Risk Factors for Normal and High-Tension Glaucoma in Poland in Connection with Polymorphisms of the Endothelial Nitric Oxide Synthase Gene

Aim

The purpose of this study was to evaluate the influence of polymorphisms of the eNOS gene on the clinical status of patients with normal and high tension glaucoma.

Methods

266 Polish Caucasian patients with primary open angle glaucoma were studied. Of the 266, 156 had normal tension glaucoma (NTG) and 110 high tension glaucoma (HTG). DNA material was isolated from peripheral venous blood using commercial kits. Real-time PCR reaction was used to amplify the promoter site of the endothelial nitric oxide synthase (eNOS) gene, including the single nucleotide polymorphism (SNP) site T-786C and part of the 7^th exon of eNOS, including G894T SNP. Genotypes were determined with TaqMan SNP Genotyping Assays.

Results

There were no significant differences in frequencies of the allelic variants of both polymorphisms. In G894T SNP, however, the wild GG form was more common in the HTG group. The SNP of the eNOS gene did not significantly influence the progression rate in either of the groups studied. There were no differences in variants of the eNOS gene regarding the necessity for and success of surgery and the progression of the disease. In the NTG group, no statistical correlation was observed between G894T, T786C polymorphism variants, and risk factors such as optic disc haemorrhages, optic disc notches, and peripapillary atrophy. Mean diastolic and systolic pressure during the day and night were lowest in NTG patients with the CC variant of the T786C polymorphism. No statistical correlation was observed between the G894T and T786C polymorphisms and capillaroscopic examination results.

Conclusions

Genotype frequencies are similar for both the eNOS G894T and T-786C polymorphisms in NTG and HTG patients. These polymorphisms do not correlate with risk factors and do not influence the state of the capillary system in NTG patients. Systolic blood pressure is lower in NTG patients with mutated alleles of both polymorphisms.     

A click chemistry approach to glycomimetics: Michael addition of 2,3,4,6-tetra-O-acetyl-1-thio-beta-D-glucopyranose to 4-deoxy-1,2-O-isopropylidene-L-glycero-pent-4-enopyranos-3-ulose--a convenient route to novel 4-deoxy-(1-->5)-5-C-thiodisaccharides

The base catalyzed conjugate Michael addition of the 1-thiosugar, 2,3,4,6-tetra-O-acetyl-beta-D-glucopyranose, 1, to a new highly reactive enone 4-deoxy-1,2-O-isopropylidene-L-glycero-pent-4-enopyranos-3-ulose, 2, proceeds steroselectively with formation of adduct 3 in 94% yield. Convenient stereoselective reduction of the C-3 keto function of 3 with L-Selectride followed by in situ acetylation produces thiodisaccharide 4 in good 82% yield. Cleavage of the 1,2-O-isopropylidene protecting group with p-toluenesulfonic acid in methanol, followed by de-O-acetylation, produced an inseparable anomeric mixture of methyl 4-deoxy-5-C-(beta-D-glucopyranosyl)-thio-alpha/beta-L-ribo-pyranoside 5 in 72% overall yield. This approach constitutes a new general two-step click chemistry route to the previously unknown class of 4-deoxy-(1-->5)-5-C-thiodisaccharides as stable and biologically important glycomimetics.     

Phosphodiesterase 4D and protein kinase a type II constitute a signaling unit in the centrosomal area

The mediation of cAMP effects by specific pools of protein kinase A (PKA) targeted to distinct subcellular domains raises the question of how inactivation of the cAMP signal is achieved locally and whether similar targeting of phosphodiesterases (PDEs) to sites of cAMP/PKA action could be observed. Here, we demonstrate that Sertoli cells of the testis contain an insoluble PDE4D3 isoform, which is shown by immunofluorescence to target to centrosomes. Staining of PDE4D and PKA shows co-localization of PDE4D with PKA-RIIalpha and RIIbeta in the centrosomal region. Co-precipitation of RII subunits and PDE4D3 from cytoskeletal extracts indicates a physical association of the two proteins. Distribution of PDE4D overlaps with that of the centrosomal PKA-anchoring protein, AKAP450, and AKAP450, PDE4D3, and PKA-RIIalpha co-immunoprecipitate. Finally, both PDE4D3 and PKA co-precipitate with a soluble fragment of AKAP450 encompassing amino acids 1710 to 2872 when co-expressed in 293T cells. Thus, a centrosomal complex that includes PDE4D and PKA constitutes a novel signaling unit that may provide accurate spatio-temporal modulation of cAMP signals.     

The induction of oxidative stress in cervix carcinoma cells by levoglucosenone derived 4-S-salicyl derivative and (1–4)-S-thio-disaccharides. Part 4

(1–4)-S-thiodisaccharides were shown to kill various cancer cell lines, including cervix, lung, mammary-gland and colon by unknown mechanisms. Here we identified two actions of levoglucosenone derived (1–4)-S-thiodisaccharides against cervix cancer cells: induction of oxidative stress and DNA damage. In consequence, (1–4)-S-thiodisaccharides lowered the cellular GSH level and changed the expression profile of genes encoding key proteins involved with oxidative stress response. We also observed that (1–4)-S-thiodisaccharides induced DNA damage and interfered with the thioredoxin (Trx) system. Both actions, as induced by FPC6, were stronger when dihedral angles of sulfur bridge were set to 110°, 100° and 109°, clearly indicating differences when compared to FPC8. These findings demonstrate that the 1–4-thio bridge of disaccharide is a powerful anticancer pharmacophore, and its potential use needs further studies.     

Viscoelastic properties of waxy maize starch and selected non-starch hydrocolloids gels

Polysaccharides play a significant role in food systems as texture forming agent. Their solutions posses some peculiar rheological behaviors. Some information about structure of these biopolymers and relaxation processes occurring in their solutions may be obtained by investigation of linear viscoelasticity properties. The aim of this research was to investigate viscoelastic properties of systems consisting of waxy maize starch–hydrocolloid–water. To achieve this aim two techniques were applied: rheological measurements in frequency domain and structural studies by means of AFM method. The results were interpreted on basis of time–temperature superposition principle and phenomenological theory of viscoelasticity. Thermal stability of analyzed systems allowed applying time-temperature superposition principle. Calculation of a_T parameter enabled to obtaine the master curves. Continuous Maxwell model was applied to analyze phase separation in examined systems. Relaxations spectra obtained by Tikhonov regularization method were heterogeneous, indicating on non-homogenous structure of system.     

Exercise training prevents Ca2+ dysregulation in coronary smooth muscle from diabetic dyslipidemic yucatan swine.

Aerobic exercise training is known to have profound cardioprotective effects in disease, yet cellular mechanisms remain largely undefined. We tested the hypothesis that increased sarcoplasmic reticulum Ca(2+) buffering and increased voltage-gated Ca(2+) channel density underlie coronary smooth muscle intracellular Ca(2+) (Ca(2+)(i)) dysregulation in diabetic dyslipidemia and that exercise training would prevent these increases. Yucatan swine were maintained in 1) control, 2) alloxan-induced hyperglycemic, 3) high fat/cholesterol fed, 4) hyperglycemic plus high fat/cholesterol fed (diabetic dyslipidemic), and 5) diabetic dyslipidemic plus exercise-trained (treadmill running) conditions. After 20 wk, the heart was removed and smooth muscle cells isolated from the right coronary artery. We utilized fura-2 imaging of Ca(2+)(i) levels to separate the functional role of the sarcoendoplasmic reticulum Ca(2+)-ATPase (SERCA) from the Na(+)-Ca(2+) exchanger and the plasmalemmal Ca(2+)-ATPase, and whole-cell patch clamp to examine voltage-gated Ca(2+) channel current density (i.e., Ca(2+) influx). Results indicated that diabetic dyslipidemia impaired plasmalemmal Ca(2+) efflux, increased basal Ca(2+)(i) levels, increased SERCA protein and sarcoplasmic reticulum Ca(2+)(i) buffering, and elicited an approximately 50% decrease in voltage-gated Ca(2+) channel current density. Exercise training concurrent with the diabetic dyslipidemic state restored plasmalemmal Ca(2+) efflux, SERCA protein, sarcoplasmic reticulum Ca(2+)(i) buffering, and voltage-gated Ca(2+) channel current density to control levels. Interestingly, basal Ca(2+)(i) levels were significantly lower in the exercise-trained group compared with control. Collectively, these results demonstrate a crucial role for exercise in the prevention of diabetic dyslipidemia-induced Ca(2+)(i) dysregulation.     

Dissociating the centrosomal matrix protein AKAP450 from centrioles impairs centriole duplication and cell cycle progression

Centrosomes provide docking sites for regulatory molecules involved in the control of the cell division cycle. The centrosomal matrix contains several proteins, which anchor kinases and phosphatases. The large A-Kinase Anchoring Protein AKAP450 is acting as a scaffolding protein for other components of the cell signaling machinery. We selectively perturbed the centrosome by modifying the cellular localization of AKAP450. We report that the expression in HeLa cells of the C terminus of AKAP450, which contains the centrosome-targeting domain of AKAP450 but not its coiled-coil domains or binding sites for signaling molecules, leads to the displacement of the endogenous centrosomal AKAP450 without removing centriolar or pericentrosomal components such as centrin, gamma-tubulin, or pericentrin. The centrosomal protein kinase A type II alpha was delocalized. We further show that this expression impairs cytokinesis and increases ploidy in HeLa cells, whereas it arrests diploid RPE1 fibroblasts in G1, thus further establishing a role of the centrosome in the regulation of the cell division cycle. Moreover, centriole duplication is interrupted. Our data show that the association between centrioles and the centrosomal matrix protein AKAP450 is critical for the integrity of the centrosome and for its reproduction.     

Myo1c regulates glucose uptake in mouse skeletal muscle

Contraction and insulin promote glucose uptake in skeletal muscle through GLUT4 translocation to cell surface membranes. Although the signaling mechanisms leading to GLUT4 translocation have been extensively studied in muscle, the cellular transport machinery is poorly understood. Myo1c is an actin-based motor protein implicated in GLUT4 translocation in adipocytes; however, the expression profile and role of Myo1c in skeletal muscle have not been investigated. Myo1c protein abundance was higher in more oxidative skeletal muscles and heart. Voluntary wheel exercise (4 weeks, 8.2 ± 0.8 km/day), which increased the oxidative profile of the triceps muscle, significantly increased Myo1c protein levels by ～2-fold versus sedentary controls. In contrast, high fat feeding (9 weeks, 60% fat) significantly reduced Myo1c by 17% in tibialis anterior muscle. To study Myo1c regulation of glucose uptake, we expressed wild-type Myo1c or Myo1c mutated at the ATPase catalytic site (K111A-Myo1c) in mouse tibialis anterior muscles in vivo and assessed glucose uptake in vivo in the basal state, in response to 15 min of in situ contraction, and 15 min following maximal insulin injection (16.6 units/kg of body weight). Expression of wild-type Myo1c or K111A-Myo1c had no effect on basal glucose uptake. However, expression of wild-type Myo1c significantly increased contraction- and insulin-stimulated glucose uptake, whereas expression of K111A-Myo1c decreased both contraction-stimulated and insulin-stimulated glucose uptake. Neither wild-type nor K111A-Myo1c expression altered GLUT4 expression, and neither affected contraction- or insulin-stimulated signaling proteins. Myo1c is a novel mediator of both insulin-stimulated and contraction-stimulated glucose uptake in skeletal muscle.     

Life cycle thinking in small and medium enterprises: the results of research on the implementation of life cycle tools in Polish SMEs—part 2: LCA related aspects

Purpose

This article is the second part of a series of articles presenting the results of research on the implementation of lifecycle management tools in small- and medium-sized companies in Poland. This work is part of a project financed by the Polish Agency for Enterprise Development (PAED), which began in February 2011. It was carried out by the Wielkopolska Quality Institute, a business environment institution associated with the Polish Centre for life cycle assessment (PCLCA). The main practical objective of the project was to support small and medium enterprises (SMEs) in their business development, e.g. by expanding their horizons beyond the sphere of their operation and identifying new areas for the improvement and promotion of the products and services they offer. The specific objective of the analysis on the environmental impact was an attempt to answer the question of whether environmental LCA is a good management tool for this type of business. Part 2 describes results of the evaluation of the implementation of LCA in SMEs conducted in 46 companies involved in the project.

Methods

In order to assess the effectiveness of the project and the effectiveness of the implementation of LCA and life cycle costing (LCC), a survey was conducted of small and medium businesses where the implementation work had been fully completed. In total, 46 organisations agreed to participate in the LCA survey, which was almost 66 % of all the companies where the LCA and LCC studies had been carried out within the project. The survey was conducted using individual in-depth interviews. Questions to the representatives of the companies referred both to aspects of their functioning in the market (characteristics of a company, its market share, management systems, environmental policy, suppliers and clients) and the operation of their environmental service (assessment of its effectiveness, motivation and difficulties in its implementation), as well as opinions on the potential applications of LCA in their current operations.

Results and discussion

The experience and observations of LCA experts resulting from their cooperation with the organisations analysed are largely supported by the results of the survey. The overall impression gained from the project is that the small- and medium-sized enterprises analysed have a problem with accepting and understanding the life cycle perspective and show limited interest in taking liability for environmental aspects beyond the mandatory legal standards and boundaries of their business operations. The survey shows that the companies rarely analyse environmental aspects appearing on many different stages of the life cycle of their products. Most of them focus on their current operations while trying to meet the mandatory legal requirements relating to environmental protection. It should be noted, however, that SMEs taking part in the studies appreciate the opportunities offered by LCA, their usefulness in business practice, recognise the potential for using life cycle techniques in the future and their impact on the management process, procedure or thinking about the products they manufacture. The result of the study is the identification of four key areas relevant to SMEs which may affect their willingness to adopt the life cycle perspective and undertake environmental measures.

Conclusions

It seems that implementing LCT in small- and medium-sized enterprises requires a special approach. These are often companies with limited human resources (often just a few people) and financial resources (often operating on the verge of survival), with a weak position in a supply chain and, therefore, having various priorities in their daily operation. The researchers also encountered awareness barriers as a result of which the idea of going beyond an organisation and making an entire LCA of a product was often simply misunderstood. The studies conducted among SMEs have shown that managers' own intuition and research on customer preferences were largely conducive to improve existing or introducing new products or services, while changes were mostly introduced due to the requirements of the market, or the desire to reduce costs. It can be assumed that their non-obligatory nature also contributed to the relatively low interest in LCA initiatives and not recognising their usefulness. It seems that it would be useful to carry out relatively simple, but integrated, LCA/LCC analyses in SMEs so that the companies would clearly see the economic effect of the proposed environmental improvements. The analyses conducted lead to the conclusion that the incentive for SMEs to take measures should come from outside, e.g. as requirements for green public procurements, or as part of assessment made by suppliers in a supply chain.