Chronic oral administration of beta-adrenoceptor agonist clenbuterol affects myosin heavy chain (MHC) expression in adult mouse heart

The aim of this study was to analyze the effects of chronic administration of the beta-adrenoceptor agonist clenbuterol (2 mg/kg body weight/day for a period of 30 days) on the major contractile protein (myosin) in the left ventricular muscle of the adult mouse heart. Separation of myosin heavy chain (MHC) isoforms on 7.5 % glycerol SDS-PAGE and subsequent quantification of the gels by laser densitometry showed a 6.5-fold increase in the beta-isoform of MHC in the clenbuterol-treated group. The alpha : beta ratio of these two isoforms in the control group was 98.16+/-0.14 %: 1.83+/-0.14 %, whereas in the treated group it was 88.05+/-1.15 % : 11.95+/-1.15 %. Actomyosin ATPase activity assay demonstrated a significant (20 %) decline in ATPase activity of the tissue in the beta-agonist-treated group. These results suggest that chronic clenbuterol treatment is capable to induced the transformation of MHC isoforms increasing the slow beta-MHC isoform, which may contribute to the altered contractile mechanics of clenbuterol-treated hearts.     

Critical evaluation of methods to incorporate entropy loss upon binding in high-throughput docking

Proper accounting of the positional/orientational/conformational entropy loss associated with protein-ligand binding is important to obtain reliable predictions of binding affinity. Herein, we critically examine two simplified statistical mechanics-based approaches, namely a constant penalty per rotor method, and a more rigorous method, referred to here as the partition function-based scoring (PFS) method, to account for such entropy losses in high-throughput docking calculations. Our results on the estrogen receptor beta and dihydrofolate reductase proteins demonstrate that, while the constant penalty method over-penalizes molecules for their conformational flexibility, the PFS method behaves in a more "DeltaG-like" manner by penalizing different rotors differently depending on their residual entropy in the bound state. Furthermore, in contrast to no entropic penalty or the constant penalty approximation, the PFS method does not exhibit any bias towards either rigid or flexible molecules in the hit list. Preliminary enrichment studies using a lead-like random molecular database suggest that an accurate representation of the "true" energy landscape of the protein-ligand complex is critical for reliable predictions of relative binding affinities by the PFS method.     

The effects of apolipoprotein F deficiency on high density lipoprotein cholesterol metabolism in mice

Apolipoprotein F (apoF) is 29 kilodalton secreted sialoglycoprotein that resides on the HDL and LDL fractions of human plasma. Human ApoF is also known as Lipid Transfer Inhibitor protein (LTIP) based on its ability to inhibit cholesteryl ester transfer protein (CETP)-mediated transfer events between lipoproteins. In contrast to other apolipoproteins, ApoF is predicted to lack strong amphipathic alpha helices and its true physiological function remains unknown. We previously showed that overexpression of Apolipoprotein F in mice reduced HDL cholesterol levels by 20-25% by accelerating clearance from the circulation. In order to investigate the effect of physiological levels of ApoF expression on HDL cholesterol metabolism, we generated ApoF deficient mice. Unexpectedly, deletion of ApoF had no substantial impact on plasma lipid concentrations, HDL size, lipid or protein composition. Sex-specific differences were observed in hepatic cholesterol content as well as serum cholesterol efflux capacity. Female ApoF KO mice had increased liver cholesteryl ester content relative to wild type controls on a chow diet (KO: 3.4+/-0.9 mg/dl vs. WT: 1.2+/-0.3 mg/dl, p<0.05). No differences were observed in ABCG1-mediated cholesterol efflux capacity in either sex. Interestingly, ApoB-depleted serum from male KO mice was less effective at promoting ABCA1-mediated cholesterol efflux from J774 macrophages relative to WT controls.     

Malignant melanoma of mediastinum misdiagnosed as a spindle cell thymoma in a fine needle aspirate: a case report

BACKGROUND: Malignant melanomas in the medastinum are extremely rare. Both primary melanomas and metastatic lesions from a primary elsewhere can occur in the mediastinum. Aspiration biopsy of a melanoma at this unusual site may pose problems in diagnosis. CASE: A 35-year-old woman presented with an anterior mediastinal mass. Cytologic smears were hemorrhaghic and revealed a loosely dispersed population of spindle cells with prominent nucleoli. In view of the location, the possibility of spindle cell thymoma was suggested on cytology. Subsequent histology revealed a malignant melanoma. CONCLUSION: This case stresses that the cytopathologist should keep in mind the remote differential diagnosis of a malignant melanoma while evaluating spindle cell neoplasms of the mediastinum, especially in tumors with prominent cell dispersal and with cells that have prominent nucleoli even without melanin pigment. Accurate diagnosis helps in evaluating patients and avoids unnecessary surgery when the lesion represents a metastasis to the mediastinum from a primary elsewhere.     

Protein Phosphatase 2A and Cdc7 Kinase Regulate the DNA Unwinding Element-binding Protein in Replication Initiation

The DNA unwinding element (DUE)-binding protein (DUE-B) binds to replication origins coordinately with the minichromosome maintenance (MCM) helicase and the helicase activator Cdc45 in vivo, and loads Cdc45 onto chromatin in Xenopus egg extracts. Human DUE-B also retains the aminoacyl-tRNA proofreading function of its shorter orthologs in lower organisms. Here we report that phosphorylation of the DUE-B unstructured C-terminal domain unique to higher organisms regulates DUE-B intermolecular binding. Gel filtration analyses show that unphosphorylated DUE-B forms multiple high molecular weight (HMW) complexes. Several aminoacyl-tRNA synthetases and Mcm2–7 proteins were identified by mass spectrometry of the HMW complexes. Aminoacyl-tRNA synthetase binding is RNase A sensitive, whereas interaction with Mcm2–7 is nuclease resistant. Unphosphorylated DUE-B HMW complex formation is decreased by PP2A inhibition or direct DUE-B phosphorylation, and increased by inhibition of Cdc7. These results indicate that the state of DUE-B phosphorylation is maintained by the equilibrium between Cdc7-dependent phosphorylation and PP2A-dependent dephosphorylation, each previously shown to regulate replication initiation. Alanine mutation of the DUE-B C-terminal phosphorylation target sites increases MCM binding but blocks Cdc45 loading in vivo and inhibits cell division. In egg extracts alanine mutation of the DUE-B C-terminal phosphorylation sites blocks Cdc45 loading and inhibits DNA replication. The effects of DUE-B C-terminal phosphorylation reveal a novel S phase kinase regulatory mechanism for Cdc45 loading and MCM helicase activation.     

An integrated diabetic index using heart rate variability signal features for diagnosis of diabetes

Electrocardiogram (ECG) signals are difficult to interpret, and clinicians must undertake a long training process to learn to diagnose diabetes from subtle abnormalities in these signals. To facilitate these diagnoses, we have developed a technique based on the heart rate variability signal obtained from ECG signals. This technique uses digital signal processing methods and, therefore, automates the detection of diabetes from ECG signals. In this paper, we describe the signal processing techniques that extract features from heart rate (HR) signals and present an analysis procedure that uses these features to diagnose diabetes. Through statistical analysis, we have identified the correlation dimension, Poincaré geometry properties (SD2), and recurrence plot properties (REC, DET, L _mean) as useful features. These features differentiate the HR data of diabetic patients from those of patients who do not have the illness, and have been validated by using the AdaBoost classifier with the perceptron weak learner (yielding a classification accuracy of 86%). We then developed a novel diabetic integrated index (DII) that is a combination of these nonlinear features. The DII indicates whether a particular HR signal was taken from a person with diabetes. This index aids the automatic detection of diabetes, thereby allowing a more objective assessment and freeing medical professionals for other tasks.     

Socioeconomic status and incidence of sudden cardiac arrest

Background:

Low socioeconomic status is associated with poor cardiovascular health. We evaluated the association between socioeconomic status and the incidence of sudden cardiac arrest, a condition that accounts for a substantial proportion of cardiovascular-related deaths, in seven large North American urban populations.

Methods:

Using a population-based registry, we collected data on out-of-hospital sudden cardiac arrests occurring at home or at a residential institution from Apr. 1, 2006, to Mar. 31, 2007. We limited the analysis to cardiac arrests in seven metropolitan areas in the United States (Dallas, Texas; Pittsburgh, Pennsylvania; Portland, Oregon; and Seattle–King County, Washington) and Canada (Ottawa and Toronto, Ontario; and Vancouver, British Columbia). Each incident was linked to a census tract; tracts were classified into quartiles of median household income.

Results:

A total of 9235 sudden cardiac arrests were included in the analysis. For all sites combined, the incidence of sudden cardiac arrestin the lowest socioeconomic quartile was nearly double that in the highest quartile (incidence rate ratio [IRR] 1.9, 95% confidence interval [CI] 1.8–2.0). This disparity was greater among people less than 65 years old (IRR 2.7, 95% CI 2.5–3.0) than among those 65 or older (IRR 1.3, 95% CI 1.2–1.4). After adjustment for study site and for population age structure of each census tract, the disparity across socioeconomic quartiles for all ages combined was greater in the United States (IRR 2.0, 95% CI 1.9–2.2) than in Canada (IRR 1.8, 95% CI 1.6–2.0) (p < 0.001 for interaction).

Interpretation:

The incidence of sudden cardiac arrest at home or at a residential institution was higher in poorer neighbourhoods of the US and Canadian sites studied, although the association was attenuated in Canada. The disparity across socioeconomic quartiles was greatest among people younger than 65. The association between socioeconomic status and incidence of sudden cardiac arrest merits consideration in the development of strategies to improve survival from sudden cardiac arrest, and possibly to identify opportunities for prevention.An estimated 250 000–300 000 sudden cardiac arrests occur each year in the United States,¹ accounting for up to 63% of cardiac-related deaths annually.² Despite advances in resuscitation, more than 95% of people who experience sudden cardiac arrest die,³ and up to 50% of sudden cardiac arrests occur in people who do not have a history of coronary artery disease.⁴Socioeconomic status has been shown to predict many health outcomes, including all-cause mortality,⁵ prevalence of risk factors for cardiovascular disease⁶ and incidence of cardiovascular disease.^7–9 Despite this substantial literature, we found only three studies that examined the potential association between socioeconomic status and sudden cardiac arrest. Although the studies were small and conducted in single communities, each showed that the incidence of sudden cardiac arrest was significantly higher in lower socioeconomic areas.^10–12 The Oregon Sudden Unexplained Death Study (Ore-SUDS) reported a 30%–80% higher incidence of sudden cardiac arrest in poorer neighbourhoods. A stronger association was observed among people less than 65 years old, a group for whom basic health care funding is not guaranteed in the United States.¹¹Low socioeconomic status may be linked to an increased risk of sudden cardiac arrest by a variety of mechanisms related to individual risk factors or health-promoting behaviours or neighbourhood characteristics. Individuals of lower socioeconomic status have been found to have a greater burden of risk factors for cardiovascular disease,¹³ poorer control of established cardiovascular risk factors¹⁴ and longer delays in seeking hospital care for acute myocardial infarction.¹⁵ Numerous studies have also shown that disparities in health outcomes are apparent across the spectrum of socioeconomic status.¹⁶A better understanding of community-level patterns in the distribution of sudden cardiac arrest may identify opportunities for improving survival, such as effective targeting of community training for cardiopulmonary resuscitation and placement of automated external defibrillators in lower-income communities. We tested the hypothesis that disparities in the incidence of sudden cardiac arrest by level of socioeconomic status would be evident in a variety of urban communities in the United States and Canada, and that this association would be most prominent among people less than 65 years old residing in US communities.