Keratins and epidermolysis bullosa simplex

Keratin intermediate filaments play an important role in maintaining the integrity of the skin structure. Understanding the importance of this subject is possible with the investigation of keratin defects in epidermolysis bullosa simplex (EBS). Nowadays, in addition to clinical criteria, new molecular diagnostic methods, such as next generation sequencing, can help to distinguish the subgroups of EBS more precisely. Because the most important and most commonly occurring molecular defects in these patients are the defects of keratins 5 and14 (KRT5 and KRT14), comprehending the nature structure of these proteins and their involved processes can be very effective in understanding the pathophysiology of this disease and providing new and effective therapeutic platforms to treat it. Here, we summarized the various aspects of the presence of KRT5 and KRT14 in the epidermis, their relation to the incidence and severity of EBS phenotypes, and the processes with which these proteins can affect them.     

Identification and natural functions of cyclic lipopeptides from Bacillus amyloliquefaciens An6

Lipopeptides constitute a structurally diverse group of metabolites produced by various bacterial and fungal genera. In the past decades, research on lipopeptides has been fueled by their surfactant activities. However, natural functions of lipopeptides compounds have received considerably less attention. The aim of this study was to isolate and identify the lipopeptides from Bacillus amyloliquefaciens An6, and further evaluate their biological activities. An6 lipopeptides were detected by PCR using degenerated primers and MALDI‐TOF‐MS. An6 strain was found to produce surfactin, fengycin, and bacillomycin. Following their purification, the in vitro antioxidant activity of An6 lipopeptides was studied through different assays. The scavenging effect on 1,1‐diphenyl‐2‐picrylhydrazyl radicals at a dosage of 0.75 mg/mL was 81%. Its reducing power was concentration‐dependant and reached a maximum of 1.07 at 2.5 mg/mL. Moreover, they showed a strong inhibition of β‐carotene bleaching. An6 lipopeptides mixture was also found to display significant antimicrobial activity against several Gram‐positive, Gram‐negative bacteria, and fungal strains. An6 lipopeptides were insensitive to proteolytic enzymes, stable between pH 4.0 and 12.0, and resistant to high temperature. Our results provided enough evidence proving that An6 lipopeptides could be used as functional‐food components.     

Preparation and testing of Sardinella peptones: application to lipase production by Staphylococcus sp

Production of lipase by Staphylococcus sp. in media containing fish peptones from sardinelle (Sardinella aurita) prepared in the laboratory was studied. Lipase production is strongly affected by lipids present in fish flours. Fish peptones prepared from dIgresed whole flesh was an excellent substrate for lipase production. A comparison of lipase production in media containing fish peptones or high quality commercial peptones indicated that fish peptones enhanced enzyme formation.     

Inflammatory process of CD8+ CD28- T cells in induced sputum from asthmatic patients

Previously unreported CD8(+) CD28(-) and CD8(+) CD28(+) T-cell subsets occur in healthy individuals and expand in patients suffering from autoimmune disease. Here we studied, for the first time, the expression of CD8(+) CD28(+) , CD8(+) CD28(-) , and CD8(+) CD56(+) subpopulations in induced sputum from asthmatics. Using sputum samples, purified CD8(+) T cells were stained for surface antigen CD28, CD56, FITC-conjugated anti-perforin, and anti-IFN-gamma. Cytotoxic activity was evaluated in a chromium releasing test. Induced sputum CD8(+) CD28(-) T cells were found to be more expanded and expressed low levels of IFN-gamma in severe asthmatics than mild asthma and age-matched healthy controls. The predominance of CD8(+) CD28(-) T cells can be in part explained by the expansion of CD8(+) CD56(+). CD8(+) CD28(-) T cells from severe asthmatics produced high intracytoplasmic perforin and exerted a potent cytotoxic activity. Considering their phenotyping and functional properties, the CD8(+) CD28(-) T-cell subset may constitute an intermediate phenotype in the process of CD8(+) T-cell differentiation of effector-type cells in severe asthmatics. Functional studies showed that CD8(+) CD28(-) T cells had cytotoxic function.     

Assessment of salt tolerance of Nasturtium officinale R. Br. using physiological and biochemical parameters

Nasturtium officinale R. Br. seedlings were treated with a range of NaCl concentrations (0, 50, 100 and 150 mM) for 21 days after seedling emergence. Physiological analysis based on growth and mineral nutrition, showed a substantial decrease in leaf dry matter with 150 mM NaCl treatment. The growth decrease was correlated with nutritional imbalance and a reduction in potassium accumulation and transport to the leaves. At the same time, we noted an increase in leaf sodium and chloride accumulation and transport. Salt tolerance of N. officinale under 100 mM NaCl was associated with osmotic adjustment via Na⁺ and Cl^? and the maintenance of high K⁺/Na⁺ selectivity. Salt decreased carotenoid content more than chlorophylls and also disturbed membrane integrity by increasing malondialdehyde content and electrolyte leakage. At 150 mM NaCl, an increase in antioxidant enzyme-specific activities for superoxide dismutase, catalase and guaiacol peroxidase occurred in concert with a decrease in ascorbic acid, polyphenol, tannin and flavonoid content. These results indicate that N. officinale can maintain growth and natural antioxidant defense compounds such as, vitamin C, carotenoids, and polyphenols, when cultivated in 100 mM NaCl, but not at higher salt levels.     

Response-surface methodology for the production and the purification of a new H2O2-tolerant alkaline protease from Bacillus invictae AH1 strain

This work deals with the optimization of the culture conditions of Bacillus invictae AH1 in order to increase the production level of the proteolytic activity. Response-surface methodology (RSM) was applied for the most significant fermentation parameters (concentration of wheat bran and K₂HPO₄/KH₂PO₄) that were earlier identified by Plackett–Burman Design from seven possible factors. A central composite design was used and the quadratic regression model of producing active protease was built. A maximum protease activity was reached and validated experimentally, using a maximum wheat bran concentration (50 g/L) with increased K₂HPO₄/KH₂PO₄ concentration (2.275 g/L). Protease production obtained experimentally coincident with the predicted value and the model was proven to be adequate. Interestingly, the use of RSM increased the protease production by four times (7,000 U/mL) using a low-cost substrate and a culture time of 40 hr, as compared to the standard culture conditions. In the second part of this study, a H₂O₂-tolerant alkaline protease produced from B. invictae AH1 with a molecular mass of about 41 kDa, noted P3, was purified by successive steps of ultrafiltration, gel filtration and ion exchange chromatography. The K _m and V_max values of the purified protease using casein, as substrate, were about 4 mg/mL and 27 μM/min, respectively. The highest enzyme activity was found at pH 9.0 and a temperature of 60°C. In addition, the enzyme showed a quasi-total stability against H₂O₂ (5% for 1 hr) and against most of the tested solid and liquid detergents, suggesting its eventual use in bio-detergent formulations.     

Impact of specific training and competition on myocardial structure and function in different age ranges of male handball players

Handball activity involves cardiac changes and demands a mixture of both eccentric and concentric remodeling within the heart. This study seeks to explore heart performance and cardiac remodeling likely to define cardiac parameters which influence specific performance in male handball players across different age ranges. Forty three players, with a regular training and competitive background in handball separated into three groups aged on average 11.78±0.41 for youth players aka “schools”, “elite juniors” 15.99±0.81 and “elite adults” 24.46±2.63 years, underwent echocardiography and ECG examinations. Incremental ergocycle and specific field (SFT) tests have also been conducted. With age and regular training and competition, myocardial remodeling in different age ranges exhibit significant differences in dilatation’s parameters between “schools” and “juniors” players, such as the end-diastolic diameter (LVEDD) and the end-systolic diameter of the left ventricle (LVESD), the root of aorta (Ao) and left atrial (LA), while significant increase is observed between “juniors” and “adults” players in the interventricular septum (IVS), the posterior wall thicknesses (PWT) and LV mass index. ECG changes are also noted but NS differences were observed in studied parameters. For incremental maximal test, players demonstrate a significant increase in duration and total work between “schools” and “juniors” and, in total work only, between “juniors” and “seniors”. The SFT shows improvement in performance which ranged between 26.17±1.83 sec to 31.23±2.34 sec respectively from “seniors” to “schools”. The cross-sectional approach used to compare groups with prior hypothesis that there would be differences in exercise performance and cardiac parameters depending on duration of prior handball practice, leads to point out the early cardiac remodeling within the heart as adaptive change. Prevalence of cardiac chamber dilation with less hypertrophy remodeling was found from “schools” to “juniors” while a prevalence of cardiac hypertrophy with less pronounced chamber dilation remodeling was noted later.     

A proteasome-dependent, TAP-independent pathway for cross-presentation of phagocytosed antigen

Major histocompatibility complex (MHC) class I cross-presentation is thought to involve two pathways, one of which depends on both the TAP transporters and the proteasome and the other on neither. We found that preincubation of TAP-deficient dendritic cells at low temperature increases the density of MHC class I at the surface and fully restores cross-presentation of phagocytosed antigen, but not of soluble antigen internalized through receptors. Restoration of cross-presentation by TAP-deficient cells requires antigen degradation by the proteasome. Thus, TAP might mainly be required for recycling cell surface class I molecules during cross-presentation of phagocytosed antigens. Furthermore, phagosomes-but not endosomes-seem to have a TAP-independent mechanism to import peptides generated by cytosolic proteasome complexes.     

Influence of oxygen supply on the stability of recombinant plasmid pTG201 in immobilized E. coli cells

Summary Cells of Escherichia coli K12, carrying the recombinant plasmid pTG201, were immobilized in -carrageenan gel in order to improve the following plasmid parameters: (i) maintenance of a high level of plasmid copy number, (ii) good plasmid stability and (iii) good expression of plasmid encoded gene. The experiments were carried out on LB medium without antibiotic selection in continuous and batch cultures supplied with air or pure oxygen. Parallel experiments with free cells were also performed. In all the cases immobilized cells presented better plasmid stability parameters than free cells. Best results were obtained with immobilized cells supplied with pure oxygen. In this case, an average plasmid copy number of 60 and a value of plasmid-carrying cells close to 100% were maintained with little change during more than 200 generations. In addition, an optical microscopy analysis is proposed to allow the quantitation of cell growth in gel beads.