Palmitate acutely raises glycogen synthesis in rat soleus muscle by a mechanism that requires its metabolization (Randle cycle)

The acute effect of palmitate on glucose metabolism in rat skeletal muscle was examined. Soleus muscles from Wistar male rats were incubated in Krebs-Ringer bicarbonate buffer, for 1 h, in the absence or presence of 10 mU/ml insulin and 0, 50 or 100 microM palmitate. Palmitate increased the insulin-stimulated [(14)C]glycogen synthesis, decreased lactate production, and did not alter D-[U-(14)C]glucose decarboxylation and 2-deoxy-D-[2,6-(3)H]glucose uptake. This fatty acid decreased the conversion of pyruvate to lactate and [1-(14)C]pyruvate decarboxylation and increased (14)CO(2) produced from [2-(14)C]pyruvate. Palmitate reduced insulin-stimulated phosphorylation of insulin receptor substrate-1/2, Akt, and p44/42 mitogen-activated protein kinases. Bromopalmitate, a non-metabolizable analogue of palmitate, reduced [(14)C]glycogen synthesis. A strong correlation was found between [U-(14)C]palmitate decarboxylation and [(14)C]glycogen synthesis (r=0.99). Also, palmitate increased intracellular content of glucose 6-phosphate in the presence of insulin. These results led us to postulate that palmitate acutely potentiates insulin-stimulated glycogen synthesis by a mechanism that requires its metabolization (Randle cycle). The inhibitory effect of palmitate on insulin-stimulated protein phosphorylation might play an important role for the development of insulin resistance in conditions of chronic exposure to high levels of fatty acids.     

Heart rate reduction with ivabradine prevents the global phenotype of left ventricular remodeling

The aim of this study was to investigate the effect of chronic heart rate (HR) reduction with the hyperpolarization-activated current inhibitor ivabradine on the global phenotype of left ventricular (LV) remodeling in a ligated rat model. Seven days after coronary artery ligation, Wistar rats received ivabradine (10 mg · kg(-1) · day(-1) administered in drinking water) [myocardial infarction + ivabradine (MI+IVA), n = 22] or vehicle only (drinking water) (MI, n = 20) for 90 days. A sham group (n = 20) was included for model validation. MI+IVA rats had 12% lower HR (P < 0.01), improved LV volumes, 15% higher LV ejection fraction (LVEF, P < 0.01) than MI rats, and 33% reductions in both plasma atrial natriuretic peptide (ANP, P = 0.052) and cardiac hydroxyproline. Using patch-clamp, action potential duration was reduced and transient outward current density increased (P < 0.05). Cardiac energy metabolism was also improved (+33% creatine phosphate, P < 0.001; +15% ATP; and +9% energy charge, P < 0.05). Significant correlations were found between HR and parameters of cardiac metabolism, ANP, and LVEF (all P < 0.05). The HR-reducing properties of ivabradine prevent changes in the global phenotype of LV remodeling in the rat, optimize energy consumption, and avoid electrophysiological and structural remodeling.     

Effect of semen preparation on casa motility results in cryopreserved bull spermatozoa

Computer-assisted sperm analyzers (CASA) have become the standard tool for evaluating sperm motility and kinetic patterns because they provide objective data for thousands of sperm tracks. However, these devices are not ready-to-use and standardization of analytical practices is a fundamental requirement. In this study, we evaluated the effects of some settings, such as frame rate and frames per field, chamber and time of analysis, and samples preparations, including thawing temperature, sperm sample concentration, and media used for dilution, on the kinetic results of bovine frozen-thawed semen using a CASA. In Experiment 1, the frame rate (30-60 frame/s) significantly affected motility parameters, whereas the number of frames per field (30 or 45) did not seem to affect sperm kinetics. In Experiment 2, the thawing protocol affects sperm motility and kinetic parameters. Sperm sample concentration significantly limited the opportunity to perform the analysis and the kinetic results. A concentration of 100 and 50 × 10⁶ sperm/mL limited the device's ability to perform the analysis or gave wrong results, whereas 5, 10, 20, and 30 × 10⁶ sperm/mL concentrations allowed the analysis to be performed, but with different results (Experiment 3). The medium used for the dilution of the sample, which is fundamental for a correct sperm head detection, affects sperm motility results (Experiment 4). In this study, Makler and Leja chambers were used to perform the semen analysis with CASA devices. The chamber used significantly affected motility results (Experiment 5). The time between chamber loading and analysis affected sperm velocities, regardless of chamber used. Based on results recorded in this study, we propose that the CASA evaluation of motility of bovine frozen-thawed semen using Hamilton-Thorne IVOS 12.3 should be performed using a frame rate of 60 frame/s and 30 frames per field. Semen should be diluted at least at 20 × 10⁶ sperm/mL using PBS. Furthermore, it is necessary to consider the type of chamber used and perform the analysis within 1 or 2 min, regardless of the chamber used.     

Nesting habits of two spider wasps: Anoplius infuscatus and Episyron sp. (Hymenoptera: Pompilidae), with a review of the literature

Many behavioural aspects of Anoplius infuscatus (Vander Linden) and Episyron sp. Schioedte were examined in detail, both on a qualitative and, when possible, on a quantitative level, during a long-term field study in Northern Italy. Both wasps dig unicellular nests on bare soil, which are filled with a single spider to feed the brood. The main differences regard the way to hunt their prey, that of burrowing, that of transporting and introducing the spider into the nest, the habit of amputating spider legs and the abdominal movements during nest closing. On the whole, starting from prey capture to nest closure, A. infuscatus is significantly slower than Episyron. The observed ethological differences are well fitted to the exploitation of the different preys consisting in wolf spiders (Lycosidae) and orb weaving spiders (Araneidae), respectively. Some flexibility in the observed behaviour is described. A critical re-examination of the literature shows that, even if most results agree with those of previous authors, two controversial points remain open regarding Episyron species: the position of the egg (on the prey or on the cell wall) and the way to grasp the spider during tranport and introdution into the nest.     

Immunohistochemical analysis of pRb2/p130, VEGF, EZH2, p53, p16(INK4A), p27(KIP1), p21(WAF1), Ki-67 expression patterns in gastric cancer

Although the considerable progress against gastric cancer, it remains a complex lethal disease defined by peculiar histological and molecular features. The purpose of the present study was to investigate pRb2/p130, VEGF, EZH2, p53, p16(INK4A), p27(KIP1), p21(WAF1), Ki-67 expressions, and analyze their possible correlations with clinicopathological factors. The expression patterns were examined by immunohistochemistry in 47 patients, 27 evaluated of intestinal-type, and 20 of diffuse-type, with a mean follow up of 56 months and by Western blot in AGS, N87, KATO-III, and YCC-2, -3, -16 gastric cell lines. Overall, stomach cancer showed EZH2 correlated with high levels of p53, Ki-67, and cytoplasmic pRb2/p130 (P < 0.05, and P < 0.01, respectively). Increased expression of EZH2 was found in the intestinal-type and correlated with the risk of distant metastasis (P < 0.05 and P < 0.01, respectively), demonstrating that this protein may have a prognostic value in this type of cancer. Interestingly, a strong inverse correlation was observed between p27(KIP1) expression levels and the risk of advanced disease and metastasis (P < 0.05), and a positive correlation between the expression levels of p21(WAF1) and low-grade (G1) gastric tumors (P < 0.05), confirming the traditionally accepted role for these tumor-suppressor genes in gastric cancer. Finally, a direct correlation was found between the expression levels of nuclear pRb2/p130 and low-grade (G1) gastric tumors that was statistically significant (P < 0.05). Altogether, these data may help shed some additional light on the pathogenetic mechanisms related to the two main gastric cancer histotypes and their invasive potentials.     

Clinical Profiles and Factors Associated with Death in Adults with Dengue Admitted to Intensive Care Units,Minas Gerais,Brazil

The purpose of our study was to describe the clinical profile of dengue-infected patients admitted to Brazilian intensive care units (ICU) and evaluate factors associated with death. A longitudinal, multicenter case series study was conducted with laboratory-confirmed dengue patients admitted to nine Brazilian ICUs situated in Minas Gerais state, southeastern Brazil from January 1, 2008, to December 31, 2013. Demographic, clinical and laboratory data; disease severity scores; and mortality were evaluated. A total of 97 patients were studied. The in-ICU and in-hospital mortality rates were 18.6% and 19.6%, respectively. Patients classified as having severe dengue according to current World Health Organization classifications showed an increased risk of death in a univariate analysis. Nonsurvivors were older, exhibited lower serum albumin concentrations and higher total leukocyte counts and serum creatinine levels. Other risk factors (vomiting, lethargy/restlessness, dyspnea/respiratory distress) were also associated with death in a univariate analysis. Multivariate analysis indicated that in-hospital mortality was significantly associated with Acute Physiology and Chronic Health Evaluation II and the Sequential Organ Failure Assessment score. The ICU and in-hospital mortality observed in this study were higher than values reported in similar studies. An increased frequency of ICU admission due to severe organ dysfunction, higher severity indices and scarcity of ICU beds may partially explain the higher mortality.     

Glycogen synthase kinase-3 beta regulates NF-kappa B1/p105 stability

A number of different kinases have been implicated in NF-kappa B regulation and survival function. Here we investigated the molecular cross-talk between glycogen synthase kinase-3 beta (GSK-3 beta) and the p105 precursor of the NF-kappa B p50 subunit. GSK-3 beta forms an in vivo complex with and specifically phosphorylates NF-kappa B1/p105 at Ser-903 and Ser-907 in vitro. In addition, the p105 phosphorylation level is reduced in fibroblasts lacking GSK-3 beta as compared with wild-type cells. GSK-3 beta has a dual effect on p105: it stabilizes p105 under resting conditions and primes p105 for degradation upon tumor necrosis factor (TNF)-alpha treatment. Indeed, constitutive processing of p105 to p50 occurs at a higher rate in cells lacking GSK-3 beta with respect to wild-type cells and can be reduced upon reintroduction of GSK-3 beta by transfection. Moreover, p105 degradation in response to TNF-alpha is prevented in GSK-3 beta-/- fibroblasts and by a Ser to Ala point mutation on p105 at positions 903 or 907. Interestingly, the increased sensitiveness to TNF-alpha-induced death occurring in GSK-3 beta-/- fibroblasts, which is coupled to a perturbation of p50/105 ratio, can be reproduced by p105 silencing in wild-type fibroblasts.