Increased Oxidative Stress and Altered Levels of Nitric Oxide and Peroxynitrite in Tunisian Patients with Chronic Obstructive Pulmonary Disease: Correlation with Disease Severity and Airflow Obstruction

This study was aimed to evaluate the oxidant–antioxidant imbalance in the pathogenesis of chronic obstructive pulmonary disease (COPD) in Tunisians. We assessed 16 parameters related to the oxidative status that include malondialdehyde (MDA), total protein carbonyls (PCs), and advanced oxidation protein products (AOPP). We also examined the activity of glutathione peroxydase (GSH-Px), catalase, and superoxide dismutase (SOD) in the plasma and erythrocytes. Levels of total thiols, reduced glutathione (GSH), total antioxidant status (TAS), hydrogen peroxide, ascorbic acid, iron, and protein sulfhydryls were determined using spectrophotometry. We also evaluated the level of nitric oxide (NO) and peroxynitrite in plasma from COPD patients and healthy controls. Estimation of DNA damage was determined using the comet assay. Pulmonary functional tests were performed by body plethysmography. Levels of MDA, PC, DNA damage, and AOPP were significantly increased while total thiols, GSH, and TAS were decreased in COPD patients. GSH-Px activity was higher in COPD patients while no difference was found for catalase and SOD. We also observed a lower level of NO and peroxynitrite in COPD patients. Decreased levels of peroxynitrite were found to correlate with disease progression, as well as with forced expiratory volume in 1 s/forced vital capacity among COPD patients. Multivariate analysis revealed that NO is associated with pathological pathways that help to predict patient outcome independently of the degree of airflow obstruction. These results indicate the presence of a systemic oxidative stress and highlight the importance of NO and peroxynitrite as major effectors in COPD development and airflow obstruction.     

Binding of thyroid hormones to human hemoglobin and localization of the binding site

Radiolabeled thyroid hormones were allowed to bind to erythrocyte cytosol and the complex was fractionated by Sephadex G-100 or by high-performance liquid chromatography (HPLC). On Sephadex G-100, four radioactive peaks (P₁P₄) were obtained, whereas HPLC gave only three radioactive peaks (P₁P₃). Chromatographic studies with human adult Hb and non-Hb cytosol protein fractions, which had been reacted with radiolabeled thyroid hormones, and immune precipitation with specific antisera for the hormones, confirmed that the first peak of Sephadex G-100 radioactivity was a mixture of Hb and non-Hb proteins, while the second peak was Hb. The third peak was free¹²⁵I and the fourth peak was unbound¹²⁵I-T₃ or¹²⁵I-T₄. The third peak of HPLC was confirmed to be a mixture of free¹²⁵I and unbound radiolabeled thyroid hormones. Scatchard analysis of the interaction between T₄ and apo-Hb, and the - and -chains of human Hb suggested the presence of the specific binding site(s) for the hormone. Interaction between T₄ and synthesized peptides, which constitute the heme pocket of the -chain of Hb (61–75, 71–85, 81–95), indicated that the T₄ binding site of Hb resides within the heme-binding cavity. It is concluded that human erythrocyte cytosol does not contain receptor for thyroid hormones and cannot be a model for studying functions of cytosol receptor for the hormones; rather, it contains binding protein with large binding capacity, including Hb and non-Hb proteins, which possibly constitute a large reservoir for the hormone in blood.     

Effect of Time of Day and Partial Sleep Deprivation on Short‐Term,High‐Power Output

The purpose of this study was to determine whether delaying bedtime or advancing rising time by 4 h affects anaerobic performance of individuals the following day in the morning and afternoon. Eleven subjects participated in the study, during which we measured the maximal, peak, and mean powers (i.e., P_max [force‐velocity test], P_peak, and P_mean [Wingate test], respectively). Measurements were performed twice daily, at 07∶00 and 18∶00 h, following a reference normal sleep night (RN), a partial sleep deprivation timed at the beginning of the night (SDB), and a partial sleep deprivation timed at the end of the night (SDE), and oral temperature was measured every 4 h. Each of the three experimental conditions was separated by a one‐week period. Our results showed a circadian rhythm in oral temperature, and analysis of variance revealed a significant sleep×test‐time effect on peak power (P_peak), mean power (P_mean), and maximal power (P_max). These variables improved significantly from the morning to the afternoon for all three experimental conditions. Whereas the morning‐afternoon improvement in the measures was similar after the RN and SDB conditions, it was smaller following the SDE condition. There was no significant difference in the effect of the two sleep‐deprivation conditions on anaerobic performances at 07∶00 and at 18∶00 h under the SDB condition in comparison with the post‐reference night. However, the performance variables were significantly lower at 18∶00 h after the SDE condition. In conclusion, a 4 h partial sleep deprivation at the end of the night appears to be more disturbing than partial sleep deprivation at the beginning of the night.     

Effects of amino acid substitutions outside an antigenic site on protein binding to monoclonal antibodies of predetermined specificity obtained by peptide immunization: demonstration with region 15-22 (antigenic site 1) of myoglobin.

Monoclonal antibodies (mAbs) of predetermined specificity were prepared by immunizing with a free (i.e., not conjugated to any carrier) synthetic peptide representing region 15–22 (site 1) of sperm whale myoglobin (SpMb). The cross-reactions of Mb variants with three mAbs were studied in order to determine whether such interactions are influenced by substitutions outsde the site. Finback whale Mb, which has no substitutions within region 15–22, showed lower cross-reactivity and relative binding affinity than the reference antigen, SpMb. Bottle-nose Atlantic dolphin myoglobin (BdMb) and badger myoglobin (BgMb), although they have identical substitutions within region 15–22 (Ala-15 to Gly and Val-21 to Leu), showed very different binding properties. The cross-reaction of BdMb was quite comparable to that of SpMb, while that of BgMb was much lower. Since the two proteins have identical structures in regions 15–22, the differences in their cross-reactivities are readily attributed to the effects of substitutions outside this region. Another pair of myoglobins, horse myoglobins (HsMb) and chicken myoglobin (ChMb), also have two identical substitutions (Ala-15 to Gly and Val-21 to Ile) within region 15–22, but possessed different cross-reactivity. The results indicate that the reaction of mAbs, whose specificity is precisely known and predetermined by the immunizing free peptide, can be markedly affected by substitutions outside the indicated binding region on the protein.     

Effects of 8-week in-season upper and lower limb heavy resistance training on the peak power, throwing velocity, and sprint performance of elite male handball players

The aims of this study were to test the potential of in-season heavy upper and lower limb strength training to enhance peak power output (Wpeak), vertical jump, and handball related field performance in elite male handball players who were apparently already well trained, and to assess any adverse effects on sprint velocity. Twenty-four competitors were divided randomly between a heavy resistance (HR) group (age 20 ± 0.7 years) and a control group (C; age 20 ± 0.1 years). Resistance training sessions were performed twice a week for 8 weeks. Performance was assessed before and after conditioning. Peak power (W(peak)) was determined by cycle ergometer; vertical squat jump (SJ) and countermovement jump (CMJ); video analyses assessed velocities during the first step (V(1S)), the first 5 m (V(5m)), and between 25 and 30 m (V(peak)) of a 30-m sprint. Upper limb bench press and pull-over exercises and lower limb back half squats were performed to 1-repetition maximum (1RM). Upper limb, leg, and thigh muscle volumes and mean thigh cross-sectional area (CSA) were assessed by anthropometry. W(peak) (W) for both limbs (p < 0.001), vertical jump height (p < 0.01 for both SJ and CMJ), 1RM (p < 0.001 for both upper and lower limbs) and sprint velocities (p < 0.01 for V(1S) and V(5m); p < 0.001 for V(peak)) improved in the HR group. Upper body, leg, and thigh muscle volumes and thigh CSA also increased significantly after strength training. We conclude that in-season biweekly heavy back half-squat, pull-over, and bench-press exercises can be commended to elite male handball players as improving many measures of handball-related performance without adverse effects upon speed of movement.     

Chemical Composition and Bioactivities of Quercus canariensis Flour Acorns Extracts Growing in Tunisia

The current study was conducted to investigate the chemical composition of Quercus canariensis flour acorns extracts as well as its biological activities in regards to the growing area using spectrophotometric and chromatographic techniques. The phenolic profile was composed of 19 compounds identified through HPLC-DAD analysis. Coumarin was the most abundant compound quantified in BniMtir, Nefza and ElGhorra and gallic (12.58–20.52 %), syringic (4.70–7.64 %) and trans-ferulic (2.28–2.94 %) acids were the abundant phenolic acids while kaempferol was the major flavonoid compounds quantified only in Quercus canariensis growing in BniMtir. On the other hand, Ain Snoussi acorn extract was characterized by its high content in luteolin-7-O-glucoside (58.46 %). The in-vitro antioxidant activities of the studied extracts were investigated and the results showed that Nefza ethanolic extract's has the highest activities. A bactericidal effect against Staphylococcus aureus was observed only by Elghorra population. On the other hand, Ain Snoussi acorn extract was efficient to inhibit growth of pathogenic bacteria, mentioned the highest activity against Escherichia coli. These results is the first study highlighted that zeen oak acorns are an excellent source of natural antioxidants and antibacterial compounds related to their lysozyme activity which could be exploited in the pharmaceutical and food sectors.     

Optimization of the expression of the HIV fusion inhibitor cyanovirin-N from the tobacco plastid genome

Plants with transgenic plastid (chloroplast) genomes represent a promising production platform in molecular farming, mainly because of the plastids' potential to accumulate foreign proteins to very high levels and the increased biosafety conferred by the maternal mode of plastid inheritance. Although some transgenes can be expressed to extraordinarily high levels, the expression of others has been unsuccessful. Lack of detectable transgene expression is usually attributable to either RNA instability or protein instability. Here, we have investigated the possibilities to improve the production of a pharmaceutical protein that is difficult to express in chloroplasts: the HIV-1 fusion inhibitor cyanovirin-N (CV-N). Testing various N-terminal and C-terminal fusions to peptide sequences from two proteins known to accumulate to high levels in transgenic plastids (GFP and the protein antibiotic PlyGBS), we show that both low mRNA stability and low protein stability contribute to the lack of detectable CV-N expression in chloroplasts. Both problems can be alleviated by N-terminal fusions to the CV-N coding region, thus highlighting a suitable strategy for optimization of plastid transgene expression.     

A novel peptide mimicking the interaction of α-neurotoxins with acetylcholine receptor

A peptide corresponding to residues 26–41 of α-bungarotoxin, and closed by a disulfide bond between two cysteine residues at the amino and C terminal ends of the peptide, was synthesized and the monomeric form was purified. The peptide, which represents the exposed part of the long central loop of the toxin molecule, was examined for binding to acetylcholine receptor. The peptide was shown by radiometric titrations to bind radiolabeled receptor, and radiolabeled peptide was bound by receptor. The specificity of the binding was confirmed by inhibition with the parent toxin. A synthetic analog of the peptide in which Trp-28 was replaced by glycine had very little (10%) of the original activity. Succinylation of the amino groups of the peptide resulted in virtually complete (98%) loss of the binding activity. These results indicate that a shortened loop peptide corresponding to the region 26–41 of α-bungarotoxin exhibits binding activities mimicking those of the parent molecule. In this region, Trp-28, and one or both of Lys-26 and Lys-38, are essential contact residues in the binding to receptor.