title: Effects of exercise training on health status in patients with chronic heart failure: HF-ACTION randomized controlled trial.

text: CONTEXT Findings from previous studies of the effects of exercise training on patient-reported health status have been inconsistent. OBJECTIVE To test the effects of exercise training on health status among patients with heart failure. DESIGN, SETTING, AND PATIENTS Multicenter, randomized controlled trial among 2331 medically stable outpatients with heart failure with left ventricular ejection fraction of 35% or less. Patients were randomized from April 2003 through February 2007. INTERVENTIONS Usual care plus aerobic exercise training (n = 1172), consisting of 36 supervised sessions followed by home-based training, vs usual care alone (n = 1159). Randomization was stratified by heart failure etiology, which was a covariate in all models. MAIN OUTCOME MEASURES Kansas City Cardiomyopathy Questionnaire (KCCQ) overall summary scale and key subscales at baseline, every 3 months for 12 months, and annually thereafter for up to 4 years. The KCCQ is scored from 0 to 100 with higher scores corresponding to better health status. Treatment group effects were estimated using linear mixed models according to the intention-to-treat principle. RESULTS Median follow-up was 2.5 years. At 3 months, usual care plus exercise training led to greater improvement in the KCCQ overall summary score (mean, 5.21; 95% confidence interval, 4.42 to 6.00) compared with usual care alone (3.28; 95% confidence interval, 2.48 to 4.09). The additional 1.93-point increase (95% confidence interval, 0.84 to 3.01) in the exercise training group was statistically significant (P < .001). After 3 months, there were no further significant changes in KCCQ score for either group (P = .85 for the difference between slopes), resulting in a sustained, greater improvement overall for the exercise group (P < .001). Results were similar on the KCCQ subscales, and no subgroup interactions were detected. CONCLUSIONS Exercise training conferred modest but statistically significant improvements in self-reported health status compared with usual care without training. Improvements occurred early and persisted over time. TRIAL REGISTRATION clinicaltrials.gov Identifier: NCT00047437.


title: Acute respiratory failure in patients with severe community-acquired pneumonia: a prospective randomized evaluation of noninvasive ventilation.

text: In uncontrolled studies, noninvasive positive pressure ventilation (NPPV) was found useful in avoiding endotracheal intubation in patients with acute respiratory failure (ARF) caused by severe community-acquired pneumonia (CAP). We conducted a prospective, randomized study comparing standard treatment plus NPPV delivered through a face mask to standard treatment alone in patients with severe CAP and ARF. Patients fitting the American Thoracic Society criteria for severe CAP were included in presence of ARF (refractory hypoxemia and/or hypercapnia with acidosis). Exclusion criteria were: severe hemodynamic instability, requirement for emergent cardiopulmonary resuscitation, home mechanical ventilation or oxygen long-term supplementation, concomitant severe disease with a low expectation of life, inability to expectorate or contraindications to the use of the mask. Fifty-six consecutive patients (28 in each arm) were enrolled, and the two groups were similar at study entry. The use of NPPV was well tolerated, safe, and associated with a significant reduction in respiratory rate, need for endotracheal intubation (21% versus 50%; p = 0.03), and duration of intensive care unit (ICU) stay (1.8 ± 0.7 d versus 6 ± 1.8 d; p = 0.04). The two groups had a similar intensity of nursing care workload, time interval from study entry to endotracheal intubation, duration of hospitalization, and hospital mortality. Among patients with chronic obstructive pulmonary disease (COPD), those randomized to NPPV had a lower intensity of nursing care workload (p = 0.04) and improved 2-mo survival (88.9% versus 37.5%; p = 0.05). We conclude that in selected patients with ARF caused by severe CAP, NPPV was associated with a significant reduction in the rate of endotracheal intubation and duration of ICU stay. A 2-mo survival advantage was seen in patients with COPD.


title: Smallest angiosperm genomes found in lentibulariaceae, with chromosomes of bacterial size.

text: Nuclear holoploid genome sizes (C-values) have been estimated to vary about 800-fold in angiosperms, with the smallest established 1C-value of 157 Mbp recorded in Arabidopsis thaliana. In the highly specialized carnivorous family Lentibulariaceae now three taxa have been found that exhibit significantly lower values: Genlisea margaretae with 63 Mbp, G. aurea with 64 Mbp, and Utricularia gibba with 88 Mbp. The smallest mitotic anaphase chromatids in G. aurea have 2.1 Mbp and are thus of bacterial size (NB: E. coli has ca. 4 Mbp). Several Utricularia species range somewhat lower than A. thaliana or are similar in genome size. The highest 1C-value known from species of Lentibulariaceae was found in Genlisea hispidula with 1510 Mbp, and results in about 24-fold variation for Genlisea and the Lentibulariaceae. Taking into account these new measurements, genome size variation in angiosperms is now almost 2000-fold. Genlisea and Utricularia are plants with terminal positions in the phylogeny of the eudicots, so that the findings are relevant for the understanding of genome miniaturization. Moreover, the Genlisea-Utricularia clade exhibits one of the highest mutational rates in several genomic regions in angiosperms, what may be linked to specialized patterns of genome evolution. Ultrasmall genomes have not been found in Pinguicula, which is the sister group of the Genlisea-Utricularia clade, and which does not show accelerated mutational rates. C-values in Pinguicula varied only 1.7-fold from 487 to 829 Mbp.


title: Parasite multiplication potential and the severity of Falciparum malaria.

text: The multiplication rates and invasiveness of Plasmodium falciparum parasites isolated from adult Thai patients hospitalized with uncomplicated malaria (n=34) were compared with those from persons with severe malaria (n=42). To simulate severe malaria and control for host effects, the in vitro cultures were adjusted to 1% parasitemia and used the same red blood cell donor. P. falciparum isolates from persons with severe malaria had initial cycle multiplication rates in vitro that were 3-fold higher than those from uncomplicated malaria (median [95% confidence interval], 8.3 [7. 1-10.5] vs. 2.8 [1.7-3.9]; P=.001). Parasites causing severe malaria exhibited unrestricted red blood cell invasion, whereas those from uncomplicated malaria were restricted to a geometric mean of 40 (31%-53%) of red blood cells. P. falciparum parasites causing severe malaria were less selective and multiplied more at high parasitemias than those causing uncomplicated malaria.


title: PHD3-dependent hydroxylation of HCLK2 promotes the DNA damage response.

text: The DNA damage response (DDR) is a complex regulatory network that is critical for maintaining genome integrity. Posttranslational modifications are widely used to ensure strict spatiotemporal control of signal flow, but how the DDR responds to environmental cues, such as changes in ambient oxygen tension, remains poorly understood. We found that an essential component of the ATR/CHK1 signaling pathway, the human homolog of the Caenorhabditis elegans biological clock protein CLK-2 (HCLK2), associated with and was hydroxylated by prolyl hydroxylase domain protein 3 (PHD3). HCLK2 hydroxylation was necessary for its interaction with ATR and the subsequent activation of ATR/CHK1/p53. Inhibiting PHD3, either with the pan-hydroxylase inhibitor dimethyloxaloylglycine (DMOG) or through hypoxia, prevented activation of the ATR/CHK1/p53 pathway and decreased apoptosis induced by DNA damage. Consistent with these observations, we found that mice lacking PHD3 were resistant to the effects of ionizing radiation and had decreased thymic apoptosis, a biomarker of genomic integrity. Our identification of HCLK2 as a substrate of PHD3 reveals the mechanism through which hypoxia inhibits the DDR, suggesting hydroxylation of HCLK2 is a potential therapeutic target for regulating the ATR/CHK1/p53 pathway.


title: Economic implications of nighttime attending intensivist coverage in a medical intensive care unit.

text: OBJECTIVE Our objective was to assess the cost implications of changing the intensive care unit staffing model from on-demand presence to mandatory 24-hr in-house critical care specialist presence. DESIGN A pre-post comparison was undertaken among the prospectively assessed cohorts of patients admitted to our medical intensive care unit 1 yr before and 1 yr after the change. Our data were stratified by Acute Physiology and Chronic Health Evaluation III quartile and whether a patient was admitted during the day or at night. Costs were modeled using a generalized linear model with log-link and γ-distributed errors. SETTING A large academic center in the Midwest. PATIENTS All patients admitted to the adult medical intensive care unit on or after January 1, 2005 and discharged on or before December 31, 2006. Patients receiving care under both staffing models were excluded. INTERVENTION Changing the intensive care unit staffing model from on-demand presence to mandatory 24-hr in-house critical care specialist presence. MEASUREMENTS AND MAIN RESULTS Total cost estimates of hospitalization were calculated for each patient starting from the day of intensive care unit admission to the day of hospital discharge. Adjusted mean total cost estimates were 61% lower in the post period relative to the pre period for patients admitted during night hours (7 pm to 7 am) who were in the highest Acute Physiology and Chronic Health Evaluation III quartile. No significant differences were seen at other severity levels. The unadjusted intensive care unit length of stay fell in the post period relative to the pre period (3.5 vs. 4.8) with no change in non-intensive care unit length of stay. CONCLUSIONS We find that 24-hr intensive care unit intensivist staffing reduces lengths of stay and cost estimates for the sickest patients admitted at night. The costs of introducing such a staffing model need to be weighed against the potential total savings generated for such patients in smaller intensive care units, especially ones that predominantly care for lower-acuity patients.


title: GENOTYPING OF THALASSEMIA IN MICROCYTIC HYPOCHROMIC ANEMIA PATIENTS FROM SOUTHWEST REGION OF IRAN

text: Objective: To evaluate the frequency of α -gene, s-gene, and hemoglobin variant numbers in subjects with Microcytic hypochromic anemia. Methodology: In total out of 850, 340 subjects with microcytic hypochromic anemia [MCV<80fl; MCH<27pg] from Southwest part of Iran, were studied in Research Center of Thalassemia and Hemoglobinopathies (RCTH) which is the only center working on hematology and oncology in Southwest (Khuzestan) region of Iran. These include 325 individuals: 171 with Beta-thalassemia trait, 88 with Alpha-thalassemia trait, 13 with thalassemia major, 11 with hemoglobin variants (HbS, HbC, and HbD Punjab ) and 42 with iron-deficiency anemia. The rest 15 patients diagnosed with no definite etiology. Results: Genotyping for -α 3.7 , -α 4.2 , – α PA , - α 5NT and - - MED was done with gap-PCR. The overall frequency of - α 3.7 deletion in 325 individuals is 20%. Genotyping for 23 most known s-gene mutations was done with direct mutation analysis by Amplification Refractory Mutation System (ARMS). The most frequent mutations were CD 36/37, IVS II-I, and IVS I-110 with 9.7%, 11.7%, and 3.5% respected frequencies in 340 patients. There was statistically significant difference between Beta-thalassemia trait and Beta-thalassemia Major in case of MCV (p- value = 0.25) and MCH (P–value =0.23) indices, and also MCH index between Beta-thalassemia trait and Hb Variants (P-value = 0.04). Conclusion: The α -gene and s-gene mutation is quite common in the Southwest part of Iran. Molecular genotyping of α -thalassemia and s-thalassemia help to diagnose unexplained microcytosis, and thus prevent unnecessary iron supplementation.


title: Systemic biomarkers in exacerbations of COPD: the evolving clinical challenge.

text: BACKGROUND Exacerbations of COPD (ECOPD) remain a major cause of mortality and morbidity. Despite advances in the understanding of their pathophysiology, their assessment relies primarily on clinical presentation, which can be variable and difficult to predict. A large number of biomarkers already have been assessed in this context, and some appear to be promising. METHODS An online search for articles published until December 2010 was conducted using three terms for ECOPD, five terms for biomarkers, and five terms for the sampling method. Biomarkers were evaluated for their potential role in the establishment and confirmation of the diagnosis of ECOPD, the evaluation of etiology and severity, the prediction of prognosis, and the guidance of treatment decisions. RESULTS Several systemic biomarkers have been measured in the context of ECOPD, and most have been found to increase at ECOPD onset and to subside during the course of exacerbations. Correlations have been reported among these biomarkers, but direct associations with clinical variables have been more difficult to establish. Although there are several limitations yet to be addressed, some of the biomarkers, most notably C-reactive protein for the identification of an ECOPD and procalcitonin for antibiotic guidance, may provide clinically relevant information. CONCLUSIONS So far , no single biomarker has been able to gain wide acceptance, but some provide clinically useful information. The evaluation of such biomarkers in large decision-making studies is expected to become an area of intense investigation in the near future.


title: Preoperative Predictors of Weight Loss Following Bariatric Surgery: Systematic Review

text: Obesity affects 32% of adults in the USA. Surgery generates substantial weight loss, but 20–30% fails to achieve successful weight loss. Our objective was to identify preoperative psychosocial factors associated with weight loss following bariatric surgery. We performed a literature search of PubMed® and the Cochrane Database of Reviews of Effectiveness between 1988 and April 2010. Articles were screened for bariatric surgery and weight loss if they included a preoperative predictor of weight loss: body mass index (BMI), preoperative weight loss, eating disorders, or psychiatric disorder/substance abuse. One thousand seven titles were reviewed, 534 articles screened, and 115 included in the review. Factors that may be positively associated with weight loss after surgery include mandatory preoperative weight loss (7 of 14 studies with positive association). Factors that may be negatively associated with weight loss include preoperative BMI (37 out of 62 studies with negative association), super-obesity (24 out of 33 studies), and personality disorders (7 out of 14 studies). Meta-analysis revealed a decrease of 10.1% excess weight loss (EWL) for super-obese patients (95% confidence interval (CI) [3.7–16.5%]), though there was significant heterogeneity in the meta-analysis, and an increase of 5.9% EWL for patients with binge eating at 12 months after surgery (95% CI [1.9–9.8%]). Further studies are necessary to investigate whether preoperative factors can predict a clinically meaningful difference in weight loss after bariatric surgery. The identification of predictive factors may improve patient selection and help develop interventions targeting specific needs of patients.


title: The tumor necrosis factor receptor stalk regions define responsiveness to soluble versus membrane-bound ligand.

text: The family of tumor necrosis factor receptors (TNFRs) and their ligands form a regulatory signaling network that controls immune responses. Various members of this receptor family respond differently to the soluble and membrane-bound forms of their respective ligands. However, the determining factors and underlying molecular mechanisms of this diversity are not yet understood. Using an established system of chimeric TNFRs and novel ligand variants mimicking the bioactivity of membrane-bound TNF (mTNF), we demonstrate that the membrane-proximal extracellular stalk regions of TNFR1 and TNFR2 are crucial in controlling responsiveness to soluble TNF (sTNF). We show that the stalk region of TNFR2, in contrast to the corresponding part of TNFR1, efficiently inhibits both the receptor's enrichment/clustering in particular cell membrane regions and ligand-independent homotypic receptor preassembly, thereby preventing sTNF-induced, but not mTNF-induced, signaling. Thus, the stalk regions of the two TNFRs not only have implications for additional TNFR family members, but also provide potential targets for therapeutic intervention.


title: A bird's-eye view of sex chromosome dosage compensation.

text: Intensive study of a few genetically tractable species with XX/XY sex chromosomes has produced generalizations about the process of sex chromosome dosage compensation that do not fare well when applied to ZZ/ZW sex chromosome systems, such as those in birds. The inherent sexual imbalance in dose of sex chromosome genes has led to the evolution of sex-chromosome-wide mechanisms for balancing gene dosage between the sexes and relative to autosomal genes. Recent advances in our knowledge of avian genomes have led to a reexamination of sex-specific dosage compensation (SSDC) in birds, which is less effective than in known XX/XY systems. Insights about the mechanisms of SSDC in birds also suggest similarities to and differences from those in XX/XY species. Birds are thus offering new opportunities for studying dosage compensation in a ZZ/ZW system, which should shed light on the evolution of SSDC more broadly.


title: The role of CHMP2BIntron5 in autophagy and frontotemporal dementia.

text: Charged multivesicular body protein 2B (CHMP2B) - a component of the endosomal complex required for transport-III (ESCRT-III) - is responsible for the vital membrane deformation functions in autophagy and endolysosomal trafficking. A dominant mutation in CHMP2B (CHMP2BIntron5) is associated with a subset of heritable frontotemporal dementia - frontotemporal dementia linked to chromosome 3 (FTD-3). ESCRT-III recruits Vps4, an AAA-ATPase that abscises the membrane during various cellular processes including autophagy and intraluminal vesicle formation. CHMP2BIntron5 results in a C-terminus truncation removing an important Vps4 binding site as well as eliminating the normal autoinhibitory resting state of CHMP2B. CHMP2B is expressed in most cell types but seems to be especially vital for proper neuronal function. CHMP2BIntron5-mediated phenotypes include misregulation of transmembrane receptors, accumulation of multilamellar structures, abnormal lysosomal morphology, down regulation of a brain-specific micro RNA (miRNA-124), abnormal dendritic spine morphology, decrease in dendritic arborization, and cell death. Currently, transgenic-fly,-mouse, and -human cell lines are being used to better understand the diverse phenotypes and develop therapeutic approaches for the CHMP2BIntron5-induced FTD-3. This article is part of a Special Issue entitled SI:Autophagy.


title: Restless legs syndrome and attention-deficit/hyperactivity disorder: a review of the literature.

text: STUDY OBJECTIVE To review evidence on the association between restless legs syndrome (RLS) and attention-deficit/hyperactivity disorder (ADHD), to discuss the hypothetical mechanisms underlying this association, and to consider the potential interest for common pharmacologic treatments of RLS and ADHD when co-occurring. METHOD A PubMed search. RESULTS In clinical samples, up to 44% of subjects with ADHD have been found to have RLS or RLS symptoms, and up to 26% of subjects with RLS have been found to have ADHD or ADHD symptoms. Several mechanisms may explain this association. Sleep disruption associated with RLS might lead to inattentiveness, moodiness, and paradoxical overactivity. Diurnal manifestations of RLS, such as restlessness and inattention, might mimic ADHD symptoms. Alternatively, RLS might be comorbid with idiopathic ADHD. Subjects with RLS and a subset of subjects with ADHD might share a common dopamine dysfunction. Limited evidence suggests that some dopaminergic agents, such as levodopa/carbidopa, pergolide, and ropinirole, may be effective in children with RLS associated with ADHD symptoms. CONCLUSIONS Although still limited, evidence from clinical studies demonstrates an association between RLS and ADHD or ADHD symptoms. Further clinical studies using standard criteria and procedures are needed to better estimate the degree of association. Epidemiologic studies are required to assess the relationship between ADHD and RLS symptoms in nonclinical samples. Further investigations should address the mechanisms underlying the relationship between RLS and ADHD. Several dopaminergic agents seem to be promising treatment for RLS associated with ADHD symptoms. To date, however, the absence of randomized and blinded controlled studies does not allow evidence-based recommendations.


title: Impact of the KU80 pathway on NHEJ-induced genome rearrangements in mammalian cells.

text: Using a substrate measuring deletion or inversion of an I-SceI-excised fragment and both accurate and inaccurate rejoining, we determined the impact of non-homologous end-joining (NHEJ) on mammalian chromosome rearrangements. Deletion is 2- to 8-fold more efficient than inversion, independent of the DNA ends structure. KU80 controls accurate rejoining, whereas in absence of KU mutagenic rejoining, particularly microhomology-mediated repair, occurs efficiently. In cells bearing both the NHEJ and a homologous recombination (HR) substrate containing a third I-SceI site, we show that NHEJ is at least 3.3-fold more efficient than HR, and translocation of the I-SceI fragment from the NHEJ substrate locus into the HR-I-SceI site can occur, but 50- to 100-fold less frequently than deletion. Deletions and translocations show both accurate and inaccurate rejoining, suggesting that they correspond to a mix of KU-dependent and KU-independent processes. Thus these processes should represent prominent pathways for DSB-induced genetic instability in mammalian cells.


title: The multi-zinc finger protein ZNF217 contacts DNA through a two-finger domain.

text: Classical C2H2 zinc finger proteins are among the most abundant transcription factors found in eukaryotes, and the mechanisms through which they recognize their target genes have been extensively investigated. In general, a tandem array of three fingers separated by characteristic TGERP links is required for sequence-specific DNA recognition. Nevertheless, a significant number of zinc finger proteins do not contain a hallmark three-finger array of this type, raising the question of whether and how they contact DNA. We have examined the multi-finger protein ZNF217, which contains eight classical zinc fingers. ZNF217 is implicated as an oncogene and in repressing the E-cadherin gene. We show that two of its zinc fingers, 6 and 7, can mediate contacts with DNA. We examine its putative recognition site in the E-cadherin promoter and demonstrate that this is a suboptimal site. NMR analysis and mutagenesis is used to define the DNA binding surface of ZNF217, and we examine the specificity of the DNA binding activity using fluorescence anisotropy titrations. Finally, sequence analysis reveals that a variety of multi-finger proteins also contain two-finger units, and our data support the idea that these may constitute a distinct subclass of DNA recognition motif.


title: Knowledge, attitudes, risk perception, and cancer screening behaviors among cancer survivors.

text: BACKGROUND Knowledge, attitudes, and risk perception in relation to second primary cancer (SPC) screening and their impact on screening practices in cancer survivors are largely unknown. METHODS A total of 326 cancer survivors who had completed primary treatment for cancer >1 year previously were recruited from 6 oncology care outpatient clinics in the Republic of Korea. Survivors' knowledge, attitudes, perceived risk, and screening practices were assessed along with sociodemographic, behavioral, and clinical characteristics. Multivariate logistic regression was used to examine behavioral factors associated with the completion of all appropriate SPC screening according to national guidelines. RESULTS Approximately 37.7% of survivors had undergone all appropriate SPC screening tests. Survivors were found to have a high perceived risk of SPC, high perceived benefits of screening, and positive attitudes toward cancer screening. However, they had limited knowledge regarding SPC screening tests and few had received a recommendation from a physician to undergo SPC screening. Although there was no association found between perceived risk and positive attitudes with screening behavior, higher knowledge was noted to be significantly associated with the completion of all appropriate SPC screening (adjusted odds ratio, 1.81; 95% confidence interval, 1.03-3.33). CONCLUSIONS In the current study, cancer survivors were found to have limited knowledge regarding second cancer screening tests, which may have resulted in lower rates of completion of screening practices in this population.


title: Ria1p (Ynl163c), a protein similar to elongation factors 2, is involved in the biogenesis of the 60S subunit of the ribosome in Saccharomyces cerevisiae

text: RIA1 (YNL163c) is a quasi-essential gene that encodes a protein with strong similarities to elongation factors 2. Small C-terminal deletions in the protein lead to a severe growth defect. In the case of a 22-residue C-terminal deletion this can be suppressed by intragenic mutations in the RIA1 gene or dominant extragenic mutations in TIF6, which is thought to be involved in the biogenesis of the 60S subunit of the ribosome. The dominant TIF6 alleles can also suppress the phenotype associated with a complete deletion of the RIA1 gene. Depletion of Ria1p has a dramatic effect on the polysome profile: there is a severe reduction in the level of the 80S monosomes, an imbalance in the 40S/60S ratio, and halfmers appear. Dissociation of the monosomes and polysomes in the Ria1p depletion mutant revealed a specific reduction in the amount of 60S subunits. Localization experiments with HA-tagged derivatives of Ria1p did not detect any stable association of Ria1p with ribosome subunits, 80S monosomes or polysomes. Cell fractionation experiments show that Ria1p is found in both the cytoplasmic fraction and the nuclear fraction. Taken together, these data suggest that Ria1p is involved in the biogenesis of the 60S subunit of the ribosome.


title: The Intestinal Immune System in Obesity and Insulin Resistance.

text: Obesity and insulin resistance are associated with chronic inflammation in metabolic tissues such as adipose tissue and the liver. Recently, growing evidence has implicated the intestinal immune system as an important contributor to metabolic disease. Obesity predisposes to altered intestinal immunity and is associated with changes to the gut microbiota, intestinal barrier function, gut-residing innate and adaptive immune cells, and oral tolerance to luminal antigens. Accordingly, the gut immune system may represent a novel therapeutic target for systemic inflammation in insulin resistance. This review discusses the emerging field of intestinal immunity in obesity-related insulin resistance and how it affects metabolic disease.


title: Human placental lactogen and unconjugated estriol concentrations in twin pregnancy: monitoring of fetal development in intrauterine growth retardation and single intrauterine fetal death.

text: Human placental lactogen and unconjugated estriol concentrations in maternal serum were evaluated in 100 uneventful twin pregnancies, and these values were compared with those observed in 16 twin pregnancies associated with intrauterine growth retardation or single intrauterine fetal death. In pregnancies associated with intrauterine growth retardation (n = 8), human placental lactogen levels were at the lower limit of normal range for singleton pregnancies, whereas estriol levels were normal in most cases. When one of the fetuses had died before week 33 of pregnancy (n = 5), both human placental lactogen and estriol levels were low and they were almost at the levels in singleton pregnancy. When intrauterine fetal death occurred after week 36 of pregnancy (n = 3), both hormone levels remained normal until term. Thus human placental lactogen rather than estriol is a good indicator of intrauterine growth retardation in twin pregnancy. Both human placental lactogen and estriol are useful for the monitoring of the surviving fetus in the case of single intrauterine fetal death.


title: Low antioxidant enzyme gene expression in pancreatic islets compared with various other mouse tissues.

text: Using a sensitive Northern blot hybridization technique, gene expression of superoxide dismutase (SOD), catalase, and glutathione peroxidase was studied in pancreatic islets and for comparison in various other mouse tissues (liver, kidney, brain, lung, skeletal muscle, heart muscle, adrenal gland, and pituitary gland). Gene expression of the antioxidant enzymes was usually in the range of +/- 50% of that in the liver. Only in pancreatic islets gene expression was substantially lower. The levels of the cytoplasmic Cu/Zn SOD and the mitochondrial Mn SOD gene expression were in the range of 30-40% of those in the liver. Glutathione peroxidase gene expression was 15%, and catalase gene expression was not at all detectable in pancreatic islets. These low levels of antioxidant enzyme gene expression may provide an explanation for the extraordinary sensitivity of pancreatic beta cells towards cytotoxic damage by diabetogenic compounds and during the development of human and animal diabetes.


title: Cyclic nucleotide phosphodiesterase isozymes expressed in mouse skeletal muscle.

text: To understand changes in cyclic nucleotide metabolism in muscle disease states, the expression of phosphodiesterase (PDE) isozymes in normal mouse leg muscle was examined. Four subcellular fractions were generated by differential centrifugation at 10,000 x g and 100,000 x g. cAMP PDE activity was found predominately in the soluble fractions, while cGMP PDE activity was more evenly distributed amongst soluble and particulate fractions. Pharmacological inhibitors demonstrate that PDE4 represents the major cAMP hydrolyzing activity and PDE2 represents the major cGMP hydrolyzing activity in mouse leg muscle. PDE1 is expressed at low levels, while PDE3 and PDE5 are intermediate. Between 20 and 40% of total PDE activity remained in the presence of inhibitors for PDE1-PDE5, indicating that other PDE families contribute to the total PDE pool. Reverse-transcription PCR with family-specific primers showed expression of mRNA for PDE7-PDE9, supporting this conclusion. Total PDE activity was found to be elevated in tissue extracts from a mouse model of Duchenne's muscular dystrophy.


title: Exercise-induced asthma.

text: The effect of exercise on ventilatory function was studied in 7 patients with asthma. Three showed appreciable decrease in forced expiratory volume in one second (FEV 1) after exercise. The degree of hyperventilation produced by exercise correlated with the postexercise decrease in FEV 1. Voluntary hyperventilation at rest produced a decrease in FEV 1 in all patients with asthma. Breathing 5.6 per cent carbon dioxide during voluntary hyperventilation instead of room air induced a larger ventilation and greater decrease in FEV1. Neither exercise nor voluntary hyperventilation had any significant effect on the FEV 1 in normal subjects and patients with chronic bronchitis. Both exercise·induced and hyperventilation-induced decrease in FEV 1 were inhibited by prior administration of isoproterenol but not by atropine. It is postulated that exercise-induced asthma is probably hyperventilation-induced asthma. Several underlying mechanisms by which airway obstruction is produced are possible.


title: An antimicrobial protein of the gut symbiont Bacteroides fragilis with a MACPF domain of host immune proteins.

text: Bacteroidales are the most abundant Gram-negative bacteria of the human intestinal microbiota comprising more than half of the bacteria in many individuals. Some of the factors that these bacteria use to establish and maintain themselves in this ecosystem are beginning to be identified. However, ecological competition, especially interference competition where one organism directly harms another, is largely unexplored. To begin to understand the relevance of this ecological principle as it applies to these abundant gut bacteria and factors that may promote such competition, we screened Bacteroides fragilis for the production of antimicrobial molecules. We found that the production of extracellularly secreted antimicrobial molecules is widespread in this species. The first identified molecule, described in this manuscript, contains a membrane attack complex/perforin (MACPF) domain present in host immune molecules that kill bacteria and virally infected cells by pore formation, and mutations affecting key residues of this domain abrogated its activity. This antimicrobial molecule, termed BSAP-1, is secreted from the cell in outer membrane vesicles and no additional proteins are required for its secretion, processing or immunity of the producing cell. This study provides the first insight into secreted molecules that promote competitive interference among Bacteroidales strains of the human gut.


title: Directional perception of distributed sound sources.

text: The perception of spatially distributed sound sources was investigated by conducting two listening experiments in anechoic conditions with 13 loudspeakers evenly distributed in the frontal horizontal plane emitting incoherent noise signals. In the first experiment, widely distributed sound sources with gaps in their distribution emitted pink noise. The results indicated that the exact loudspeaker distribution could not be perceived accurately and that the width of the distribution was perceived to be narrower than it was in reality. Up to three spatially distributed loudspeakers that were simultaneously emitting sound could be individually perceived. In addition, the number of loudspeakers that were indicated as emitting sound was smaller than the actual number. In the second experiment, a reference with 13 loudspeakers and test cases with fewer loudspeakers were presented and their perceived spatial difference was rated. The effect of the noise bandwidth was of particular interest. Noise with different bandwidths centered around 500 and 4000 Hz was used. The results indicated that when the number of loudspeakers was increased from four to seven, the perceived auditory event was very similar to that perceived with 13 loudspeakers at all bandwidths. The perceived differences were larger in wideband noise than in narrow-band noise.


title: Adoptive T cell transfer for cancer immunotherapy in the era of synthetic biology.

text: Adoptive T cell transfer for cancer and chronic infection is an emerging field that shows promise in recent trials. Synthetic-biology-based engineering of T lymphocytes to express high-affinity antigen receptors can overcome immune tolerance, which has been a major limitation of immunotherapy-based strategies. Advances in cell engineering and culture approaches to enable efficient gene transfer and ex vivo cell expansion have facilitated broader evaluation of this technology, moving adoptive transfer from a "boutique" application to the cusp of a mainstream technology. The major challenge currently facing the field is to increase the specificity of engineered T cells for tumors, because targeting shared antigens has the potential to lead to on-target off-tumor toxicities, as observed in recent trials. As the field of adoptive transfer technology matures, the major engineering challenge is the development of automated cell culture systems, so that the approach can extend beyond specialized academic centers and become widely available.


title: Bruce treadmill test in children: normal values in a clinic population.

text: The Bruce treadmill protocol is suitable for children as young as age 4 years. Maximal endurance time may be used as the sole criterion of exercise capacity, and normal values were established with 327 children having an innocent heart murmur. Mean endurance time in boys increased from 10.4 minutes at age 4 to 5 years, to 14.1 minutes at age 13 to 15 years. Mean endurance time in girls increased from 9.5 minutes at age 4 to 5 years to 12.3 minutes at age 10 to 12 years. Mean maximal heart rate ranged from 193 to 206 beats/min. Age differences in mean maximal and submaximal heart rates were small. There were negative correlations between endurance time and the ratio of weight to height. There were negative correlations between heart rates at treadmill stages 1 to 3 and the endurance times. The correlation coefficient of endurance time with maximal oxygen uptake was 0.88, but for clinical purposes endurance time alone is a satisfactory indicator of exercise performance.


title: Immunoproteasome and LMP2 polymorphism in aged and Alzheimer's disease brains.

text: In this study, we investigated the presence and role of immunoproteasome and its LMP2 subunit polymorphism at codon 60 in Alzheimer's disease (AD). Immunoproteasome was present in brain areas such as hippocampus and cerebellum and localized in neurons, astrocytes and endothelial cells. A higher expression of immunoproteasome was found in brain of AD patients than in brain of non-demented elderly, being its expression in young brain negligible or absent. Furthermore, AD affected regions showed a partial decrease in proteasome trypsin-like activity. The study of LMP2 polymorphism (R/H) showed that it does not influence LMP2 expression (neither the mRNA nor mature protein) in brain tissue. However, control brain areas of AD patients carrying the RR genotype showed an increased proteasome activity in comparison with RH carriers. To test whether this effect of the genotype might be related to AD onset we performed a genetic study, which allowed us to exclude an association of LMP2 codon 60 polymorphism with AD onset, despite its influence on the proteasome activity in human brain.


title: The centrosome and its mode of inheritance: the reduction of the centrosome during gametogenesis and its restoration during fertilization.

text: Neither the restoration of the centrosome during fertilization nor its reduction during gametogenesis is fully understood, but both are pivotal events in development. During each somatic cell cycle, the chromosomes, cytoplasm, and centrosomes duplicate in interphase, and all three split in two during each cell division. While it has long been recognized that both the sperm and the egg contribute equal haploid genomes during fertilization and that the vast majority of the cytoplasm is contributed by the egg, the relative contributions of the centrosome by each gamete are still in question. This article explores centrosome inheritance patterns and considers nine integral and secondarily derived activities of the centrosome. Boveri once hypothesized that "The ripe egg possesses all of the elements necessary for development save an active division-center. The sperm, on the other hand, possesses such a center but lacks the protoplasmic substratum in which to operate. In this respect the egg and sperm are complementary structures; their union in syngamy thus restores to each the missing element necessary to further development. " This article reviews the evidence gathered from 11 experimental strategies used to test this theory. While the majority of these approaches supports the hypothesis that the sperm introduces the centrosome at fertilization, the pattern did not reveal itself as universal, since parthenogenesis occurs in nature and can be induced artificially, since centrosome and centriole form de novo in extracts from unfertilized eggs and since the centrosome is derived from maternal sources during fertilization in some systems--notably, in mice. Models of the centrosome are proposed, along with speculative mechanisms which might lead to the cloaking of the reproducing element of the maternal centrosome during oogenesis and the retention of this structure by the paternal centrosome during spermatogenesis. Proteins essential for microtubule nucleation, like gamma-tubulin, are retained in the cytoplasm during oogenesis, but are largely lost during spermatogenesis. It is further postulated that the restoration of the zygotic centrosome at fertilization requires the attraction of maternal centrosomal components (in particular, gamma-tubulin and the 25S "gamma-some" particle) to the paternal reproducing element; this, along with post-translational modifications (including phosphorylation, disulfide reduction, and calcium ion binding), creates a functional zygote centrosome by blending both maternal and paternal constituents.(ABSTRACT TRUNCATED AT 400 WORDS)


title: Aggregation of vascular risk factors and risk of incident Alzheimer disease.

text: BACKGROUND The prevalence of Alzheimer disease (AD) is increasing in the elderly, and vascular risk factors may increase its risk. OBJECTIVE To explore the association of the aggregation of vascular risk factors with AD. METHODS The authors followed 1,138 individuals without dementia at baseline (mean age 76.2) for a mean of 5.5 years. The presence of vascular risk factors was related to incident possible and probable AD. RESULTS Four risk factors (diabetes, hypertension, heart disease, and current smoking) were associated with a higher risk of AD (p < 0.10) when analyzed individually. The risk of AD increased with the number of risk factors (diabetes + hypertension + heart disease + current smoking). The adjusted hazards ratio of probable AD for the presence of three or more risk factors was 3.4 (95% CI: 1.8, 6.3; p for trend < 0.0001) compared with no risk factors. Diabetes and current smoking were the strongest risk factors in isolation or in clusters, but hypertension and heart disease were also related to a higher risk of AD when clustered with diabetes, smoking, or each other. CONCLUSIONS The risk of Alzheimer disease (AD) increased with the number of vascular risk factors. Diabetes and current smoking were the strongest risk factors, but clusters including hypertension and heart disease also increased the risk of AD. These associations are unlikely to be explained by misclassification of the outcome, given strong associations when only probable AD is considered.


title: Hotspot mutations in H3F3A and IDH1 define distinct epigenetic and biological subgroups of glioblastoma.

text: Glioblastoma (GBM) is a brain tumor that carries a dismal prognosis and displays considerable heterogeneity. We have recently identified recurrent H3F3A mutations affecting two critical amino acids (K27 and G34) of histone H3.3 in one-third of pediatric GBM. Here, we show that each H3F3A mutation defines an epigenetic subgroup of GBM with a distinct global methylation pattern, and that they are mutually exclusive with IDH1 mutations, which characterize a third mutation-defined subgroup. Three further epigenetic subgroups were enriched for hallmark genetic events of adult GBM and/or established transcriptomic signatures. We also demonstrate that the two H3F3A mutations give rise to GBMs in separate anatomic compartments, with differential regulation of transcription factors OLIG1, OLIG2, and FOXG1, possibly reflecting different cellular origins.


title: Glucocorticoid with cyclophosphamide for paraquat-induced lung fibrosis.

text: BACKGROUND Paraquat is an effective and widely used herbicide but is also a lethal poison. In many developing countries paraquat is widely available and inexpensive, making poisoning prevention difficult. However most of the people who become poisoned from paraquat have taken it as a means of suicide. Standard treatment for paraquat poisoning both prevents further absorption and reduces the load of paraquat in the blood through haemoperfusion or haemodialysis. The effectiveness of standard treatments is extremely limited. The immune system plays an important role in exacerbating paraquat-induced lung fibrosis. Immunosuppressive treatment using glucocorticoid and cyclophosphamide in combination is being developed and studied. OBJECTIVES To assess the effects of glucocorticoid with cyclophosphamide on mortality in patients with paraquat-induced lung fibrosis. SEARCH METHODS To identify randomised controlled trials (RCTs) on this topic, we searched the Cochrane Injuries Group's Specialised Register (searched 1 February 2012), the Cochrane Central Register of Controlled Trials (CENTRAL) (The Cochrane Library 2012, Issue 1), MEDLINE (Ovid SP) (1946 January Week 3 2012), EMBASE (Ovid SP) (1947 to Week 4 2012), ISI Web of Science: Science Citation Index Expanded (SCI-EXPANDED) (1970 to January 2012), ISI Web of Science: Conference Proceedings Citation Index - Science (CPCI-S) (1990 to January 2012), Chinese Biomedical Literature and Retrieval System (CBM) (1978 to April 2012), Chinese Medical Current Contents (CMCC) (1995 to April 2012), and Chinese Medical Academic Conference (CMAC) (1994 to April 2012). Searches were completed on English language databases on 1 February 2012 and on Chinese language databases on 12 April 2012. SELECTION CRITERIA RCTs were included in this review. All patients were to receive standard care, plus the intervention or control. The intervention was glucocorticoid with cyclophosphamide in combination versus a control of a placebo, standard care alone or any other therapy in addition to standard care. DATA COLLECTION AND ANALYSIS The mortality risk ratio (RR) and 95% confidence interval (CI) was calculated for each study on an intention-to-treat basis. Data for all-cause mortality at final follow-up were summarised in a meta-analysis using a fixed-effect model. MAIN RESULTS This systematic review includes three trials with a combined total of 164 participants who had moderate to severe paraquat poisoning. Patients who received glucocorticoid with cyclophosphamide in addition to standard care had a lower risk of death at final follow-up than those receiving standard care only (RR 0.72; 95% CI 0.59 to 0.89). AUTHORS' CONCLUSIONS Based on the findings of three small RCTs of moderate to severely poisoned patients, glucocorticoid with cyclophosphamide in addition to standard care may be a beneficial treatment for patients with paraquat-induced lung fibrosis. To enable further study of the effects of glucocorticoid with cyclophosphamide for patients with moderate to severe paraquat poisoning, hospitals may provide this treatment as part of an RCT with allocation concealment.


title: Hydroxyethyl starch (HES) versus other fluid therapies: effects on kidney function.

text: BACKGROUND Hydroxyethyl starches (HES) are synthetic colloids commonly used for fluid resuscitation, yet controversy exists about their impact on kidney function. OBJECTIVES To examine the effects of HES on kidney function compared to other fluid resuscitation therapies in different patient populations. SEARCH STRATEGY We searched the Cochrane Renal Group's specialised register, the Cochrane Central Register of Controlled Trials (CENTRAL, in The Cochrane Library), MEDLINE, EMBASE, MetaRegister and reference lists of articles. SELECTION CRITERIA Randomised controlled trials (RCTs) and quasi-RCTs in which HES was compared to an alternate fluid therapy for the prevention or treatment of effective intravascular volume depletion. Primary outcomes were renal replacement therapy (RRT), author-defined kidney failure and acute kidney injury (AKI) as defined by the RIFLE criteria. Secondary outcomes included serum creatinine and creatinine clearance. DATA COLLECTION AND ANALYSIS Screening, selection, data extraction and quality assessments for each retrieved article were carried out by two authors using standardised forms. Authors were contacted when published data were incomplete. Preplanned sensitivity and subgroup analyses were performed after data were analysed with a random effects model. MAIN RESULTS The review included 34 studies (2607 patients). Overall, the RR of author-defined kidney failure was 1.50 (95% CI 1.20 to 1.87; n = 1199) and 1.38 for requiring RRT (95% CI 0.89 to 2.16; n = 1236) in HES treated individuals compared with other fluid therapies. Subgroup analyses suggested increased risk in septic patients compared to non-septic (surgical/trauma) patients. Non-septic patient studies were smaller and had lower event rates, so subgroup differences may have been due to lack of statistical power in these studies. Only limited data was obtained for analysis of kidney outcomes by the RIFLE criteria. Overall, methodological quality of studies was good but subjective outcomes were potentially biased because most studies were unblinded. AUTHORS' CONCLUSIONS Potential for increased risk of AKI should be considered when weighing the risks and benefits of HES for volume resuscitation, particularly in septic patients. Large studies with adequate follow-up are required to evaluate the renal safety of HES products in non-septic patient populations. RIFLE criteria should be applied to evaluate kidney function in future studies of HES and, where data is available, to re-analyse those studies already published. There is inadequate clinical data to address the claim that safety differences exist between different HES products.


title: Dietary fat and obesity: evidence from epidemiology.

text: The epidemiological evidence that a high-fat diet promotes the development of obesity is considered suggestive but not definitive. The purpose of this paper is to provide a review of various epidemiological methods that have been used to address this issue as well as an updated summary of the existing evidence. Ecological studies describing dietary fat intake and obesity at the population level provide mixed results and are likely to be biased by both confounding and unknown data quality factors that differ systematically across the populations studied. Cross-sectional studies are generally in agreement that the concentration of fat in the diet is positively associated with relative weight. Prospective studies of diet in relation to subsequent weight change give inconsistent results. This may be due to behavioural factors such as dieting in response to weight gain; in addition, this type of study rarely takes into account the possible interaction between genetic predisposition and dietary fat in promoting weight gain. Finally, intervention studies in free-living subjects are considered, providing evidence of a consistent but short-lived period of active weight loss on low-fat diets. The experimental evidence on this relationship is more conclusive than the epidemiological evidence, although biological mechanisms remain controversial. Some areas for future epidemiological research involve: longitudinal studies of dietary fat intake as a predictor of growth in children; observational studies relating total dietary fat and specific types of fat to overall as well as regional adiposity; and randomized intervention studies of the effect of low-fat diets with particular emphasis on and familial predisposition to obesity and other possible modifying factors.


title: Uncoupling of leading- and lagging-strand DNA replication during lesion bypass in vivo.

text: Numerous agents attack DNA, forming lesions that impair normal replication. Specialized DNA polymerases transiently replace the replicative polymerase and copy past lesions, thus generating mutations, the major initiating cause of cancer. We monitored, in Escherichia coli, the kinetics of replication of both strands of DNA molecules containing a single replication block in either the leading or lagging strand. Despite a block in the leading strand, lagging-strand synthesis proceeded further, implying transient uncoupling of concurrent strand synthesis. Replication through the lesion requires specialized DNA polymerases and is achieved with similar kinetics and efficiencies in both strands.


title: Phosphoinositide 3-kinase signalling--which way to target?

text: Abstract Phosphoinositide 3-kinases (PI3Ks) are central to the control of cell growth, proliferation and survival, and drive the progression of tumours by activating phosphoinositide-dependent kinase, protein kinase B and the target of rapamycin. Other downstream effectors link PI3K to cell motility and the control of cardiovascular parameters. Current knowledge indicates that PI3Ks might qualify as drug targets for the treatment of cancer, chronic inflammation, allergy and cardiovascular failure. However, PI3Ks also modulate vital processes such as metabolic control and nutrient uptake. Here, mechanistic data and mouse phenotypic analyses are summarised, and the possible success of therapeutic inhibition of distinct PI3K isoforms is discussed.


title: Determination of the time course of capacitation in mouse spermatozoa using a chlortetracycline fluorescence assay.

text: The heads of mouse spermatozoa obtained 5 min after release from the excised caudae epididymides showed a characteristic fluorescence pattern in the presence of the fluorophore chlortetracycline (CTC). There was uniform fluorescence over the entire head with about half the sperm population showing a brighter line of fluorescence across the equatorial segment; this fluorescence pattern was designated "F". After 90-min incubation in culture medium (CM) containing 2% (w/v) bovine serum albumin, most of the sperm heads showed a dark band of nonfluorescence over the equatorial and postequatorial segment, while the anterior portion of the head showed bright fluorescence. This fluorescence pattern was designated "B." The time course for the disappearance of pattern F matched the time course of the appearance of pattern B, with a half-time of 30 min. The transformation was complete in 90 min. At longer times of incubation in CM, the percentage of spermatozoa showing pattern B declined; fluorescence over the entire head was lost, characteristic of the pattern for acrosome-reacted sperm (P. M. Saling and B. T. Storey (1979). J. Cell Biol. 83, 544-555). Mouse sperm showing pattern B were able to undergo the acrosome reaction, either spontaneously or by induction with acid-solubilized zonae pellucidae from mouse eggs (H.M. Florman and B. T. Storey (1982). Dev. Biol. 91, 121-130). The latter reaction was blocked by its specific inhibitor 3-quinuclidinyl benzilate (QNB). Mouse sperm showing pattern F could not be induced to undergo the acrosome reaction by exposure to solubilized zonae. This implies that the change from fluorescence pattern F to fluorescence pattern B corresponds with changes in the sperm which make them susceptible to undergo the acrosome reaction. This change occurs during the time interval previously determined to be needed for capacitation of mouse sperm in vitro in CM (M. Inoue and D. P. Wolf (1975). Biol. Reprod. 13, 340-346). These results imply that spermatozoa showing CTC fluorescence pattern B can be considered to be capacitated and that a functional definition for capacitation is the acquired ability to undergo the acrosome reaction rapidly when treated with acid-solubilized zonae pellucidae. The CTC fluorescence assay provides for the first time a means to monitor the time course of epididymal mouse sperm capacitation in vitro.


title: [Detection of CRISPR and its relationship to drug resistance in Shigella].

text: OBJECTIVE To detect clustered regularly interspaced short palindromic repeats (CRISPR) in Shigella, and to analyze its relationship to drug resistance. METHODS Four pairs of primers were used for the detection of convincing CRISPR structures CRISPR-S2 and CRISPR-S4, questionable CRISPR structures CRISPR-S1 and CRISPR-S3 in 60 Shigella strains. All primers were designed using sequences in CRISPR database. CRISPR Finder was used to analyze CRISPR and susceptibilities of Shigella strains were tested by agar diffusion method. Furthermore, we analyzed the relationship between drug resistance and CRISPR-S4. RESULTS The positive rate of convincing CRISPR structures was 95%. The four CRISPR loci formed 12 spectral patterns (A-L), all of which contained convincing CRISPR structures except type K. We found one new repeat and 12 new spacers. The multi-drug resistance rate was 53. 33% . We found no significant difference between CRISPR-S4 and drug resistant. However, the repeat sequence of CRISPR-S4 in multi- or TE-resistance strains was mainly R4.1 with AC deletions in the 3' end, and the spacer sequences of CRISPR-S4 in multi-drug resistance strains were mainly Sp5.1, Sp6.1 and Sp7. CONCLUSION CRISPR was common in Shigella. Variations df repeat sequences and diversities of spacer sequences might be related to drug resistance in Shigella.


title: The helicase DDX41 senses intracellular DNA mediated by the adaptor STING in dendritic cells

text: The recognition of pathogenic DNA is important to the initiation of antiviral responses. Here we report the identification of DDX41, a member of the DEXDc family of helicases, as an intracellular DNA sensor in myeloid dendritic cells (mDCs). Knockdown of DDX41 expression by short hairpin RNA blocked the ability of mDCs to mount type I interferon and cytokine responses to DNA and DNA viruses. Overexpression of both DDX41 and the membrane-associated adaptor STING together had a synergistic effect in promoting Ifnb promoter activity. DDX41 bound both DNA and STING and localized together with STING in the cytosol. Knockdown of DDX41 expression blocked activation of the mitogen-activated protein kinase TBK1 and the transcription factors NF-κB and IRF3 by B-form DNA. Our results suggest that DDX41 is an additional DNA sensor that depends on STING to sense pathogenic DNA.


title: Radiotherapy plus concomitant and adjuvant temozolomide for glioblastoma.

text: BACKGROUND Glioblastoma, the most common primary brain tumor in adults, is usually rapidly fatal. The current standard of care for newly diagnosed glioblastoma is surgical resection to the extent feasible, followed by adjuvant radiotherapy. In this trial we compared radiotherapy alone with radiotherapy plus temozolomide, given concomitantly with and after radiotherapy, in terms of efficacy and safety. METHODS Patients with newly diagnosed, histologically confirmed glioblastoma were randomly assigned to receive radiotherapy alone (fractionated focal irradiation in daily fractions of 2 Gy given 5 days per week for 6 weeks, for a total of 60 Gy) or radiotherapy plus continuous daily temozolomide (75 mg per square meter of body-surface area per day, 7 days per week from the first to the last day of radiotherapy), followed by six cycles of adjuvant temozolomide (150 to 200 mg per square meter for 5 days during each 28-day cycle). The primary end point was overall survival. RESULTS A total of 573 patients from 85 centers underwent randomization. The median age was 56 years, and 84 percent of patients had undergone debulking surgery. At a median follow-up of 28 months, the median survival was 14.6 months with radiotherapy plus temozolomide and 12.1 months with radiotherapy alone. The unadjusted hazard ratio for death in the radiotherapy-plus-temozolomide group was 0.63 (95 percent confidence interval, 0.52 to 0.75; P<0.001 by the log-rank test). The two-year survival rate was 26.5 percent with radiotherapy plus temozolomide and 10.4 percent with radiotherapy alone. Concomitant treatment with radiotherapy plus temozolomide resulted in grade 3 or 4 hematologic toxic effects in 7 percent of patients. CONCLUSIONS The addition of temozolomide to radiotherapy for newly diagnosed glioblastoma resulted in a clinically meaningful and statistically significant survival benefit with minimal additional toxicity.


title: Drug interactions between oral contraceptives and antibiotics.

text: OBJECTIVE To evaluate the evidence on possible drug interactions between antibiotics and oral contraceptives (OCs) that may lead to OC failure. DATA SOURCES MEDLINE and Lexis/Nexis Medical Library searches for 1966-1999 using the key word "oral contraceptives," cross-indexed with the terms "antibiotics," "adverse effects," and "pregnancy," and MEDLINE search using the additional MeSH term "drug interactions. " No language restrictions were used. METHODS OF STUDY SELECTION A total of 167 articles were retrieved for analysis. Another 32 articles were identified by review of the references cited in these publications. Articles were selected based on their ability to provide information on the relationship between antibiotic therapy and OC efficacy in otherwise compliant users (defined as women with unplanned pregnancies who reported compliance with their OC regimen). Additionally, studies that either directly measured the effects of antibiotics on the pharmacokinetics of OC components, or that analyzed the effects of antibiotics on measures of ovulation in OC users were accepted. TABULATION, INTEGRATION, AND RESULTS At least 30 cases have been reported of pregnancies occurring in women taking OCs and antibiotics, particularly rifampin. Approximately 20% of pregnant women reporting to family planning or abortion clinics reported concomitant OC and antibiotic use. Information from adverse event reporting databases generally mirrors the types of information gleaned from these case reports and clinical surveys and accounts for approximately one-third of reported cases. Retrospective surveys, primarily from dermatology-based practices, also have reported 24 pregnancies in OC users who concomitantly received therapy with antibiotics, most commonly tetracyclines and penicillins. Apparent OC failure rates in clinical surveys were within the usual range expected for patterns of typical use. In pooled results obtained from relatively small populations, oral antibiotics, with the exception of rifampin, have not significantly affected the pharmacokinetics of ethinyl estradiol, levonorgestrel, and norethindrone or reduced the serum concentrations of gonadotropins. However, individual patients have been identified who experienced significant decreases in the plasma concentration of these components of OCs and who appeared to ovulate. CONCLUSION Rifampin impairs the effectiveness of OCs. Pharmacokinetic studies of other antibiotics have not shown any systematic interaction between antibiotics and OC steroids. However, individual patients do show large decreases in the plasma concentrations of ethinyl estradiol when they take certain other antibiotics, notably tetracycline and penicillin derivatives. Because it is not possible to identify these women in advance, a cautious approach is advised.


title: Male Japanese quails with female brains do not show male sexual behaviors.

text: During embryonic development, gonadal steroid hormones (androgens and estrogens) are thought to organize the sexual differentiation of the brain in the heterogametic sexes of higher vertebrates (males in mammals, females in birds). Brain differentiation of the homogametic sexes is thought to proceed by default, not requiring sex hormones for sex-specific organization. In gallinaceous birds such as the Japanese quail, female brain organization is thought to develop via estrogen-dependent demasculinization of a default male brain phenotype. We performed male donor-to-female host (MF), female-to-male (FM), male-to-male (MM), and female-to-female (FF) isotopic, isochronic transplantation of the forebrain primordium in Japanese quail embryos before gonadal differentiation had occurred; brain chimeras had a forebrain (including the hypothalamus) originating exclusively from donor cells. MM, FF, and MF chimeras all showed sexual behavior governed by the genetic sex of the host. In contrast, FM chimeras (genetically female forebrain, all other tissues genetically male) showed no mounting and only rudimentary crowing behavior. Although MM, FF, MF, and FM chimeras all showed host-typical production of steroid hormones during embryonic life, only FM chimeras were hypogonadal, had atypical low levels of circulating testosterone in adulthood, and showed reduction (crowing) or absence (mounting) of reproductive behaviors. Morphological features of the medial preoptic nucleus (a sexually dimorphic brain area) also were not male-like in FM males. These data demonstrate a brain-intrinsic, genetically determined component that organizes the sex-typical production of gonadal hormones in adulthood and call for a reevaluation of the mechanisms underlying brain sexual differentiation in other higher-vertebrate species.


title: DNA joint dependence of pol X family polymerase action in nonhomologous end joining.

text: DNA double strand breaks (DSBs) can be rejoined directly by the nonhomologous end-joining (NHEJ) pathway of repair. Nucleases and polymerases are required to promote accurate NHEJ when the terminal bases of the DSB are damaged. The same enzymes also participate in imprecise rejoining and joining of incompatible ends, important mutagenic events. Previous work has shown that the Pol X family polymerase Pol4 is required for some but not all NHEJ events that require gap filling in Saccharomyces cerevisiae. Here, we systematically analyzed DSB end configurations and found that gaps on both strands and overhang polarity are the principal factors that determine whether a joint requires Pol4. DSBs with 3'-overhangs and a gap on each strand strongly depended on Pol4 for repair, DSBs with 5'-overhangs of the same sequence did not. Pol4 was not required when 3'-overhangs contained a gap on only one strand, however. Pol4 was equally required at 3'-overhangs of all lengths within the NHEJ-dependent range but was dispensable outside of this range, indicating that Pol4 is specific to NHEJ. Loss of Pol4 did not affect the rejoining of DSBs that utilized a recessed microhomology or DSBs bearing 5'-hydroxyls but no gap. Finally, mammalian Pol X polymerases were able to differentially complement a pol4 mutation depending on the joint structure, demonstrating that these polymerases can participate in yeast NHEJ but with distinct properties.


title: ABC of burns. Introduction.

text: Burns are one of the most devastating conditions encountered in medicine. The injury represents an assault on all aspects of the patient, from the physical to the psychological. It affects all ages, from babies to elderly people, and is a problem in both the developed and developing world. All of us have experienced the severe pain that even a small burn can bring. However the pain and distress caused by a large burn are not limited to the immediate event. The visible physical and the invisible psychological scars are long lasting and often lead to chronic disability. Burn injuries represent a diverse and varied challenge to medical and paramedical staff. Correct management requires a skilled multidisciplinary approach that addresses all the problems facing a burn patient. This series provides an overview of the most important aspects of burn injuries for hospital and non-hospital healthcare workers.workers. Figure 1    Top: Child with 70% full thickness burns, which required resuscitation, intensive care support, and extensive debridement and skin grafting. Left: The same child one year later at a burns camp, having made a good recovery. A reasonable outcome is possible ...


title: DNA methylation is a critical cell-intrinsic determinant of astrocyte differentiation in the fetal brain.

text: Astrocyte differentiation, which occurs late in brain development, is largely dependent on the activation of a transcription factor, STAT3. We show that astrocytes, as judged by glial fibrillary acidic protein (GFAP) expression, never emerge from neuroepithelial cells on embryonic day (E) 11.5 even when STAT3 is activated, in contrast to E14.5 neuroepithelial cells. A CpG dinucleotide within a STAT3 binding element in the GFAP promoter is highly methylated in E11.5 neuroepithelial cells, but is demethylated in cells responsive to the STAT3 activation signal to express GFAP. This CpG methylation leads to inaccessibility of STAT3 to the binding element. We suggest that methylation of a cell type-specific gene promoter is a pivotal event in regulating lineage specification in the developing brain.


title: DNMT1 Inhibition Reprograms Pancreatic Cancer Stem Cells via Upregulation of the miR-17-92 Cluster.

text: Pancreatic ductal adenocarcinoma (PDAC) and other carcinomas are hierarchically organized, with cancer stem cells (CSC) residing at the top of the hierarchy, where they drive tumor progression, metastasis, and chemoresistance. As CSC and non-CSC share an identical genetic background, we hypothesize that differences in epigenetics account for the striking functional differences between these two cell populations. Epigenetic mechanisms, such as DNA methylation, play an important role in maintaining pluripotency and regulating the differentiation of stem cells, but the role of DNA methylation in pancreatic CSC is obscure. In this study, we investigated the genome-wide DNA methylation profile of PDAC CSC, and we determined the importance of DNA methyltransferases for CSC maintenance and tumorigenicity. Using high-throughput methylation analysis, we discovered that sorted CSCs have a higher level of DNA methylation, regardless of the heterogeneity or polyclonality of the CSC populations present in the tumors analyzed. Mechanistically, CSC expressed higher DNMT1 levels than non-CSC. Pharmacologic or genetic targeting of DNMT1 in CSCs reduced their self-renewal and in vivo tumorigenic potential, defining DNMT1 as a candidate CSC therapeutic target. The inhibitory effect we observed was mediated in part through epigenetic reactivation of previously silenced miRNAs, in particular the miR-17-92 cluster. Together, our findings indicate that DNA methylation plays an important role in CSC biology and also provide a rationale to develop epigenetic modulators to target CSC plasticity and improve the poor outcome of PDAC patients. Cancer Res; 76(15); 4546-58. ©2016 AACR.


title: Mutation spectrum in the nephrin gene (NPHS1) in congenital nephrotic syndrome.

text: Congenital nephrotic syndrome, Finnish type (CNF or NPHS1), is an autosomal recessive disease characterized by massive proteinuria and development of nephrotic syndrome shortly after birth. The disease is most common in Finland, but many patients have been identified in other populations. The disease is caused by mutations in the gene for nephrin which is a key component of the glomerual ultrafilter, the podocyte slit diaphragm. A total of 30 mutations have been reported in the nephrin gene in patients with congenital nephrotic syndrome worldwide. In the Finnish population, two main mutations have been found. These two nonsense mutations account for over 94% of all mutations in Finland. Most mutations found in non-Finnish patients are missense mutations, but they include also nonsense and splice site mutations, as well as deletions and insertions. This mutation update summarizes the nature of all previously reported nephrin mutations and, additionally, describes 20 novel mutations recently identified in our laboratory.


title: The HSA domain binds nuclear actin-related proteins to regulate chromatin-remodeling ATPases

text: We identify the helicase-SANT–associated (HSA) domain as the primary binding platform for nuclear actin-related proteins (ARPs) and actin. Individual HSA domains from chromatin remodelers (RSC, yeast SWI-SNF, human SWI-SNF, SWR1 and INO80) or modifiers (NuA4) reconstitute their respective ARP–ARP or ARP–actin modules. In RSC, the HSA domain resides on the catalytic ATPase subunit Sth1. The Sth1 HSA is essential in vivo, and its omission causes the specific loss of ARPs and a moderate reduction in ATPase activity. Genetic selections for arp suppressors yielded specific gain-of-function mutations in two new domains in Sth1, the post-HSA domain and protrusion 1, which are essential for RSC function in vivo but not ARP association. Taken together, we define the role of the HSA domain and provide evidence for a regulatory relationship involving the ARP–HSA module and two new functional domains conserved in remodeler ATPases that contain ARPs.


title: Increased dendritic complexity and axonal length in cultured mouse cortical neurons overexpressing methyl-CpG-binding protein MeCP2

text: Rett syndrome is caused by loss-of-function mutations in the gene encoding the methyl DNA-binding factor MeCP2. As brain mass and neuronal complexity tend to be diminished in Rett patients, we tested whether MeCP2 directly influences the morphological complexity of developing neurons. Our results show that cultured mouse neurons overexpressing MeCP2beta (MECP2A) develop more complex morphologies, having longer axonal and dendritic processes, and an increased number of axonal and dendritic terminal endings. We then tested whether overexpressing a mutant form of MeCP2beta lacking its carboxyl terminus would elicit the same effects. Interestingly, while neurons overexpressing this mutant failed to enhance axonal and dendritic process elongation, the complexity of their axonal and dendritic processes remained significantly elevated. Taken together, these data support the hypothesis that MeCP2 directly regulates neuronal maturation and/or synaptogenesis, and provides evidence that MeCP2 may influence neuritic elongation and process branching through different mechanisms.


title: Folate affects apoptosis in human trophoblastic cells.

text: Effects of folate deficiency on the rate of apoptosis in human cytotrophoblastic cells has been investigated. Apoptosis was determined in cytotrophoblastic cells after culture in 1. control medium, 2. folate-free medium and 3. folate-free medium plus 10% fetal calf serum. Apoptosis rates in cells cultured in mediums 2 and 3 were significantly higher than those cultured in the control medium (P<0.02 and P<0.03, respectively). In conclusion, human cytotrophoblastic cells show a significantly increased rate of apoptosis in vitro after culture in a folate-free medium. Possible explanations for the association between folate deficiency and pregnancy complications are suggested.


title: Role of KRAS and EGFR as biomarkers of response to erlotinib in National Cancer Institute of Canada Clinical Trials Group Study BR.21.

text: PURPOSE To evaluate the effect of KRAS and epidermal growth factor receptor (EGFR) genotype on the response to erlotinib treatment in the BR.21, placebo-controlled trial. PATIENTS AND METHODS We analyzed 206 tumors for KRAS mutation, 204 tumors for EGFR mutation, and 159 tumors for EGFR gene copy by fluorescent in situ hybridization (FISH). We reanalyzed EGFR deletion/mutation using two highly sensitive techniques that detect abnormalities in samples with 5% to 10% tumor cellularity. KRAS mutation was analyzed by direct sequencing. RESULTS Thirty patients (15%) had KRAS mutations, 34 (17%) had EGFR exon 19 deletion or exon 21 L858R mutations, and 61 (38%) had high EGFR gene copy (FISH positive). Response rates were 10% for wild-type and 5% for mutant KRAS (P = .69), 7% for wild-type and 27% for mutant EGFR (P = .03), and 5% for EGFR FISH-negative and 21% for FISH-positive patients (P = .02). Significant survival benefit from erlotinib therapy was observed for patients with wild-type KRAS (hazard ratio [HR] = 0.69, P = .03) and EGFR FISH positivity (HR = 0.43, P = .004) but not for patients with mutant KRAS (HR = 1.67, P = .31), wild-type EGFR (HR = 0.74, P = .09), mutant EGFR (HR = 0.55, P = .12), and EGFR FISH negativity (HR = 0.80, P = .35). In multivariate analysis, only EGFR FISH-positive status was prognostic for poorer survival (P = .025) and predictive of differential survival benefit from erlotinib (P = .005). CONCLUSION EGFR mutations and high copy number are predictive of response to erlotinib. EGFR FISH is the strongest prognostic marker and a significant predictive marker of differential survival benefit from erlotinib.


title: Immune dysregulation in major depression: a critical review of existing evidence.

text: It is now well established that depression is associated with immune dysregulation. It is not, however, known whether this immune dysregulation plays a role in the pathophysiology of major depression or whether it increases the susceptibility of the depressed patient to immune-related disorders. This article presents a critical review of existing evidence for immune dysregulation in major depression, including changes in leucocyte trafficking, lymphocyte function, and markers of immune activation. Possible mediators of immune dysregulation in major depression are briefly discussed. Finally, the relationship between major depression and several medical conditions such as infection, allergy and autoimmune disorders, cardiovascular diseases, cancer and AIDS is critically reviewed.


title: Regulation of E2Fs and senescence by PML nuclear bodies.

text: The tumor suppressor PML (promyelocytic leukemia protein) regulates cellular senescence and terminal differentiation, two processes that implicate a permanent exit from the cell cycle. Here, we show that the mechanism by which PML induces a permanent cell cycle exit and activates p53 and senescence involves a recruitment of E2F transcription factors bound to their promoters and the retinoblastoma (Rb) proteins to PML nuclear bodies enriched in heterochromatin proteins and protein phosphatase 1α. Blocking the functions of the Rb protein family or adding back E2Fs to PML-expressing cells can rescue their defects in E2F-dependent gene expression and cell proliferation, inhibiting the senescent phenotype. In benign prostatic hyperplasia, a neoplastic disease that displays features of senescence, PML was found to be up-regulated and forming nuclear bodies. In contrast, PML bodies were rarely visualized in prostate cancers. The newly defined PML/Rb/E2F pathway may help to distinguish benign tumors from cancers, and suggest E2F target genes as potential targets to induce senescence in human tumors.


title: The International Classification of Headache Disorders.

text: A set of related medical disorders that lack a proper classification system and diagnostic criteria is like a society without laws. The result is incoherence at best, chaos at worst. For this reason, the International Classification of Headache Disorders (ICHD) is arguably the single most important breakthrough in headache medicine over the last 50 years. The ICHD identifies and categorizes more than a hundred different kinds of headache in a logical, hierarchal system. Even more important, it has provided explicit diagnostic criteria for all of the headache disorders listed. The ICHD quickly became universally accepted, and criticism of the classification has been minor relative to that directed at other disease classification systems. Over the 20 years following publication of the first edition of the ICHD, headache research has rapidly accelerated despite sparse allocation of resources to that effort. In summary, the ICHD has attained widespread acceptance at the international level and has substantially facilitated both clinical research and clinical care in the field of headache medicine.


title: Call to Develop a Standard Acquisition Charge Model for Kidney Paired Donation

text: We propose a Medicare Demonstration Project to develop a standard acquisition charge for kidney paired donation. A new payment strategy is required because Medicare and commercial insurance companies may not directly pay living donor costs intended to lead to transplantation of a beneficiary of a different insurance provider. Until the 1970s, when organ procurement organizations were empowered to serve as financial intermediaries to pay the upfront recovery expenses for deceased donor kidneys before knowing the identity of the recipient, there existed similar limitations in the recovery and placement of deceased donor organs. Analogous to the recovery of deceased donor kidneys, kidney paired donation requires the evaluation of living donors before identifying their recipient. Tissue typing, crossmatching and transportation of living donors or their kidneys represent additional financial barriers. Finally, the administrative expenses of the organizations that identify and coordinate kidney paired donation transplantation require reimbursement akin to that necessary for organ procurement organizations. To expand access to kidney paired donation for more patients, we propose a model to reimburse paired donation expenses analogous to the proven strategy used for over 30 years to pay for deceased donor solid organ transplantation in America.


title: Oxidant stress promotes disease by activating CaMKII.

text: CaMKII is activated by oxidation of methionine residues residing in the regulatory domain. Oxidized CaMKII (ox-CaMKII) is now thought to participate in cardiovascular and pulmonary diseases and cancer. This invited review summarizes current evidence for the role of ox-CaMKII in disease, considers critical knowledge gaps and suggests new areas for inquiry.


title: Genetic diagnosis and prognosis of Alzheimer's disease: challenges and opportunities.

text: Alzheimer's disease (AD), the most common form of dementia in western societies, is a pathologically and clinically heterogeneous disease with a strong genetic component. The recent advances in high-throughput genome technologies allowing for the rapid analysis of millions of polymorphisms in thousands of subjects has significantly advanced our understanding of the genomic underpinnings of AD susceptibility. During the last 5 years, genome-wide association and whole-exome- and whole-genome sequencing studies have mapped more than 20 disease-associated loci, providing insights into the molecular pathways involved in AD pathogenesis and hinting at potential novel therapeutic targets. This review article summarizes the challenges and opportunities of when using genomic information for the diagnosis and prognosis of AD.


title: WNT5A regulates adipose tissue angiogenesis via antiangiogenic VEGF-A165b in obese humans.

text: Experimental studies have suggested that Wingless-related integration site 5A (WNT5A) is a proinflammatory secreted protein that is associated with metabolic dysfunction in obesity. Impaired angiogenesis in fat depots has been implicated in the development of adipose tissue capillary rarefaction, hypoxia, inflammation, and metabolic dysfunction. We have recently demonstrated that impaired adipose tissue angiogenesis is associated with overexpression of antiangiogenic factor VEGF-A165b in human fat and the systemic circulation. In the present study, we postulated that upregulation of WNT5A is associated with angiogenic dysfunction and examined its role in regulating VEGF-A165b expression in human obesity. We biopsied subcutaneous and visceral adipose tissue from 38 obese individuals (body mass index: 44 ± 7 kg/m2, age: 37 ± 11 yr) during planned bariatric surgery and characterized depot-specific protein expression of VEGF-A165b and WNT5A using Western blot analysis. In both subcutaneous and visceral fat, VEGF-A165b expression correlated strongly with WNT5A protein (r = 0.9, P < 0.001). In subcutaneous adipose tissue where angiogenic capacity is greater than in the visceral depot, exogenous human recombinant WNT5A increased VEGF-A165b expression in both whole adipose tissue and isolated vascular endothelial cell fractions (P < 0.01 and P < 0.05, respectively). This was associated with markedly blunted angiogenic capillary sprout formation in human fat pad explants. Moreover, recombinant WNT5A increased secretion of soluble fms-like tyrosine kinase-1, a negative regulator of angiogenesis, in the sprout media (P < 0.01). Both VEGF-A165b-neutralizing antibody and secreted frizzled-related protein 5, which acts as a decoy receptor for WNT5A, significantly improved capillary sprout formation and reduced soluble fms-like tyrosine kinase-1 production (P < 0.05). We demonstrated a significant regulatory nexus between WNT5A and antiangiogenic VEGF-A165b in the adipose tissue of obese subjects that was linked to angiogenic dysfunction. Elevated WNT5A expression in obesity may function as a negative regulator of angiogenesis. NEW & NOTEWORTHY Wingless-related integration site 5a (WNT5A) negatively regulates adipose tissue angiogenesis via VEGF-A165b in human obesity.


title: At least 1400 base pairs of 5'-flanking DNA is required for the correct expression of the HO gene in yeast.

text: Homothallic yeast cells undergo a specific pattern of mating-type switching initiated by an endonuclease encoded by the HO gene. HO transcription is affected by cell type (a, alpha, and a/alpha), by cell age (mother or daughter), and by the cell cycle. This paper investigates the sequences involved in HO transcription by replacing genomic DNA with copies mutated in vitro. A region between -1000 and 1400 (called URS1) is necessary for transcription in addition to a "TATA"-like region at -90. The 900 bp of DNA separating URS1 from the "TATA" box is not necessary for transcription nor for a/alpha repression and some measure of mother/daughter control, but it is necessary for correct cell cycle control.


title: Inflammation and its role in neuroprotection, axonal regeneration and functional recovery after spinal cord injury.

text: Trauma to the central nervous system (CNS) triggers intraparenchymal inflammation and activation of systemic immunity with the capacity to exacerbate neuropathology and stimulate mechanisms of tissue repair. Despite our incomplete understanding of the mechanisms that control these divergent functions, immune-based therapies are becoming a therapeutic focus. This review will address the complexities and controversies of post-traumatic neuroinflammation, particularly in spinal cord. In addition, current therapies designed to target neuroinflammatory cascades will be discussed.


title: Order of intron removal influences multiple splice outcomes, including a two-exon skip, in a COL5A1 acceptor-site mutation that results in abnormal pro-alpha1(V) N-propeptides and Ehlers-Danlos syndrome type I.

text: Ehlers-Danlos syndrome (EDS) type I (the classical variety) is a dominantly inherited, genetically heterogeneous connective-tissue disorder. Mutations in the COL5A1 and COL5A2 genes, which encode type V collagen, have been identified in several individuals. Most mutations affect either the triple-helical domain of the protein or the expression of one COL5A1 allele. We identified a novel splice-acceptor mutation (IVS4-2A-->G) in the N-propeptide-encoding region of COL5A1, in one patient with EDS type I. The outcome of this mutation was complex: In the major product, both exons 5 and 6 were skipped; other products included a small amount in which only exon 5 was skipped and an even smaller amount in which cryptic acceptor sites within exon 5 were used. All products were in frame. Pro-alpha1(V) chains with abnormal N-propeptides were secreted and were incorporated into extracellular matrix, and the mutation resulted in dramatic alterations in collagen fibril structure. The two-exon skip occurred in transcripts in which intron 5 was removed rapidly relative to introns 4 and 6, leaving a large (270 nt) composite exon that can be skipped in its entirety. The transcripts in which only exon 5 was skipped were derived from those in which intron 6 was removed prior to intron 5. The use of cryptic acceptor sites in exon 5 occurred in transcripts in which intron 4 was removed subsequent to introns 5 and 6. These findings suggest that the order of intron removal plays an important role in the outcome of splice-site mutations and provide a model that explains why multiple products derive from a mutation at a single splice site.


title: Kruppel-like factor KLF10 targets transforming growth factor-beta1 to regulate CD4(+)CD25(-) T cells and T regulatory cells.

text: CD4(+)CD25(+) regulatory T cells (T regs) play a major role in the maintenance of self-tolerance and immune suppression, although the mechanisms controlling T reg development and suppressor function remain incompletely understood. Herein, we provide evidence that Kruppel-like factor 10 (KLF10/TIEG1) constitutes an important regulator of T regulatory cell suppressor function and CD4(+)CD25(-) T cell activation through distinct mechanisms involving transforming growth factor (TGF)-beta1 and Foxp3. KLF10 overexpressing CD4(+)CD25(-) T cells induced both TGF-beta1 and Foxp3 expression, an effect associated with reduced T-Bet (Th1 marker) and Gata3 (Th2 marker) mRNA expression. Consistently, KLF10(-/-) CD4(+)CD25(-) T cells have enhanced differentiation along both Th1 and Th2 pathways and elaborate higher levels of Th1 and Th2 cytokines. Furthermore, KLF10(-/-) CD4(+)CD25(-) T cell effectors cannot be appropriately suppressed by wild-type T regs. Surprisingly, KLF10(-/-) T reg cells have reduced suppressor function, independent of Foxp3 expression, with decreased expression and elaboration of TGF-beta1, an effect completely rescued by exogenous treatment with TGF-beta1. Mechanistic studies demonstrate that in response to TGF-beta1, KLF10 can transactivate both TGF-beta1 and Foxp3 promoters, implicating KLF10 in a positive feedback loop that may promote cell-intrinsic control of T cell activation. Finally, KLF10(-/-) CD4(+)CD25(-) T cells promoted atherosclerosis by approximately 2-fold in ApoE(-/-)/scid/scid mice with increased leukocyte accumulation and peripheral pro-inflammatory cytokines. Thus, KLF10 is a critical regulator in the transcriptional network controlling TGF-beta1 in both CD4(+)CD25(-) T cells and T regs and plays an important role in regulating atherosclerotic lesion formation in mice.


title: Synchrotron radiation solution X-ray scattering study of the pH dependence of the quaternary structure of yeast pyruvate decarboxylase.

text: The pH dependence of the quaternary structure of pyruvate decarboxylase from yeast was studied in the range 6.2 less than pH less than 8.4. There is an equilibrium with a midpoint around pH 7.5 between tetramers and dimers, and the catalytic activity of the enzyme depends on the volume fraction of tetramer. This equilibrium may provide an additional regulating mechanism besides substrate activation since accumulation of pyruvate would lead to a reduction in pH and hence an increase of the concentration of the catalytically active tetramer. Radiation damage during the X-ray scattering experiments results in a shift of this equilibrium and in the formation of octamers. These effects could be circumvented and analyzed using experimental and data processing methods which can be readily applied to other radiation-sensitive systems. The low-resolution shapes of the dimers and tetramers were determined from the scattering curves using spherical harmonics. The results indicate that a conformational change must occur in the dimers upon formation of the tetramers, in agreement with earlier circular dichroism measurements.


title: Enhancement of anxiety-like responsiveness to the cannabinoid CB(1) receptor agonist HU-210 following chronic stress.

text: The effect that chronic unpredictable stress had on the anxiety-like response elicited by the cannabinoid receptor agonist HU-210 [3-(1,1-dimethylheptyl)-(-)-11-hydroxy-delta 8-tetrahydrocannabinol] in the elevated plus maze was investigated here. Male Long-Evans rats were either unstressed or were subjected to a 21-day regimen of chronic unpredictable stress, and subsequently were subdivided into three testing groups (vehicle, 10 and 50 microg/kg of HU-210) and tested on the elevated plus maze. Results demonstrated that in unstressed animals, a low dose of HU-210 induced an anxiolytic response, whereas a high dose induced an anxiogenic response. Further, in stressed animals both the low and the high doses of HU-210 induced anxiogenic responses. These findings suggest that chronic stress enhances either cannabinoid receptor responsivity or one of the interacting systems implicated in emotional states.


title: Cardiac glycosides inhibit TNF-α/NF-κB signaling by blocking recruitment of TNF receptor-associated death domain to the TNF receptor

text: Digitoxin and structurally related cardiac glycoside drugs potently block activation of the TNF-α/NF-κB signaling pathway. We have hypothesized that the mechanism might be discovered by searching systematically for selective inhibitory action through the entire pathway. We report that the common action of these drugs is to block the TNF-α-dependent binding of TNF receptor 1 to TNF receptor-associated death domain. This drug action can be observed with native cells, such as HeLa, and reconstituted systems prepared in HEK293 cells. All other antiinflammatory effects of digitoxin on NF-κB and c-Jun N-terminal kinase pathways appear to follow from the blockade of this initial upstream signaling event.


title: Zinc supplementation during lactation: effects on maternal status and milk zinc concentrations.

text: The effects of a zinc supplement on maternal zinc status and milk zinc concentrations through > or = 7 mo of lactation were examined. Seventy-one lactating women received either a daily 15-mg zinc supplement (ZS, n = 40) or placebo (NZS, n = 31) started by 2 wk postpartum in a double-blind, randomized design. Overall mean zinc intakes were 13.0 +/- 3.4 mg/d for the NZS group and 25.7 +/- 3.9 mg/d (including supplement) for the ZS group. Plasma zinc concentrations of the ZS group were significantly higher than those of the NZS group (P = 0.05). Milk zinc concentrations declined significantly over the course of the study for all subjects but were not affected by zinc supplementation. The mean dietary zinc intake observed in the nonsupplemented group was adequate to maintain normal maternal zinc status and milk zinc concentrations through > or = 7 mo lactation. Similar controlled intervention trials in less well-nourished populations will be required to assess the impact of lower zinc intakes on milk zinc concentrations.


title: DMSO as a vehicle for central injections: tests with feeding elicited by norepinephrine injected into the paraventricular nucleus.

text: Dimethyl sulfoxide (DMSO) is becoming increasingly popular as a vehicle in studies employing central injections. The aim of the present study was to determine whether the vehicle required for solubilization of substances for central injection [75% DMSO and 25% artificial CSF (aCSF)] would alter the well-characterized stimulatory response to norepinephrine (NE) injected into the paraventricular nucleus (PVN) on short-term food intake. To evaluate its suitability, we compared the effects of repeated unilateral injections of NE dissolved in two different vehicles (100% aCSF or 75% DMSO, 25% aCSF), in separate groups of animals every 48 h over a 30-day period. NE (40 nmol) stimulated food intake by approximately sevenfold compared to either vehicle alone, and the stimulatory effect was similar whether aCSF or 75% DMSO was used as a vehicle. Furthermore, the NE-induced feeding did not vary in magnitude across a series of 13 tests. These results suggest that 75% DMSO is a suitable vehicle for administering NE (and likely other water-insoluble substances)in small volumes of 0.3 microl into specific brain regions.


title: Dodecameric structure and ATPase activity of the human TIP48/TIP49 complex.

text: TIP48 and TIP49 are two related and highly conserved eukaryotic AAA(+) proteins with an essential biological function and a critical role in major pathways that are closely linked to cancer. They are found together as components of several highly conserved chromatin-modifying complexes. Both proteins show sequence homology to bacterial RuvB but the nature and mechanism of their biochemical role remain unknown. Recombinant human TIP48 and TIP49 were assembled into a stable high molecular mass equimolar complex and tested for activity in vitro. TIP48/TIP49 complex formation resulted in synergistic increase in ATPase activity but ATP hydrolysis was not stimulated in the presence of single-stranded, double-stranded or four-way junction DNA and no DNA helicase or branch migration activity could be detected. Complexes with catalytic defects in either TIP48 or TIP49 had no ATPase activity showing that both proteins within the TIP48/TIP49 complex are required for ATP hydrolysis. The structure of the TIP48/TIP49 complex was examined by negative stain electron microscopy. Three-dimensional reconstruction at 20 A resolution revealed that the TIP48/TIP49 complex consisted of two stacked hexameric rings with C6 symmetry. The top and bottom rings showed substantial structural differences. Interestingly, TIP48 formed oligomers in the presence of adenine nucleotides, whilst TIP49 did not. The results point to biochemical differences between TIP48 and TIP49, which may explain the structural differences between the two hexameric rings and could be significant for specialised functions that the proteins perform individually.