EGCG, a green tea polyphenol, improves endothelial function and insulin sensitivity, reduces blood pressure, and protects against myocardial I/R injury in SHR

Epigallocatechin gallate (EGCG), a bioactive polyphenol in green tea, may augment metabolic and vascular actions of insulin. Therefore, we investigated effects of EGCG treatment to simultaneously improve cardiovascular and metabolic function in spontaneously hypertensive rats (SHR; model of metabolic syndrome with hypertension, insulin resistance, and overweight). In acute studies, EGCG (1-100 microM) elicited dose-dependent vasodilation in mesenteric vascular beds (MVB) isolated from SHR ex vivo that was inhibitable by N(omega)-nitro-L-arginine methyl ester (L-NAME; nitric oxide synthase antagonist) or wortmannin [phosphatidylinositol (PI) 3-kinase inhibitor]. In chronic studies, 9-wk-old SHR were treated by gavage for 3 wk with EGCG (200 mg.kg(-1).day(-1)), enalapril (30 mg.kg(-1).day(-1)), or vehicle. A separate group of SHR receiving L-NAME (80 mg/l in drinking water) was treated for 3 wk with either EGCG or vehicle. Vasodilator actions of insulin were significantly improved in MVB from EGCG- or enalapril-treated SHR (when compared with vehicle-treated SHR). Both EGCG and enalapril therapy significantly lowered systolic blood pressure (SBP) in SHR. EGCG therapy of SHR significantly reduced infarct size and improved cardiac function in Langendorff-perfused hearts exposed to ischemia-reperfusion (I/R) injury. In SHR given L-NAME, beneficial effects of EGCG on SBP and I/R were not observed. Both enalapril and EGCG treatment of SHR improved insulin sensitivity and raised plasma adiponectin levels. We conclude that acute actions of EGCG to stimulate production of nitric oxide from endothelium using PI 3-kinase-dependent pathways may explain, in part, beneficial effects of EGCG therapy to simultaneously improve metabolic and cardiovascular pathophysiology in SHR. These findings may be relevant to understanding potential benefits of green tea consumption in patients with the metabolic syndrome.     

High prevalence of drug resistance in animal trypanosomes without a history of drug exposure

Background

Trypanosomosis caused by Trypanosoma congolense is a major constraint to animal health in sub-Saharan Africa. Unfortunately, the treatment of the disease is impaired by the spread of drug resistance. Resistance to diminazene aceturate (DA) in T. congolense is linked to a mutation modifying the functioning of a P2-type purine-transporter responsible for the uptake of the drug. Our objective was to verify if the mutation was linked or not to drug pressure.

Methodology/Principal Findings

Thirty-four T. congolense isolates sampled from tsetse or wildlife were screened for the DA-resistance linked mutation using DpnII-PCR-RFLP. The results showed 1 sensitive, 12 resistant and 21 mixed DpnII-PCR-RFLP profiles. This suggests that the mutation is present on at least one allele of each of the 33 isolates. For twelve of the isolates, a standard screening method in mice was used by (i) microscopic examination, (ii) trypanosome-specific 18S-PCR after 2 months of observation and (iii) weekly trypanosome-specific 18S-PCR for 8 weeks. The results showed that all mice remained microscopically trypanosome-positive after treatment with 5 mg/kg DA. With 10 and 20 mg/kg, 8.3% (n = 72) and 0% (n = 72) of the mice became parasitologically positive after treatment. However, in these latter groups the trypanosome-specific 18S-PCR indicated a higher degree of trypanosome-positivity, i.e., with a unique test, 51.4% (n = 72) and 38.9% (n = 72) and with the weekly tests 79.2% (n = 24) and 66.7% (n = 24) for 10 and 20 mg/kg respectively.

Conclusion/Significance

The widespread presence of the DA-resistance linked mutation in T. congolense isolated from wildlife suggests that this mutation is favourable to parasite survival and/or its dissemination in the host population independent from the presence of drug. After treatment with DA, those T. congolense isolates cause persisting low parasitaemias even after complete elimination of the drug and with little impact on the host''s health.     

Inter-Annual Variability of Fledgling Sex Ratio in King Penguins

As the number of breeding pairs depends on the adult sex ratio in a monogamous species with biparental care, investigating sex-ratio variability in natural populations is essential to understand population dynamics. Using 10 years of data (2000–2009) in a seasonally monogamous seabird, the king penguin (Aptenodytes patagonicus), we investigated the annual sex ratio at fledging, and the potential environmental causes for its variation. Over more than 4000 birds, the annual sex ratio at fledging was highly variable (ranging from 44.4% to 58.3% of males), and on average slightly biased towards males (51.6%). Yearly variation in sex-ratio bias was neither related to density within the colony, nor to global or local oceanographic conditions known to affect both the productivity and accessibility of penguin foraging areas. However, rising sea surface temperature coincided with an increase in fledging sex-ratio variability. Fledging sex ratio was also correlated with difference in body condition between male and female fledglings. When more males were produced in a given year, their body condition was higher (and reciprocally), suggesting that parents might adopt a sex-biased allocation strategy depending on yearly environmental conditions and/or that the effect of environmental parameters on chick condition and survival may be sex-dependent. The initial bias in sex ratio observed at the juvenile stage tended to return to 1∶1 equilibrium upon first breeding attempts, as would be expected from Fisher’s classic theory of offspring sex-ratio variation.     

Insight into the bind-lock mechanism of the yeast mitochondrial ATP synthase inhibitory peptide

The mechanism of yeast mitochondrial F1-ATPase inhibition by its regulatory peptide IF1 was investigated with the noncatalytic sites frozen by pyrophosphate pretreatment that mimics filling by ATP. This allowed for confirmation of the mismatch between catalytic site occupancy and IF1 binding rate without the kinetic restriction due to slow ATP binding to the noncatalytic sites. These data strengthen the previously proposed two-step mechanism, where IF1 loose binding is determined by the catalytic state and IF1 locking is turnover-dependent and competes with IF1 release (Corvest, V., Sigalat, C., Venard, R., Falson, P., Mueller, D. M., and Haraux, F. (2005) J. Biol. Chem. 280, 9927-9936). They also demonstrate that noncatalytic sites, which slightly modulate IF1 access to the enzyme, play a minor role in its binding. It is also shown that loose binding of IF1 to MgADP-loaded F1-ATPase is very slow and that IF1 binding to ATP-hydrolyzing F1-ATPase decreases nucleotide binding severely in the micromolar range and moderately in the submillimolar range. Taken together, these observations suggest an outline of the total inhibition process. During the first catalytic cycle, IF1 loosely binds to a catalytic site with newly bound ATP and is locked when ATP is hydrolyzed at a second site. During the second cycle, blocking of ATP hydrolysis by IF1 inhibits ATP from becoming entrapped on the third site and, at high ATP concentrations, also inhibits ADP release from the second site. This model also provides a clue for understanding why IF1 does not bind ATP synthase during ATP synthesis.     

Regulation of phospholipase D by P2X7 receptors in submandibular ductal cells

ATP (1 mM) increased the phospholipase D (PLD) activity of rat submandibular gland (RSMG) ductal cells in a concentration-dependent and calcium-sensitive manner. The response to ATP was reproduced by benzoylbenzoyl-ATP (Bz-ATP, 100 microM) and also partly by adenosine 5'-(gamma-thio)triphosphate (ATPgammaS, 1 mM). A similar stimulation was observed in control mice (P2X7R+/+ mice) but not in mice lacking the P2X7 receptors (P2X7R-/- mice). Oxidized ATP and Coomassie blue or the addition of magnesium or nickel to the incubation medium inhibited the response to ATP. The stimulation of PLD by purinergic agonist was inhibited by about 50% by calphostin C and chelerythrine, two protein kinase C (PKC) inhibitors. The stimulation of PLD by Bz-ATP and by o-tetradecanoylphorbol 13-acetate (TPA), a phorbol ester which activates PKC, were not additive.From these results we can conclude that the activation of P2X7 receptors in RSMG ductal cells is coupled to the activation of a PLD. This activation is partly mediated by protein kinase C.