In vitro activity of temocillin (BRL 17421), a novel beta-lactam antibiotic.
The minimal inhibitory concentration of temocillin (BRL 17421) against 476 clinical isolates was determined by an agar dilution method. Temocillin was active against most of the Enterobacteriaceae, Haemophilus, and Neisseria strains tested. The compound showed low activity or was inactive against Bacteroides, Campylobacter, Acinetobacter, Pseudomonas, and Staphylococcus aureus strains. Within each species, individual strains showed a high degree of uniformity in their susceptibility to temocillin; the drug concentrations that inhibited the growth of 90% of organisms were the same or close to those which inhibited the growth of 50% of organisms. In contrast, the same strains showed a very wide range of susceptibility to the other antibiotics tested, including third-generation cephalosporins. Against strains of Enterobacteriaceae highly susceptible to third-generation cephalosporins, temocillin was considerably less active than cefotaxime, ceftazidime, and moxalactam, although it was more active than cefazolin and piperacillin. Against certain strains of Enterobacter and Citrobacter resistant to third-generation cephalosporins, temocillin was more active than cefotaxime and ceftazidime. An increase in the inoculum size did not alter the activity of temocillin, indicating that the compound has high stability to beta-lactamases. The minimal lethal concentration was also very similar to the minimal inhibitory concentration when an inoculum of 10(5) colony-forming units was used.


Role of Energy Metabolism in the Brown Fat Gene Program.
In murine and human brown adipose tissue (BAT), mitochondria are powerful generators of heat that safely metabolize fat, a feature that has great promise in the fight against obesity and diabetes. Recent studies suggest that the actions of mitochondria extend beyond their conventional role as generators of heat. There is mounting evidence that impaired mitochondrial respiratory capacity is accompanied by attenuated expression of Ucp1 and other BAT-selective genes, implying that mitochondria exert transcriptional control over the brown fat gene program. In this review, we discuss the current understanding of brown fat mitochondria, their potential role in transcriptional control of the brown fat gene program, and potential strategies to treat obesity in humans by leveraging thermogenesis in brown adipocytes.


Reduced MIP-1β as a Trait Marker and Reduced IL-7 and IL-12 as State Markers of Anorexia Nervosa.
Alterations in certain inflammatory markers have been found in individuals with anorexia nervosa (AN). However, their relation to clinical characteristics has not been extensively explored, nor is it clear whether they are trait or state features of the disorder. This cross-sectional study measured serum concentrations of 36 inflammatory markers in people with acute AN (n = 56), recovered AN (rec-AN; n = 24) and healthy controls (HC; n = 51). The relationship between body mass index (BMI), eating disorder psychopathology, depression symptoms and inflammatory markers was assessed. Statistical models controlled for variables known to influence cytokine concentrations (i.e., age, ethnicity, smoking status and medication usage). Overall, most inflammatory markers including pro-inflammatory cytokines were unchanged in AN and rec-AN. However, in AN and rec-AN, concentrations of macrophage inflammatory protein (MIP)-1β were lower than HCs. Interleukin (IL)-7 and IL-12/IL-23p40 were reduced in AN, and concentrations of macrophage-derived chemokine, MIP-1α and tumor necrosis factor-α were reduced in rec-AN compared to HC. In conclusion, a reduction in MIP-1β may be a trait marker of the illness, whereas reductions in IL-7 and IL-12/IL-23p40 may be state markers. The absence of increased pro-inflammatory cytokines in AN is contradictory to the wider literature, although the inclusion of covariates may explain our differing findings.


Nuclear diacylglycerol kinase-theta is activated in response to alpha-thrombin.
Currently, there is substantial evidence that nuclear lipid metabolism plays a critical role in a number of signal transduction cascades. Previous work from our laboratory showed that stimulation of quiescent fibroblasts with alpha-thrombin leads to the production of two lipid second messengers in the nucleus: an increase in nuclear diacylglycerol mass and an activation of phospholipase D, which catalyzes the hydrolysis of phosphatidylcholine to generate phosphatidic acid. Diacylglycerol kinase (DGK) catalyzes the conversion of diacylglycerol to phosphatidic acid, making it an attractive candidate for a signal transduction component. There is substantial evidence that this activity is indeed regulated in a number of signaling cascades (reviewed by van Blitterswijk, W. J., and Houssa, B. (1999) Chem. Phys. Lipids 98, 95-108). In this report, we show that the addition of alpha-thrombin to quiescent IIC9 fibroblasts results in an increase in nuclear DGK activity. The examination of nuclei isolated from quiescent IIC9 cells indicates that DGK-theta and DGK-delta are both present. We took advantage of the previous observations that phosphatidylserine inhibits DGK-delta (reviewed by Sakane, F., Imai, S., Kai, M., Wada, I., and Kanoh, H. (1996) J. Biol. Chem. 271, 8394-8401), and constitutively active RhoA inhibits DGK-theta (reviewed by Houssa, B., de Widt, J., Kranenburg, O., Moolenaar, W. H., and van Blitterswijk, W. J. (1999) J. Biol. Chem. 274, 6820-6822) to identify the activity induced by alpha-thrombin. Constitutively active RhoA inhibited the nuclear stimulated activity, whereas phosphatidylserine did not have an inhibitory effect. In addition, a monoclonal anti-DGK-theta antibody inhibited the alpha-thrombin-stimulated nuclear activity in vitro. These results demonstrate that DGK-theta is the isoform responsive to alpha-thrombin stimulation. Western blot and immunofluorescence microscopy analyses showed that alpha-thrombin induced the translocation of DGK-theta to the nucleus, implicating that this translocation is at least partly responsible for the increased nuclear activity. Taken together, these data are the first to demonstrate an agonist-induced activity of nuclear DGK-theta activity and a nuclear localization of DGK-delta.


Temporal analysis of individual ethanol consumption in socially housed mice and the effects of oxytocin.
The majority of preclinical studies assessing treatments for alcohol use disorder use singly housed animals. Because social factors affect ethanol intake, studies investigating such treatments in group-housed animals are needed. We investigated the effects of repeated oxytocin treatment on ethanol intake in socially housed male and female C57BL/6J mice. We used the novel "Herdsman" system implementing radiotracking technology to measure individual ethanol intake in group-housed animals. Mice were housed in same-sex groups of 4 per cage and exposed to 3 and 6% ethanol solutions. After baseline drinking was established, half of the animals in each cage received repeated intraperitoneal injections of 3 mg/kg oxytocin. During baseline, females consumed more ethanol than males partly due to greater number of ethanol drinks taken by females. We also observed a gradual development of two peaks of ethanol consumption during the dark phase of the circadian cycle. The effects of oxytocin treatment were short-acting and varied across treatment days. Oxytocin significantly decreased ethanol intake on three out the four treatment days. On the fourth treatment day, oxytocin decreased ethanol intake and water intake. The greater intake of ethanol in female mice is associated with the number of drinks taken. Oxytocin treatments not only cause an acute decrease in ethanol consumption, but can also change in efficacy over time. While the oxytocin system remains a promising therapeutic target for alcoholism, studies investigating longer periods of repeated oxytocin treatment and those using additional oxytocin receptor agonists are warranted.


Novel semantic similarity measure improves an integrative approach to predicting gene functional associations.
Elucidation of the direct/indirect protein interactions and gene associations is required to fully understand the workings of the cell. This can be achieved through the use of both low- and high-throughput biological experiments and in silico methods. We present GAP (Gene functional Association Predictor), an integrative method for predicting and characterizing gene functional associations. GAP integrates different biological features using a novel taxonomy-based semantic similarity measure in predicting and prioritizing high-quality putative gene associations. The proposed similarity measure increases information gain from the available gene annotations. The annotation information is incorporated from several public pathway databases, Gene Ontology annotations as well as drug and disease associations from the scientific literature. We evaluated GAP by comparing its prediction performance with several other well-known functional interaction prediction tools over a comprehensive dataset of known direct and indirect interactions, and observed significantly better prediction performance. We also selected a small set of GAP's highly-scored novel predicted pairs (i.e., currently not found in any known database or dataset), and by manually searching the literature for experimental evidence accessible in the public domain, we confirmed different categories of predicted functional associations with available evidence of interaction. We also provided extra supporting evidence for subset of the predicted functionally-associated pairs using an expert curated database of genes associated to autism spectrum disorders. GAP's predicted "functional interactome" contains ≈1M highly-scored predicted functional associations out of which about 90% are novel (i.e., not experimentally validated). GAP's novel predictions connect disconnected components and singletons to the main connected component of the known interactome. It can, therefore, be a valuable resource for biologists by providing corroborating evidence for and facilitating the prioritization of potential direct or indirect interactions for experimental validation. GAP is freely accessible through a web portal: http://ophid.utoronto.ca/gap.


Treatment of acute myeloid leukemia in children: experience from a tertiary care hematology centre in India.
To assess the treatment and outcome of children with acute myeloid leukemia. The Primary objectives were to assess remission rates, treatment related toxicity and disease free survival. Secondary objective was to assess prognostic factors associated with poor outcome. A retrospective analysis of all treated patients with acute myeloid leukemia, less than 18 year of age from Sept 2005 to Aug 2009 was done. Clinical laboratory, treatment and follow up records retrieved to calculate remission rate, treatment related toxicity, disease free survival and poor prognostic factors. This analysis included thirty five patients (male : female; 23:12), twenty seven (77.1%) achieved remission after one 3 + 7 induction and seven required two inductions. High dose cytosine arabinoside consolidation was given in thirty one patients while one underwent allogenic stem cell transplantation. Two patients died during chemotherapy (TRM- 5.7%), two did not complete the therapy, seventeen relapsed (48.5%) with 80% of relapses occurring within first year of remission and no relapse occurred after 2 years. Fourteen patients are in remission (40%, follow up 5-54 months) and cumulative median disease free survival is of 13 months. The present data suggests that 3 + 7 induction, followed by high dose cytarabine consolidation has low treatment related toxicity and resource utilization; however, relapse free survival is inferior to more intensive regimens, highlighting the need to intensify chemotherapy regimen once the treatment related mortality has been minimized.


New insights into the association between lameness, behavior, and performance in Simmental cows.
In a herd of 100 milking Simmental cows, data of performance and behavior parameters were collected automatically with different systems such as pedometers, an automatic milking system, and automatic weighing troughs for 1 yr. Performance measures were several milking-related parameters, live weight, as well as feed intake. Behavior-associated measures were feeding behavior (e.g. feeding duration, number of visits to the trough, and feeding pace) as well as activity such as lying duration, number of lying bouts, and overall activity. In the same time, lameness status of every cow was assessed with weekly locomotion scoring. According to the score animals were then classified lame (score 4 or 5) or nonlame (score 1, 2, or 3). From these data in total, 25 parameters summarized to daily values were evaluated for their ability to determine the lameness status of a cow. Data were analyzed with a regularized regression method called elastic net with the outcome lame or nonlame. The final model had a high prediction accuracy with an area under the curve of 0.91 [95% confidence interval (CI) = 0.88-0.94]. Specificity was 0.81 (95% CI = 0.73-0.85) and sensitivity was 0.94 (95% CI = 0.88-1.00). The most important factors associated with a cow being lame were number of meals, average feed intake per meal, and average duration of a meal. Lame cows fed in fewer and shorter meals with a decreased intake per meal. Milk yield and lying-behavior-associated parameters were relevant in the model, too, but only as parts of interaction terms demonstrating their strong dependence on other factors. A higher milk yield only resulted in higher risk of being lame if feed intake was decreased. The same accounts for lying duration: only if lying time was below the 50% quantile did an increased milk yield result in a higher risk of being lame. The association of lameness and daily lying duration was influenced by daily feeding duration and feeding duration at daytime. The results of the study give deeper insights on how the association between behavior and performance parameters and lameness is influenced by intrinsic factors in particular and that many of these have to be considered when trying to predict lameness based on such data. The findings lead to a better understanding why, for instance, lying duration or milk yield seem to be highly correlated with lameness in cows but still have not been overly useful as parameters in other lameness detection models.


Extended Time Accommodation and the Academic, Behavioral, and Psychological Outcomes of Students With Learning Disabilities.
This study analyzed performance, process, and survey data of eighth graders with learning disabilities (LDs) who took the 2017 National Assessment of Educational Progress (NAEP) digital math test. Compared with students with LDs who did not receive extended time accommodations (ETAs), students with LDs who received and used ETA scored significantly higher on the test, whereas students with LDs who received but did not use ETA scored significantly lower on the test. In addition, students with LDs in the two ETA groups reported a lower level of perceived time pressure and a higher level of math interest and enjoyment than their peers who did not receive ETA. For students with LDs who received ETA, optimal performance was achieved with 50% additional time, while their peers who did not receive ETA typically performed best when utilizing most of their allotted time. The analysis of process data revealed that students with LDs who used ETA performed more actions, had a higher number of revisits, used universal design digital tools more frequently, and performed better on time-consuming items than their peers who did not receive ETA at the same level of math performance.


18F-Choline PET/CT Identifies High-Grade Prostate Cancer Lesions Expressing Bone Biomarkers.
The main aim of this study was to investigate the possible association between 18F-choline uptake and histopathological features of prostate biopsies such as the Gleason Group and the expression of both epithelial to mesenchymal transition (vimentin) and bone mineralization (bone morphogenetics protein (BMP)-2, runt-related transcription factor 2 (RUNX2), receptor activator of nuclear factor-κB ligand (RANKL), vitamin D receptor (VDR), and pentraxin 3 (PTX3) in situ biomarkers. To this end, we enrolled 79 consecutive prostate cancer patients that underwent both the 18F-choline PET/CT analysis and the prostate bioptic procedure. The standardized uptake value (SUV) average values were collected from 18F-choline PET/CT analysis whereas Gleason Group and immunostaining data were collected from paraffin-embedded sections. Histological classification showed a heterogenous population including both low/intermediate and high-grade prostate cancers. A significant increase of 18F-choline uptake in high-grade prostate lesions (Gleason Score ≥8) was found. Also, linear regression analysis showed a significant correlation between 18F-choline uptake and the number of vimentin, RANKL, VDR, or PTX3 positive prostate cancer cells. Conversely, we observed no significant association between 18F-choline uptake and the expression of bone biomarkers involved in the early phases of osteoblast differentiation (BMP-2, RUNX2). In conclusion, results here reported can lay the foundation for the use of 18F-choline positron emission tomography (PET)/computed tomography (CT) as a diagnostic tool capable of identifying high-grade prostate cancer lesions expressing bone biomarkers.


Differential gene expression in murine large cell B-cell lymphoma metastatic variants.
Previous studies from this laboratory have characterized RAW117-P murine large cell B-cell lymphoma and its in vivo selected highly malignant and liver metastatic RAW117-H10 subline for their biological and biochemical properties. In this study, to understand the molecular basis of low and high metastatic behavior of these variant sublines, we have investigated the molecular phenotypes of these cells using differential display techniques and cDNA array analysis. Differential display analysis indicated a significant difference in expression of several genes between these two metastatic variant lymphoma cells. Further analyses of these cells using microarray showed an increased expression of several genes including uPAR1, CRE-BP1, Chop-10, IGF, insulin-like growth factor-IA, STAT6, Cyclin-D1, Cyclin-E, ERBB-3, Alpha NGF, Kruppel-like factor LKLF, (P)19INK4 in metastatic RAW117-H10 cells compared to parental RAW117-P cells. On the other hand, MIP1beta, CD14 antigen, Cathepsin B and MOD are expressed more in RAW117-P cells compared to RAW117-H10 cells. Differential expression of the selected genes was confirmed using semiquantitative RT-PCR techniques. The combination of plasminogen activator and its receptor and IGF-like growth factors, cell cycle regulatory molecules and transcription factors might provide an ideal environment for RAW117-H10 cells to metastasize to distant organs and colonize. Thus these results identify certain differentially expressed genes that are involved in the metastatic properties of these lymphoma cells and lay foundation for further in depth analyses to use this information to develop therapy for metastatic lymphoma.


Duplication as a source of genetic innovation for «antiviral genes».
The antiviral proteins, also known as restriction factors, are the primary cellular defense against viral pathogens. These proteins from the innate immune system are in direct interactions with viral proteins. These antagonistic interactions come in two flavors: restriction factors are able to directly target the virus to restrict its replication and/or they may be the target of viral antagonists. Such long-term antagonistic virus-host interactions have set up an evolutionary "arms-race" between the two adversarial entities. This genetic conflict leads to a rapid evolution of viruses and antiviral genes. In particular, amino acids at the virus-host interface change more frequently than expected over time. On the other hand, more drastic host genome modifications have also been selected over time to counteract the rapid evolution of viruses. Reflecting the fact that numerous restriction factors belong to a gene family, it appears that duplication of antiviral genes has occurred frequently during the course of evolution. Here, we will review how duplication of antiviral genes has been selected in the host and how the divergence and plasticity of the duplicated genes may have been advantageous in the virus-host genetic conflict. We will also briefly expose the limits to such innate immune gene expansion.


Treatment Course and Outcomes of Intracranial Teratomas in Pediatric Patients: A Retrospective 15-Year Case Series Study.
There is no standard treatment paradigm for intracranial teratomas, a rare subset of primary intracranial non-germinomatous germ cell tumors (NGGCT), which comprise less than 1% of pediatric brain tumors. This case series retrospectively analyzes treatment and outcomes of pediatric intracranial teratomas from a single institution. Authors reviewed a comprehensive pathology database at Stanford's Lucile Packard Children's Hospital for intracranial teratomas in pediatric patients treated from 2006 to 2021; their demographics, treatment, and clinical course were analyzed. Among 14 patients, median follow-up time was 4.6 years and mean age at diagnosis was 10.5 years. Ten had elevated tumor markers and underwent chemotherapy as initial treatment for NGGCT. Ultimately, these patients all required surgery for progressive or residual disease. Two patients did not undergo radiation. After biopsy or resection, 8 patients had pure mature teratoma, five had mixed germ cell tumor with teratoma component, and one had immature teratoma. The patient with immature teratoma died during chemotherapy from septic shock. No patients experienced recurrence. Common sequelae were endocrine (42.8%) and eye movement (50.0%) abnormalities. We highlight the variable treatment course and outcome for pediatric patients with intracranial teratomas. Elevated tumor markers at presentation, along with imaging findings, favor chemotherapy initiation for presumed NGGCT. Resection of residual tumor is recommended even if tumor markers return to normal. Prognosis remains excellent; no patients had recurrence with a median follow-up of 4.6 years.


Cardiac electrical conduction, autonomic activity and biomarker release during recovery from prolonged strenuous exercise in trained male cyclists.
Although markers of myocyte injury, electrolyte disturbances and an autonomic imbalance have been reported following exercise, the effect of prolonged strenuous activity on cardiac electrical conduction is not well understood. This study examined atrial and ventricular conduction dynamics during recovery from exercise. Electrocardiographic intervals were obtained from eight highly-trained males before, during recovery (15, 30, 45 and 60 min post-exercise) and 24 h after a prolonged bout of strenuous exercise. Time-domain, frequency-domain and non-linear analyses of the RR, PR and QT intervals were analysed to investigate the effect of exercise on autonomic modulation and cardiac electrical conduction. Serum electrolyte and high-sensitivity cardiac troponin T (hs-cTnT) concentrations were measured before exercise, and after 60 min and 24 h of recovery. The root mean square of the successive differences of RR, PR and QT intervals was significantly reduced during recovery (p < 0.05). Normalised low- and high-frequency power of RR intervals significantly increased and decreased, respectively, during recovery. Approximate entropy of PR and QT intervals, and the QT-variability index significantly increased during recovery. All measures except mean QT interval (pre 422 ± 10 ms vs 24 h post 442 ± 11 ms, p = 0.013) returned to pre-exercise values after 24 h. Serum hs-cTnT was significantly elevated 60 min after exercise (pre 5.2 ± 0.7 ng L(-1) vs 60 min post 27.4 ± 6.2 ng L(-1), p = 0.01) and correlated with exercising heart rate (R(2) = 0.89, p < 0.001). Serum electrolyte concentrations were unchanged (p > 0.05). The results suggest suppressed parasympathetic and/or sustained sympathetic modulation of heart rate during recovery, concomitant with perturbations in atrial and ventricular conduction dynamics. Exercise-induced hs-cTnT release was heart rate dependent.


Tai Chi Chuan optimizes the functional organization of the intrinsic human brain architecture in older adults.
Whether Tai Chi Chuan (TCC) can influence the intrinsic functional architecture of the human brain remains unclear. To examine TCC-associated changes in functional connectomes, resting-state functional magnetic resonance images were acquired from 40 older individuals including 22 experienced TCC practitioners (experts) and 18 demographically matched TCC-naïve healthy controls, and their local functional homogeneities across the cortical mantle were compared. Compared to the controls, the TCC experts had significantly greater and more experience-dependent functional homogeneity in the right post-central gyrus (PosCG) and less functional homogeneity in the left anterior cingulate cortex (ACC) and the right dorsal lateral prefrontal cortex. Increased functional homogeneity in the PosCG was correlated with TCC experience. Intriguingly, decreases in functional homogeneity (improved functional specialization) in the left ACC and increases in functional homogeneity (improved functional integration) in the right PosCG both predicted performance gains on attention network behavior tests. These findings provide evidence for the functional plasticity of the brain's intrinsic architecture toward optimizing locally functional organization, with great implications for understanding the effects of TCC on cognition, behavior and health in aging population.


Advances on electrochemical disinfection research: Mechanisms, influencing factors and applications.
Disinfection, a vital barrier against pathogenic microorganisms, is crucial in halting the spread of waterborne diseases. Electrochemical methods have been extensively researched and implemented for the inactivation of pathogenic microorganisms from water and wastewater, primarily owing to their simplicity, efficiency, and eco-friendliness. This review succinctly outlined the core mechanisms of electrochemical disinfection (ED) and systematically examined the factors influencing its efficacy, including anode materials, system conditions, and target species. Additionally, the practical application of ED in water and wastewater treatment was comprehensively reviewed. Case studies involving various scenarios such as drinking water, hospital wastewater, black water, rainwater, and ballast water provided concrete instances of the expansive utility of ED. Finally, coupling ED with other technologies and the resulting synergies were introduced as pivotal foundations for subsequent engineering advancements.


Plant and microbial features governing an endophytic lifestyle.
Beneficial microorganisms colonizing internal plant tissues, the endophytes, support their host through plant growth promotion, pathogen protection, and abiotic stress alleviation. Their efficient application in agriculture requires the understanding of the molecular mechanisms and environmental conditions that facilitate in planta accommodation. Accumulating evidence reveals that commensal microorganisms employ similar colonization strategies as their pathogenic counterparts. Fine-tuning of immune response, motility, and metabolic crosstalk accounts for their differentiation. For a holistic perspective, in planta experiments with microbial collections and comprehensive genome data exploration are crucial. This review describes the most recent findings on factors involved in endophytic colonization processes, focusing on bacteria and fungi, and discusses required methodological approaches to unravel their relevance within a community context.


Impact of ADHD and its treatment on substance abuse in adults.
Attention-deficit/hyperactivity disorder (ADHD) is a risk factor for substance abuse in adults. Additional psychiatric comorbidity increases this risk. ADHD is associated with different characteristics of substance abuse: substance abuse transitions more rapidly to dependence, and lasts longer in adults with ADHD than those without ADHD. Self-medication may be a factor in the high rate of substance abuse in adults with ADHD. While previous concerns arose whether stimulant therapy would increase the ultimate risk for substance abuse, recent studies have indicated that pharmacologic treatment appears to reduce the risk of substance abuse in individuals with ADHD. When treating adults with ADHD and substance abuse, clinicians should assess the relative severity of the substance abuse, the symptoms of ADHD, and any other comorbid disorders. Generally, stabilizing or addressing the substance abuse should be the first priority when treating an adult with substance abuse and ADHD. Treatment for adults with ADHD and substance abuse should include a combination of addiction treatment/psychotherapy and pharmacotherapy. The clinician should begin pharmacotherapy with medications that have little likelihood of diversion or low liability, such as bupropion and atomoxetine, and, if necessary, progress to the stimulants. Careful monitoring of patients during treatment is necessary to ensure compliance with the treatment plan.


KDM2A Deficiency in the Liver Promotes Abnormal Liver Function and Potential Liver Damage.
Dysregulation of metabolic functions in the liver impacts the development of diabetes and metabolic disorders. Normal liver function can be compromised by increased inflammation via the activation of signaling such as nuclear factor (NF)-κB signaling. Notably, we have previously identified lysine demethylase 2A (KDM2A)-as a critical negative regulator of NF-κB. However, there are no studies demonstrating the effect of KDM2A on liver function. Here, we established a novel liver-specific Kdm2a knockout mouse model to evaluate KDM2A's role in liver functions. An inducible hepatic deletion of Kdm2a, Alb-Cre-Kdm2afl/fl (Kdm2a KO), was generated by crossing the Kdm2a floxed mice (Kdm2afl/fl) we established with commercial albumin-Cre transgenic mice (B6.Cg-Tg(Alb-cre)21Mgn/J). We show that under a normal diet, Kdm2a KO mice exhibited increased serum alanine aminotransferase (ALT) activity, L-type triglycerides (TG) levels, and liver glycogen levels vs. WT (Kdm2afl/fl) animals. These changes were further enhanced in Kdm2a liver KO mice in high-fat diet (HFD) conditions. We also observed a significant increase in NF-κB target gene expression in Kdm2a liver KO mice under HFD conditions. Similarly, the KO mice exhibited increased immune cell infiltration. Collectively, these data suggest liver-specific KDM2A deficiency may enhance inflammation in the liver, potentially through NF-κB activation, and lead to liver dysfunction. Our study also suggests that the established Kdm2afl/fl mouse model may serve as a powerful tool for studying liver-related metabolic diseases.


31P-NMR study of dog submandibular gland in vivo and in vitro using the topical magnetic resonance.
The topical magnetic resonance (TMR) was used to study the phosphorous metabolism of a dog's submandibular gland during acetylcholine stimulation. The experiments were carried out under two conditions. One was a normal blood supply (in vivo) and the other was vascular perfusion with the artificial perfusate (in vitro). The levels of phosphorous compounds in the gland were obtained for 31P-NMR spectra of the TMR. The saliva was secreted in the both conditions, though the salivary flow under the in vivo condition was higher than in vitro. During salivary secretion, only the levels of creatine phosphate decreased under in vivo condition, while the levels of creatine phosphate and ATP decreased and the levels of inorganic phosphate increased in in vitro cases. It was concluded that the decrement of creatine phosphate during the salivary secretion indicates the acceleration of the energy metabolism in the submandibular gland.


Non-invasive vagus nerve stimulation in healthy humans reduces sympathetic nerve activity.
Vagus nerve stimulation (VNS) is currently used to treat refractory epilepsy and is being investigated as a potential therapy for a range of conditions, including heart failure, tinnitus, obesity and Alzheimer's disease. However, the invasive nature and expense limits the use of VNS in patient populations and hinders the exploration of the mechanisms involved. We investigated a non-invasive method of VNS through electrical stimulation of the auricular branch of the vagus nerve distributed to the skin of the ear--transcutaneous VNS (tVNS) and measured the autonomic effects. The effects of tVNS parameters on autonomic function in 48 healthy participants were investigated using heart rate variability (HRV) and microneurography. tVNS was performed using a transcutaneous electrical nerve stimulation (TENS) machine and modified surface electrodes. Participants visited the laboratory once and received either active (200 μs, 30 Hz; n = 34) or sham (n = 14) stimulation. Active tVNS significantly increased HRV in healthy participants (P = 0.026) indicating a shift in cardiac autonomic function toward parasympathetic predominance. Microneurographic recordings revealed a significant decrease in frequency (P = 0.0001) and incidence (P = 0.0002) of muscle sympathetic nerve activity during tVNS. tVNS can increase HRV and reduce sympathetic nerve outflow, which is desirable in conditions characterized by enhanced sympathetic nerve activity, such as heart failure. tVNS can therefore influence human physiology and provide a simple and inexpensive alternative to invasive VNS.


Aggregation and microparticle production through toll-like receptor 4 activation in platelets from recently menopausal women.
Bacterial infection may increase risk for thrombosis and atherosclerosis. Human platelets express toll-like receptor 4 (TLR4), the receptor for gram-negative bacterial lipopolysaccharide (LPS). Experiments were designed to evaluate direct, acute effects of TLR4 activation on aggregation, secretion, and generation of prothrombogenic microparticles in vitro on platelets derived from healthy women at risk for development of cardiovascular disease because of their hormonal status. Platelet-rich plasma from recently menopausal women was incubated with ultrapure Escherichia coli LPS in the absence or presence of antibodies that neutralize the human TLR4. Incubating platelets with LPS (100 ng/mL) for 5 minutes decreased aggregation and dense granule adenosine triphosphate secretion induced by thrombin receptor agonist peptide (TRAP) but not by adenosine diphosphate or collagen. The antibody to TLR4 blocked this effect of LPS. TLR4 activation increased phosphorylation of p38 mitogen-activated protein kinase and decreased production of prothrombotic phosphatidylserine and P-selectin-positive microparticles in response to TRAP. Therefore, acute, direct activation of TLR4 reduces platelet reactivity to TRAP stimulation in vitro. Increased thrombotic and cardiovascular risk with bacterial infection most likely reflects the sum of TLR4 activation on other blood and vascular cells to release proinflammatory cytokines/chemokines, which indirectly affect platelet reactivity.


Performing a percutaneous liver biopsy in parenchymal liver diseases.
There are many ways of obtaining a liver biopsy sample but the percutaneous method is deemed as one of the simplest and safest methods. Percutaneous liver biopsy (PLB) is a commonly performed procedure carried out for the diagnosis and management of patients with parenchymal liver diseases. It plays a central role in providing histological assessment for either diagnostic or therapeutic purposes. This article describes how PLB is performed at the Royal Free Hospital radiology department, London, under ultrasound guidance, along with indications, contraindications, complications and advantages/disadvantages this method offers. Nursing implications are discussed in terms of assisting, caring and managing for these patients safely. This article aims to raise awareness of PLB and inform the reader how this tissue sample is taken, thus enabling further understanding of this procedure. It is concluded that the percutaneous route of obtaining a liver biopsy enables a good size and quality of sample to be taken in a safe and effective manner, with usually one pass being required with minimal associated complications.


Leyla loop: a time-saving suture technique for robotic atrial closure.
The longer durations of cardiopulmonary bypass and aortic cross-clamp times remain the disadvantages of robotic or minimally invasive cardiac surgery. For this reason, every small contribution to speeding up these procedures is of the utmost importance. Here, we present a practical, easy and time-saving suture technique for atrial closure. It consists of a hand-made loop at one end of the suture and saves the time otherwise consumed by knotting. It may also be used during conventional or minimally invasive cardiac surgery.


Sexual isolation with and without ecological isolation in marine isopods Jaera albifrons and J. praehirsuta.
Sexual barriers associated with mate choice are often found to be associated with some level of ecological isolation between species. The independence and relative strength of sexual isolation are thus difficult to assess. Here, we take advantage of a pair of marine isopod species (Jaera albifrons and J. praehirsuta) that show sexual isolation and coexist in populations where they share the same microhabitat or not (i.e. without or with ecological isolation). We estimated the strength of sexual isolation between J. albifrons and J. praehirsuta using no-choice trials and a multiple-choice experimental population. We found that sexual isolation is strong in both the presence and the absence of ecological isolation, but that it is asymmetric and fails to prevent interspecific gene flow entirely. First-generation intrinsic post-zygotic barriers were low, and there was no sexual isolation within J. praehirsuta across habitats. The J. albifrons/J. praehirsuta species pair thus provides an example where the role of sexual isolation as a barrier to gene flow (a) does not depend upon current ecological isolation, (b) seems to have evolved independently of local ecological conditions, but (c) is insufficient to complete speciation entirely on its own.


Individualization of long-term enzyme replacement therapy for Gaucher disease.
Gaucher disease, the most common lysosomal storage disorder, is a heterogeneous condition affecting multiple organ systems. Patients with nonneuronopathic (type 1) Gaucher disease may suffer from hepatomegaly, splenomegaly, thrombocytopenia, bleeding tendencies, anemia, hypermetabolism, skeletal pathology, growth retardation, pulmonary disease, and decreased quality of life. Enzyme replacement therapy (ERT) with mannose-terminated glucocerebrosidase (imiglucerase, Cerezyme, Genzyme Corporation, Cambridge, MA) reverses or ameliorates many of the manifestations of type 1 Gaucher disease. However, due to the variable pattern and severity of disease, and the uncertain manner of progression, implementation of treatment, choice of initial and maintenance imiglucerase dose, and evaluation of the therapeutic response must be tailored to the individual patient. For the past 14 years, the US Regional Coordinators of the International Collaborative Gaucher Group have individually and collectively developed extensive clinical experience in managing patients with Gaucher disease. In this review, we present recommendations for initial imiglucerase treatment and subsequent dose adjustments based on a schedule of regular assessment and monitoring, and achievement and maintenance of defined therapeutic goals.


[Secondary uroliths after surgical correction of urinary bladder extrophy].
The analysis of long-term results of complex surgical correction of urinary bladder extrophy in 83 patients has shown that 21 (25.3%) patients had concrements in the urethra. Multiple bacteriological tests detected urease-producing flora which was managed with specific antibacterial therapy. Two patients were exposed to extracorporeal shock-wave lithotripsy. Low-invasive lithotripsy was low effective and all the patients were operated. Chemical structure of the removed concrements was determined with x-ray difractometry. Recurrent concrements were found in 7 (33.3%) of 21 patients who had not received specific antibacterial therapy. All the stones detected were struvite. Thus, struvite concrements' risk in patients after surgical correction of urinary bladder extrophy can be reduced under the following conditions: suppression of urease-producing flora activity, catheterization regime compliance, systemic sanation of the artificial urinary bladder.


A study of the interaction between bovine cardiac-muscle cyclic AMP-dependent protein kinase and cyclic AMP using fluorescence-polarization spectroscopy.
The C-subunit of type II cyclic AMP-dependent protein kinase from bovine heart was labelled with the fluorophore fluorescamine (FAM). The association of the dye-labelled subunit (CFAM) with the R-subunit isolated from the same source was monitored by fluorescence polarization spectroscopy. The stoichiometry of C to R in the final complex was close to 1:1. The affinity of the two subunits could be described by a dissociation constant in the nanomolar range. Holoenzyme (formed from CFAM and R) was titrated with cyclic AMP, and the changes in fluorescence anisotropy, due to dissociation of the holoenzyme, recorded. The titration curves were analysed in terms of a model which required computer simulation. Cyclic AMP-induced dissociation proceeds via one or more ternary complexes, and all four cyclic AMP-binding sites on the R-dimer are accessible in the holoenzyme. The dissociation constants describing the release of the C-subunits from the two ternary complexes containing four cyclic AMP molecules were both approx. 9 microM. The binding of two cyclic AMP molecules to protein kinase is necessary and sufficient to cause the dissociation of both C-subunits. The state of association at 'in vivo' concentrations of protein and cyclic AMP is discussed.


Clinical practice: heart failure in children. Part II: current maintenance therapy and new therapeutic approaches.
The current maintenance treatment for children with heart failure remains controversial: To a large extent, it is based on extrapolation of data derived from trials in adult populations. There are only a few randomized trials focused on the treatment of children with cardiac disease, especially in the subgroup with cardiomyopathy and heart failure. The goals of therapy are to maintain circulatory and end-organ function and to allow for recovery and reverse remodeling to occur. When maintenance therapy fails and medical treatment does not result in clinical improvement, the alternative of device therapy must be considered: In that case, the usual aim is to stabilize circulatory status, as a bridge to either recovery or to cardiac transplantation. Recently, carefully selected patients with electromechanical dyssynchrony of ventricular systolic function have demonstrated a benefit from biventricular pacing devices (cardiac resynchronization therapy), with improved functional capacity and quality of life and, in some patients, avoidance of the need for transplantation.


Primary culture of adult mouse olfactory receptor neurons.
Olfactory receptor neurons (ORNs) are unique because they can be replaced by stem cells throughout life. Previous studies have demonstrated that adult mouse olfactory epithelium (OE) injured by exposure to ZnSO4 through nasal irrigation can stimulate stem cell mitotic activity in situ, which continues when placed in culture. We report on an improved ZnSO4 delivery method, mist inhalation, which produces more consistent and greater yields of OE cells. Cultures established following this method contained bipolar, nest, fusiform, and giant cells. The bipolar cells usually underwent asymmetric process development. Some bipolar cells reacted positively to neuron-specific antibodies and were immunonegative for keratin and glia-specific proteins, suggesting that they were ORNs. Those that were negative for the neuron-specific proteins may represent either neuron progenitors or olfactory ensheathing cells. The fusiform cells were relatively small and undifferentiated, exposure to brain-derived neurotrophic factor resulted in their decrease and an increase in bipolar cells. Therefore, they might be the stem cells. The nest cells had morphological characteristics of epithelia and bound keratin antibodies. The giant cells had the morphology of epithelial cells but were negative for keratin; they may represent a unique cell population induced by the ZnSO4. These results indicate that the major cell types of intact OE are present in our cultures, and each retains characteristics found in situ. The mist inhalation method provides an in vitro population of adult mitotically active neurons for study.


Forkhead box F1 induces columnar phenotype and epithelial-to-mesenchymal transition in esophageal squamous cells to initiate Barrett's like metaplasia.
Multiple genome-wide association studies (GWAS) have linked Forkhead Box F1 (FOXF1) to Barrett's esophagus (BE). Understanding whether FOXF1 is involved in initiation of Barrett's metaplasia could allow FOXF1 to be used for risk stratification and for therapy. Two-dimensional cell cultures and three-dimensional organoid cultures and well-annotated human biopsies were used to determine the role of FOXF1 in BE pathogenesis. Multiple established esophageal squamous and BE cell lines were tested in gain- and loss-of-function studies. Initiation of a BE-like metaplastic change was evaluated by measuring characteristic cytokeratins and global gene expression profiling and by culturing organoids. Epithelial-mesenchymal transition (EMT) was evaluated by immunostaining for E-cadherin, vimentin and Snail, and by cell motility assay. Columnar esophageal epithelium of BE patients exhibited higher expression of FOXF1 compared to normal squamous esophageal epithelium of GERD patients (P < 0.001). Acidic bile salts induced nuclear FOXF1 in esophageal squamous cells. FOXF1 overexpression in normal esophageal squamous cells: (a) increased columnar cytokeratins and decreased squamous cytokeratins, (b) converted squamous organoids to glandular organoids, and (c) switched global gene profiles to resemble that of human BE epithelium (P = 2.1685e - 06 for upregulated genes and P = 8.3378e - 09 for downregulated genes). FOXF1 inhibition in BE cell lines led to loss of BE differentiation markers, CK7, and mucin 2. Also, FOXF1 induced EMT and promoted cell motility in normal esophageal squamous epithelial cells. FOXF1-induced genes mapped to pathways such as Cancer, Cellular Assembly and Organization, DNA Replication, Recombination, and Repair. In conclusion, FOXF1 promotes a BE-like columnar phenotype and cell motility in esophageal squamous epithelial cells, which may have a critical role in BE development. FOXF1 should be studied further as a biomarker for BE and as a target for BE treatment.


Inhibition of nitric oxide synthase attenuates NNMU-induced alveolar injury in vivo.
The purpose of this study was to determine if the acute alveolar injury induced by subcutaneous injections of N-nitroso-N-methylurethane (NNMU) in rats is mediated by nitric oxide (NO.). We show that intraperitoneal injections of the NO. synthase (NOS) inhibitor N omega-nitro-L-arginine methyl ester (L-NAME) or aminoguanidine significantly attenuate the NNMU-induced alveolar injury as assessed by 1) normalization of the alveolar-arterial O2 difference, 2) attenuation of the lowered phospholipid-to-protein ratio in the crude surfactant pellet (CSP), 3) attenuation of the elevated minimal surface tension of the CSP, and 4) attenuation of polymorphonuclear neutrophilic infiltration into the alveolar space. Injections of N omega-nitro-D-arginine methyl ester, the inactive stereoisoform of L-NAME, did not affect the acute lung injury. Western blot analysis of whole lung homogenates demonstrate an elevated expression of transcriptionally inducible, Ca(2+)-independent NOS (iNOS) in NNMU-injected rats compared with control saline-injected rats. NOS inhibitors did not affect NNMU-induced iNOS expression. These investigations demonstrate that the inhibition of NOS attenuates NNMU-induced acute lung injury, suggesting a role for NO. in the progression of acute respiratory distress syndrome.


Fidelity of frequency and phase entrainment of circuit-level spike activity during DBS.
High-frequency stimulation is known to entrain spike activity downstream and upstream of several clinical deep brain stimulation (DBS) targets, including the cerebellar-receiving area of thalamus (VPLo), subthalamic nucleus (STN), and globus pallidus (GP). Less understood are the fidelity of entrainment to each stimulus pulse, whether entrainment patterns are stationary over time, and how responses differ among DBS targets. In this study, three rhesus macaques were implanted with a single DBS lead in VPLo, STN, or GP. Single-unit spike activity was recorded in the resting state in motor cortex during VPLo DBS, in GP during STN DBS, and in STN and pallidal-receiving area of motor thalamus (VLo) during GP DBS. VPLo DBS induced time-locked spike activity in 25% (n = 15/61) of motor cortex cells, with entrained cells following 7.5 ± 7.4% of delivered pulses. STN DBS entrained spike activity in 26% (n = 8/27) of GP cells, which yielded time-locked spike activity for 8.7 ± 8.4% of stimulus pulses. GP DBS entrained 67% (n = 14/21) of STN cells and 32% (n = 19/59) of VLo cells, which showed a higher fraction of pulses effectively inhibiting spike activity (82.0 ± 9.6% and 86.1 ± 16.6%, respectively). Latency of phase-locked spike activity increased over time in motor cortex (58%, VPLo DBS) and to a lesser extent in GP (25%, STN DBS). In contrast, the initial inhibitory phase observed in VLo and STN during GP DBS remained stable following stimulation onset. Together, these data suggest that circuit-level entrainment is low-pass filtered during high-frequency stimulation, most notably for glutamatergic pathways. Moreover, phase entrainment is not stationary or consistent at the circuit level for all DBS targets.


[Intracranial meningiomas in the elderly].
Since the introduction of CT and then MRI in the clinical practice, the number of elderly patients with intracranial meningiomas observed in neurosurgical departments has increased to three/four times. Considering that some patients die because of their meningioma and others die with their meningioma, the neurosurgeon should not only be able to eradicate surgically the lesion, but also to decide if that benign tumor should be treated or not. Forty-three patients with intracranial meningioma, aged over 65 were operated upon between 1989 and 1995. They constitute about one fourth of all the meningioma-patients operated upon in that period at our Department. Main symptoms were focal deficits, psychorganic syndrome, and seizures. Mean duration of symptoms was 15 months, with 30% of the histories lasting less than 3 months. Associate pathologies were those typical of the elderly population with hypertension and senile cardiopathy at the first places. Two patients had insulin-dependent diabetes, 2 prostatic carcinoma, and 8 were obese. MRI was useful either in depicting the location of the tumor or in helping the surgeon to give a correct operative indication and risk evaluation. MRI criteria for risk evaluation were severe edema and venous vascular infiltration. Forty-five operations were performed. Their duration was not a risk factor by itself, it only reflected the difficulty of the removal, due to the location of the tumor. Dissection of tumor remnants from main arteries or veins was never attempted in this group of patients; total removal was achieved in 75.5% of the cases. Immediate postoperative results were good in 71% of cases. Four patients died (9%). Long term results were evaluated in 29 patients (mean follow-up period: 35 months). Twenty-two patients (76%) had excellent or good results, 5 (17%) fair, and 2 (7%) had severe neurological dysfunctions and needed continuous assistance. Surgical risk factors typical of this age group were identified: cranio-spinal location, damage to the cortical or deep venous system, severe peritumoral edema, and poor neurological preoperative conditions. Systemic risk factors were insulin-dependent diabetes and obesity.


Healthy volunteers data bank: where and how?
In order to improve safety for healthy volunteers to be included in phase I clinical trials, a national data bank will be set up and controlled by the French Ministry of Health. Each investigator responsible for an agreed phase I center will have a personal access code to the central computer in order to consult the national registry before including any healthy volunteer in a study. If the volunteer has already participated in a recent study, the investigator will find 2 important pieces of information in the data bank: the duration of the safety period following the preceding trial and during which the subject is not allowed to participate in another study; the total amount of money received by the subject during the last 12 months in grants for phase I studies; this amount will be limited by the Ministry of Health. This healthy volunteer data bank will decrease the risk of drug interaction and will improve safety in phase I trials. However, many practical and ethical problems remain to be solved.


Decreased Bilateral FDG-PET Uptake and Inter-Hemispheric Connectivity in Multi-Domain Amnestic Mild Cognitive Impairment Patients: A Preliminary Study.
Background: Amnestic mild cognitive impairment (aMCI) is a heterogeneous condition. Based on clinical symptoms, aMCI could be categorized into single-domain aMCI (SD-aMCI, only memory deficit) and multi-domain aMCI (MD-aMCI, one or more cognitive domain deficit). As core intrinsic functional architecture, inter-hemispheric connectivity maintains many cognitive abilities. However, few studies investigated whether SD-aMCI and MD-aMCI have different inter-hemispheric connectivity pattern. Methods: We evaluated inter-hemispheric connection pattern using fluorine-18 positron emission tomography - fluorodeoxyglucose (18F PET-FDG), resting-state functional MRI and structural T1 in 49 controls, 32 SD-aMCI, and 32 MD-aMCI patients. Specifically, we analyzed the 18F PET-FDG (intensity normalized by cerebellar vermis) in a voxel-wise manner. Then, we estimated inter-hemispheric functional and structural connectivity by calculating the voxel-mirrored homotopic connectivity (VMHC) and corpus callosum (CC) subregions volume. Further, we correlated inter-hemispheric indices with the behavioral score and pathological biomarkers. Results: We found that MD-aMCI exhibited more several inter-hemispheric connectivity damages than SD-aMCI. Specifically, MD-aMCI displayed hypometabolism in the bilateral middle temporal gyrus (MTG), inferior parietal lobe, and left precuneus (PCu) (p < 0.001, corrected). Correspondingly, MD-aMCI showed decreased VMHC in MTG, PCu, calcarine gyrus, and postcentral gyrus, as well as smaller mid-posterior CC than the SD-aMCI and controls (p < 0.05, corrected). Contrary to MD-aMCI, there were no neuroimaging indices with significant differences between SD-aMCI and controls, except reduced hypometabolism in bilateral MTG. Within aMCI patients, hypometabolism and reduced inter-hemispheric connectivity correlated with worse executive ability. Moreover, hypometabolism indices correlated to increased amyloid deposition. Conclusion: In conclusion, patients with MD-aMCI exhibited the more severe deficit in inter-hemispheric communication than SD-aMCI. This long-range connectivity deficit may contribute to cognitive profiles and potentially serve as a biomarker to estimate disease progression of aMCI patients.


Water-mediated hydrogen-bonded network on the cytoplasmic side of the Schiff base of the L photointermediate of bacteriorhodopsin.
The L intermediate in the proton-motive photocycle of bacteriorhodopsin is the starting state for the first proton transfer, from the Schiff base to Asp85, in the formation of the M intermediate. Previous FTIR studies of L have identified unique vibration bands caused by the perturbation of several polar amino acid side chains and several internal water molecules located on the cytoplasmic side of the retinylidene chromophore. In the present FTIR study we describe spectral features of the L intermediate in D(2)O in the frequency region which includes the N-D stretching vibrations of the backbone amides. We show that a broad band in the 2220-2080 cm(-1) region appears in L. By use of appropriate (15)N labeling and mutants, the lower frequency side of this band in L is assigned to the amides of Lys216 and Gly220. These amides are coupled to each other, and interact with Thr46 and Val49 in helix B and Asp96 in helix C via weakly H-bonding water molecules that exhibit O-D stretching vibrations at 2621 and 2605 cm(-1). These water molecules are part of a hydrogen-bonded network characteristic of L which includes other water molecules located closer to the chromophore that exhibit an O-D stretching vibration at 2589 cm(-1). This structure, extending from the Schiff base to the internal proton donor Asp96, stabilizes L and affects the L-to-M transition.


How do SET-domain protein lysine methyltransferases achieve the methylation state specificity? Revisited by Ab initio QM/MM molecular dynamics simulations.
A distinct protein lysine methyltransferase (PKMT) only transfers a certain number of methyl group(s) to its target lysine residue in spite of the fact that a lysine residue can be either mono-, di-, or tri-methylated. In order to elucidate how such a remarkable product specificity is achieved, we have carried out ab initio quantum mechanical/molecular mechanical (QM/MM) molecular dynamics simulations on two SET-domain PKMTs: SET7/9 and Rubisco large subunit methyltransferase (LSMT). The results indicate that the methylation state specificity is mainly controlled by the methyl-transfer reaction step, and confirm that SET7/9 is a mono-methyltransferase while LSMT has both mono-and di-methylation activities. It is found that the binding of the methylated lysine substrate in the active site of SET7/ 9 opens up the cofactor AdoMet binding channel so that solvent water molecules get access to the active site. This disrupts the catalytic machinery of SET7/9 for the di-methylation reaction, which leads to a higher activation barrier, whereas for the LSMT, its active site is more spacious than that of SET7/9, so that the methylated lysine substrate can be accommodated without interfering with its catalytic power. These detailed insights take account of protein dynamics and are consistent with available experimental results as well as recent theoretical findings regarding the catalytic power of SET7/9.


The accuracy of colposcopic biopsy: analyses from the placebo arm of the Gardasil clinical trials.
We evaluated the overall agreement between colposcopically directed biopsies and the definitive excisional specimens within the context of three clinical trials. A total of 737 women aged 16-45 who had a cervical biopsy taken within 6 months before their definitive therapy were included. Per-protocol, colposcopists were to also obtain a representative cervical biopsy immediately before definitive therapy. Using adjudicated histological diagnoses, the initial biopsies and the same day biopsies were correlated with the surgically excised specimens. The overall agreement between the biopsies taken within 6 months of definitive therapy, and the definitive therapy diagnoses was 42% (weighted kappa = 0.34) (95% CI: 0.29-0.39). The overall underestimation of cervical intraepithelial neoplasia grade 2/3 or adenocarcinoma in situ (CIN2-3/AIS) and CIN3/AIS was 26 and 42%, respectively. When allowing for one degree of variance in the correlation, the overall agreement was 92% for CIN2-3/AIS. The overall agreement between the same day biopsy and definitive therapy specimen was 56% (weighted kappa = 0.41) (95% CI: 0.36-0.47), and the underestimation of CIN2-3/AIS was 57%. There were significant associations in the agreement between biopsies and excisional specimen diagnoses when patients were stratified by age, number of biopsies, lesion size, presence of human papillomavirus (HPV)16/18 and region. Of 178 diagnostic endocervical curettages performed, 14 (7.9%) found any HPV disease. Colposcopic accuracy improved when CIN2 and CIN3/AIS were grouped as a single predictive measure of high-grade disease. Colposcopy functioned well when allowed a one-degree difference between the biopsy and the surgical histologic interpretations, as done in clinical practice. Taking more than one biopsy improved colposcopic accuracy and could improve patient management.


Effect of isometric handgrip exercise on the size of cephalic veins in patients with stage 3 and 4 chronic kidney disease: A randomized controlled trial.
Proper vascular access is essential for the long-term survival of chronic haemodialysis patients. The preferred vascular access in terms of long-term function is the native arteriovenous fistula. The success of native arteriovenous fistula depends mainly on a sufficient vein diameter. Thus, any intervention that could increase vein diameter before arteriovenous fistula creation could improve its patency. We conduct a study to investigate the effect of local physical training, namely handgrip exercise, on the distal forearm cephalic vein diameter in patients with chronic renal disease. A total of 34 chronic renal disease patients (stage 3 and 4) were recruited in a randomized controlled trial. Handgrip exercise was performed for 8 weeks in the intervention group. Handgrip-strength measurement and distal forearm cephalic vein diameter of a non-dominant hand with and without tourniquet was recorded (measurement is taken 1 cm proximal to the radial styloid). After 8 weeks, the mean cephalic vein diameter in the intervention group increased from 1.77 and 1.97 mm to 2.15 and 2.43 mm, without and with a tourniquet, respectively (p < 0.05). There is also a significant change in the mean diameter of distal forearm cephalic vein (p < 0.05) in the intervention group when measured in both the absence (mean change 0.39 ± 0.06 mm vs 0.01 ± 0.02 mm) and the presence of tourniquet (mean change 0.47 ± 0.07 mm vs 0.01 ± 0.01 mm). These findings suggest that non-invasive handgrip exercise can increase in the diameter of the distal forearm cephalic vein, thereby increasing the rate of successful arteriovenous fistula creation.


Clinical Cirrhosis Dilemmas: Survey of Practice from the 7th International Coagulation in Liver Disease Conference.
Hemostatic disorders in chronic liver disease and cirrhosis show continued expansion of research efforts. However, clinical decision making is often practiced on an individual patient level as consensus guidelines are lacking. We aimed to better assess individual day-to-day clinical practice through gauging clinicians' responses to common clinical scenarios. A series of ten clinical scenarios (seven procedural coagulation and three thrombosis management) were posed to conference attendees utilizing real-time polling software (Poll Everywhere). Responses were binomial and were submitted as "Agree" or "Disagree." Results were displayed real time following a standardized response period and an open-forum discussion ensued between conference faculty and attendees following response submission. Twenty conference attendees participated in the clinical scenario plenary session. In general, agreement rates were high. All but one of the ten clinical scenarios had ≥ 70% agreement. Agreement was based both on procedural risk, with greatest agreement seen for low-risk procedures (80-93%), and on peri-procedural coagulation parameters of platelet count and fibrinogen level where > 50,000μ/L and 120 mg/dL were the most agreed upon thresholds, respectively. 75-95% agreement was reached when surveying the need for anticoagulation for mesenteric vein thrombosis in liver transplant candidates; slightly less (71%) agreement was found when deciding to proceed with anticoagulation in non-liver transplant candidates with mesenteric vein thrombosis. While large-scale, methodologically rigorous randomized controlled trials are lacking to guide clinical decision making in patients with coagulation disorders and chronic liver disease, consensus expert opinion regarding mitigating peri-procedural bleeding risk and treatment of thrombosis appears consistent and strong.


Rotational diffusion of human lipoproteins and their receptors as determined by time-resolved phosphorescence anisotropy.
Time-resolved phosphorescence anisotropy has been used to assess the rotational dynamics of human serum lipoproteins labeled with phosphorescent probes of high triplet yield. Labeling the lipid phase of low density, very low density, and high density lipoproteins with an eosinyl fatty acid revealed the existence of two motions. The shorter time constant was attributed to motion of the chromophore within the lipoprotein particle, while the longer time constant represented the global tumbling of the particles in solution. The measured correlation times for this global motion were about twice those predicted from the Stokes-Einstein relationship. Covalent labeling of the apolipoproteins of the low and high density lipoproteins with erythrosin revealed the existence of segmental motion of labeled domains of the apolipoprotein within their respective particles. The correlation times for this motion were within the range 10-50 microseconds. The binding of low density lipoproteins to receptors on membranes isolated from the adrenal cortex resulted in a freezing of the global motion, but maintenance of the faster segmental motion of the labeled domains of the apolipoprotein. The experiments imply that in these membranes there is no global motion of the low density lipoprotein-receptor complex on the phosphorescence time scale. Similar results were found for the binding of high density lipoproteins to liver plasma membranes. The contributions of nonspecific binding of the labeled lipoproteins to the measured phosphorescence anisotropy were carefully assessed.


Tearable and Fillable Composite Sponges Capable of Heat Generation and Drug Release in Response to Alternating Magnetic Field.
Tearable and fillable implants are used to facilitate surgery. The use of implants that can generate heat and release a drug in response to an exogenous trigger, such as an alternating magnetic field (AMF), can facilitate on-demand combined thermal treatment and chemotherapy via remote operation. In this study, we fabricated tearable sponges composed of collagen, magnetite nanoparticles, and anticancer drugs. Crosslinking of the sponges by heating for 6 h completely suppressed undesirable drug release in saline at 37 °C but allowed drug release at 45 °C. The sponges generated heat immediately after AMF application and raised the cell culture medium temperature from 37 to 45 °C within 15 min. Heat generation was controlled by switching the AMF on and off. Furthermore, in response to heat generation, drug release from the sponges could be induced and moderated. Thus, remote-controlled heat generation and drug release were achieved by switching the AMF on and off. The sponges destroyed tumor cells when AMF was applied for 15 min but not when AMF was absent. The tearing and filling properties of the sponges may be useful for the surgical repair of bone and tissue defects. Moreover, these sponges, along with AMF application, can facilitate combined thermal therapy and chemotherapy.


Correlates of mercury in female river otters (Lontra canadensis) from Nova Scotia, Canada.
Mercury (Hg) can reach toxic concentrations in aquatic habitats, sometimes as a consequence of human activity. Mercury can have deleterious effects, particularly in piscivorous mammals in which it bioaccumulates. Furs from trapper-provided female otter (Lontra canadensis) carcasses in Nova Scotia were analyzed for total Hg. Concentrations of total Hg in fur samples averaged 25 µg/g dry weight, ranging from 1.4 to 137 µg/g; 20 µg/g is the fur concentration at which toxic effects are expected. Mercury concentrations were greater in otters from watersheds with bedrock substrates known to contain more available Hg, from otters trapped farther inland, and from otters trapped on watersheds with hydroelectric dams. Otter reproductive potential was measured by counting the number of blastocysts in reproductive tracts. Tooth annuli were used to age otters. Reproductive potential was not related to Hg concentration, nor was Hg concentration related to age. In a general linear model, 53% of variation in fur Hg was explained by underlying bedrock, distance from the coast at which otters were trapped, and presence/absence of a hydroelectric dam. The proportion of juveniles in a population did not differ relative to bedrock Hg concentration, but was lower on watersheds with hydroelectric dams. Because we found no evidence of reduced reproductive potential from greater Hg concentrations, the low proportion of juveniles suggests that Hg reduced juvenile survival, although our evidence is circumstantial.


Rituximab therapy for follicular lymphoma: a comprehensive review of its efficacy as primary treatment, treatment for relapsed disease, re-treatment and maintenance.
Advances in the treatment of follicular lymphoma (FL) have been achieved through the development of newer combination regimens, mostly based upon purine analogs and/ or monoclonal antibodies, as well as recent progress in the area of stem cell transplantation. These advances have increased the need to obtain consensus regarding treatment priorities from the various options available, ranging from the traditional palliative approach through to the novel regimens available including stem cell transplantation. The apparent synergism between chemotherapy and rituximab, which facilitates the achievement of complete clinical (CR) and molecular remission (MR), together with the possible feasibility of maintenance and re-treatment with rituximab have increased the interest in the use of this drug as primary treatment for FL. This review summarizes the available literature and deals with the role of rituximab in refractory/relapsed FL as well as in previously untreated FL patients. The improvement in remission rates and response duration associated with immunotherapy are contrasted with the potential risks, such as the development of rituximab resistance, as well as other less recognized complications related to altered humoral immunity. Despite all the advances reported, treatment might still have to be individualized for patients with FL, until evidence of an important survival advantage for rituximab over chemotherapy is established. The possible role of autotransplantation for FL and the use of rituximab is also reviewed.


Ribosomal DNA sequence divergence and group I introns within the Leucostoma species L. cinctum, L. persoonii, and L. parapersoonii sp. nov., ascomycetes that cause Cytospora canker of fruit trees.
Leucostoma species that are the causal agents of Cytospora canker of stone and pome fruit trees were studied in detail. DNA sequence of the internal transcribed spacer regions and the 5.8S of the nuclear ribosomal DNA operon (ITS rDNA) supplied sufficient characters to assess the phylogenetic relationships among species of Leucostoma, Valsa, Valsella, and related anamorphs in Cytospora. Parsimony analysis of the aligned sequence divided Cytospora isolates from fruit trees into clades that generally agreed with the morphological species concepts, and with some of the phenetic groupings (PG 1-6) identified previously by isozyme analysis and cultural characteristics. Phylogenetic analysis inferred that isolates of L. persoonii formed two well-resolved clades distinct from isolates of L. cinctum. Phylogenetic analysis of the ITS rDNA, isozyme analysis, and cultural characteristics supported the inference that L. persoonii groups PG 2 and PG 3 were populations of a new species apparently more genetically different from L. persoonii PG 1 than from isolates representative of L. massariana, L. niveum, L. translucens, and Valsella melastoma. The new species, L. parapersoonii, was described. A diverse collection of isolates of L. cinctum, L. persoonii, and L. parapersoonii were examined for genetic variation using restriction fragment length polymorphism (RFLP) analysis of the ITS rDNA and the five prime end of the large subunit of the rDNA (LSU rDNA). HinfI and HpaII endonucleases were each useful in dividing the Leucostoma isolates into RFLP profiles corresponding to the isozyme phenetic groups, PG 1-6. RFLP analysis was more effective than isozyme analysis in uncovering variation among isolates of L. persoonii PG 1, but less effective within L. cinctum populations. Isolates representative of seven of the L. persoonii formae speciales proposed by G. Défago in 1935 were found to be genetically diverse isolates of PG 1. Two large insertions, 415 and 309 nucleotides long, in the small subunit (SSU) of the nuclear rDNA of L. cinctum were identified as Group 1 introns; intron 1 at position 943 and intron 2 at position 1199. The two introns were found to be consistently present in isolates of L. cinctum PG 4 and PG 5 and absent from L. cinctum PG 6 isolates, despite the similarity of the ITS sequence and teleomorph morphology. Intron 1 was of subgroup 1C1 whereas intron 2 was of an unknown subgroup. RFLP patterns and presence/absence of introns were useful characters for expediting the identification of cultures of Leucostoma isolated from stone and pome fruit cankers. RFLP patterns from 13 endonucleases provided an effective method for selecting an array of diverse PG 1 isolates useful in screening plant germplasm for disease-resistance.


Upstream sequences required for efficient expression of a soybean heat shock gene.
A soybean gene (Gmhsp17.5-E) encoding a small heat shock protein was introduced into primary sunflower tumors via T-DNA-mediated transformation. RNA blot hybridizations and S1-nuclease hybrid protection studies indicated that the heat shock gene containing 3.25 kilobases of 5'-flanking sequences was strongly transcribed in a thermoinducible (40 degrees C) manner. Transcriptional induction also occurred to a lesser extent upon treatment of whole tumors with sodium arsenite and CdCl2. Basal (26 degrees C) transcription was not detected in soybean seedlings, but it was quite evident in transformed tumor tissue. A 5' deletion to -1,175 base pairs with respect to the CAP site had no effect on the levels of thermoinducible transcription, but it resulted in a large increase in basal transcription. Further removal of DNA sequences (including the TATA-distal heat shock consensus element) to -95 base pairs reduced thermoinducible transcription by 95% and also greatly decreased basal transcription. The termini of the Gmhsp17.5-E RNA in the tumor were generally the same as those present in soybean RNA, with the exception of several additional 3' termini.


trans-hydroalumination/alkylation: one-pot synthesis of trisubstituted allylic alcohols.
[reaction: see text] Described herein is a method of stereoselective synthesis of trisubstituted allylic alcohols by alkylation of alkenyl alanates, formed in situ through treatment of propargyl alcohols with Vitride (Red-Al). This technique represents the first of its kind to feature a trans-hydrometalation, and is particularly effective for the formation of 1,4-dienes. Applications involving primary, secondary, and tertiary alcohols are discussed, as well as limitations regarding both alkyne and electrophile components.


Thermal inactivation of Newcastle disease virus.
The rate of destruction of hemagglutinins and infectivity of Newcastle disease virus was determined over a temperature range of 37.8 to 60 C. From the calculated values of deltaH and deltaS, it was concluded that inactivation of the hemagglutinating activity and viral infectivity was due to protein denaturation.


A critical role for human caspase-4 in endotoxin sensitivity.
Response to endotoxins is an important part of the organismal reaction to Gram-negative bacteria and plays a critical role in sepsis and septic shock, as well as other conditions such as metabolic endotoxemia. Humans are generally more sensitive to endotoxins when compared with experimental animals such as mice. Inflammatory caspases mediate endotoxin-induced IL-1β secretion and lethality in mice, and caspase-4 is an inflammatory caspase that is found in the human, and not mouse, genome. To test whether caspase-4 is involved in endotoxin sensitivity, we developed a transgenic mouse expressing human caspase-4 in its genomic context. Caspase-4 transgenic mice exhibited significantly higher endotoxin sensitivity, as measured by enhanced cytokine secretion and lethality following LPS challenge. Using bone marrow-derived macrophages, we then observed that caspase-4 can support activation of caspase-1 and secretion of IL-1β and IL-18 in response to priming signals (LPS or Pam3CSK4) alone, without the need for second signals to stimulate the assembly of the inflammasome. These findings indicate that the regulation of caspase-1 activity by human caspase-4 could represent a unique mechanism in humans, as compared with laboratory rodents, and may partially explain the higher sensitivity to endotoxins observed in humans. Regulation of the expression, activation, or activity of caspase-4 therefore represents targets for systemic inflammatory response syndrome, sepsis, septic shock, and related disorders.


Induction of apoptosis by [8]-shogaol via reactive oxygen species generation, glutathione depletion, and caspase activation in human leukemia cells.
Ginger, the rhizome of Zingiber officinale , is a traditional medicine with a carminative effect and antinausea, anti-inflammatory, and anticarcinogenic properties. This study examined the growth inhibitory effects of [8]-shogaol, one of the pungent phenolic compounds in ginger, on human leukemia HL-60 cells. It demonstrated that [8]-shogaol was able to induce apoptosis in a time- and concentration-dependent manner. Treatment with [8]-shogaol caused a rapid loss of mitochondrial transmembrane potential, stimulation of reactive oxygen species (ROS) production, release of mitochondrial cytochrome c into cytosol, and subsequent induction of procaspase-9 and procaspase-3 processing. Taken together, these results suggest for the first time that ROS production and depletion of glutathione that contributed to [8]-shogaol-induced apoptosis in HL-60 cells.


An evaluation of the 10-item vision core measure 1 (VCM1) scale (the Core Module of the Vision-Related Quality of Life scale) using Rasch analysis.
To assess and re-engineer the Vision Core Measure 1 (VCM1) questionnaire in low vision (LV) and cataract participants using Rasch analysis. 295 participants drawn from a low vision clinic and 181 from a cataract surgery waiting list completed the 10-item VCM1. Unidimensionality, item fit to the model, response category performance, differential item functioning (DIF) and targeting of items to patients were assessed. Category collapsing and item removal were considered to improve the questionnaire. The initial fit of the VCM1 (combined populations) to the Rasch model showed lack of fit (chi2 = 83.3, df = 50, p = 0.002). There was evidence of DIF between the two populations which could not be resolved. Consequently, each population was assessed separately. Irrespective of the population, disordering of response category thresholds was evident. However, collapsing categories produced ordered thresholds and resulted in fit to the Rasch model for the LV (Total chi2 = 41.6, df = 30; p = 0.08) and cataract population (Total chi2 = 17.9, df = 20, p = 0.59). Overall, the VCM1 behaved as a unidimensional scale for each population and no item showed evidence of DIF. Item targeting to patients was however sub-optimal particularly for the cataract population. The VCM1 questionnaire could be improved by shortening the response scale, although different response categories are required for cataract and LV populations. Calibration of items also differed across populations. While the VCM1 performs well within the Rasch model, in line with its initial purpose, it requires the addition of items to satisfactorily target low vision and cataract populations.


Mechanisms of neurodegeneration shared between multiple sclerosis and Alzheimer's disease.
Multiple sclerosis and Alzheimer's disease are fundamentally different diseases. However, recent data suggest that certain mechanisms of neurodegeneration may be shared between the two diseases. Inflammation drives the disease in multiple sclerosis. It is also present in Alzheimer's disease lesions, where it may have dual functions in amyloid clearance as well as in the propagation of neurodegeneration. In both diseases, degeneration of neurons, axons, and synapses occur on the background of profound mitochondrial injury. Reactive oxygen and nitric oxide intermediates are major candidates for the induction of mitochondrial injury. Radicals are produced through the induction of the respiratory burst in activated microglia, which are present in the lesions of both diseases. In addition, liberation of toxic iron from intracellular stores may augment radical formation. Finally reactive oxygen species are also produced in the course of mitochondrial injury itself. Anti-oxidant and mitochondria protective therapeutic strategies may be beneficial both in multiple sclerosis and Alzheimer's disease in particular in early stages of the disease.


Linker Flexibility Facilitates Module Exchange in Fungal Hybrid PKS-NRPS Engineering.
Polyketide synthases (PKSs) and nonribosomal peptide synthetases (NRPSs) each give rise to a vast array of complex bioactive molecules with further complexity added by the existence of natural PKS-NRPS fusions. Rational genetic engineering for the production of natural product derivatives is desirable for the purpose of incorporating new functionalities into pre-existing molecules, or for optimization of known bioactivities. We sought to expand the range of natural product diversity by combining modules of PKS-NRPS hybrids from different hosts, hereby producing novel synthetic natural products. We succeeded in the construction of a functional cross-species chimeric PKS-NRPS expressed in Aspergillus nidulans. Module swapping of the two PKS-NRPS natural hybrids CcsA from Aspergillus clavatus involved in the biosynthesis of cytochalasin E and related Syn2 from rice plant pathogen Magnaporthe oryzae lead to production of novel hybrid products, demonstrating that the rational re-design of these fungal natural product enzymes is feasible. We also report the structure of four novel pseudo pre-cytochalasin intermediates, niduclavin and niduporthin along with the chimeric compounds niduchimaeralin A and B, all indicating that PKS-NRPS activity alone is insufficient for proper assembly of the cytochalasin core structure. Future success in the field of biocombinatorial synthesis of hybrid polyketide-nonribosomal peptides relies on the understanding of the fundamental mechanisms of inter-modular polyketide chain transfer. Therefore, we expressed several PKS-NRPS linker-modified variants. Intriguingly, the linker anatomy is less complex than expected, as these variants displayed great tolerance with regards to content and length, showing a hitherto unreported flexibility in PKS-NRPS hybrids, with great potential for synthetic biology-driven biocombinatorial chemistry.


Computed tomography and blunt abdominal injury: patient selection based on examination, haematocrit and haematuria.
The criteria for ordering abdominal CT scans in the secondary survey of stable bluntly injured patients was examined. A patient population at high risk for having intra-abdominal injury (IAI) was identified by physical examination, a fall in haematocrit, and haematuria. A total of 444 patients receiving abdominal CT scans at a large urban trauma centre were reviewed. IAI was diagnosed in 49 (11 per cent), by radiographic and/or intra-operative findings. Abdominal tenderness was present in all 17 patients who underwent surgery. The sensitivity and specificity of abdominal CT scanning was 90 per cent and 99 per cent, respectively. The abdominal exam had a sensitivity of 63 per cent and a specificity of 65 per cent. A fall in haematocrit > or = 5 was not statistically significant. The combined abdominal exam and haematuria yielded a specificity of 93 per cent with a negative predictive value (NPV) of 93 per cent. Early CT scanning of stable patients who had sustained blunt injuries is an effective screen for IAI. The benefit of a CT scan for patients without abdominal tenderness or with an isolated fall in haematocrit is questionable. Serial abdominal examinations should remain the most timely and cost-effective method for identifying IAI in stable patients. The specificity and NPV of abdominal tenderness combined with haematuria approaches that of CT.


Developmental regulation of erythropoietin and erythropoiesis.
It is well established that erythropoiesis occurs first in the yolk sac, then in the liver, subsequently moving to the bone marrow and, in rodents, the spleen during development. The origin of the erythropoietic precursors and some factors suggested to be important for the changing location of erythropoiesis are discussed in this review. Until recently, the major site of erythropoietin (Epo) production in the fetus was thought to be the liver, but studies have shown now that the Epo gene is expressed strongly in the fetal kidney, even in the temporary mesonephros. The metanephric Epo mRNA is upregulated by anemia, downregulated by glucocorticoids, and contributes substantially to circulating hormone levels in hemorrhaged ovine fetuses. Other sites of Epo and Epo receptor production, likely to have important actions during development, are the placenta and the brain.


Distortion Product Otoacoustic Emissions in acute acoustic trauma.
Acute acoustic traumas are caused by exposure to extremely high noise levels ranging from milliseconds to several hours' duration. In pure tone audiometry they range from the C5 dip to basomediocochlear sensorineural hearing loss. Their pathogenesis is assumed to consist of micromechanical-traumatic and biochemical-metabolic damage to the outer hair cells. In order to establish the changes to the DPOAE (distortion products of otoacoustic emissions), 17 patients were examined after sustaining acute acoustic trauma. The causes included firework explosions, anti-tank rocket launchers, vehicle tyre bursting, rock concerts, hand-gun shots, sub-machine gun fire, hand grenade explosion, exploding car battery. The pure tone audiogram, tympanogram, tinnitus maskability and DPOAE (both DP-gram and growth rate in various frequencies) were determined in all patients. If the event had occurred some time ago, measurements were taken only once; in acute cases measurements were repeated at different times. In nine patients with persistent hearing impairment, clear DPs were found in the unaffected frequencies but were completely absent in the affected frequency range. Four of these patients were unilaterally and two patients were bilaterally affected; three patients had a different (not noise-induced) hearing loss on the opposite side. In eight patients with regressive hearing loss, DPs were by contrast detectable throughout the entire frequency range, their amplitudes only rising slightly as hearing recovered. Of these eight patients, three were unilaterally and five bilaterally affected. DPOAE seem to indicate the likelihood of recovery of hearing threshold after an acute acoustic trauma. In cases with DPs completely absent in the affected frequency range, the prognosis seems to be much worse than in cases with present DPs in the frequency range of hearing.


Zebrafish ambra1a and ambra1b knockdown impairs skeletal muscle development.
The essential role of autophagy in muscle homeostasis has been clearly demonstrated by phenotype analysis of mice with muscle-specific inactivation of genes encoding autophagy-related proteins. Ambra1 is a key component of the Beclin 1 complex and, in zebrafish, it is encoded by two paralogous genes, ambra1a and ambra1b, both required for normal embryogenesis and larval development. In this study we focused on the function of Ambra1, a positive regulator of the autophagic process, during skeletal muscle development by means of morpholino (MO)-mediated knockdown and compared the phenotype of zebrafish Ambra1-depleted embryos with that of Ambra1gt/gt mouse embryos. Morphological analysis of zebrafish morphant embryos revealed that silencing of ambra1 impairs locomotor activity and muscle development, as well as myoD1 expression. Skeletal muscles in ATG-morphant embryos displayed severe histopathological changes and contained only small areas of organized myofibrils that were widely dispersed throughout the cell. Double knockdown of ambra1a and ambra1b resulted in a more severe phenotype whereas defects were much less evident in splice-morphants. The morphants phenotypes were effectively rescued by co-injection with human AMBRA1 mRNA. Together, these results indicate that ambra1a and ambra1b are required for the correct development and morphogenesis of skeletal muscle.


Within-family plasticity of nervous system architecture in Syllidae (Annelida, Errantia).
The ground pattern underlying the nervous system of the last common ancestor in annelids was long thought to be settled, consisting of a dorsal brain, circumoesophageal connectives and a subepithelial, ladder-like ventral nerve cord with segmental ganglia connected by paired connectives. With the advent of immunocytochemical stainings and confocal laser scanning microscopy, it becomes evident that its architecture is extremely diverse, which makes the reconstruction of a ground pattern in annelida challenging. Whereas the nervous systems of many different families has already been described, only very few studies looked at the diversity of nervous systems within such clades to give a closer estimate on how plastic the annelid nervous system really is. So far, little is known on syllid nervous system architecture, one of the largest and most diverse groups of marine annelids. The position of the brain, the circumoesophageal connectives, the stomatogastric nervous system, the longitudinal nerves that traverse each segment and the innervation of appendages are relatively uniform within the clade. Both the number of connectives within the ventral nerve cord and the number of segmental nerves, which in earlier studies were used to infer phylogenetic relationships and to reconstruct an annelid ground pattern, are highly diverse and differ between genera or even within a given genus. Differences in the distribution of somata of the brain, the nuchal innervation and its associated cell bodies were found between Syllinae and Exogoninae and may be subfamily-specific. The nervous system morphology of syllids very likely depends on the taxon-specific ecological requirements. Thus, it is not surprising that in a clade, which occupies such diverse niches as the Annelida, we find similar patterns in phylogenetically widely separated species in similar niches and a high degree of modularity within a family. Only standardized protocols and staining methods can lead to comparable results, but so far different approaches have been taken to describe annelid nervous systems, making homologization of certain structures difficult. This study provides the first thorough description of the nervous system in the family Syllidae, allowing more detailed comparisons between annelid families in the future.


Molecular mechanisms of binding and single-strand scission of deoxyribonucleic acid by the antitumor antibiotics saframycins A and C.
The quinone antitumor antibiotics saframycins A and C bind to duplex DNA and G,C containing complementary oligodeoxyribonucleotides at pHs below 5.5 via selective protonation of the antibiotics at the N-12 positions in an equilibrium process. This binding shows minor groove specificity to T4 DNA for both antibiotics. Saframycin A exhibits an additional mode of covalent binding to DNA, which is reversible by heating, via an animal linkage with the NH2 group of guanine. Covalent binding of saframycin A is enhanced by prior reduction of the quinone moiety via a different mechanism with concomitant elimination of the nitrile group and plausibly involves an intermediate quinone methide. Both saframycin A and saframycin C when reduced in situ with NADPH or sodium borohydride cause single-strand breaks, but not double-strand breaks, in PM2 covalently closer circular DNA. This latter reaction proceeds via intermediate semiquinones, the identity and conformation of which are established by EPR. The DNA scission process which requires oxygen is mediated by the generation of O2-., H2O2 and OH., the latter of which species was identified by spin trapping. The strand scission of DNA is pH dependent and, unlike the binding of the antibiotics, is strongly inhibited by Mg2+ and Zn2+ ions.


Relationship between obesity and physical fitness and periodontitis.
Obesity and exercise are important elements associated with lifestyle-related diseases, and studies suggested that these factors may also be related to periodontitis. This study investigates the relationship between obesity and physical fitness and periodontitis. One thousand, one hundred and sixty Japanese subjects, aged 20 to 77 years, who participated in health examinations at Fukuoka Health Promotion Center were analyzed. Periodontal conditions were evaluated using the Community Periodontal Index (CPI), and subjects with > or =3 sextants of CPI code 3 or 4 were defined as having severe periodontitis. We used the body mass index (BMI) and percentage of body fat as indicators of obesity and estimated the maximal oxygen consumption (VO(2max)) during exercise as an indicator of physical fitness. We divided these variables into quintiles. We examined the single effect and interactions of the obesity index and VO(2max) on severe periodontitis. The lowest quintile in BMI and the highest quintile in VO(2max) were inversely associated with severe periodontitis, singly, in multivariate logistic regression analyses. Subjects with the combined lowest quintile in BMI and the highest quintile in VO(2max) had a significantly lower risk of severe periodontitis compared to subjects with other combined quintiles in BMI and in VO(2max) (odds ratio: 0.17; 95% confidence interval: 0.05 to 0.55). This study suggests that obesity and physical fitness may have some interactive effect on periodontal health status.


Induction of conduction block by Campylobacter jejuni lipopolysaccharides and focal neural insult.
A systemic exposure to gram negative LPS have caused transient conduction abnormalities in a certain strain of rats probably associated with the action of cytokines secreted by macrophages. Our previous studies demonstrated that anti-GM1 antibodies induced in rats by the cross-reactive Cj-LPS, caused no conduction abnormalities. We designed the present study to evaluate the effect of systemic exposure to Cj-LPS on nerve conduction after a focal minor neural trauma. Female Lewis rats were sensitized against KLH by repetitive subcutaneous injections. After 28 days rats were intraneurally injected with saline in the right sciatic nerve and concomitantly with intraperitoneal Cj-LPS. Sciatic nerve conduction studies were performed on days 0, 1, 2, 3, and 7 after injections. Nerve conduction blocks developed in all the rats (n=10) which received an intraneural injection of saline concomitantly with the systemic Cj-LPS exposure, before titers of anti-ganglioside antibodies were detected. We conclude that humoral factors (possibly cytokines), other than antibodies are secreted by lymphocytes and macrophages stimulated by gram negative LPS, and cause functional conduction abnormalities when the blood-nerve barrier is disrupted.


Latency competence of thirteen HSV-1 temperature-sensitive mutants.
Thirteen temperature-sensitive (ts) mutants of HSV-1 were analysed for their capacity to establish latent infections in the brains of mice. Eleven of the mutants could be classified as latency-positive or -negative; two could not be assigned to either group. Leakiness of mutants in the brain and differences in particle/infectivity ratios were found not to play a role in the results. Ts+ revertants of selected latency-negative mutants regained the capacity to establish latent infections, indicating that it was the ts lesion in these agents which was involved in latency. Ultrastructural studies of neuroblastoma cells infected with various mutants and maintained at the restrictive temperature showed that no absolute correlations could be made between capacity to establish latent infection and synthesis of various morphologically identifiable virus products. Finally, from a comparison of latency characteristics with previously established polypeptide phenotypes of mutants it was concluded that one immediate early and one or more later virus functions are necessary for establishment and/or maintenance of the latent state.