular component in patients at high risk of instability.MethodsThis was a retrospective review of 65 primary total hip arthroplasties in 55 patients (15 men, 40 women), performed between October 2005 and November 2009. The majority (80%) of patients had at least two and 26% had at least three risk factors for instability. The mean age was 76 years (range: 44–92 years). The patients were followed up for a mean duration of 60 months (range: 36–85 months).ResultsFourteen patients died and one was lost to follow-up, leaving fifty hips for final assessment. Until the final follow-up appointment, no patients had dislocation and none required revision surgery. The mean Oxford hip score improved from 45.0 to 26.5 (p<0.0001). The mean Merle d’Aubigné pain score improved from 1.4 to 4.9 (p<0.0001), the walking score from 2.3 to 3.1 (p<0.07) and the absolute hip function score from 5.4 to 10.8 (p<0.0001). There were no clinical or radiographic signs of loosening.ConclusionsThe double mobility acetabular component was successful at preventing dislocation during early to medium-term follow-up. However, as data are still lacking with regard to polyethylene wear rates at the additional bearing surface, it would be prudent to restrict the use of this implant to selected patients at high risk of instability.