The effect of tobacco and marijuana use on dental health status in Nevada adolescents: a trend analysis.
Tobacco use is the leading cause of preventable death worldwide. If current trends persist, tobacco will kill more than 8 million people worldwide by 2030 and 1 billion by the end of the century. The purpose of this study was to determine trends in tobacco/marijuana use in Nevada adolescents and their effect on dental health status. Relative comparative data were compared with nationally reported data. Retrospective data in this cohort study was from an ongoing statewide, school-based, dental health screening initiative that was conducted across 8 years (2002-2010) in public/private middle/high schools in Nevada. A total of 66,941 dental health screenings of adolescents between ages 13-18 were conducted. Self-reported data were collected on tobacco/marijuana use. Descriptive statistics and trends were reported. Means (SE) were computed for caries prevalence and severity. Effect size was reported on dental caries and use of tobacco/marijuana. Overall, percentage prevalence of tobacco use was approximately the same as the national average; however, there were significantly higher rates of marijuana use (12.0% vs. 3.3%). Prevalence and severity of dental caries was significantly higher in those who used tobacco/marijuana than those who did not across all variables and across all 8 years controlling for gender, race/ethnicity, where they lived, and exposure to secondhand smoke. Tobacco use negatively affected dental health status with marijuana having the largest negative effect. The findings from this study identified the need for tobacco/marijuana prevention services targeting adolescents residing in the geographic areas most at risk.


Sequestration of Mycobacterium tuberculosis in tight vacuoles in vivo in lung macrophages of mice infected by the respiratory route.
Following aerosol infection of mice with Mycobacterium tuberculosis, single mycobacteria or pairs of bacilli were observed within individual phagocytic vacuoles bound by tightly apposed vacuolar membranes. The virulent organism was not observed free in the cytoplasm of the parasitized cells or in the extracellular space of the lung granulomata. This study indicates that in vivo, virulent mycobacteria survive and probably replicate within a unique tight vacuole in the infected phagocyte within the lung.


Inhibitors of the AAA+ chaperone p97.
It is remarkable that a pathway as ubiquitous as protein quality control can be targeted to treat cancer. Bortezomib, an inhibitor of the proteasome, was first approved by the US Food and Drug Administration (FDA) more than 10 years ago to treat refractory myeloma and later extended to lymphoma. Its use has increased the survival rate of myeloma patients by as much as three years. This success was followed with the recent accelerated approval of the natural product derived proteasome inhibitor carfilzomib (Kyprolis®), which is used to treat patients with bortezomib-resistant multiple myeloma. The success of these two drugs has validated protein quality control as a viable target to fight select cancers, but begs the question why are proteasome inhibitors limited to lymphoma and myeloma? More recently, these limitations have encouraged the search for additional targets within the protein quality control system that might offer heightened cancer cell specificity, enhanced clinical utility, a lower rate of resistance, reduced toxicity, and mitigated side effects. One promising target is p97, an ATPase associated with various cellular activities (AAA+) chaperone. p97 figures prominently in protein quality control as well as serving a variety of other cellular functions associated with cancer. More than a decade ago, it was determined that up-regulation of p97 in many forms of cancer correlates with a poor clinical outcome. Since these initial discoveries, a mechanistic explanation for this observation has been partially illuminated, but details are lacking. Understandably, given this clinical correlation, myriad roles within the cell, and its importance in protein quality control, p97 has emerged as a potential therapeutic target. This review provides an overview of efforts towards the discovery of small molecule inhibitors of p97, offering a synopsis of efforts that parallel the excellent reviews that currently exist on p97 structure, function, and physiology.


Untouchability of natural spa waters: Perspectives for treatments within a personalized water safety plan.
Natural SPA waters and their environments were known since ancient times and used for health or recreational purposes in different societies, worldwide. The composition and uses of these spring waters may not allow standard disinfection in pools, representing a challenge for hygiene management. Several safety and quality procedures were proposed, but a systematic approach is still needed. Here, we focus on alternative strategies to provide hints for developing a sustainable Water Safety Plan, based on intrinsic water properties and photocatalytic materials. The antimicrobial activity of four different SPA waters with high mineral content and one drinkable spring water with a low mineral content, was assessed and then tested for the additional bactericidal activity of Titanium Dioxide (TiO2) nanomaterials and/or light exposure at different wavelengths (200-635 nm). A native antibacterial activity was observed in all high mineral content waters, with a CFU reduction of 75-80%. The bactericidal action of TiO2 showed an additional incremental effect, with a reduction of over 99% within 2-5 h. Interestingly, the antibacterial photocatalytic effect was detected also in the visible light range, with a possible pick around 450-455 nm, blue-light. Based on observed results, we propose a model for developing a water safety plan, considering water properties and bather exposure. This candidate approach is personalized on water composition and pool use, trying to avoid chemical disinfectants. Photocatalytic nanotechnologies represent one of the promising alternative treatments and can provide novel perspectives for a sustainable managing of natural SPA water hygiene.


Clustering of main orthologs for multiple genomes.
The identification of orthologous genes shared by multiple genomes is critical for both functional and evolutionary studies in comparative genomics. While it is usually done by sequence similarity search and reconciled tree construction in practice, recently a new combinatorial approach and high-throughput system MSOAR for ortholog identification between closely related genomes based on genome rearrangement and gene duplication has been proposed in Fu et al. MSOAR assumes that orthologous genes correspond to each other in the most parsimonious evolutionary scenario, minimizing the number of genome rearrangement and (postspeciation) gene duplication events. However, the parsimony approach used by MSOAR limits it to pairwise genome comparisons. In this paper, we extend MSOAR to multiple (closely related) genomes and propose an ortholog clustering method, called MultiMSOAR, to infer main orthologs in multiple genomes. As a preliminary experiment, we apply MultiMSOAR to rat, mouse, and human genomes, and validate our results using gene annotations and gene function classifications in the public databases. We further compare our results to the ortholog clusters predicted by MultiParanoid, which is an extension of the well-known program InParanoid for pairwise genome comparisons. The comparison reveals that MultiMSOAR gives more detailed and accurate orthology information, since it can effectively distinguish main orthologs from inparalogs.


Distinct Life Histories Impact Dikaryotic Genome Evolution in the Rust Fungus Puccinia striiformis Causing Stripe Rust in Wheat.
Stripe rust of wheat, caused by the obligate biotrophic fungus Puccinia striiformis f.sp. tritici, is a major threat to wheat production worldwide with an estimated yearly loss of US $1 billion. The recent advances in long-read sequencing technologies and tailored-assembly algorithms enabled us to disentangle the two haploid genomes of Pst. This provides us with haplotype-specific information at a whole-genome level. Exploiting this novel information, we perform whole-genome comparative genomics of two P. striiformis f.sp. tritici isolates with contrasting life histories. We compare one isolate of the old European lineage (PstS0), which has been asexual for over 50 years, and a Warrior isolate (PstS7 lineage) from a novel incursion into Europe in 2011 from a sexual population in the Himalayan region. This comparison provides evidence that long-term asexual evolution leads to genome expansion, accumulation of transposable elements, and increased heterozygosity at the single nucleotide, structural, and allele levels. At the whole-genome level, candidate effectors are not compartmentalized and do not exhibit reduced levels of synteny. Yet we were able to identify two subsets of candidate effector populations. About 70% of candidate effectors are invariant between the two isolates, whereas 30% are hypervariable. The latter might be involved in host adaptation on wheat and explain the different phenotypes of the two isolates. Overall, this detailed comparative analysis of two haplotype-aware assemblies of P. striiformis f.sp. tritici is the first step in understanding the evolution of dikaryotic rust fungi at a whole-genome level.


The epidemiology of epidemic parotitis in the USSR.
Morbidity from epidemic parotitis in the USSR in the period from 1958 to 1972 ranged within the limits of 266.6 and 521.7 cases per 100 000 inhabitants. A 3--4 years cyclic recurrence of rises and falls in morbidity was observed. The bulk of cases of disease occur in the winter-spring period. In towns, parotitis is registered 3 times more frequently, in the mean, than in rural districts. More than 95% of patients in the USSR consisted of children under 15, while in Moscow, children aged 3--5 years were the most affected group. In Moscow in the period from February 1, 1972 to January 31, 1973, the morbidity in men was 493.2 per 100 000 and in women 339.5 per 100 000 (ratio 1:1.45).


Impact of combination therapies on HIV risk perceptions and sexual risk among HIV-positive and HIV-negative gay and bisexual men.
The availability of improved HIV treatments may prompt reduced concern about HIV and sexual risk. Gay and bisexual men (N = 554, 17% HIV-positive) completed measures of treatment attitudes, sexual risk, and assumptions regarding the infectiousness of sexual partners. A substantial minority reported reduced HIV concern related to treatment advances. Reduced HIV concern was an independent predictor of sexual risk, particularly among HIV-positive men. In response to hypothetical scenarios describing sex with an HIV-positive partner, participants rated the risk of unprotected sex to be lower if the partner was taking combination treatments and had an undetectable viral load, relative to scenarios with a seropositive partner not taking combination treatments. Prevention efforts must address attitudinal shifts prompted by recent treatment successes, stressing the continued importance of safer sex, and that an undetectable viral load does not eliminate infection risks.


Inhibition of rotavirus replication by downregulation of fatty acid synthesis.
Recently the recruitment of lipid droplets (LDs) to sites of rotavirus (RV) replication was reported. LDs are polymorphic organelles that store triacylglycerols, cholesterol and cholesterol esters. The neutral fats are derived from palmitoyl-CoA, synthesized via the fatty acid biosynthetic pathway. RV-infected cells were treated with chemical inhibitors of the fatty acid biosynthetic pathway, and the effects on viral replication kinetics were assessed. Treatment with compound C75, an inhibitor of the fatty acid synthase enzyme complex (FASN), reduced RV infectivity 3.2-fold (P = 0.07) and modestly reduced viral RNA synthesis (1.2-fold). Acting earlier in the fatty acid synthesis pathway, TOFA [5-(Tetradecyloxy)-2-furoic acid] inhibits the enzyme acetyl-CoA carboxylase 1 (ACC1). TOFA reduced the infectivity of progeny RV 31-fold and viral RNA production 6-fold. The effect of TOFA on RV infectivity and RNA replication was dose-dependent, and infectivity was reduced by administering TOFA up to 4 h post-infection. Co-treatment of RV-infected cells with C75 and TOFA synergistically reduced viral infectivity. Knockdown by siRNA of FASN and ACC1 produced findings similar to those observed by inhibiting these proteins with the chemical compounds. Inhibition of fatty acid synthesis using a range of approaches uniformly had a more marked impact on viral infectivity than on viral RNA yield, inferring a role for LDs in virus assembly and/or egress. Specific inhibitors of fatty acid metabolism may help pinpoint the critical structural and biochemical features of LDs that are essential for RV replication, and facilitate the development of antiviral therapies.


Severity of Staphylococcus aureus infection of the skin is associated with inducibility of human beta-defensin 3 but not human beta-defensin 2.
Gram-positive bacteria are the predominant cause of skin infections. Antimicrobial peptides (AMPs) are believed to be of major importance in skin's innate defense against these pathogens. This study aimed at providing clinical evidence for the contribution of AMP inducibility to determining the severity of Gram-positive skin infection. Using real-time PCR, we determined the induction of human beta-defensin 2 (HBD-2), HBD-3, and RNase 7 by comparing healthy and lesional mRNA levels in 32 patients with Gram-positive skin infection. We then examined whether AMP induction differed by disease severity, as measured by number of recurrences and need for surgical drainage in patients with Staphylococcus aureus-positive lesions. We found that HBD-2 and -3, but not RNase 7, mRNA expression was highly induced by Gram-positive bacterial infection in otherwise healthy skin. Less induction of HBD-3, but not HBD-2, was associated with more-severe S. aureus skin infection: HBD-3 mRNA levels were 11.4 times lower in patients with more than 6 recurrences (P = 0.01) and 8.8 times lower in patients reporting surgical drainage (P = 0.01) than in the respective baseline groups. This suggests that inducibility of HBD-3 influences the severity of Gram-positive skin infection in vivo. The physiological function of HBD-2 induction in this context remains unclear.


Recent Progress on the Fabrication and Properties of Silver Nanowire-Based Transparent Electrodes.
Transparent electrodes (TEs) made of metallic nanowires, such as Ag, Au, Cu, and Ni, are attracting increasing attention for several reasons: (1) they can act as a substitute for tin oxide-based TEs such as indium-tin oxide (ITO) and fluorine-doped tin oxide (FTO); (2) various methods exist for fabricating such TEs such as filtration, spraying, and Meyer bar coating; (3) greater compatibility with different substrates can be achieved due to the variety of fabrication methods; and (4) extra functions in addition to serving as electrodes, such as catalytic abilities, can be obtained due to the metals of which the TEs are composed. There are a large number of applications for TEs, ranging from electronics and sensors to biomedical devices. This short review is a summary of recent progress, mainly over the past five years, on silver nanowire-based TEs. The focus of the review is on theory development, mechanical, chemical, and thermal stability as well as optical properties. The many applications of TEs are outside the scope of this review.


Comparative pathogenicity of Actinomyces species in mice.
The comparative pathogenicity of different species of Actinomyces was studied in a susceptible weanling-mouse model. After the intraperitoneal injection of strains of Actinomyces israeli, A. naeslundi, A. viscosus and Arachnia propionica, numerous abscesses developed in the intestine, mesentery, liver, and at the site of injection. Lesions were not produced by A. odontolyticus. A. naeslundi and A. viscosus produced acute lesions that resolved after a few weeks. Abscesses produced by rough strains of A. israeli and Arach. propionica persisted and led to a slowly progressive chronic infection. Viable organisms were always recovered from the lesions. Spread of the lesions by extension into other areas, including the thoracic cavity, led to the death of the animal after approximately 1 year. This study demonstrated a clear difference in the pattern of infection produced by the different species of Actinomyces as well as Arach. propionica.


Inactivation of monazomycin-induced voltage-dependent conductance in thin lipid membranes. I. Inactivation produced by long chain quaternary ammonium ions.
The voltage-dependent conductance induced in thin lipid membranes by monazomycin undergoes inactivation upon the introduction of quaternary ammonium ions (QA) having a long alkyl chain (e.g. dodecyltrimethylammonium [C12]) to the side containing monazomycin. That is, in response to a step of voltage the conductance rises to a peak and then falls to a much lower steady-state value. We demonstrate that the basis of this phenomenon is the ability of QA to pass through the stimulated membrane and bind to the opposite surface. As a consequence, the surface potential on that side becomes more positive, thus reducing the voltage across the membrane proper and turning off the monazomycin-induced conductance. Because the flux of QA through the membrane increases linearly with conductance, we believe that these ions pass through the monazomycin channels. QA permeability increases with alkyl chain length; remarkably, in spite of its much larger size, C12 is about 150 times more permeant than K+. It appears, therefore, that there is a hydrophobic region of the cahnnel that favors the alkyl chain; we propose that this region is formed by the hydrophobic faces of the monazomycin channels in lipid bilayers to QA inactivation of potassium channels in the squid giant azon, and suggest that there may be a common structural feature for the two channels. It is possible that some of the inactivation phenomena in excitable cells may arise from local field changes not measurable by the recording electrodes.


Herbal medicines for treating HIV infection and AIDS.
HIV-infected people and AIDS patients often seek complementary therapies including herbal medicines due to reasons such as unsatisfactory effects, high cost, non-availability, or adverse effects of conventional medicines. To assess beneficial effects and risks of herbal medicines in patients with HIV infection and AIDS. Electronic searches included the Cochrane Central Register of Controlled Trials, MEDLINE, EMBASE, LILACS, Science Citation Index, the Chinese Biomedical Database, TCMLARS; plus CISCOM, AMED, and NAPRALERT; combined with manual searches. The search ended in December 2004. Randomized clinical trials on herbal medicines compared with no intervention, placebo, or antiretroviral drugs in patients with HIV infection, HIV-related disease, or AIDS. The outcomes included mortality, HIV disease progression, new AIDS-defining event, CD4 cell counts, viral load, psychological status, quality of life, and adverse effects. Two authors extracted data independently and assessed the methodological quality of trials according to randomization, allocation concealment, double blinding, and drop-out. Nine randomized placebo-controlled trials involving 499 individuals with HIV infection and AIDS met the inclusion criteria. Methodological quality of trials was assessed as adequate in five full publications and unclear in other trials. Eight different herbal medicines were tested.A compound of Chinese herbs (IGM-1) showed significantly better effect than placebo in improvement of health-related quality of life in 30 symptomatic HIV-infected patients (WMD 0.66, 95% CI 0.05 to 1.27). IGM-1 appeared not to affect overall health perception, symptom severity, CD4 count, anxiety or depression (Burack 1996a). An herbal formulation of 35 Chinese herbs did not affect CD4 cell counts, viral load, AIDS events, symptoms, psychosocial measure, or quality of life (Weber 1999). There was no statistical difference between SPV30 and placebo in new AIDS-defining events, CD4 cell counts, or viral load (Durant 1998) although an earlier pilot trial showed positive effect of SPV30 on CD4 cell count (Durant 1997). Combined treatment of Chinese herbal compound SH and antiretroviral agents showed increased antiviral benefit compared with antiretrovirals alone (Sangkitporn 2004). SP-303 appeared to reduce stool weight (p = 0.008) and abnormal stool frequency (p = 0.04) in 51 patients with AIDS and diarrhoea (Holodniy 1999). Qiankunning appeared not to affect HIV-1 RNA levels (Shi 2003), Curcumin ineffective in reducing viral load or improving CD4 cell counts (Hellinger 1996), and Capsaicin ineffective in relieving pain associated with HIV-related peripheral neuropathy (Paice 2000). The occurrence of adverse effects was higher in the 35 Chinese herbs preparation (19/24) than in placebo (11/29) (79% versus 38%, p = 0.003) (Weber 1999). Qiankunning was associated with stomach discomfort and diarrhoea (Shi 2003). There is insufficient evidence to support the use of herbal medicines in HIV-infected individuals and AIDS patients. Potential beneficial effects need to be confirmed in large, rigorous trials.


The DUF81 protein TauE in Cupriavidus necator H16, a sulfite exporter in the metabolism of C2 sulfonates.
The degradation of taurine, isethionate and sulfoacetate in Cupriavidus necator (Ralstonia eutropha) H16 was shown by enzyme assays to be inducible, and each pathway involved sulfoacetaldehyde, which was subject to phosphatolysis by a common sulfoacetaldehyde acetyltransferase (Xsc, H16_B1870) to yield acetyl phosphate and sulfite. The neighbouring genes encoded phosphate acetyltransferase (Pta, H16_B1871) and a hypothetical protein [domain of unknown function (DUF)81, H16_B1872], with eight derived transmembrane helices. RT-PCR showed inducible transcription of these three genes, and led to the hypothesis that H16_B1872 and orthologous proteins represent a sulfite exporter, which was named TauE.


Outcomes of open and endovascular lower extremity revascularization in active smokers with advanced peripheral arterial disease.
Concern over perioperative and long-term durability of lower extremity revascularizations among active smokers is a frequent deterrent for vascular surgeons to perform elective lower extremity revascularization. In this study, we examined perioperative outcomes of lower extremity endovascular (LEE) revascularization and open lower extremity bypass (LEB) in active smokers with intermittent claudication (IC) and critical limb ischemia (CLI). Active smokers undergoing LEE or LEB from 2011 to 2014 were identified in the American College of Surgeons (ACS) National Surgical Quality Improvement Program (NSQIP) targeted vascular data set. Patient demographics, comorbidities, anatomic features, and perioperative outcomes were compared between LEE and LEB procedures. Subgroup analysis was performed for patients undergoing revascularization for IC and CLI independently. From 2011 to 2014, 4706 lower extremity revascularizations were performed in active smokers (37% of all revascularizations). In this group, 1497 were LEE (55.6% for CLI, 13.4% for below-knee pathology) and 3209 were LEB (68.9% CLI, 34.7% below-knee). Patients undergoing LEE had higher rates of female gender, hypertension, end-stage renal disease, and diabetes (all P ≤ .02). LEE patients also had a higher frequency of prior percutaneous interventions (22.7% vs 17.2%; P < .01) and preoperative antiplatelet therapy (82.3% vs 78.7%; P = .02). On risk-adjusted multivariate analysis, LEE patients had higher need for reintervention on the treated arterial segment than LEB (5.1% vs 5.2%; odds ratio [OR], 1.52; 95% confidence interval [CI], 1.08-2.13; P = .02) but had lower wound complications (3.1% vs 13.2%; OR, 0.32; 95% CI, 0.23-0.45; P < .01) and no statistically significant difference in 30-day mortality (0.6% vs 0.9%), myocardial infarction or stroke (1.1% vs 2.6%), or major amputation (3.2% vs 2.1%) in the overall cohort of active smokers. In the IC subgroup, myocardial infarction or stroke was significantly higher in the LEB group (1.9% vs 0.6%; OR, 1.83; 95% CI, 1.17-1.97; P = .03), although no difference was found in the CLI subgroup (2.8% vs 1.4%; OR, 0.75; 95% CI, 0.37-1.52; P = .42,). Also in IC group, there was a trend for lower major amputation rates ≤30 days in the LEE group, whereas in the CLI group, LEE had a trend toward higher risk of early amputation compared with LEB. In active smokers, LEB for IC and CLI requires fewer reinterventions but is associated with a higher rate of postoperative wound complications compared with LEE revascularization. However, the risk for limb amputation is higher in actively smoking patients when treated by LEE compared with LEB for CLI. Importantly, cardiovascular complications are significantly higher in actively smoking patients with IC undergoing LEB compared with LEE. This additional cardiovascular risk should be carefully weighed when proposing LEB for actively smoking patients with nonlimb-threatening IC.


[Nphe(1)]NC(1-13)NH(2) selectively antagonizes nociceptin/orphanin FQ-stimulated G-protein activation in rat brain.
[Phe(1)psi(CH(2)-NH)Gly(2)]noc/OFQ(1-13)-amide ([F/G]NC(1-13)NH(2)) and acetyl-RYYRIK-amide (Ac-RYYRIK-NH(2)), two peptidic ligands of the nociceptin/orphanin FQ (noc/OFQ) receptor, have been shown to exert both agonist and antagonist activity in different in vitro and in vivo systems. This is despite the observation that both peptides competitively antagonized the coupling of the activated receptor to G-proteins in brain preparations, measured in GTPgamma(35)S binding assays. In this study, [Nphe(1)]NC(1-13)-amide ([Nphe(1)]NC(1-13)NH(2)), a new noc/OFQ analog recently characterized as a pure and selective noc/OFQ receptor antagonist in several in vitro and in vivo assay systems, was shown to competitively inhibit the noc/OFQ-stimulated GTPgamma(35)S binding to rat cerebral cortex membranes with pA(2) of 7.76 (Schild analysis). This antagonism of noc/OFQ receptor G-protein coupling was selective because the peptide inhibited the noc/OFQ-evoked GTPgamma(35)S binding to rat brain membranes but not that evoked by selective agonists of the mu-, delta-, and kappa-opioid receptors. In rat cortical membranes, the effects of [F/G]NC(1-13)NH(2) and Ac-RYYRIK-NH(2) on the binding of GTPgamma(35)S were clearly differentiated from the effect of [Nphe(1)]NC(1-13)NH(2) when the concentration of GDP, competing with GTPgammaS for binding, was lowered from 100 microM (assay optimum) to 5 microM. At 5 microM GDP, the former peptides showed clear partial agonist activity, whereas [Nphe(1)]NC(1-13)NH(2) did not. These data indicate that only [Nphe(1)]NC(1-13)NH(2) was a pure antagonist of noc/OFQ receptor G-protein coupling. Furthermore, it is suggested that the variable behavior of [F/G]NC(1-13)NH(2) and Ac-RYYRIK-NH(2) (agonist, partial agonist, and antagonist) in different in vitro and in vivo systems may be explained by different partial GTP binding agonism and the existence of a GTP binding stimulus/response reserve (coupling reserve).


Reorganization of the DNA replication landscape during adipogenesis is closely linked with adipogenic gene expression.
The temporal order of DNA replication along the chromosomes is thought to reflect the transcriptional competence of the genome. During differentiation of mouse 3T3-L1 cells into adipocytes, cells undergo one or two rounds of cell division called mitotic clonal expansion (MCE). MCE is an essential step for adipogenesis; however, little is known about the regulation of DNA replication during this period. Here, we performed genome-wide mapping of replication timing (RT) in mouse 3T3-L1 cells before and during MCE, and identified a number of chromosomal regions shifting toward either earlier or later replication through two rounds of replication. These RT changes were confirmed in individual cells by single-cell DNA-replication sequencing. Coordinate changes between a shift toward earlier replication and transcriptional activation of adipogenesis-associated genes were observed. RT changes occurred before the full expression of these genes, indicating that RT reorganization might contribute to the mature adipocyte phenotype. To support this, cells undergoing two rounds of DNA replication during MCE had a higher potential to differentiate into lipid droplet-accumulating adipocytes, compared with cells undergoing a single round of DNA replication and non-replicating cells.


Upper airway obstruction in neonates and infants with CHARGE syndrome.
Upper airway obstruction can be life-threatening in neonates and infants with CHARGE syndrome, many of whom undergo intratracheal intubation early in life. Although some of these patients are successfully extubated, others require tracheotomy. Deciding whether to complete tracheotomy is challenging since there are no clear criteria upon which to base this decision. We assessed 10 infants with CHARGE syndrome, 5 of whom required tracheotomy. Fiberoptic laryngoscopy showed that all of these patients shared certain features: anteroposterior flattening of the larynx; short vocal cords; anteriorly positioned, tall and hypertrophic arytenoids obscuring the glottis; uncoordinated movement of the vocal cords, epiglottis and arytenoids; salivary pooling. In addition, we observed only in those requiring tracheotomy an obstructive supraglottis that prevented visualization of the vocal cords throughout respiration. Salivary retention was much more severe in this group. These findings might be helpful for predicting the need for an early tracheotomy in situations where the vocal cords are not visible throughout the entire respiratory cycle. Given the high prevalence of malformed larynx and abnormal cranial nerve function, which are not alleviated by supraglottoplasty, we suggest that a thorough investigation of upper airway obstructive entities other than laryngomalacia be performed before embarking on supraglottoplasty in patients with CHARGE syndrome.


Geometry of the soluble methane monooxygenase catalytic diiron center in two oxidation states.
The hydroxylase component of soluble methane monooxygenase (sMMO) contains a dinuclear iron center responsible for the oxidation of methane to methanol. As isolated, the center is in the oxidized, diiron(III) state. The 2.2 A resolution X-ray structure of the oxidized hydroxylase, Hox, from Methylococcus capsulatus (Bath) was previously determined at 4 degrees C. In this structure the two iron atoms are bridged by a glutamate, a hydroxide ion, and an acetate ion, and additionally coordinated to two His residues, three Glu residues, and a water molecule. The 1.7 A resolution crystal structures of the sMMO hydroxylase from Methylococcus capsulatus (Bath) in both its oxidized diiron(III), Hox, and dithionite-treated, reduced diiron(II), Hred, oxidation states were determined at -160 degrees C. The structure of the diiron center in Hox differs from that previously reported at 2.2 A resolution and 4 degrees C. Although the hydroxide bridge is retained, the bidentate, bridging ligand assigned as acetate is replaced by a weakly coordinating monoatomic water bridge. In the resulting four-membered Fe(OH)Fe(OH2) ring, the Fe ... Fe distance is shortened from 3.4 A to 3.1 A. In protomer A of Hred, the hydroxide bridge is displaced by an oxygen atom of Glu243, which undergoes a carboxylate shift from its terminal monodentate binding mode in Hox to a mode in which the carboxylate is both monoatomic bridging and bidentate chelating. We therefore conclude that the center has been reduced to the diiron(II) oxidation state. Both iron atoms are coordinated to five ligands and weakly to a sixth water molecule in the resulting structure. The diiron center in protomer B of Hred has the same composition as those in Hox. In both the oxidized and reduced structures, the diiron core is connected through hydrogen bonds involving exogenous species to Thr213 in the active site cavity. The diiron center in Hox can change its exogenous ligand coordination and geometry, a property that could be important in the catalytic cycle of sMMO. In Hred, a carboxylate shift occurs, extruding hydroxide ion and opening coordination sites for reaction with O2 to form the diiron(III) peroxo intermediate, Hperoxo. Residue Thr213 may function in catalysis.


Antimicrobial and Wound-Healing Activities of Graphene-Reinforced Electrospun Chitosan/Gelatin Nanofibrous Nanocomposite Scaffolds.
This study aims at preparing electrospun chitosan/gelatin nanofiber scaffolds reinforced with different amounts of graphene nanosheets to be used as antibacterial and wound-healing scaffolds. Full characterization was carried out for the different fabricated scaffolds before being assessed for their antimicrobial activity against Escherichia coli and Staphylococcus aureus, cytotoxicity, and cell migration capacity. Raman and transmission electron microscopies confirmed the successful reinforcement of nanofibers with graphene nanosheets. Scanning electron microscopy and porosity revealed that nanofibers reinforced with 0.15% graphene nanosheets produced the least diameter (106 ± 30 nm) and the highest porosity (90%), in addition to their good biodegradability and swellability. However, the excessive increase in graphene nanosheet amount produced beaded nanofibers with decreased porosity, swellability, and biodegradability. Interestingly, nanofibers reinforced with 0.15% graphene nanosheets showed E. coli and S. aureus growth inhibition percents of 50 and 80%, respectively. The cell viability assay showed no cytotoxicity on human fibroblasts when cultured with either unreinforced or reinforced nanofibers. The cell migration was higher in the case of reinforced nanofibers when compared to the unreinforced nanofibers after 24 and 48 h, which is substantially associated with the great effect of the graphene nanosheets on the cell migration capability. Unreinforced and reinforced nanofibers showed cell migration results up to 93.69 and 97%, respectively, after 48 h.


[Cutaneous regressing/regressed malignant melanoma: a clinicopathologic analysis of 8 cases].
To study the clinicopathologic features and differential diagnosis of cutaneous regressing/regressed melanoma. Histopathologic evaluation and immunohistochemical study by EnVision method were performed in 8 cases of cutaneous regressing/regressed melanoma. The clinical presentation, treatment and follow-up data were analyzed. The age of the patients ranged from 40 to 69 years (mean 58 years). The male-to-female ratio was 3: 1. Tumors were located on the back (4 cases), sole of the foot (2 cases), ventral aspect of the toes (1 case), and the forearm (1 case). Clinically, 6 patients presented with progressive black mole of the skin, followed by subsequent focal hypopigmentation, even scarring. Two patients presented with multiple foci of dark-brown pigmentation. Microscopically, 3 cases were completely regressed malignant melanoma. Tumoral melanosis was found in 1 of 3 cases. The other 5 cases were melanoma with severe regression. The extent of regression ranged from 75% to 90%. The Breslow depth of the tumors ranged from 0.5 to 1.0 mm. Immunohistochemically, both metastatic and primary tumor cells were diffusely positive for S-100, HMB45 and Melan A, while melanophages were positive for CD68. Follow-up data were available in 8 patients, ranging from 8 to 27 months. Five patients were alive with no evidence of disease, 1 patient was alive with stable disease and 2 patients died of metastatic melanoma. Correlation between clinical presentation and pathologic features is important for diagnosis of cutaneous regressing/regressed melanoma. Thin melanoma with extensive regression ( ≥ 75%) should not been regarded as low metastatic risk and wide excision combined with sentinel lymph node biopsy is recommended.


[Dementia and hypothyroidism].
The clinical presentation of hypothyroidism varies considerably and may be dominated by cognitive deficits and psychological symptoms. Hence, when diagnosing dementia it is important to rule out hypothyroidism. Hypothyroidism has also been considered a classic reversible dementia; however, the aetiological association between dementia and hypothyroidism is by no means indisputable. A review of available literature concerning hypothyroidism, cognitive functioning and dementia. The importance of thyroxin for the development of the brain and for intellectual development is well known. Early reports suggested that thyroxin improved or reversed dementia in patients with hypothyroidism. Later studies have shown contradictory results; however, several prospective intervention and follow-up studies of overt and subclinical hypothyroidism do not provide for support the belief that there is an aetiological association between dementia and hypothyroidism. The lack of association between dementia and hypothyroidism has consequences for the treatment, and for information to patients and relatives of patients with concomitant dementia symptoms and hypothyroidism. The introduction of acetylcholinesterase inhibitors in the treatment of patients with Alzheimer's disease has brought new interest to this matter. When hypothyroidism is detected in a patient with suspected dementia, thyroxin should be tried first of all. If there is no obvious improvement of symptoms and there still is evidence of dementia, one should consider whether the patient is suitable for a trial with an acetylcholine inhibitor.


A prospective study of volunteerism and hypertension risk in older adults.
The purpose of the current study was to determine whether volunteerism is prospectively associated with hypertension risk among older adults. Participants provided data during the 2006 and 2010 waves of the Health and Retirement Study, a longitudinal panel survey using a nationally representative sample of community-dwelling older adults (age > 50 years). Volunteerism and blood pressure were measured at baseline and again 4 years later. Analyses excluded individuals hypertensive at baseline and controlled for age, race, sex, education, baseline systolic/diastolic blood pressure, and major chronic illnesses. Those who had volunteered at least 200 hr in the 12 months prior to baseline were less likely to develop hypertension (OR = 0.60; 95% CI [0.40, 0.90]) than nonvolunteers. There was no association between volunteerism and hypertension risk at lower levels of volunteer participation. Volunteering at least 200 hr was also associated with greater increases in psychological well-being (B = 0.99, β = .05, p = .006) and physical activity (B = 0.21, β = .05, p = .04) compared with nonvolunteers; however, these factors did not explain the association of volunteerism with hypertension risk.


Metformin decreases hepatocellular carcinoma risk in a dose-dependent manner: population-based and in vitro studies.
Type 2 diabetes mellitus is associated with a higher risk of hepatocellular carcinoma (HCC), which is attenuated by the use of metformin. However, there are no studies addressing the effect of metformin on hepatocarcinoma cells from the antitumoural perspective. In the nationwide case-control study, the authors recruited 97,430 HCC patients and 19,860 age-, gender- and physician visit date-matched controls. The chemopreventive effects of metformin were examined by multivariate analysis and stratified analysis. The in vitro effects of metformin on cell proliferation and cell cycle were studied in HepG2 and Hep3B hepatoma cell lines. The OR of diabetes in HCC patients was 2.29 (95% CI 2.25 to 2.35, p<0.001). Each incremental year increase in metformin use resulted in 7% reduction in the risk of HCC in diabetic patients (adjusted OR=0.93, 95% CI 0.91 to 0.94, p<0.0001). In the multivariate stratified analysis, metformin use was associated with a reduced risk of HCC in diabetic patients in nearly all subgroups. Cell line studies showed that metformin inhibits hepatocyte proliferation and induces cell cycle arrest at G0/G1 phase via AMP-activated protein kinase and its upstream kinase LKB1 to upregulate p21/Cip1 and p27/Kip1 and downregulate cyclin D1 in a dose-dependent manner, but independent of p53. Combined treatment of oral metformin with doxorubicin functioned more efficiently than either agent alone, in vivo. Use of metformin is associated with a decreased risk of HCC in diabetic patients in a dose-dependent manner, via inhibition of hepatoma cells proliferation and induction of cell cycle arrest at G0/G1 phase.


Rat testicular testibumin is a protein responsive to follicle stimulating hormone and testosterone that shares immunodeterminants with albumin.
During a search for hormonally responsive products in media from Sertoli cell enriched cultures, a major follicle stimulating hormone responsive and testosterone-responsive protein was identified and designated CMB-1. The results of the present study indicate that this protein is related immunologically to rat albumin and rat alpha-fetoprotein (AFP) and is concentrated in the testis of the adult rat. CMB-1 was therefore termed testibumin. Testibumin was purified from Sertoli cell enriched cultures to apparent homogeneity by sequential high-performance liquid chromatography on anion-exchange, chromatofocusing, gel permeation, and hydroxylapatite columns. The purified protein consists of two concanavalin A (Con A) reactive forms: one which does not interact with Con A and the other which binds to this lectin and is eluted with methyl alpha-mannoside. Testibumin is a monomer with an apparent molecular weight of 69,000 and a pI ranging between 4.5 and 4.85. The heterogeneity of this protein was further demonstrated by crossed-immunoelectrophoresis and two-dimensional gel electrophoresis. A monospecific antiserum and highly purified testibumin were used to develop a specific radioimmunoassay which permitted studies of the hormonal responsiveness of Sertoli cell enriched culture and of the content of testibumin in the reproductive tract fluids in vivo. Even though testibumin was found in serum of both sexes, it was highly concentrated in the testicular and epididymal compartments in adult rats. This protein was compared to rat serum albumin and rat AFP immunologically. With the use of immunoblots, antiserum developed against testibumin showed partial cross-reactivity with albumin and AFP when these latter proteins were denatured and were present in amounts several orders of magnitude greater than testibumin. The extent of this cross-reactivity was then examined by comparing the ability of native and S-carboxymethylated albumin to compete with 125I-testibumin for binding to a monospecific testibumin antiserum. It was shown that the unfolded derivative of albumin showed partial cross-reactivity with testibumin. We conclude that testibumin is immunologically related to albumin and AFP as these latter proteins are related to one another and that testibumin is possibly the homologue of albumin in the seminiferous tubular compartment.


The effects of physical activity on functional MRI activation associated with cognitive control in children: a randomized controlled intervention.
This study used functional magnetic resonance imaging (fMRI) to examine the influence of a 9-month physical activity program on task-evoked brain activation during childhood. The results demonstrated that 8- to 9-year-old children who participated in 60+ min of physical activity, 5 days per week, for 9 months, showed decreases in fMRI brain activation in the right anterior prefrontal cortex coupled with within-group improvements in performance on a task of attentional and interference control. Children assigned to a wait-list control group did not show changes in brain function. Furthermore, at post-test, children in the physical activity group showed similar anterior frontal brain patterns and incongruent accuracy rates to a group of college-aged young adults. Children in the wait-list control group still differed from the young adults in terms of anterior prefrontal activation and performance at post-test. There were no significant changes in fMRI activation in the anterior cingulate cortex (ACC) for either group. These results suggest that physical activity during childhood may enhance specific elements of prefrontal cortex function involved in cognitive control.


Diversity matters: Effects of density compensation in pollination service during rainfall shift.
Extreme weather events are increasing in frequency due to the warming climate. Such extremities can jeopardize ecosystem services and create economic imbalances. Tropical developing countries are predicted to suffer the maximum consequences of such events.We examined the impact of such an event-extreme rainfall fluctuation-on a critical ecosystem service-pollination, which can be intricately linked to a country's economy. We performed this study in a dominant peri-urban vegetable hub of an agriculture-dependent developing country.We found that the yield of all pollinator-dependent crops grown across a large spatial scale (district) over multiple years (six) drastically declined with the decrease in rainfall.At the local scale, we found that the dominant crop (representative horticultural crop) had a significant drop in yield during drought, likely due to the production of fewer female flowers and a significant shift in the pollinator community.We found that Trigona sp. (one of the four pollinators) was the critical pollinator positively influencing fruit-to-flower ratio (FFR) (an indicator of pollination service) in the normal rainfall year. However, despite its sharp decline during drought, the FFR remained unaffected. We found that during drought, Apis dorsata was crucial in maintaining FFR and compensated for the decline of the critical pollinator across 67% farmlands.Our study demonstrates the role of ecosystem stabilizing mechanism rescuing the crucial ecosystem service during climatic variability over the temporal scale.


Efficacy and safety of single-agent pertuzumab (rhuMAb 2C4), a human epidermal growth factor receptor dimerization inhibitor, in castration-resistant prostate cancer after progression from taxane-based therapy.
Pertuzumab represents a new class of targeted anticancer agents, human epidermal growth factor receptor (HER) dimerization inhibitors. The aim of this single-arm phase II clinical study was to assess the efficacy and safety of single-agent pertuzumab in castration-resistant prostate cancer (CRPC) patients who had experienced progression after prior chemotherapy. Patients received pertuzumab every 3 weeks. All castration-resistant patients had experienced progression after at least one taxane-based regimen. Patients received a loading dose of 840 mg pertuzumab (cycle 1) followed by 420 mg for subsequent cycles. The primary end point was overall response and safety. A separate retrospective analysis of actual survival time versus predicted survival time for a patient population with comparable prognostic features was performed. Patients were enrolled (N = 42) and treated (n = 41). No patients had complete or partial response (as defined by Response Evaluation Criteria in Solid Tumors Group or 50% decline in prostate-specific antigen). Of 30 efficacy-assessable patients, five had stable disease (SD) for at least 23 weeks; one of five had SD for 36 weeks. Pertuzumab was well tolerated; diarrhea was the most common adverse effect (61.0%, grades 1 to 3). Retrospective analysis of survival using a validated nomogram suggested that survival was prolonged with pertuzumab treatment, compared with historic controls with similar baseline prognostic features. Pertuzumab was well tolerated and resulted in no objective responses, but several patients had SD more than 23 weeks from a heavily pretreated population. Retrospective analysis suggested prolonged median survival time with pertuzumab compared with historical controls. Thus, inhibition of HER dimerization may have clinical utility in CRPC patients.


Spongiosum-combined glanuloplasty reduces glans complications after proximal hypospadias repair.
We aim to design a new glanuloplasty procedure combined with spongiosum to reduce the incidence of glans dehiscence and coronal fistula after proximal hypospadias repair. Patients who underwent urethroplasty by dorsal preputial island flap for proximal hypospadias between January 2014 and December 2016 were reviewed in this retrospective cohort study. Those who underwent spongiosum-combined glanuloplasty comprised the new-maneuver group, whereas those who underwent conventional glanuloplasty comprised the control group. The incidence of complications was then compared. In the new-maneuver group, dysplastic corpus spongiosum alongside lateral Buck's fascia (0.3-0.4 cm wide) on both sides of the urethral plate was separated from the proximal normal spongy tissue, joining into the glans wings to increase tissue volume and covering the neourethra in the glans penis. In the control group, the neourethra was covered with superficial fascia under the coronal sulcus. As a result, the new-maneuver and control groups comprised 47 and 28 patients, respectively. In the new-maneuver group, no glans dehiscence was detected; however, two (4.3%) patients had coronal fistula, two (4.3%) had urethral stricture, and four (8.5%) had diverticulum. In the control group, two (7.1%) patients had glans dehiscence, eight (28.6%) had coronal fistula, four (14.3%) had urethral stricture, one (3.6%) had diverticulum, and one (3.6%) had penile curvature recurrence. The new-maneuver group had less incidences of coronal fistula (P < 0.001), glans dehiscence (P = 0.033), and urethral stricture (P = 0.008) but had a higher incidence of diverticulum than the control group (P = 0.040). It clearly demonstrates that spongiosum-combined glanuloplasty can significantly reduce the incidences of coronal fistula and glans dehisce.


Metabolic effects of bacitracin in isolated rat hepatocytes.
Bacitracin is a proteolytic inhibitor which interacts with the intracellular processing of insulin. Its effects on pyruvate, fatty acid and amino acid metabolism were examined in rat hepatocyte suspensions. Bacitracin (0.25-1.0 mM) increased the oxidation of [1-14C]pyruvate by 50-70% and presumably therefore increased the flux through pyruvate dehydrogenase. This was found both in the presence of extracellular Ca2+ and in its absence, but not in the presence of 2 mM-2-chloropropionate, which inhibits pyruvate dehydrogenase kinase. Insulin did not further stimulate [1-14C]pyruvate oxidation in the presence of 1 mM-bacitracin. Bacitracin decreased 14CO2 formation from [2-14C]pyruvate (20-40%) and [U-14C]palmitate (30-70%), suggesting a decreased flux through the tricarboxylic acid cycle. Fatty acid oxidation before acetyl-CoA formation was also decreased. Bacitracin decreased the incorporation of label from [3H]leucine into protein in the absence of insulin, but not in its presence. Bacitracin is commonly used in studies on insulin action. Our results suggest that in such studies the effects noted may be related not only to an interaction of bacitracin with the intracellular processing of insulin but also to direct metabolic effects of bacitracin independent of insulin.


Antimicrobial resistance among Shigella in New Zealand.
We undertook a national survey to provide current information on antimicrobial resistance among Shigella isolated in New Zealand. Diagnostic laboratories are requested to refer all Shigella isolates to the Institute of Environmental Science and Research (ESR) for epidemiological typing as part of the national surveillance of shigellosis. The antimicrobial susceptibility of 263 non-duplicate Shigella isolates referred to ESR in 2015 and 2016 was tested. The 263 Shigella comprised 141 (53.6%) S. sonnei, 113 (43.0%) S. flexneri, 7 (2.7%) S. boydii and 2 (0.8%) S. dysenteriae. Among the 141 S. sonnei, the majority were either biotype g (90) or biotype a (50). Rates of resistance to the two currently recommended first-line antibiotics, co-trimoxazole and fluoroquinolones, were relatively high at 56.7% and 22.8%, respectively. Azithromycin is considered a second-line treatment option, but 11.0% of Shigella were categorised as having a non-wildtype (NWT) azithromycin phenotype (ie, having some mechanism of azithromycin resistance although not necessarily clinically resistant). There were several significant differences in resistance between the two most prevalent S. sonnei biotypes, with resistance being significantly more prevalent among biotype g isolates. Shigella from patients who had not travelled overseas were significantly more likely to be azithromycin NWT than isolates from patients who had recently travelled (20.7 vs 5.6%). Azithromycin NWT was more prevalent among Shigella from males than females (13.9 vs 7.7%). These results suggest there is an immediate need to revise the currently recommended first-line treatment for shigellosis, especially when treatment is given on an empirical basis. Equally concerning is the fact that resistance to the second-line antibiotic for shigellosis, azithromycin, appears to be emerging in New Zealand. As diagnostic laboratories increase their use of culture-independent testing, it is recommended that they should continue to culture specimens from all shigellosis cases so that isolates are available for susceptibility testing and epidemiological typing.


Peptide YY: a potential proabsorptive hormone for the treatment of malabsorptive disorders.
Peptide YY (PYY) is a 36 amino acid peptide that is released from the endocrine cells of the distal ileum, colon, and rectum following a meal. PYY is strongly proabsorptive in the small intestine. We studied the effects of intravenous PYY on colonic water and electrolyte transport in awake dogs. Dogs had 20 cm neurovascularly intact colon Thiry-Vella fistulas (TVS) surgically constructed. Colonic transport was studied in three experimental groups. Group 1 animals received a standard mixed meal. Group 2 animals were unfed and received intravenous PYY and 100 pmol/kg/hr for two hours. This dose of PYY has previously been shown to simulate the plasma levels of PYY normally seen after a meal. Group 3 received intravenous PYY at the same dose in addition to a mixed meal. Our study shows an increase in colonic water, Na+, and Cl- absorption after a meal (P < 0.05). Infusion of PYY at a 100 pmol/kg/hr was significantly proabsorptive beginning at 60 minutes (P < 0.01). Infusion of PYY in addition to a meal further increased absorption (P < 0.05). PYY is a potent proabsorptive agent in the colon of the conscious dog. PYY, or its analogs, may be useful clinical agents in intestinal malabsorptive disorders or after bowel resection.


Efficacy of aerobic exercise for treatment of chronic low back pain: a meta-analysis.
A meta-analysis of relevant cohort studies was performed to investigate the efficacy of aerobic exercise for the treatment of patients with chronic low back pain (CLBP). A range of electronic databases were searched: MEDLINE (1966-2013), the Cochrane Library Database (issue 12, 2013), EMBASE (1980-2013), CINAHL (1982-2013), Web of Science (1945~2013), and the Chinese Biomedical Database (1982-2013), without language restrictions. The Roland-Morris Disability Questionnaire, the Oswestry Disability Questionnaire, the McGill Pain Questionnaire, visual analog scale, and heart rate, sit-and-reach test, and maximum oxygen consumption were used to evaluate the efficacy of aerobic exercise. Meta-analysis was performed with the use of the STATA statistical software. The standardized mean difference (SMD) with its corresponding 95% confidence interval (CI) was calculated. Eight clinical cohort studies with a total of 310 CLBP patients were included in the meta-analysis. The results of this meta-analysis indicated that CLBP patients exhibited positive decreases in scores on the Roland-Morris Disability Questionnaire (SMD, 0.44; 95% CI, 0.20-0.68; P < 0.001), Oswestry Disability Questionnaire (SMD, 1.03; 95% CI, 0.67-1.39; P < 0.001), Hospital Anxiety and Depression Scale (SMD, 1.35; 95% CI, 0.34-2.37; P = 0.009), and McGill Pain Questionnaire (SMD, 0.32; 95% CI, 0.07-0.56; P = 0.011) after aerobic exercise. It was also observed that aerobic exercise could markedly reduce the visual analog scale score for pain of CLBP patients (SMD, 0.75; 95% CI, 0.48-1.02; P < 0.001). Nevertheless, this study's findings showed that aerobic exercise had no effects on heart rate, sit-and-reach test, and maximum oxygen consumption of CLBP patients (all P > 0.05). The current meta-analysis provides reliable evidence that aerobic exercise could effectively diminish pain intensity and improve the physical and psychologic functioning of CLBP patients. Thus, aerobic exercise may be a good choice in the treatment for CLBP.


Yoga training and motor speed based on a finger tapping task.
A finger tapping task was used to assess motor speed (MS) of both hands in 53 adults and 152 children before and after yoga training and in 38 adults of a non-yoga (control) group. All subjects were right hand dominant. The 30-second tapping speed (TS) test was considered as three time intervals, i.e. 0-10 second (TS1), 10-20 seconds (TS2) and 20-30 seconds (TS3). There was a significant (Student's t-test) increase in all three TS values following 10 days of yoga in children and 30 days of yoga in adults. However for both groups at baseline and final assessments, TS2 and TS3 were significantly lower than TS1. Hence the TS was increased after yoga training during the first 10-seconds of the test but not during the next 20 seconds. These results suggest an increase in motor speed for repetitive finger movements following yoga training, but not in strength or endurance, as the increase was not sustained over 30 sec.


Anti-inflammatory protein TSG-6 secreted by activated MSCs attenuates zymosan-induced mouse peritonitis by decreasing TLR2/NF-κB signaling in resident macrophages.
Human mesenchymal stem/progenitor cells (hMSCs) repair tissues and modulate immune systems but the mechanisms are not fully understood. We demonstrated that hMSCs are activated by inflammatory signals to secrete the anti-inflammatory protein, TNF-α-stimulated gene 6 protein (TSG-6) and thereby create a negative feedback loop that reduces inflammation in zymosan-induced peritonitis. The results demonstrate for the first time that TSG-6 interacts through the CD44 receptor on resident macrophages to decrease zymosan/TLR2-mediated nuclear translocation of the NF-κB. The negative feedback loop created by MSCs through TSG-6 attenuates the inflammatory cascade that is initiated by resident macrophages and then amplified by mesothelial cells and probably other cells of the peritoneum. Because inflammation underlies many pathologic processes, including immune responses, the results may explain the beneficial effects of MSCs and TSG-6 in several disease models.


[Early prognostic factors of the surgery of aneurysms of the abdominal aorta with renal artery clamping].
The aneurysms of the abdominal aorta requiring suprarenal clamping are rare. Suprarenal clamping was required for only 43 of 544 aneurysms operated electively from 1981 to 1989. Twenty-five patients had a juxtarenal aneurysm, without any normal aortic segment under the renal arteries, and suprarenal clamping was therefore necessary while the upper anastomosis was being established (group I). Eighteen patients had an aneurysm enclosing the root of at least one renal artery (group II). Several prognostic factors have been assessed: patient's age, presence of preoperative renal insufficiency, of arterial hypertension or of coronary insufficiency, and revascularization method. Five patients died. Four of them belonged to group II and were over 75 years old. All presented with a preoperative renal insufficiency. Two of these deaths were caused by mesenteric infarction. Four cases of regressive postoperative renal insufficiency were observed in patients for whom renal clamping had lasted longer than 45 minutes. This study allowed outlining three prognostic factors: the patient's age, preoperative renal insufficiency, a period of renal ischemia exceeding 40 minutes. On the other hand, the severity of hypertension had no predictive value. Coronary insufficiency requires a strict hemodynamic surveillance, but is not a contraindication for revascularization.


Elk-1 associates with the mitochondrial permeability transition pore complex in neurons.
The nuclear transcription factor E-26-like protein 1 (Elk-1) is thought to impact neuronal differentiation [Sharrocks, A. D. (2001) Nat. Rev. Mol. Cell Biol. 2, 827-837], cell proliferation [Sharrocks, A. D. (2002) Biochem. Soc. Trans. 30, 1-9], tumorigenesis [Chai, Y. L., Chipitsyna, G., Cui, J., Liao, B., Liu, S., Aysola, K., Yezdani, M., Reddy, E. S. P. & Rao, V. N. (2001) Oncogene 20, 1357-1367], and apoptosis [Shao, N., Chai, Y., Cui, J., Wang, N., Aysola, K., Reddy, E. S. P. & Rao, V. N. (1998) Oncogene 17, 527-532]. In addition to its nuclear localization, Elk-1 is found throughout the cytoplasm, including localization in neuronal dendrites [Sgambato, V., Vanhoutte, P., Pages, C., Rogard, M., Hipskind, R., Besson, M. J. & Caboche, J. (1998) J. Neurosci. 18, 214-226], raising the possibility that Elk-1 may have alternative extranuclear functions in neurons. Using coimmunoprecipitation and reciprocal coimmunoprecipitation from adult rat brain, we found an association between Elk-1 protein and the mitochondrial permeability transition pore complex (PTP), a structure involved in both apoptotic and necrotic cell death. Electron microscopy in adult rat brain sections confirmed this association with mitochondria. Elk-1 was also identified from purified mitochondrial fractions by using Western blotting, and Elk-1 increased its association with mitochondria following proapoptotic stimuli. Consistent with a role for Elk-1 in neuron viability, overexpression of Elk-1 in primary neurons decreased cell viability, whereas Elk-1 siRNA-mediated knockdown increased cell viability. This decrease in viability induced by Elk-1 overexpression was blocked with application of a PTP inhibitor. These results show an association of the nuclear transcription factor Elk-1 with the mitochondrial PTP and suggest an additional extranuclear function for Elk-1 in neurons.


A phase II trial of isotretinoin and alpha interferon in patients with recurrent squamous cell carcinoma of the cervix: a Gynecologic Oncology Group study.
From January 1993 through January 1996, 37 patients with unresectable squamous carcinoma of the cervix were entered on study and scheduled to receive oral isotretinoin 1 mg/kg per day with subcutaneous alpha interferon 6,000,000 units/day. A course was defined as 4 continuous weeks of therapy. The mean number of four-course cycles delivered was 1.8. One patient was ineligible because of wrong cell type and two were never treated. Thus, 34 patients were evaluable for toxicity. Eight patients were inevaluable for response. Five did not receive a complete 4-week course and three did not have additional tumor measurements; thus 26 were evaluable for response. Prior radiotherapy had been given to 25 patients and prior chemotherapy to 23 patients. There was no grade 4 neutropenia. The incidence of Gynecologic Oncology Group (GOG) grade 3 granulocytopenia and thrombocytopenia was 8.8% and 5.8%, respectively. Six patients (17.6%) developed grade 3 or worse nausea and vomiting. Four (11.7%) patients developed grade 3 neurologic symptoms. There were no complete responses and one partial response. The overall response rate was 3.8% (95% confidence interval, 0.1-19.6%). In this pretreated population, isotretinoin and alpha interferon in the dose and schedule employed exhibit minimal activity.


Amlodipine and atorvastatin exert protective and additive effects via antiapoptotic and antiautophagic mechanisms after transient middle cerebral artery occlusion in Zucker metabolic syndrome rats.
We examined the neuroprotective effects amlodipine and/or atorvastatin in metabolic syndrome (MetS) Zucker fatty rats against transient (90 min) middle cerebral artery occlusion (MCAO). The rats were pretreated with vehicle, amlodipine, atorvastatin, or amlodipine plus atorvastatin for 28 days, and 24 hr after transient MCAO the infarct size was assessed via hematoxylin and eosin staining, and terminal deoxynucleotidyl transferase-mediated dUTP-biotin in situ nick end labeling (TUNEL) and microtubule-associated protein 1 light chain 3 (LC3) expression were examined by immunohistochemistry to evaluate apoptosis and autophagy, respectively. Compared with the vehicle group, rats treated with amlodipine or atorvastatin alone showed a significant decrease in infarct volume (P < 0.01), which was further decreased in the amlodipine plus atorvastatin group (P < 0.001). Compared with the vehicle group, the numbers of TUNEL- and LC3-positive cells were markedly reduced by amlodipine or atorvastatin alone (P < 0.01) and further decreased by amlodipine plus atorvastatin (P < 0.001). The number of apoptotic TUNEL/autophagic LC3 double-positive cells was also significantly decreased with amlodipine or atorvastatin alone compared with vehicle (P < 0.01) and was further decreased by amlodipine plus atorvastatin (P < 0.001). These data suggest additive neuroprotective effects of combination amlodipine and atorvastatin treatment after acute ischemic stroke in MetS model Zucker rats. These effects are mediated, at least in part, via antiapoptotic and antiautophagic mechanisms. Further studies are now needed to expand these preliminary results to understand fully the mechanisms involved in the protective effects of amlodipine and atorvastatin against ischemic stroke.


Universal hepatitis B vaccination in Taiwan and the incidence of hepatocellular carcinoma in children. Taiwan Childhood Hepatoma Study Group.
A nationwide hepatitis B vaccination program was implemented in Taiwan in July 1984. To assess the effect of the program on the development of hepatocellular carcinoma, we studied the incidence of this cancer in children in Taiwan from 1981 to 1994. We collected data on liver cancer in children from Taiwan's National Cancer Registry, which receives reports from each of the country's 142 hospitals with more than 50 beds. Data on childhood liver cancer were also obtained from Taiwan's 17 major medical centers. To prevent the inclusion of cases of hepatoblastoma, the primary analysis was confined to liver cancers in children six years of age or older. Data were also obtained on mortality from liver cancer among children. The average annual incidence of hepatocellular carcinoma in children 6 to 14 years of age declined from 0.70 per 100,000 children between 1981 and 1986 to 0.57 between 1986 and 1990, and to 0.36 between 1990 and 1994 (P<0.01). The corresponding rates of mortality from hepatocellular carcinoma also decreased. The incidence of hepatocellular carcinoma in children 6 to 9 years of age declined from 0.52 for those born between 1974 and 1984 to 0.13 for those born between 1984 and 1986 (P<0.001). Since the institution of Taiwan's program of universal hepatitis B vaccination, the incidence of hepatocellular carcinoma in children has declined.


Influence of tooth length on the accuracy of the Root ZX electronic apical foramen locator: an ex vivo study.
Electronic apical foramen locators are now widely used to determine working length. This study was designed to determine whether tooth length influenced the accuracy of the Root ZX device. Forty extracted maxillary canine teeth with a length range of 27-29 mm were selected. Access cavities were prepared and coronal flaring of canals performed. The teeth were mounted in self-polymerizing acrylic resin to facilitate horizontal sectioning except for the apical 3-4-mm portion of the root and embedded in alginate as the electronic medium. Electronic measurements were taken at the major foramen, 'zero' reading using the Root ZX and compared with the actual root canal length. The teeth were sectioned 3 mm from the coronal reference point to create a second group with shorter length; these reductions in the length continued six times in all to create seven groups of 40 specimens each. The actual and electronic lengths of specimens in each group were measured. Data were analyzed by Pearson's correlation coefficient. Identical measurements between the actual and electronic root canal length from the longest to the shortest groups were 12.5%, 10.0%, 20.0%, 27.5%, 37.5%, 35.0% and 45.0%, respectively. There was a mild negative correlation between the precise measurements of the Root ZX and root canal lengths in the seven groups (r = -0.964, p < 0.001). Under the conditions of the study, the Root ZX device was more accurate in shorter teeth compared to longer ones.


Minimally invasive surgery for atrial fibrillation.
Atrial fibrillation (AF) remains the most common cardiac arrhythmia, affecting nearly 2% of the general population worldwide. Minimally invasive surgical ablation remains one of the most dynamically evolving fields of modern cardiac surgery. While there are more than a dozen issues driving this development, two seem to play the most important role: first, there is lack of evidence supporting percutaneous catheter based approach to treat patients with persistent and long-standing persistent AF. Paucity of this data offers surgical community unparalleled opportunity to challenge guidelines and change indications for surgical intervention. Large, multicenter prospective clinical studies are therefore of utmost importance, as well as honest, clear data reporting. Second, a collaborative methodology started a long-awaited debate on a Heart Team approach to AF, similar to the debate on coronary artery disease and transcatheter valves. Appropriate patient selection and tailored treatment options will most certainly result in better outcomes and patient satisfaction, coupled with appropriate use of always-limited institutional resources. The aim of this review, unlike other reviews of minimally invasive surgical ablation, is to present medical professionals with two distinctly different, approaches. The first one is purely surgical, Standalone surgical isolation of the pulmonary veins using bipolar energy source with concomitant amputation of the left atrial appendage-a method of choice in one of the most important clinical trials on AF-The Atrial Fibrillation Catheter Ablation Versus Surgical Ablation Treatment (FAST) Trial. The second one represents the most complex approach to this problem: a multidisciplinary, combined effort of a cardiac surgeon and electrophysiologist. The Convergent Procedure, which includes both endocardial and epicardial unipolar ablation bonds together minimally invasive endoscopic surgery with electroanatomical mapping, to deliver best of the two worlds. One goal remains: to help those in urgent need for everlasting relief.


Technological devices for deaf-blind children: needs and potential impact.
The authors provide a glimpse into the world of congenitally deaf-blind children and how the application of technology might enable them to lead more productive lives. They describe this population and then summarize the present status of efforts to educate and communicate with them. A review of technological devices presently usable and potentially usable to complement these efforts is given. The authors conclude with a list of technological tools that are needed for communication with deaf-blind persons.


Integration of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry in blood culture diagnostics: a fast and effective approach.
Sepsis is a major cause of mortality in hospitalized patients worldwide, with lethality rates ranging from 30 to 70 %. Sepsis is caused by a variety of different pathogens, and rapid diagnosis is of outstanding importance, as early and adequate antimicrobial therapy correlates with positive clinical outcome. In recent years, matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) fingerprinting has become a powerful tool in microbiological diagnostics. The direct identification of micro-organisms in a positive blood culture by MALDI-TOF MS can shorten the diagnostic procedure significantly. Therefore, the aim of the present study was to evaluate whether identification rates could be improved by using the new Sepsityper kit from Bruker Daltonics for direct isolation and identification of bacteria from positive blood cultures by MALDI-TOF MS compared with the use of conventional separator gel columns, and to integrate the MALDI-TOF MS-based identification method into the routine course of blood culture diagnostics in the setting of a microbiological laboratory at a university hospital in Germany. The identification of Gram-negative bacteria by MALDI-TOF MS was significantly better using the Sepsityper kit compared with a separator gel tube-based method (99 and 68 % correct identification, respectively). For Gram-positive bacteria, only 73 % were correctly identified by MALDI-TOF with the Sepsityper kit and 59 % with the separator gel tube assay. A major problem of both methods was the poor identification of Gram-positive grape-like clustered cocci. As differentiation of Staphylococcus aureus from coagulase-negative staphylococci is of clinical importance, a PCR was additionally established that was capable of identifying S. aureus directly from positive blood cultures, thus closing this diagnostic gap. Another benefit of the PCR approach is the possibility of directly detecting the genes responsible for meticillin resistance in staphylococci and for vancomycin resistance in enterococci, which is of high importance for early adequate treatment. Both of the described methods were finally integrated into a protocol for fast and effective identification of bacteria from positive blood cultures.


Direct Exports of Phosphorus from Fertilizers Applied to Grazed Pastures.
Since its discovery in 1669, phosphorus (P) in the form of fertilizer has become an essential input for many agroecosystems. By introducing a concentrated P source, fertilizers increase short-term P export potential soon after their application and longer-term export potential by increasing soil fertility (legacy P). The 4R concept was developed to help mitigate P exports from the fertilizers that sustain agricultural productivity. This review investigates the factors affecting P exports soon after the application of mineral fertilizers to pasture-based grazing systems and studies quantifying its potential impact in different systems, with an emphasis on Australasia. Initially, P fertilizers and reactions that might affect their short-term P export potential are reviewed, along with P transport pathways, the forms of P exported from grazing systems, factors affecting P mobilization into water, and studies demonstrating the possible short-term effects of fertilizer application on P exports. Using that foundation, we review studies quantifying the short-term impact of fertilizer application in different regions; they show that under poor management, recently applied fertilizer can contribute a considerable proportion (30-80%) of total farm P exports in drainage, but when fertilizer is well-managed, that figure is expected to be <10%. We then use three model systems of varying hydrology that are common to Australasia to demonstrate the principles for selecting fertilizers that are likely to minimize P exports soon after their application.


New semiphysiological absorption model to assess the pharmacodynamic profile of cefuroxime axetil using nonparametric and parametric population pharmacokinetics.
Cefuroxime axetil is widely used to treat respiratory tract infections. We are not aware of a population pharmacokinetic (PK) model for cefuroxime axetil. Our objectives were to develop a semiphysiological population PK model and evaluate the pharmacodynamic profile for cefuroxime axetil. Twenty-four healthy volunteers received 250 mg oral cefuroxime as a suspension after a standardized breakfast. Liquid chromatography-tandem mass spectrometry was used for drug analysis, NONMEM and S-ADAPT (results reported) were used for parametric population PK modeling, and NPAG was used for nonparametric population PK modeling. Monte Carlo simulations were used to predict the duration for which the non-protein-bound-plasma concentration was above the MIC (fT(>MIC)). A model with one disposition compartment, a saturable and time-dependent drug release from the stomach, and fast drug absorption from the intestine yielded precise (r > 0.992) and unbiased curve fits and an excellent predictive performance. The apparent clearance was 21.7 liters/h (19.8% coefficient of variation [CV]) and the volume of distribution 38.7 liters (18.3% CV). Robust (>or=90%) probabilities of target attainment (PTAs) were achieved by 250 mg cefuroxime given every 12 h (q12h) or q8h for MICs of <or=0.375 mg/liter or <or=0.5 mg/liter, respectively, for the bacteriostasis target fT(>MIC) of >or=40% and for MICs of <or=0.094 mg/liter or <or=0.375 mg/liter, respectively, for the near-maximal-killing target fT(>MIC) of >or=65%. For the >or=40% fT(>MIC) target, the PTAs for 250 mg cefuroxime q12h were >or=97.8% for Streptococcus pyogenes and penicillin-susceptible Streptococcus pneumoniae. Cefuroxime at 250 mg q12h or q8h achieved PTAs below 73% or 92%, respectively, for Haemophilus influenzae, Moraxella catarrhalis, and penicillin-intermediate S. pneumoniae for susceptibility data from various countries. Depending on the MIC distribution, 250 mg oral cefuroxime q8h instead of q12h should be considered, especially for more-severe infections that require near-maximal killing by cefuroxime.


Delayed Clinical Manifestation of Parkinson's Disease Among Physically Active: Do Participants in a Long-Distance Ski Race Have a Motor Reserve?
Physical activity is associated with reduced risk of Parkinson's disease (PD). The explanations for this association are not completely elucidated. We use long-term PD-incidence data from long-distance skiers to study the relationship between exercise and PD. We aimed to investigate if physical activity is associated with long-term lower risk of PD and if this association could be explained by physically active people being able to sustain more PD neuropathology before clinical symptoms, a motor reserve. Using a prospective observational design, we studied whether long-distance skiers of the Swedish Vasaloppet (n = 197,685), exhibited reduced incidence of PD compared to matched individuals from the general population (n = 197,684) during 21 years of follow-up (median 10, interquartile range (IQR) 5-15 years). Vasaloppet skiers (median age 36.0 years [IQR 29.0-46.0], 38% women) had lower incidence of PD (HR: 0.71; 95 % CI 0.56-0.90) compared to non-skiers. When reducing risk for reverse causation by excluding PD cases within the first five years from race participation, there was still a trend for lower risk of PD (HR: 0.80; 95 % CI 0.62-1.03). Further, the PD prevalence converged between skiers and non-skiers after 15 years of follow-up, which is more consistent with a motor reserve in the physically active rather than neuroprotection. A physical active lifestyle is associated with reduced risk for PD. This association weakens with time and might be explained by a motor reserve among the physically active.


Single nucleotide extension technology for quantitative site-specific evaluation of metC/C in GC-rich regions.
The development and use of high throughput technologies for detailed mapping of methylated cytosines (metC) is becoming of increasing importance for the expanding field of epigenetics. The single nucleotide primer extension reaction used for genotyping of single nucleotide polymorphisms has been recently adapted to interrogate the bisulfite modification induced 'quantitative' C/T polymorphism that corresponds to metC/C in the native DNA. In this study, we explored the opportunity to investigate C/T (and G/A) ratios using the Applied Biosystems (ABI) SNaPshot technology. The main effort of this study was dedicated to addressing the complexities in the analysis of DNA methylation in GC-rich regions where interrogation of the target cytosine can be confounded by variable degrees of methylation in other cytosines (resulting in variable C/T or G/A ratios after treatment with bisulfite) in the annealing site of the interrogating primer. In our studies, the mismatches of the SNaPshot primer with the target DNA sequence resulted in a biasing effect of up to 70% while these effects decreased as the location of the polymorphic site moved upstream of the target cytosine. We demonstrated that the biasing effect can be corrected with the SNaPshot primers containing degenerative C/T and G/A nucleotides. A series of experiments using various permutations of quantitative C/T and G/A polymorphisms at various positions of the target DNA sequence demonstrated that SNaPshot is able to accurately report cytosine methylation levels with <5% average SD from the true values. Given the relative simplicity of the method and the possibility to multiplex C/T and G/A interrogations, the SNaPshot approach may become a useful tool for large-scale mapping of metC.


Comparative immunohistochemical and biochemical analysis of pancreatic polypeptide-like peptides with special reference to presence of neuropeptide Y in central and peripheral neurons.
Antisera raised against porcine neuropeptide Y (NPY) and peptide YY (PYY) were characterized with regard to immunohistochemical staining, cross-reactivity to several pancreatic polypeptide (PP)-related peptides, and radioimmunoassayable tissue levels in the rat and pig. The NPY antiserum (102B) reacted with nerves in many areas of both the central and peripheral nervous systems, but it did not stain endocrine cells of the pancreas or intestine. No evidence for any cross-reactivity of the NPY antiserum with related peptides of the PP family, such as avian PP, bovine PP, PYY, gamma-MSH, FMRF-amide, or avian PP (31-36), was obtained. The NPY antiserum was N-terminally directed, and regional levels of NPY as seen by radioimmunoassay paralleled well the occurrence of NPY-immunoreactive structures seen in the immunohistochemical study. High pressure liquid chromatography analysis revealed that the NPY-immunoreactive material from cerebral cortex and vas deferens had elution profiles similar to those of standard porcine NPY. The PYY antiserum mainly stained endocrine cells in the pancreas and intestine as well as a small neuron system in the brainstem of the rat. Although this antiserum had a slight cross-reactivity to NPY in radioimmunoassay, the neuronal PYY staining was separate from that of NPY. High levels of PYY were found in the intestine, and levels above the threshold were also seen in the dorsal vagal complex of the rat. The other antisera investigated (raised against avian PP, bovine PP, gamma-MSH, and FMRF-amide) caused neuronal staining that was abolished by preabsorption with NPY. This was also seen even if no detectable cross-reactivity with NPY was found in radioimmunoassay. These latter antisera also stained endocrine cells in the pancreas and intestine with complex cross-reactivity relationships, suggesting the presence of intestinal PP-like peptides in addition to PYY and NPY.


Conformal radiotherapy for hepatocellular carcinoma.
Technical advancements in radiation therapy (RT) have facilitated the safe delivery of conformal, dose-escalated radiation to a wide spectrum of hepatocellular carcinoma (HCC) patients. A variety of doses and RT fractionation schemes have been used, and RT has been used in combination with transarterial chemoembolization (TACE). Compared to untreated historical controls or those treated with TACE alone, outcomes following RT alone or TACE and RT are better. Despite advances in RT delivery, liver toxicity following RT remains a dose-limiting factor, and investigations to better understand the pathophysiology of RT-induced liver toxicity are warranted. For most tumors, RT can provide sustained local control. However, HCC tends to recur within the liver away from the irradiated volume, providing rationale for combining RT with systemic or regional therapies. There is a particular interest in combining RT with anti-VEGF-targeted agents for their independent activity in HCC as well as their radiation sensitization properties. Randomized trials of RT are warranted.


Refined crystal structure of the complex of subtilisin BPN' and Streptomyces subtilisin inhibitor at 1.8 A resolution.
The crystal structure of subtilisin BPN' complexed with a proteinaceous inhibitor SSI (Streptomyces subtilisin inhibitor) was refined at 1.8 A resolution to an R-factor of 0.177 with a root-mean-square deviation from ideal bond lengths of 0.014 A. The work finally established that the SSI-subtilisin complex is a Michaelis complex with a distance between the O gamma of active Ser221 and the carbonyl carbon of the scissile peptide bond being an intermediate value between a covalent bond and a van der Waals' contact, 2.7 A. This feature, as well as the geometry of the catalytic triad and the oxyanion hole, is coincident with that found in other highly refined crystal structures of the complex of subtilisin Novo, subtilisin Carlsberg, bovine trypsin or Streptomyces griseus protease B with their proteinaceous inhibitors. The enzyme-inhibitor beta-sheet interaction is composed of two separate parts: that between the P1-P3 residues of SSI and the 125-127 chain segment (the "S1-3 site") of subtilisin and that between the P4-P6 residues of SSI and th 102-104 chain segment (the "S4-6 site") of subtilisin. The latter beta-interaction is unique to subtilisin. In contrast, the beta-sheet interaction previously found in the complex of subtilisin Novo and chymotrypsin inhibitor 2 or in the complex of subtilisin Carlsberg and Eglin C is distinct from the present complex in that the two types of beta-interactions are not separate. As for the flexibility of the molecules comprising the present complex, the following observations were made by comparing the B-factors for free and complexed SSI and comparing those for free and complexed subtilisin BPN'. The rigidification of the component molecules upon complex formation occurs in a very localized region: in SSI, the "primary" and "secondary" contact regions and the flanking region; in subtilisin BPN', the S1-3 and S4-6 sites and the flanking region.


Valvular heart disease: shifting the focus to the myocardium.
Adverse cardiac remodelling is the main determinant of patient prognosis in degenerative valvular heart disease (VHD). However, to give an indication for valvular intervention, current guidelines include parameters of cardiac chamber dilatation or function which are subject to variability, do not directly reflect myocardial structural changes, and, more importantly, seem to be not sensitive enough in depicting early signs of myocardial dysfunction before irreversible myocardial damage has occurred. To avoid irreversible myocardial dysfunction, novel biomarkers are advocated to help refining indications for intervention and risk stratification. Advanced echocardiographic modalities, including strain analysis, and magnetic resonance imaging have shown to be promising in providing new tools to depict the important switch from adaptive to maladaptive myocardial changes in response to severe VHD. This review, therefore, summarizes the current available evidence on the role of these new imaging biomarkers in degenerative VHD, aiming at shifting the clinical perspective from a valve-centred to a myocardium-focused approach for patient management and therapeutic decision-making.


Retroviral transduction of hematopoietic progenitors derived from human embryonic stem cells.
It has been recently identified that cytokines and BMP-4 promote hematopoiesis from human embryonic stem cells (hESC) and that, before hematopoietic commitment, a rare subpopulation of cells lacking CD45, but expressing PECAM-1, Flk-1, and VE-cadherin (hereinafter termed CD45(neg)PFV precursors), are exclusively responsible for hematopoietic cell fate on cytokine stimulation. Efficient strategies to stably transduce these hematopoietic precursors specifically generated from hESCs would provide a novel and desirable tool to study hematopoietic development through the introduction and characterization of candidate genes suspected to regulate self-renewal processes of hESC-derived hematopoietic cells or dynamically track hESC-derived hematopoietic stem cells in vivo. To date, only transient transfection and stable transduction using lentiviral vectors have been reported in undifferentiated hESC followed by random and spontaneous differentiation into different cell types. However, protocols for stable transduction of hematopoietic progenitors prospectively derived from hESC need to be developed yet. In the present chapter, we described detailed methods on the recently characterized and optimized GALV-pseudotyped retroviral gene transfer strategy to stably transduce the hematopoietic progenitor cells prospectively derived from CD45(neg)PFV hemogenic precursors as a vital tool to study hematopoietic development and to characterize candidate genes suspected to eventually confer robust and sustained repopulating ability to hESC-derived hematopoietic cells.


Direct Visualisation of the Depth-Dependent Mechanical Properties of Full-Thickness Articular Cartilage.
The structural anisotropy of articular cartilage controls its deformation response. As proteoglycans and collagen vary with depth, simple uniaxial compression results in inhomogeneous deformation with distinct depth-dependent mechanical properties. Investigations into depth-dependent mechanical properties of articular cartilage have previously required tissue modification after specimen isolation. Such modifications include histological processes, freezing, subchondral bone removal, and fluorescent staining that may alter the tissue, limiting in vivo applicability. Using a custom tissue-sectioning device, 0.1 mm thick unfixed, unstained, osetochondral samples were obtained. A customized apparatus loaded samples to 12.5, 24, and 29% compression in under a microscope with 10× magnification. Equilibrium load was measured after stress relaxation. Intra-tissue displacement was measured by tracing groups of cells between the different compression levels using a digital imaging program. Cell distance from the subchondral bone was measured to identify intra-tissue displacement and calculate strain. The results reveal that stress levels and intra-tissue displacement increased with greater tissue compression (p <0.05). Intra-tissue displacement decreased as depth from the articular surface increased (p<0.01). This occurred for each level of tissue compression. Overall compressive resistance is seen to increase with depth from the articular surface. The current study identifies a method directly visualising and assessing the depth-dependent structural response to compression. The ability to avoid tissue modification after specimen isolation, allows this procedure to more closely approximate in vivo conditions and may provide an important method for analyzing the coordinated changes in cartilage composition and function due to ageing and disease.


Lipid distribution and transport across cellular membranes.
In eukaryotic cells, the membranes of different intracellular organelles have different lipid composition, and various biomembranes show an asymmetric distribution of lipid types across the membrane bilayer. Membrane lipid organization reflects a dynamic equilibrium of lipids moving across the bilayer in both directions. In this review, we summarize data supporting the role of specific membrane proteins in catalyzing transbilayer lipid movement, thereby controlling and regulating the distribution of lipids over the leaflets of biomembranes.


Titrating the Rig: How Paramedics Work in and on Their Ambulance.
In this article, I take readers inside of an ambulance and explore how paramedics work in and on their "apparatus unit" to make it a workable fit. This taken-for-granted work is important because much is at stake in the back of the ambulance, particularly in relation to quality of care and safety. I draw on data from an institutional ethnography into the socially organized work and work settings of paramedics, which included more than 200 hr of observations and more than 100 interviews with paramedics. The findings shed light on the situated work processes of paramedics as they orient and respond to their "apparatus unit" and enact quality and safety in practice. This article adds to the sociological literature on work and occupations as well as safety and quality in health care of an increasingly important group of health care and emergency services professional.


Imatinib front-line therapy is safe and effective in patients with chronic myelogenous leukemia with pre-existing liver and/or renal dysfunction.
Imatinib 400 mg daily is the standard treatment for patients with chronic myelogenous leukemia (CML). The safety and efficacy of imatinib in CML patients with pre-existing liver and/or renal dysfunction has not been analyzed. The authors analyzed the outcome of 259 patients with early chronic phase CML treated with imatinib (starting dose 400 mg in 50, 800 mg in 209). Pre-existing liver and/or renal dysfunction was seen in 38 (15%) and 11 (4%) patients, respectively. Dose reductions were required in 91 (43%) of 210 patients with normal organ function, compared with 8 (73%) of 11 (P = .065) with renal dysfunction, and 19 (50%) of 38 (P = .271) with liver dysfunction. Grade 3-4 hematologic toxicities including anemia (29%, 10%, and 7% of patients with renal dysfunction, liver dysfunction, and normal organ function, respectively), neutropenia (57%, 30%, and 30%), and thrombocytopenia (43%, 30%, and 26%) were more frequent in patients with pre-existing renal dysfunction treated with high-dose imatinib. Grade 3-4 nonhematologic toxicities were observed at similar frequencies. Complete cytogenetic response rates, event-free survival, and overall survival were similar in all groups. Although patients with pre-existing liver and/or renal dysfunction might have a higher rate of hematologic toxicity and require more frequent dose reductions, most patients can be adequately managed, resulting in response rates and survival similar to those without pre-existing organ dysfunction.


Radioimmunoassay determination of formononetin in murine plasma and mammary glandular tissue.
Formononetin is an isoflavonoid phytoestrogen found in animal fodder and in certain human foodstuffs. Since high consumption of formononetin has been implicated in infertility among ruminants and may relate to human breast cancer, we developed a radioimmunoassay method and applied it to quantitate formononetin in murine plasma and mammary glandular tissue for animal model studies. The radioimmunoassay utilized an antiserum raised in rabbits following immunization with formononetin-7-O(carboxymethyl)ether coupled to bovine serum albumin. The tracer was an [3H]leucine derivative of formononetin synthesized by mixed anhydride reaction between formononetin-7-O(carboxymethyl)ether and [3H]leucine. The bound and free forms of formononetin were separated by adding dextran-coated charcoal. This radioimmunoassay procedure enabled the quantification of 4 ng/ml of plasma or 50 pg/mg of mammary tissue, and the antiserum showed no marked cross-reaction with the reactants tested. The reliability and reproducibility of the assay were demonstrated by intra- and inter-assay variation that was 6.5% and 11.9%, respectively. This radioimmunoassay was compared with a high-performance liquid chromatographic method by determining concentrations of formononetin in ethanol extracts of red clover. Good correlation existed between the radioimmunoassay and high-performance liquid chromatographic method (r = 0.980), but the radioimmunoassay values were on the average 5% higher than high-performance liquid chromatographic values. Furthermore, we used the radioimmunoassay to assess the pharmacokinetics of formononetin in castrated female BALB/c mice after a single subcutaneous injection at 40 mg/kg dose. The sensitivity of the radioimmunoassay permitted the detection of a prolonged elimination phase for formononetin in both plasma and mammary glandular tissues, with mean elimination half-lives of 2 and 2.5 hr, respectively. A specific, sensitive, and fast radioimmunoassay procedure has been developed for the determination of formononetin in murine plasma and mammary glandular tissue. We conclude that the presented radioimmunoassay is a valid alternative method to the quantification of formononetin in biological fluids and/or plants.


Chromatin modification by olive phenolics: In silico molecular docking studies utilising the phenolic groups categorised in the OliveNet™ database against lysine specific demethylase enzymes.
Extra virgin olive oil is the principal source of dietary fat in the Mediterranean diet and is considered to have beneficial health effects. There is evidence to suggest that the phenolic compounds within Olea europaea have the ability to inhibit lysine-specific demethylase 1 (LSD1). This is an epigenetic enzyme that removes methyl groups from histone proteins and regulates gene transcription. Conversely, SET domain-containing protein 7 (SETD7) has opposing enzymatic activity and is a histone methyltransferase. Due to the involvement of these proteins in a number of pathological processes, including cancer and diabetes, further research needs to be conducted into the way in which they can be targeted. A large number of phenolic compounds (>200) have been identified in Olea europaea. To help expedite the discovery of promising lead compounds, in this study, in silico molecular docking methods were used to investigate the molecular binding properties of the phenolic compounds obtained from the OliveNet™ database to LSD1 and its variants, LSD2, and SETD7. Numerous Olea europaea phenolic compounds were predicted to bind to the epigenetic enzymes and several had stronger binding affinities than the LSD1 and SETD7 positive control inhibitors. The protein-ligand interactions of the phenolic compounds were also compared to known inhibitors and the molecular docking results suggest that the flavonoids, secoiridoids and glucosides may bind particularly strongly to the epigenetic regulators. Overall, several ligands were identified as lead compounds from this research and their potential inhibitory activity could be validated further in the laboratory.