Method for monitoring of the protein amino group changes during carbonylation

Objectives

Carbonylation of the protein amino, guanidino and thiol groups is one of the important causes of vascular complications in diabetes. We developed a simple spectrophotometric method for monitoring of the changes in the protein amino group contents during carbonylation.

Design and methods

The method is based on the reaction of amino group with p-benzoquinone in the slightly alkaline media. It was applied during carbonylation in vitro with methylglyoxal and in vivo in 13 patients with type 2 diabetes and 20 healthy persons.

Results

The method is simple, fast, precise (RSD in the range of 1.2–1.8%) and accurate (recovery about 100%). The content of amino groups in human serum albumin isolated from diabetics was significantly lower (p < 0.01) in comparison with a control group.

Conclusion

The method developed is suitable for quantification of protein amino groups during in vitro carbonylation as well as for clinical practice.     

Enucleation of feeder cells and egg cells with psoralens

The cell nucleus must be inactivated or destroyed in order to generate feeder layers for cultured cells or to prepare recipient egg cells for nuclear transfer. Existing enucleation techniques are either cumbersome or employ toxic chemicals. Here we report a new method to enucleate cells by treatment with a psoralen and long‐wave ultraviolet light. The technique is >90% efficient and causes little cytoplasmic damage to the treated cell. We have used psoralen treatment to enucleate a wide variety of cells, including eggs, sperm, HeLa cells, and fibroblasts. Colonies of human embryonic stem cells (hESCs) and human keratinocyte precursors grown on psoralen‐treated feeders are indistinguishable from those grown on gamma‐irradiated or mitomycin C‐treated cells. Psoralen enucleation provides a rapid, simple, and non‐toxic method to generate feeder cells. The technique is also useful for nuclear transfer studies in species with large eggs whose cleavage divisions are not regulated by cell‐cycle checkpoints. Developmental Dynamics 238:2614–2621, 2009. © 2009 Wiley‐Liss, Inc.     

Relation between QRS amplitude and left ventricular mass in the initial stage of exercise-induced left ventricular hypertrophy in rats

The aim of the study was to analyze the relationship between QRS amplitude and left ventricular mass (LVM) in early stages of two different experimental models of left ventricular hypertrophy (LVH) in rats: in exercise-induced hypertrophy and pathological hypertrophy due to genetically conditioned pressure overload. Three groups of experimental animals were studied: healthy control Wistar-Kyoto rats (WKYs), spontaneously hypertensive rats (SHRs), and WKY rats exposed to training by intermittent swimming (SWIM). Orthogonal electrocardiograms were recorded in each group at the age of 12 and 20 weeks, and the maximum spatial QRS vector (QRSmax) was calculated. Then the animals were sacrificed and LVM was measured. The specific potential of myocardium (SP) was calculated as a ratio of QRSmax to LVM. The QRSmax values did not follow the changes in LVM. At the end of the follow-up period, the highest values of QRSmax were recorded in the control WKY rats (0.80 +/- 0.05 mV). The QRSmax values in both groups with experimental LVH were significantly lower as compared with control animals (SHR 0.44 +/- 0.02 mV, p < 0.001; SWIM 0.53 +/- 0.04 mV, p < 0.001). Similarly, the SP values were significantly lower in both groups with experimental LVH as compared with control animals (SHR 0.42 +/- 0.02 mV/g, p < 0.001; SWIM 0.55 +/- 0.05 mV/g, p < 0.001). A decrease in QRSmax and SP was observed in both models of experimental LVH. We attributed these findings to the changes in electrogenetic properties of myocardium in the early stage of developing LVH. In other words, it is changes of nonspatial determinants that influence the resultant QRS voltage in terms of the solid angle theory.     

Performance implications of Porter's generic strategies in Slovak hospitals.

The aim of the study was to examine the use of Porter's generic strategies and their effect on performance in the context of the Slovak hospital industry. Using mail survey the study first identified the natural taxonomy of four strategic types of Slovak hospitals, based on their use of Porter's generic strategies in pure form and in combination. Next the study examined whether different strategic types were associated with different levels of organisational performance, while controlling for such variables as size and location, which have been argued to influence the hospital performance. The findings indicate that hospitals which follow a "stuck-in-the-middle" strategy, in general, have superior performance on all used performance measures, while hospitals that place only low emphasis on cost leadership, differentiation and focus, labelled "wait and see" in this study, perform the poorest. The study concludes that the research provided body of knowledge relevant for the Slovak hospital industry, that may be used by hospital managers in the strategy formulation process as well as by the researches in exploring the influence of different contingencies on hospitals' strategic orientation.     

Change in the iso-enzyme profiles of urinary N-acetyl-beta-D-glucosoaminidase in workers exposed to mercury

The iso-enzyme profiles of urinary N-acetyl-beta-D-glucosoaminidase (NAG) were studied in the workers from a chloro-alkaline electrolysis plant that had been continuously exposed to elemental mercury. The same workers then had a work break of four months and were later re-exposed to mercury for another five months. The activities of urinary NAG in workers exposed to mercury before and after the work break and in the control group were 2.09 +/- 1.03 IU/L; 0.90 +/- 0.52 IU/L and 1.13 +/- 0.35 IU/L, respectively. The fraction of the dominant A form activity in the total activity, in the workers exposed to mercury after a work break (85.31 +/- 5.32%), was almost equal to the fraction of A in the control group (84.64 +/- 2.75%). The percent fraction of A forms (A and A2) during continuous exposure was almost equal to the sum of the fractions A and A2 in the control group and for exposure after the work break. The fraction of the B form activity in the total activity during exposure to elemental mercury for five months after the work break (7.41 +/- 3.45%) was lower (P < 0.1) than the fraction of the B form in the control group (8.62 +/- 2.19%). The decrease in the fraction of the B form compared with the control group was more significant (P < 0.001) in the case of continuously exposed workers (5.25 +/- 2.55%). Beside the major A and B iso-enzymes of NAG, the A2 form was also isolated, and its fraction in the control group (6.73 +/- 2.15%) was not negligible. For workers exposed to mercury for five months, it was 7.28 +/- 3.52%. It is concluded that mercury affected the increased exocytosis of iso-enzyme A and the inhibition of B iso-enzyme of NAG. The increase in activity of urinary NAG in subjects exposed to mercury was a consequence of the increased excretion of A form. It was established also that the four months work break was sufficient for the repair of renal damages that occurred during continuous exposure, and which were monitored by the activity of NAG. Thus, the effects of mercury seem to have been reversible.     

Hospital human resource planning in Slovakia

The study is an attempt to provide empirical evidence, in the context of acute hospital care, of the current human resource practices in the health sector of the Slovak Republic. Using a sample of 72 acute care hospitals the research explored the perceived functions, typical customers and priorities of hospital human resource departments, ownership of a workforce plan, and the relationships between ownership of a workforce plan and type of hospital, as well as the degree to which different human resource activities are given priority. Cross-tabulation procedure revealed statistically significant relationships between ownership of a workforce plan and the degree of priority given to having a quick, efficient and cost-effective recruitment and selection system and, not surprisingly, the degree of priority given to ensuring that the human resource department has a workforce plan. The study evidence also indicates that, although the human resource staff in hospitals seem to be aware of their role in assisting hospital management in decision making, the human resource function in the Slovak hospitals still rather resembles that of a personnel administration than that of an important strategic human resource activity.     

Discrepancy between increased left ventricular mass and "normal" QRS voltage is associated with decreased connexin 43 expression in early stage of left ventricular hypertrophy in spontaneously hypertensive rats

Background and Purpose

On the basis of our previous results of animal and human studies, we assume that the discrepancies between increased left ventricular mass (LVM) and electrocardiographic (ECG) findings not exceeding the upper normal limits in left ventricular hypertrophy (LVH) are conditioned by the electrical remodeling of hypertrophied myocardium. We assumed that these discrepancies observed in the early stage of LVH in spontaneously hypertensive rats (SHR) are associated with a decreased expression of connexin 43.

Methods

Standard 12-lead ECG was recorded in 20-week-old male SHR and age-matched and sex-matched normotensive Wistar rats (Institute of Experimental Pharmacology SAV, Dobra Voda, Slovakia). The approximated maximum QRS spatial vector magnitude (QRSmax) was calculated from leads V₂, aVF, and V₅. Left ventricular mass was weighed, and the specific potential (SP) of myocardium was calculated as the QRSmax-to-LVM ratio. Left ventricular protein levels of connexin 43 were analyzed with sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting.

Results

The LVM values were significantly higher in SHR than in normotensive controls (0.96 ± 0.03 g and 0.680 ± 0.07 g, respectively; P < .001). The QRSmax values in SHR did not follow the increase either in systolic blood pressure or in LVM. The SP values in SHR were significantly lower than those in control rats (0.92 ± 0.11 mV/g and 1.358 ± 0.06 mV/g, respectively; P < .01). A 37% decrease in connexin 43 level was observed in SHR.

Conclusions

The QRS voltage did not follow the increase in the LVM in 20-week-old SHR, and the values of connexin 43 were lower in SHR than in normotensive controls. We believe that the discrepant findings between ECG voltage and LVM can be caused by the electrical remodeling in the early stages of LVH.     

Alteration of adenoid tissue alkaline and acid phosphatase in children with secretory otitis media

OBJECTIVE: The role of pharyngeal lymphoid tissue in etiopathogenesis of secretory otitis is not yet defined. The influence of tonsillar and adenoid mass, weight, obstruction of naspharyngeal orrifitium, bacterial reservoire or some immunological events are of scientific interest. Tissue nonspecific alkaline phosphatase (TNAP) and acid phosphatase (ACP) are enzymes detected in lymphoid tissue, TNAP as characteristic of B cells, ACP as a characteristic of macrophages and folucullardentritic cells. These enzymes interfere in cell metabolism by removing 5' phosphate group from nucleotides and proteins. Specific activity and kinetic properties were studied in palatinal tonsils and adenoids of children with secretory otitis (OME) and compared with children with recurrent tonsillitis without ear involvement. METHOD: Adenoid and tonsillar tissue of l7 children with OME and 30 children with recurrent tonsillitis were subjected to biochemical investigation using method of releasing of p-nitrophenol from p-nitrophenylphosphate (pNPP). Kinetic parameters as Michaelis-Menten constant were calculated by non-linear regression estimation method. RESULTS: Specific activity of adenoid alkaline phosphatase was lower in children with OME in relation to children with recurrent tonsillitis (t=5.733507, p<0.01). Specific activity of adenoid acid phosphatase was also lower in children with OME (t=3.655456, p<0.01). pH optimum for both enzymes was the same in these two groups of children. Michaelis-Menten constant for both enzymes was significantly higher in adenoid of children with OME than in children with recurrent tonsillitis suggesting lower enzyme affinity for the substrate. CONCLUSION: Differences in specific activities and kinetic properties of adenoid alkaline and acid phosphatases between children with OME and children with recurrent tonsillitis without OME were verified in this study. The results of the study are not able to explain the alteration of alkaline and acid phosphatase characteristics but could point to some possible and specific role of nasopharyngeal lymphoid tissue in pathogenesis of secretary otitis.