High-intensity endurance training in 20- to 30- and 60- to 70-yr-old healthy men

Factors contributing to maximal incremental and short-term exercise capacity were measured before and after 12 wk of high-intensity endurance training in 12 old (60-70 yr) and 10 young (20-30 yr) sedentary healthy males. Peak O2 uptake in incremental cycle ergometer exercise increased from 1.60 +/- 0.073 to 2.21 +/- 0.073 (SE) l/min (38% increase) in the old subjects and from 2.54 +/- 0.141 to 3.26 +/- 0.181 l/min (29%) in the young subjects. Peak cardiac output, estimated by extrapolation from a series of submaximal measurements by the CO2 rebreathing method, increased by 30% (from 12.7 to 16.5 l/min) in the old subjects, associated with a 6% increase (from 126 to 135 ml/l) in arteriovenous O2 difference; in the young subjects there were equal 14% increases in both variables (18.0 to 20.5 l/min and 140 to 159 ml/l, respectively). Submaximal mean arterial pressure and cardiac output were lower posttraining in the old subjects; total vascular conductance and cardiac stroke volume increased. Although peak power at the start of a short-term maximal isokinetic test did not change, total work accomplished in 30 s at a pedaling frequency of 110 revolutions/min increased in both groups, from 11.2 to 12.6 kJ and from 15.7 to 16.9 kJ in the old and young, respectively; fatigue during the 30-s test was less, and postexercise plasma lactate concentrations were lower. In older subjects, increases in aerobic power after high-intensity endurance training are at least as large as in younger subjects and are associated with increases in vascular conductance, maximal cardiac output, and stroke volume.     

Facile entry to (-)-(R)- and (+)-(S)-mexiletine

The title compounds, 1a and 1b, have been synthesized in a three-step sequence starting from (-)-(S) and (+)-(R)-propylene oxide, respectively, in acceptable overall yields. The enantiomeric excess values for 1a and 1b were 96% and 93% respectively, as assessed by HPLC analysis on a chiral stationary phase of the corresponding N-acetyl derivatives. The synthetic route herein presented may represent a facile entry to highly enriched mexiletine enantiomers, alternative to those previously reported in the literature.     

Behavioral responses of Vipr2-/- mice to light

Vasoactive intestinal polypeptide and its receptor, VPAC2, play important roles in the functioning of the dominant circadian pacemaker, located in the hypothalamic suprachiasmatic nuclei (SCN). Mice lacking VPAC2 receptors (Vipr2-/-) show altered circadian rhythms and impaired synchronization to environmental lighting cues. However, light can increase phosphoprotein and immediate early gene expression in the Vipr2-/- SCN demonstrating that the circadian clock is readily responsive to light in these mice. It is not clear whether these neurochemical responses to light can be transduced to behavioral changes as seen in wild-type (WT) animals. In this study we investigated the diurnal and circadian wheel-running profile of WT (C57BL/6J) and Vipr2-/- mice under a 12-h light:12-h complete darkness (LD) lighting schedule and in constant darkness (DD) and used 1-h light pulses to shift the activity of mice in DD. Unlike WT mice, Vipr2-/- mice show grossly altered locomotor patterns making the analysis of behavioral responses to light problematic. However, analyses of both the onset and the offset of locomotor activity reveal that in a subset of these mice, light can reset the offset of behavioral rhythms during the subjective night. This suggests that the SCN clock of Vipr2-/- mice and the rhythms it generates are responsive to photic stimulation and that these responses can be integrated to whole animal behavioral changes.     

Multivalency - a way to enhance binding avidities and bioactivity - preliminary applications to EPO.

Multivalency has advantages over monovalency for binding interactions and even for activity. In particular, avidity is higher since the off-rate of a multivalent species is much slower than that of a monomer. This is particularly profitable for ligand-binding receptors that require dimerization for activity, such as the receptor of erythropoietin (EPOR). Peptides that mimic the action of erythropoietin (EPO) have been described with no sequence similarity with the human hormone: erythropoietin mimetic peptide (EMP) and EPO receptor peptide (ERP). These two peptides have similar activity but interact through different sites on the EPOR. Here, we describe the construction of several new synthetic homo- and hetero-dimers based on EMP-ERP sequences. To link the monomeric molecules together, several monodisperse polyamide linkers of different lengths were synthesized with dialdehyde functionalities. Chemoselective oxime chemistry was used to obtain homogeneous constructs. Certain chemical incompatibilities were dealt with via a protection approach. The oximes are stable under normal conditions and so lend themselves to biological testing.     

Gallstones - approach to medical management

Between 10% and 15% of individuals in the industrialized world have gallstones. The standard treatment is laparoscopic cholecystectomy, making gallstone disease the second most costly digestive disorder in most Western countries. Despite a rapid convalescence, the procedure is not devoid of morbidity or even mortality. Bile duct injury is particularly troublesome, occurring in 0.1% to 0.5% of cases, even in the most experienced hands. Moreover, some 20% of patients continue to suffer from pain (the main indication for treatment) after cholecystectomy. In patients with mild symptoms, surgical treatment has been associated with a higher morbidity than the natural course of the disease. Medical dissolution therapy with bile acids is an alternative for patients with mild-to-moderate symptoms due to cholesterol gallstones. Chenodeoxycholic acid (CDCA, chenodiol) has been largely replaced by the safer and more efficient ursodeoxycholic acid (UDCA). The main drawbacks of UDCA treatment are its low efficacy (approximately 40%), slowness in action, and the possibility of stone recurrence. However, this treatment is extremely safe, and the efficacy and slowness can be somewhat improved by stricter patient selection. Moreover, patient symptoms may respond to this therapy even without complete stone dissolution. New strategies employing more efficient bile acids or related compounds may increase the efficacy of medical dissolution. Furthermore, recent advances in the understanding of biliary lipid secretion and regulation should offer novel opportunities to further improve the prospects of medical treatment of gallstones.     

I - insulin transfer to mitochondria

The aim of this study was to determine if insulin is transferred to mitoplasts by insulin-degrading enzyme (IDE). Hepatic mitochondria were isolated and controlled by electron microscopy. IDE was obtained from rats muscle by successive chromatography steps. Insulin accumulation in mitoplasts and outer membrane + intermembrane space (OM + IMS) was studied with ¹²⁵I-insulin. Mitochondrial insulin accumulation and degradation was assayed with Sephadex G50 chromatography, insulin antibody and 5 % TCA. Mitoplasts and OM + IMS were isolated with digitonin. Insulin accumulation was studied at 25 °C at different times, without or with IDE, Bacitracin, 2,4-dinitrophenol, apyrase or sodium succinate + adenosine diphosphate. Insulin accumulation in mitoplasts and OM + IMS after mitochondrial cross-linking was studied with electrophoresis in SDS-PAGE, immunoblots of IDE, insulin or TIM23 (inner mitochondrial transporter) and autoradiography. The studies showed that addition of IDE increased insulin transfer from OM + IMS to mitoplasts, and the insulin accumulation in mitoplast was IDE dependent. Bacitracin and 2,4-dinitrophenol decreased this transfer. The [Insulin-IDE] complex and [Mitoplasts] was studied as a bimolecular reaction following a second order reaction. The constant “k” (liter.mol^?1 s^?1) showed that IDE increased and Bacitracin or 2,4-dinitrophenol decreased the velocity of insulin transfer. SDS-PAGE and immunoblots studies showed bands and radioactivity coincident with IDE, insulin and TIM23. Non degraded insulin was demonstrated in immunoblot after IDE immunoprecipitation from mitoplasts. Confocal studies showed mitochondrial colocalization of IDE and insulin. The results showed that insulin at 25 °C were transferred from OM + IMS to mitoplasts by IDE or that the enzyme facilitates this transfer, and they reach the matrix together.     

14-3-3 Binding to Cyclin Y contributes to cyclin Y/CDK14 association

Cyclin Y is a highly conserved cyclin among eumetazoans, yet its function and regulation are poorly understood. To search for Cyclin Y-interacting proteins, we screened a yeast two-hybrid library using human Cyclin Y （CCNY） as a bait and identified the following interactors： CDK14 and four members of the 14-3-3 family （ε,β,η,τ）. The interaction between CCNY and 14-3-3 proteins was confirmed both in vitro and in vivo. The results showed that Ser-100 and Ser-326 residues in CCNY were crucial for 14-3-3 binding. Interestingly, binding of CCNY to 14-3-3 significantly enhanced the association between CCNY and CDK14. Our findings may add a new layer of regulation of CCNY binding to its kinase partner.     

14-3-3 antagonizes Ras-mediated Raf-1 recruitment to the plasma membrane to maintain signaling fidelity

We have investigated the role that S259 phosphorylation, S621 phosphorylation, and 14-3-3 binding play in regulating Raf-1 activity. We show that 14-3-3 binding, rather than Raf-1 phosphorylation, is required for the correct regulation of kinase activity. Phosphorylation of S621 is not required for activity, but 14-3-3 binding is essential. When 14-3-3 binding to conserved region 2 (CR2) was disrupted, Raf-1 basal kinase activity was elevated and it could be further activated by (V12,G37)Ras, (V23)TC21, and (V38)R-Ras. Disruption of 14-3-3 binding at CR2 did not recover binding of Raf-1 to (V12,G37)Ras but allowed more efficient recruitment of Raf-1 to the plasma membrane and stimulated its phosphorylation on S338. Finally, (V12)Ras, but not (V12,G37)Ras, displaced 14-3-3 from full-length Raf-1 and the Raf-1 bound to Ras. GTP was still phosphorylated on S259. Our data suggest that stable association of Raf-1 with the plasma membrane requires Ras-mediated displacement of 14-3-3 from CR2. Small G proteins that cannot displace 14-3-3 fail to recruit Raf-1 to the membrane efficiently and so fail to stimulate kinase activity.     

Functional polymorphisms in matrix metalloproteinases -1, -3, -9 and -12 in relation to cervical artery dissection

Background  

Cervical artery dissection is a leading cause of cerebral ischemia in young adults. Morphological investigations have shown alterations in the extracellular matrix (ECM) of affected vessel walls. As matrix metalloproteinases (MMP) play a central role in the regulation of the ECM, an increased expression of these enzymes might lead to the endothelial damage in spontaneous cervical artery dissection (sCAD). Five different DNA polymorphisms in MMP-1, -3, -9 and -12 were tested for their frequency in patients with sCAD and compared with those of a control population.     

Molecular cloning and cellular distribution of two 14-3-3 isoforms from Hydra: 14-3-3 proteins respond to starvation and bind to phosphorylated targets

In the simple metazoan Hydra a clear link between food supply and cell survival has been established. Whilst in plants 14-3-3 proteins are found to be involved in signalling cascades that regulate metabolism, in animals they have been shown to participate in cell survival pathways. In order to explore the possibility that 14-3-3 proteins in Hydra could be involved in regulating metabolism under different conditions of food supply, we have cloned two isoforms of 14-3-3 proteins. We show here that 14-3-3 proteins bind to phosphorylated targets in Hydra and form homo- and heterodimers in vitro. 14-3-3 proteins are localised in the cytoplasm of all cells and also in the nuclei of some epithelial cells. This nuclear localisation becomes more prominent during starvation. Moreover, 14-3-3 protein is present in large amounts in food granules and from this we conclude that it performs functions which are associated with metabolism and food storage in Hydra.     

14-3-3 Family Members Act Coordinately to Regulate Mitotic Progression

CARP1 and CARP2 proteins (CARPs) are E3 ligases that target p53 as well as phospho-p53 for degradation. Because MDM2 is a critical regulator of p53 turnover, we investigated and found that CARPs associate with MDM2. We provide evidence that CARPs stabilize MDM2 by inhibiting MDM2 self-ubiquitination. CARPs together with MDM2 enhance p53 degradation, thereby inhibiting p53-mediated cell death. CARP protein levels correlate with MDM2 levels including under hypoxia where both are reduced. CARP2 was found to target 14-3-3σ for degradation, leading to MDM2 stabilization. MDMX, a homolog of MDM2, is not absolutely required for MDM2 stabilization by CARPs, although overexpression of CARP2 enhances MDM2/MDMX interaction. Taken together, our study identifies novel mechanisms by which CARP proteins regulate the p53 signaling pathway.     

MicroRNA Targets - How to predict?

A number of web tools are available for the prediction and identification of target microRNAs (miRNAs). The choice, availability, validity and selection of an optimal yet appropriate tool are a challenge for the design of high throughput assays with promising miRNA targets. The current trends and challenges for target microRNAs (miRNAs) prediction, identification and selection is described in this review.     

Odefy - From discrete to continuous models

Background  

Phenomenological information about regulatory interactions is frequently available and can be readily converted to Boolean models. Fully quantitative models, on the other hand, provide detailed insights into the precise dynamics of the underlying system. In order to connect discrete and continuous modeling approaches, methods for the conversion of Boolean systems into systems of ordinary differential equations have been developed recently. As biological interaction networks have steadily grown in size and complexity, a fully automated framework for the conversion process is desirable.     

Nocturnal acid breakthrough - approach to management

Nocturnal acid breakthrough is defined as the presence of intragastric pH < 4 during the overnight period for at least 60 continuous minutes in patients taking a proton-pump inhibitor (PPI). Nocturnal acid breakthrough occurs in more than 70% of Helicobacter pylori-negative patients on PPI therapy and has clinical consequences in particular in patients with complicated gastroesophageal reflux disease (GERD), Barrett's esophagus, and esophageal motility abnormalities. The clinical importance of nocturnal acid breakthrough and the benefit of adding histamine-2 receptor antagonists (H2RAs) to PPI therapy have been debated ever since these concepts were introduced. In our experience, the addition of bedtime H2RAs is clinically effective in controlling nocturnal acid breakthrough and GERD symptoms.