Effect of pH on infectivity and morphology of ecotropic moloney murine leukemia virus

A number of factors affect the infectivity of retroviruses. The effect of pH on infectivity and morphology of ecotropic moloney murine leukemia virus (MoMuLV) was determined in this work. The ecotropic MoMuLVs were found to remain infectious at a narrow pH range from 5.5 to 8.0. Our experiments indicated that the viruses were inactivated swiftly at lower or higher pH. Within 5 min of exposure to pH 4 about 95% of the viruses lost infectiousness. The viruses were completely inactivated after exposure to pH < 3 or pH >11 for 5 min. The inactivation of MoMuLV was irreversible. Electron microscopy revealed that ecotropic MoMuLV remained round-shaped at pH between 7.0 and 5. They became irregular with a convex head at pH < 4. At pH 2, virtually all virion particles were penetrated by stains, causing the accumulation of heavy metals inside the particles. The penetration of heavy metal inside the particles indicated the disassociation of the lipid bilayer of the viruses at low pH. A FACS-based screening strategy for selecting high-titer retrovirus producing cell lines is also presented in this report.     

Weight lifting belt use patterns among a population of health club members

The purpose of this study was to identify patterns of weight belt use in a population of recreational weight trainers. We hypothesized that the majority of weight belt users utilized weight belts to reduce injury risk and/or improve performance while lifting submaximal loads. Three hundred fifty-two strength training health center members (189 men, 156 women, 7 gender not indicated) aged 20-72 years (36.5 +/- 10.5 years) completed a voluntary weight belt use survey. Overall, 27% (94/352) were weight belt users and 73% (258/352) were nonusers. Ninety percent (85/94) of belt users and 63% (55/88) of former belt users utilized belts to prevent injury, whereas 22% (21/94) of belt users and 28% (25/88) of former belt users utilized belts to improve performance. Many belt users were found to utilize belts during inappropriate situations such as lifting light loads or during exercises that do not typically stress the trunk musculature. Based on these findings we suggest that specific educational interventions be developed at health and fitness facilities to help provide a foundation for more informed decision-making regarding weight belt use.     

Comparison between linear and nonlinear in-season training programs in freshman football players

The purpose of this study was to compare linear (LT) with nonlinear (NL) in-season training programs in freshman football players during the course of 2 separate seasons. During the first year (n = 14, mean +/- SD = 177.3 +/- 4.8 cm, 88.0 +/- 9.7 kg), the LT program was employed 2 days per week. In the second year (n = 14, 175.0 +/- 7.1 cm, 94.2 +/- 20.5 kg), a 2 days per week LT was used. Subjects were tested for maximal strength in the squat (1 repetition maximum [1RM]) and bench press (1RM) exercises. A significant improvement in 1RM squat was seen in LT, but not in NL. No significant improvement in 1RM bench press was seen in either group. A significant difference between LT and NL was observed in Delta1RM squat (13.8 +/- 7.4 kg compared with 1.6 +/- 2.6 kg, respectively). Results of this study suggest that LT may be more effective in eliciting strength gains than NL in freshman football players during an in-season training program.     

Dynamics of herpes simplex virus capsid maturation visualized by time-lapse cryo-electron microscopy

The capsid of the herpes simplex virus initially assembles as a procapsid that matures through a massive conformational change of its 182 MDa surface shell. This transition, which stabilizes the fragile procapsid, is facilitated by the viral protease that releases the interaction between the shell and the underlying scaffold; however, protease-deficient procapsids mature slowly in vitro. To study procapsid maturation as a time-resolved process, we monitored this reaction by cryo-electron microscopy (cryo-EM). The resulting images were sorted into 17 distinct classes, and three-dimensional density maps were calculated for each. When arranged in a chronological series, these maps yielded molecular movies of procapsid maturation. A single major switching event takes place at stages 8-9, preceded by relatively subtle adjustments in the pattern of interactions and followed by similarly small 'aftershocks'. The primary mechanism underlying maturation is relative rotations of domains of VP5, the major capsid protein.     

The thrombomodulin-protein C system is essential for the maintenance of pregnancy

Disruption of the mouse gene encoding the blood coagulation inhibitor thrombomodulin (Thbd) leads to embryonic lethality caused by an unknown defect in the placenta. We show that the abortion of thrombomodulin-deficient embryos is caused by tissue factor-initiated activation of the blood coagulation cascade at the feto-maternal interface. Activated coagulation factors induce cell death and growth inhibition of placental trophoblast cells by two distinct mechanisms. The death of giant trophoblast cells is caused by conversion of the thrombin substrate fibrinogen to fibrin and subsequent formation of fibrin degradation products. In contrast, the growth arrest of trophoblast cells is not mediated by fibrin, but is a likely result of engagement of protease-activated receptors (PAR)-2 and PAR-4 by coagulation factors. These findings show a new function for the thrombomodulin-protein C system in controlling the growth and survival of trophoblast cells in the placenta. This function is essential for the maintenance of pregnancy.     

Engineering a mevalonate pathway in Escherichia coli for production of terpenoids

Isoprenoids are the most numerous and structurally diverse family of natural products. Terpenoids, a class of isoprenoids often isolated from plants, are used as commercial flavor and fragrance compounds and antimalarial or anticancer drugs. Because plant tissue extractions typically yield low terpenoid concentrations, we sought an alternative method to produce high-value terpenoid compounds, such as the antimalarial drug artemisinin, in a microbial host. We engineered the expression of a synthetic amorpha-4,11-diene synthase gene and the mevalonate isoprenoid pathway from Saccharomyces cerevisiae in Escherichia coli. Concentrations of amorphadiene, the sesquiterpene olefin precursor to artemisinin, reached 24 microg caryophyllene equivalent/ml. Because isopentenyl and dimethylallyl pyrophosphates are the universal precursors to all isoprenoids, the strains developed in this study can serve as platform hosts for the production of any terpenoid compound for which a terpene synthase gene is available.     

Vertebrate gene predictions and the problem of large genes

To find unknown protein-coding genes, annotation pipelines use a combination of ab initio gene prediction and similarity to experimentally confirmed genes or proteins. Here, we show that although the ab initio predictions have an intrinsically high false-positive rate, they also have a consistently low false-negative rate. The incorporation of similarity information is meant to reduce the false-positive rate, but in doing so it increases the false-negative rate. The crucial variable is gene size (including introns)--genes of the most extreme sizes, especially very large genes, are most likely to be incorrectly predicted.     

Convenient synthesis of 2'-deoxy-2-fluoroadenosine from 2-fluoroadenine

A convenient synthesis of 2'-deoxy-2-fluoroadenosine from commercially available 2-fluoroadenine is described. The coupling reaction of silylated 2-fluoroadenine with phenyl 3,5-bis[O-(t-butyldimethylsilyl)]-2-deoxy-1-thio-D-erythro-pentofuranoside gave the corresponding 2-fluoro-2'-deoxyadenosine derivative (alpha/beta = 1:1) in good yield. The alpha- and beta-anomers were separated by chromatography, and then desilylated to give compounds 1a and 1b.     

Preparation and characterization of novel coenzyme Q<Subscript>10</Subscript> nanoparticles engineered from microemulsion precursors

The purpose of these studies was to prepare and characterize nanoparticles into which Coenzyme Q₁₀ (CoQ₁₀) had been incorporated (CoQ₁₀-NPs) using a simple and potentially scalable method. CoQ₁₀-NPs were prepared by cooling warm microemulsion precursors composed of emulsifying wax, CoQ₁₀, Brij 78, and/or Tween 20. The nanoparticles were lyophilized, and the stability of CoQ₁₀-NPs in both lyophilized form and aqueous suspension was monitored over 7 days. The release of CoQ₁₀ from the nanoparticles was investigated at 37°C. Finally, an in vitro study of the uptake of CoQ₁₀-NPs by mouse macrophage, J774A.1, was completed. The incorporation efficiency of CoQ₁₀ was approximately 74%±5%. Differential Scanning Calorimetry (DSC) showed that the nanoparticle was not a physical mixture of its individual components. The size of the nanoparticles increased over time if stored in aqueous suspension. However, enhanced stability was observed when the nanoparticles were stored at 4°C. Storage in lyophilized form demonstrated the highest stability. The in vitro release profile of CoQ₁₀ from the nanoparticles showed an initial period of rapid release in the first 9 hours followed by a period of slower and extended release. The uptake of CoQ₁₀-NPs by the J774A.1 cells was over 4-fold higher than that of the CoQ₁₀-free nanoparticles (P<.05). In conclusion, CoQ₁₀-NPs with potential application for oral CoQ₁₀ delivery were engineered readily from microemulsion precursors.