Cytokinesis in bloodstream stage Trypanosoma brucei requires a family of katanins and spastin

Microtubule severing enzymes regulate microtubule dynamics in a wide range of organisms and are implicated in important cell cycle processes such as mitotic spindle assembly and disassembly, chromosome movement and cytokinesis. Here we explore the function of several microtubule severing enzyme homologues, the katanins (KAT80, KAT60a, KAT60b and KAT60c), spastin (SPA) and fidgetin (FID) in the bloodstream stage of the African trypanosome parasite, Trypanosoma brucei. The trypanosome cytoskeleton is microtubule based and remains assembled throughout the cell cycle, necessitating its remodelling during cytokinesis. Using RNA interference to deplete individual proteins, we show that the trypanosome katanin and spastin homologues are non-redundant and essential for bloodstream form proliferation. Further, cell cycle analysis revealed that these proteins play essential but discrete roles in cytokinesis. The KAT60 proteins each appear to be important during the early stages of cytokinesis, while downregulation of KAT80 specifically inhibited furrow ingression and SPA depletion prevented completion of abscission. In contrast, RNA interference of FID did not result in any discernible effects. We propose that the stable microtubule cytoskeleton of T. brucei necessitates the coordinated action of a family of katanins and spastin to bring about the cytoskeletal remodelling necessary to complete cell divisio

Deficiency of FLCN in Mouse Kidney Led to Development of Polycystic Kidneys and Renal Neoplasia

The Birt–Hogg–Dubé (BHD) disease is a genetic cancer syndrome. The responsible gene, BHD, has been identified by positional cloning and thought to be a novel tumor suppressor gene. BHD mutations cause many types of diseases including renal cell carcinomas, fibrofolliculomas, spontaneous pneumothorax, lung cysts, and colonic polyps/cancers. By combining Gateway Technology with the Ksp-Cre gene knockout system, we have developed a kidney-specific BHD knockout mouse model. BHDflox/flox/Ksp-Cre mice developed enlarged kidneys characterized by polycystic kidneys, hyperplasia, and cystic renal cell carcinoma. The affected BHDflox/flox/Ksp-Cre mice died of renal failure at approximate three weeks of age, having blood urea nitrogen levels over tenfold higher than those of BHD flox/+/Ksp-Cre and wild-type littermate controls. We further demonstrated that these phenotypes were caused by inactivation of BHD and subsequent activation of the mTOR pathway. Application of rapamycin, which inhibits mTOR activity, to the affected mice led to extended survival and inhibited further progression of cystogenesis. These results provide a correlation of kidney-targeted gene inactivation with renal carcinoma, and they suggest that the BHD product FLCN, functioning as a cyst and tumor suppressor, like other hamartoma syndrome–related proteins such as PTEN, LKB1, and TSC1/2, is a component of the mTOR pathway, constituting a novel FLCN-mTOR signaling branch that regulates cell growth/proliferation

Bioassay guided purification of the antimicrobial fraction of a Brazilian propolis from Bahia state

<p>Abstract</p> <p>Background</p> <p>Brazilian propolis type 6 (Atlantic forest, Bahia) is distinct from the other types of propolis especially due to absence of flavonoids and presence of other non-polar, long chain compounds, but presenting good <it>in vitro </it>and <it>in vivo </it>antimicrobial activity. Several authors have suggested that fatty acids found in this propolis might be responsible for its antimicrobial activity; however, so far no evidence concerning this finding has been reported in the literature. The goals of this study were to evaluate the antibacterial activity of the main pure fatty acids in the ethanolic extract and fractions and elucidate the chemical nature of the bioactive compounds isolated from Brazilian propolis type 6.</p> <p>Methods</p> <p>Brazilian propolis type 6 ethanolic extract (EEP), hexane fraction (H-Fr), major fatty acids, and isolated sub-fractions were analyzed using high performance liquid chromatography (HPLC), high resolution gas chromatography with flame ionization detection (HRGC-FID), and gas chromatography-mass spectrometry (GC-MS). Three sub-fractions of H-Fr were obtained through preparative HPLC. Antimicrobial activity of EEP, H-Fr, sub-fractions, and fatty acids were tested against <it>Staphyloccus aureus </it>ATCC 25923 and <it>Streptococcus mutans </it>Ingbritt 1600 using minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC).</p> <p>Results</p> <p>EEP and H-Fr inhibited the growth of the microorganisms tested; nevertheless, no antimicrobial activity was found for the major fatty acids. The three sub-fractions (1, 2, and 3) were isolated from H-Fr by preparative HPLC and only sub-fraction 1 showed antimicrobial activity.</p> <p>Conclusion</p> <p>a) The major fatty acids tested were not responsible for the antimicrobial activity of propolis type 6; b) Sub-fraction 1, belonging to the benzophenone class, was responsible for the antimicrobial activity observed in the present study. The identification of the bioactive compound will improve the development of more efficient uses of this natural product.</p

Adenovirus-Vectored Drug-Vaccine Duo as a Rapid-Response Tool for Conferring Seamless Protection against Influenza

Few other diseases exert such a huge toll of suffering as influenza. We report here that intranasal (i.n.) administration of E1/E3-defective (ΔE1E3) adenovirus serotype 5 (Ad5) particles rapidly induced an anti-influenza state as a means of prophylactic therapy which persisted for several weeks in mice. By encoding an influenza virus (IFV) hemagglutinin (HA) HA1 domain, an Ad5-HA1 vector conferred rapid protection as a prophylactic drug followed by elicitation of sustained protective immunity as a vaccine for inducing seamless protection against influenza as a drug-vaccine duo (DVD) in a single package. Since Ad5 particles induce a complex web of host responses, which could arrest influenza by activating a specific arm of innate immunity to impede IFV growth in the airway, it is conceivable that this multi-pronged influenza DVD may escape the fate of drug resistance that impairs the current influenza drugs

Estimating the number of children exposed to parental psychiatric disorders through a national health survey

<p>Abstract</p> <p>Objective</p> <p>Children whose parents have psychiatric disorders experience an increased risk of developing psychiatric disorders, and have higher rates of developmental problems and mortality. Assessing the size of this population is important for planning of preventive strategies which target these children.</p> <p>Methods</p> <p>National survey data (CCHS 1.2) was used to estimate the number of children exposed to parental psychiatric disorders. Disorders were diagnosed using the World Psychiatric Health Composite International Diagnostic Interview (WMH-CIDI) (12 month prevalence). Data on the number of children below 12 years of age in the home, and the relationship of the respondents with the children, was used to estimate exposure. Parent-child relations were identified, as was single parenthood. Using a design-based analysis, the number of children exposed to parental psychiatric disorders was calculated.</p> <p>Results</p> <p>Almost 570,000 children under 12 live in households where the survey respondent met criteria for one or more mood, anxiety or substance use disorders in the previous 12 months, corresponding to 12.1% of Canadian children under the age of 12. Almost 3/4 of these children have parents that report receiving no mental health care in the 12 months preceding the survey. For 17% of all Canadian children under age 12, the individual experiencing a psychiatric disorder is the only parent in the household.</p> <p>Conclusion</p> <p>The high number of children exposed causes major concern and has important implications. Although these children will not necessarily experience adversities, they possess an elevated risk of accidents, mortality, and of developing psychiatric disorders. We expect these estimates will promote further research and stimulate discussion at both health policy and planning tables.</p

Tai Chi for treating knee osteoarthritis: Designing a long-term follow up randomized controlled trial

<p>Abstract</p> <p>Background</p> <p>Knee Osteoarthritis (KOA) is a major cause of pain and functional impairment among elders. Currently, there are neither feasible preventive intervention strategies nor effective medical remedies for the management of KOA. Tai Chi, an ancient Chinese mind-body exercise that is reported to enhance muscle function, balance and flexibility, and to reduce pain, depression and anxiety, may safely and effectively be used to treat KOA. However, current evidence is inconclusive. Our study examines the effects of a 12-week Tai Chi program compared with an attention control (wellness education and stretching) on pain, functional capacity, psychosocial variables, joint proprioception and health status in elderly people with KOA. The study will be completed by July 2009.</p> <p>Methods/Design</p> <p>Forty eligible patients, age > 55 yr, BMI ≤ 40 kg/m²with tibiofemoral osteoarthritis (American College of Rheumatology criteria) are identified and randomly allocated to either Tai Chi (10 modified forms from classical Yang style Tai Chi) or attention control (wellness education and stretching). The 60-minute intervention sessions take place twice weekly for 12 weeks. The study is conducted at an urban tertiary medical center in Boston, Massachusetts. The primary outcome measure is the Western Ontario and McMaster Universities (WOMAC) pain subscale at 12 weeks. Secondary outcomes include weekly WOMAC pain, function and stiffness scores, patient and physician global assessments, lower-extremity function, knee proprioception, depression, self-efficacy, social support, health-related quality of life, adherence and occurrence of adverse events after 12, 24 and 48 weeks.</p> <p>Discussion</p> <p>In this article, we present the challenges of designing a randomized controlled trial with long-term follow up. The challenges encountered in this design are: strategies for recruitment, avoidance of selection bias, the actual practice of Tai Chi, and the maximization of adherence/follow-up while conducting the clinical trial for the evaluation of the effectiveness of Tai Chi on KOA.</p> <p>Trial registration</p> <p>ClinicalTrials.gov identifier: NCT00362453</p

Three-Dimensional Geometric Analysis of Felid Limb Bone Allometry

Studies of bone allometry typically use simple measurements taken in a small number of locations per bone; often the midshaft diameter or joint surface area is compared to body mass or bone length. However, bones must fulfil multiple roles simultaneously with minimum cost to the animal while meeting the structural requirements imposed by behaviour and locomotion, and not exceeding its capacity for adaptation and repair. We use entire bone volumes from the forelimbs and hindlimbs of Felidae (cats) to investigate regional complexities in bone allometry.Computed tomographic (CT) images (16435 slices in 116 stacks) were made of 9 limb bones from each of 13 individuals of 9 feline species ranging in size from domestic cat (Felis catus) to tiger (Panthera tigris). Eleven geometric parameters were calculated for every CT slice and scaling exponents calculated at 5% increments along the entire length of each bone. Three-dimensional moments of inertia were calculated for each bone volume, and spherical radii were measured in the glenoid cavity, humeral head and femoral head. Allometry of the midshaft, moments of inertia and joint radii were determined. Allometry was highly variable and related to local bone function, with joint surfaces and muscle attachment sites generally showing stronger positive allometry than the midshaft.Examining whole bones revealed that bone allometry is strongly affected by regional variations in bone function, presumably through mechanical effects on bone modelling. Bone's phenotypic plasticity may be an advantage during rapid evolutionary divergence by allowing exploitation of the full size range that a morphotype can occupy. Felids show bone allometry rather than postural change across their size range, unlike similar-sized animals

Myosin IIA Modulates T Cell Receptor Transport and CasL Phosphorylation during Early Immunological Synapse Formation

Activation of T cell receptor (TCR) by antigens occurs in concert with an elaborate multi-scale spatial reorganization of proteins at the immunological synapse, the junction between a T cell and an antigen-presenting cell (APC). The directed movement of molecules, which intrinsically requires physical forces, is known to modulate biochemical signaling. It remains unclear, however, if mechanical forces exert any direct influence on the signaling cascades. We use T cells from AND transgenic mice expressing TCRs specific to the moth cytochrome c 88–103 peptide, and replace the APC with a synthetic supported lipid membrane. Through a series of high spatiotemporal molecular tracking studies in live T cells, we demonstrate that the molecular motor, non-muscle myosin IIA, transiently drives TCR transport during the first one to two minutes of immunological synapse formation. Myosin inhibition reduces calcium influx and colocalization of active ZAP-70 (zeta-chain associated protein kinase 70) with TCR, revealing an influence on signaling activity. More tellingly, its inhibition also significantly reduces phosphorylation of the mechanosensing protein CasL (Crk-associated substrate the lymphocyte type), raising the possibility of a direct mechanical mechanism of signal modulation involving CasL

Nasal Delivery of an Adenovirus-Based Vaccine Bypasses Pre-Existing Immunity to the Vaccine Carrier and Improves the Immune Response in Mice

Pre-existing immunity to human adenovirus serotype 5 (Ad5) is common in the general population. Bypassing pre-existing immunity could maximize Ad5 vaccine efficacy. Vaccination by the intramuscular (I.M.), nasal (I.N.) or oral (P.O.) route with Ad5 expressing Ebola Zaire glycoprotein (Ad5-ZGP) fully protected naïve mice against lethal challenge with Ebola. In the presence of pre-existing immunity, only mice vaccinated I.N. survived. The frequency of IFN-γ+ CD8+ T cells was reduced by 80% and by 15% in animals vaccinated by the I.M. and P.O. routes respectively. Neutralizing antibodies could not be detected in serum from either treatment group. Pre-existing immunity did not compromise the frequency of IFN-γ+ CD8+ T cells (3.9±1% naïve vs. 3.6±1% pre-existing immunity, PEI) nor anti-Ebola neutralizing antibody (NAB, 40±10 reciprocal dilution, both groups). The number of INF-γ+ CD8+ cells detected in bronchioalveolar lavage fluid (BAL) after I.N. immunization was not compromised by pre-existing immunity to Ad5 (146±14, naïve vs. 120±16 SFC/million MNCs, PEI). However, pre-existing immunity reduced NAB levels in BAL by ∼25% in this group. To improve the immune response after oral vaccination, the Ad5-based vaccine was PEGylated. Mice given the modified vaccine did not survive challenge and had reduced levels of IFN-γ+ CD8+ T cells 10 days after administration (0.3±0.3% PEG vs. 1.7±0.5% unmodified). PEGylation did increase NAB levels 2-fold. These results provide some insight about the degree of T and B cell mediated immunity necessary for protection against Ebola virus and suggest that modification of the virus capsid can influence the type of immune response elicited by an Ad5-based vaccine