Novel antihypertensive hexa- and heptapeptides with ACE-inhibiting properties: from the in vitro ACE assay to the spontaneously hypertensive rat

Bioactive ACE inhibiting peptides are gaining interest in hypertension treatment. We have designed and screened six synthetic heptapeptides (PACEI48 to PACEI53) based on two hexapeptide leads (PACEI32 and PACEI34) to improve ACE inhibitory properties and assess their antihypertensive effects. ACE activity was assayed in vitro and ex vivo. Selected peptides were administered to spontaneously hypertensive rats (SHRs) and normotensive Wistar-Kyoto (WKY) rats. In vitro cytotoxicity was assessed with the MTT reduction test. The six heptapeptides at low micromolar concentration produced different degrees of in vitro inhibition of ACE activity using the synthetic substrate HHL or the natural substrate angiotensin I; and ex vivo inhibition of ACE-dependent, angiotensin I-induced vasoconstriction, but not angiotensin II-induced vasoconstriction. Oral administration of the hexapeptide PACEI32L, and the heptapeptides PACEI50L and PACEI52L, induced reductions in systolic blood pressure lasting up to 3 h in SHRs but not in WKY rats. Intravenous injection of PACEI32L and PACEI50L, but not PACEI52L, induced acute transient reductions in mean blood pressure of SHRs. d-Amino acid peptides showed five-fold less ACE inhibitory potency, no inhibitory effect on angiotensin I-induced vasoconstriction, and antihypertensive effect in SHRs after i.v. injection, but not after oral administration. The toxicity of peptides to reduce the viability of cultured cells was in the millimolar range. In conclusion, we have obtained novel rationally designed heptapeptides with improved ACE inhibitory properties when compared to lead hexapeptides. One selected hexapeptide and two heptapeptides show oral antihypertensive effects in SHRs and appear safe in cytotoxicity assays.     

Daily profile in ghrelin and hypothalamic ghrelin receptors in obese and lean mice Neotomodon alstoni

Mice Neotomodon alstoni develops, in some individuals, overweight and obesity when kept in vivarium conditions and fed on standard rodent diet. In the present study, we explored the possible differences between lean and obese mice, on the daily expression of the hormone ghrelin, its total and active form, as well as the relative expression of ghrelin receptors (GSHR) in hypothalamic tissue. Plasma hormones were detected and quantified by immunological techniques by ELISA; the relative presence of GSHR was assessed by western blot. Mice were sacrificed at different times throughout the day. The results obtained indicate that there is a rhythm of presence of total and acylated ghrelin in lean mice, which dampers and changes its peak’s phase in obese mice. The GHSR expression tend to be more abundant in obese mice than in lean mice. The results indicate that, in the obese mice there is a possible resistance to ghrelin.     

Quantitative determination of the protein content of citrus leaf extracts: A comparative study

Three spectrophotometric methods for the quantitative determination of proteins were compared taking as a reference the method of Kjeldahl. The abnormally high protein values obtained when these methods were applied to soluble extracts were decreased by using protective and precipitating agents. The method of M. M. Bradford (Anal. Biochem. 72, 248–254 (1976)) gave values identical to the method of Kjeldahl when ethylenediaminetetracetic acid, 2-mercaptoethanol, and polyvinylpolypyrrolidone were added to the extraction medium and the soluble proteins were precipitated with trichloroacetic acid prior to protein determination.     

A Bayesian analysis of the effect of selection for growth rate on growth curves in rabbits

Gompertz growth curves were fitted to the data of 137 rabbits from control (C) and selected (S) lines. The animals came from a synthetic rabbit line selected for an increased growth rate. The embryos from generations 3 and 4 were frozen and thawed to be contemporary of rabbits born in generation 10. Group C was the offspring of generations 3 and 4, and group S was the contemporary offspring of generation 10. The animals were weighed individually twice a week during the first four weeks of life, and once a week thereafter, until 20 weeks of age. Subsequently, the males were weighed weekly until 40 weeks of age. The random samples of the posterior distributions of the growth curve parameters were drawn by using Markov Chain Monte Carlo (MCMC) methods. As a consequence of selection, the selected animals were heavier than the C animals throughout the entire growth curve. Adult body weight, estimated as a parameter of the Gompertz curve, was 7% higher in the selected line. The other parameters of the Gompertz curve were scarcely affected by selection. When selected and control growth curves are represented in a metabolic scale, all differences disappear.     

Concurrent Akt,ERK1/2 and AMPK Activation by Obestatin Inhibits Apoptotic Signaling Cascades on Nutrient-Deprived PC12 Cells

Targeting apoptosis in the ischemic penumbra is a rational therapeutic approach for restricting cerebral infarct volume after clinical stroke. The present work explored the capability of the obestatin peptide, as a novel approach to inhibit apoptotic signaling cascades on PC12 cells. According to the results, obestatin treatment significantly reduced nutrient deprivation-induced apoptotic cell death. The protective effects were related to the regulation of the anti-apoptotic protein, BCL-2, and the apoptotic protein caspase-3. This encompasses the control of apoptosis by the interplay between Akt, ERK1/2 and AMPK signaling pathways. The activation of Akt and AMPK was concomitant with the phosphorylation of their downstream targets, GSK3 and ACC, respectively. Besides, obestatin also causes FoxO1 nuclear export supporting the prevention of the apoptosome formation. The concurrent activation of Akt and AMPK by obestatin via the GPR39 receptor, supports a role for this system in the balance concerning the catabolic and the anabolic signaling to sustain cellular function and viability. Furthermore, these results provide both an insight into how the obestatin/GPR39 system regulates anti-apoptotic pathways, and a framework for ascertaining how this system can be optimally targeted in treatment of brain cell death after stroke.

     

Generation of Surface Plasmons at Waveguide Surfaces in the Mid-Infrared Region

A technique is proposed to extend the application of surface-plasmon-based spectroscopy into the mid-infrared spectral regime, which is of substantial interest in the field of chemical analysis and biosensing. Surface plasmons can be excited for wavelengths of the order of 6???m at corrugated waveguides for a given combination of materials and thicknesses, and for refractive indices of the surrounding medium corresponding to those of organic solvents. This approach can easily be extrapolated to other values of any of these parameters. Based on these considerations, a new generation of mid-IR SPR sensors can be developed with a diverse range of potential applications in chem/bio sensing.