Synthesis and structural study of highly constrained hybrid cyclobutane-proline γ,γ-peptides

Two diastereomeric series of hybrid γ,γ-peptides derived from conveniently protected derivatives of (1R,2S)- and (1S,2R)-3-amino-2,2-dimethylcyclobutane-1-carboxylic acid and cis-4-amino-l-proline joined in alternation have efficiently been prepared through convergent synthesis. High-resolution NMR experiments show that these compounds present defined conformations in solution affording very compact structures as the result of intra and inter residue hydrogen-bonded ring formation. (R,S)-cyclobutane containing peptides adopt more twisted conformations than (S,R) diastereomers. In addition, all these γ-peptides have high tendency to aggregation providing vesicles of nanometric size, which were stable when allowed to stand for several days, as verified by transmission electron microscopy.     

Testosterone and year‐round territoriality in tropical and non‐tropical songbirds

Past studies have suggested a fundamental difference in testosterone concentrations between tropical and northern latitude male birds, with the convention being that males in the tropics express much lower levels of testosterone. However, recent comparative studies have shown that tropical males with a short and synchronous breeding season (i.e. a breeding season typical of northern species) express maximum testosterone levels similar to those of northern latitude birds. Here, we ask the converse: do northern latitude songbirds that express a defining life‐history characteristic typical of the tropics, i.e. year‐round territoriality, have an annual testosterone profile similar to that of tropical songbirds? For the few year‐round territorial species for which data are available, we found that seasonal testosterone profiles and seasonal maxima in plasma testosterone were similar between males of tropical and non‐tropical species. For example, males of both groups expressed seasonal maxima during the period when females were fertile, and testosterone levels at this time were similar. In contrast, this and other studies show that species with seasonal territories typically express maximum testosterone levels earlier in the breeding cycle, when territories are first being established. Taken together, we suggest that specific life‐history traits may play a more important role in determining testosterone profiles of tropical and non‐tropical birds than breeding latitude and encourage further studies to allow for more formal comparisons.     

Uncovering genomic causes of co-morbidity in epilepsy: gene-driven phenotypic characterization of rare microdeletions

Background

Patients with epilepsy often suffer from other important conditions. The existence of such co-morbidities is frequently not recognized and their relationship with epilepsy usually remains unexplained.

Methodology/Principal Findings

We describe three patients with common, sporadic, non-syndromic epilepsies in whom large genomic microdeletions were found during a study of genetic susceptibility to epilepsy. We performed detailed gene-driven clinical investigations in each patient. Disruption of the function of genes in the deleted regions can explain co-morbidities in these patients.

Conclusions/Significance

Co-morbidities in patients with epilepsy can be part of a genomic abnormality even in the absence of (known) congenital malformations or intellectual disabilities. Gene-driven phenotype examination can also reveal clinically significant unsuspected condition.     

Growth and indole-3-acetic acid biosynthesis of Azospirillum brasilense Sp245 is environmentally controlled

Batch and fed batch cultures of Azospirillum brasilense Sp245 were conducted in a bioreactor. Growth response, IAA biosynthesis and the expression of the ipdC gene were monitored in relation to the environmental conditions (temperature, availability of a carbon source and aeration). A. brasilense can grow and produce IAA in batch cultures between 20 and 38 degrees C in a standard minimal medium (MMAB) containing 2.5 gl(-1)l-malate and 50 microgml(-1) tryptophan. IAA synthesis requires depletion of the carbon source from the growth medium in batch culture, causing growth arrest. No significant amount of IAA can be detected in a fed batch culture. Varying the concentration of tryptophan in batch experiments has an effect on both growth and IAA synthesis. Finally we confirmed that aerobic growth inhibits IAA synthesis. The obtained profile for IAA synthesis coincides with the expression of the indole-3-pyruvate decarboxylase gene (ipdC), encoding a key enzyme in the IAA biosynthesis of A. brasilense.     

Mitochondrial reactive oxygen species generation triggers inflammatory response and tissue injury associated with hepatic ischemia-reperfusion: Therapeutic potential of mitochondrially targeted antioxidants

Mitochondrial reactive oxygen species generation has been implicated in the pathophysiology of ischemia-reperfusion (I/R) injury; however, its exact role and its spatial-temporal relationship with inflammation are elusive. Herein we explore the spatial-temporal relationship of oxidative/nitrative stress and inflammatory response during the course of hepatic I/R and the possible therapeutic potential of mitochondrial-targeted antioxidants, using a mouse model of segmental hepatic ischemia-reperfusion injury. Hepatic I/R was characterized by early (at 2h of reperfusion) mitochondrial injury, decreased complex I activity, increased oxidant generation in the liver or liver mitochondria, and profound hepatocellular injury/dysfunction with acute proinflammatory response (TNF-α, MIP-1α/CCL3, MIP-2/CXCL2) without inflammatory cell infiltration, followed by marked neutrophil infiltration and a more pronounced secondary wave of oxidative/nitrative stress in the liver (starting from 6h of reperfusion and peaking at 24h). Mitochondrially targeted antioxidants, MitoQ or Mito-CP, dose-dependently attenuated I/R-induced liver dysfunction, the early and delayed oxidative and nitrative stress response (HNE/carbonyl adducts, malondialdehyde, 8-OHdG, and 3-nitrotyrosine formation), and mitochondrial and histopathological injury/dysfunction, as well as delayed inflammatory cell infiltration and cell death. Mitochondrially generated oxidants play a central role in triggering the deleterious cascade of events associated with hepatic I/R, which may be targeted by novel antioxidants for therapeutic advantage.     

Size of the pores created by an electric pulse: Microsecond vs millisecond pulses

Here, the sizes of the pores created by square-wave electric pulses with the duration of 100μs and 2ms are compared for pulses with the amplitudes close to the threshold of electroporation. Experiments were carried out with three types of cells: mouse hepatoma MH-22A cells, Chinese hamster ovary (CHO) cells, and human erythrocytes. In the case of a short pulse (square-wave with the duration of 100μs or exponential with the time constant of 22μs), in the large portion (30-60%) of electroporated (permeable to potassium ions) cells, an electric pulse created only the pores, which were smaller than the molecule of bleomycin (molecular mass of 1450Da, r≈0.8nm) or sucrose (molecular mass of 342.3Da, radius-0.44-0.52nm). In the case of a long 2-ms duration pulse, in almost all cells, which were electroporated, there were the pores larger than the molecules of bleomycin and/or sucrose. Kinetics of pore resealing depended on the pulse duration and was faster after the shorter pulse. After a short 100-μs duration pulse, the disappearance of the pores permeable to bleomycin was completed after 6-7min at 24-26°C, while after a long 2-ms duration pulse, this process was slower and lasted 15-20min. Thus, it can be concluded that a short 100-μs duration pulse created smaller pores than the longer 2-ms duration pulse. This could be attributed to the time inadequacy for pores to grow and expand during the pulse, in the case of short pulses.     

Oxidative effects of nanosecond pulsed electric field exposure in cells and cell-free media

Nanosecond pulsed electric field (nsPEF) is a novel modality for permeabilization of membranous structures and intracellular delivery of xenobiotics. We hypothesized that oxidative effects of nsPEF could be a separate primary mechanism responsible for bioeffects. ROS production in cultured cells and media exposed to 300-ns PEF (1-13kV/cm) was assessed by oxidation of 2',7'-dichlorodihydrofluoresein (H(2)DCF), dihidroethidium (DHE), or Amplex Red. When a suspension of H(2)DCF-loaded cells was subjected to nsPEF, the yield of fluorescent 2',7'-dichlorofluorescein (DCF) increased proportionally to the pulse number and cell density. DCF emission increased with time after exposure in nsPEF-sensitive Jurkat cells, but remained stable in nsPEF-resistant U937 cells. In cell-free media, nsPEF facilitated the conversion of H(2)DCF into DCF. This effect was not related to heating and was reduced by catalase, but not by mannitol or superoxide dismutase. Formation of H(2)O(2) in nsPEF-treated media was confirmed by increased oxidation of Amplex Red. ROS increase within individual cells exposed to nsPEF was visualized by oxidation of DHE. We conclude that nsPEF can generate both extracellular (electrochemical) and intracellular ROS, including H(2)O(2) and possibly other species. Therefore, bioeffects of nsPEF are not limited to electropermeabilization; concurrent ROS formation may lead to cell stimulation and/or oxidative cell damage.     

The impacts of heavy metals on oxidative stress and growth of spring barley

Oxidative stress is accepted to play a significant role in stress symptoms, caused by different stressors in a variety of organisms. In this study seedlings of spring barley (Hordeum vulgare L.) were exposed to a wide range of copper, zinc, chromium, nickel, lead and cadmium concentrations in order to determine the relationships between heavy metals-induced oxidative stress and plant growth inhibition. All investigated heavy metals induced an essential increase in lipid peroxidation and a reduction of dry biomass along with an increase in metal concentration in the nutrient solution. A very close and statistically significant exponential relationship between lipid peroxidation and growth inhibition was detected in this study. According to the results of analysis of variance (ANOVA), the intensity of nonspecific oxidative stress is identified as the main factor of barley growth inhibition, explaining 75% of total variance. Almost 10% of growth inhibition is attributed to the specific impact of heavy metals. The most pronounced increase of malondialdehyde content and growth inhibition was observed in Cu and Cd treatments, whereas the lowest changes in observed indicators were detected after exposure to Zn and Pb.