Martin Gajdoš

Total plasma Nε-(carboxymethyl)lysine and sRAGE levels are inversely associated with a number of metabolic syndrome risk factors in non-diabetic young-to-middle-aged medication-free subjects

Abstract Background: Interaction of advanced glycation end products (AGEs) with their specific cell-surface receptor for AGEs (RAGE) induces production of reactive oxygen species, pro-diabetic, pro-inflammatory, and pro-atherogenic responses. The metabolic syndrome (Metsy) imposes a high risk of development of cardiovascular disease and unequivocally predisposes the non-diabetics to type 2 diabetes mellitus. The aim of the study was to investigate the association between circulating soluble RAGE (sRAGE), Nε-(carboxymethyl)lysine (CML) or AGE-associated fluorescence of plasma (AGE-Fl) with the number of manifested Metsy risk factors in young-to-middle-aged medication-free non-diabetic subjects. Methods: Metsy was classified according to NCEP/ATP III criteria; plasma sRAGE and total CML were determined by ELISA methods and AGE-Fl fluorimetrically. Results: From among 437 participants aged 33±11 years, 58% were females. In total 174 subjects were Metsy risk factors-free, 142 presented one, 59 presented two risk factors, and 62 suffered from Metsy. Plasma sRAGE and CML/albumin levels decreased with increasing number of Metsy risk factors (p<0.01, both), while AGE-Fl/albumin levels remained similar. Multivariate analysis selected waist circumference as a main determinant of plasma sRAGE as well as CML/albumin levels. Conclusions: In young-to-middle-aged non-diabetic medication-free subjects plasma total CML/albumin and sRAGE levels decrease prior to the manifestation of Metsy. With regards to RAGE-mediated CML trapping into adipose tissue inducing dysregulation of pro-inflammatory cytokines, adipokines, and the development of obesity-related insulin resistance, and the potential involvement of sRAGE in feedback regulation of the toxic effects of AGE/RAGE-mediated signaling, this early decline might be of clinical impact in development of type 2 diabetes and its complications.

Alterations in the QRS complex in the offspring of patients with metabolic syndrome and diabetes mellitus: early evidence of cardiovascular pathology

OBJECTIVE This study was undertaken to evaluate the nature and onset of changes in the QRS complex in the offspring of patients with diabetes mellitus (DM) and metabolic syndrome (MetS). METHODS AND METHODS A total of 529 subjects, divided into 5 groups, were included in the study: (i) group DM (n = 92), patients with DM; (ii) group MetS (n = 125), patients with MetS; (iii) group O-DM (n = 109), offspring of patients with DM; (iv) group O-MetS (n = 122), offspring of patients with MetS; and (v) group HO (n = 81), offspring of healthy subjects. QRS parameters analyzed included amplitude, maximum QRS spatial vector magnitude, electrical axis (EA), and 3 electrocardiogram (ECG) criteria for left ventricular hypertrophy based on amplitude criteria: Sokolow-Lyon index, Cornell voltage, and Gubner criterion. RESULTS Patients with DM and MetS showed a significant leftward shift of the EA when compared with the control group. A modest but significant leftward shift of EA was also observed in both offspring groups. These EA and maximum QRS spatial vector magnitude changes were reflected in the individual leads of the 12-lead ECG. The prevalence of a positive diagnosis by accepted electrocardiographic criteria (ECG left ventricular hypertrophy) was low. CONCLUSION Patients with DM and MetS displayed significant changes in QRS complex that suggest depolarization sequence deterioration. Similar changes were observed also in the offspring of patients with DM and MetS, which suggests early subclinical cardiovascular damage. These findings have implications for prevention, early diagnosis, and treatment in the offspring of patients with DM and MetS.

Impact of ovariectomy, high fat diet, and lifestyle modifications on oxidative/antioxidative status in the rat liver

Aim To estimate the impact of high fat diet and estrogen deficiency on the oxidative and antioxidative status in the liver of the ovariectomized rats, as well as the ameliorating effect of physical activity or consumption of functional food containing bioactive compounds with antioxidative properties on oxidative damage in the rat liver. Methods The study was conducted from November 2012 to April 2013. Liver oxidative damage was determined by lipid peroxidation levels expressed in terms of thiobarbituric acid reactive substances (TBARS), while liver antioxidative status was determined by catalase (CAT), glutathione peroxidase (GPx), glutathione S-transferase (GST), glutathione reductase (GR) activities, and glutathione (GSH) content. Sixty-four female Wistar rats were divided into eight groups: sham operated and ovariectomized rats that received either standard diet, high fat diet, or high fat diet supplemented with cereal selenized onion biscuits or high fat diet together with introduction of physical exercise of animals. Results High fat diet significantly increased TBARS content in the liver compared to standard diet (P = 0.032, P = 0.030). Furthermore, high fat diet decreased the activities of CAT, GR, and GST, as well as the content of GSH (P < 0.050). GPx activity remained unchanged in all groups. Physical activity and consumption of cereal selenized onion biscuits showed protective effect through increased GR activity in sham operated rats (P = 0.026, P = 0.009), while in ovariectomized group CAT activity was increased (P = 0.018) in rats that received cereal selenized onion biscuits. Conclusion Feeding rats with high fat diet was accompanied by decreased antioxidative enzyme activities and increased lipid peroxidation. Bioactive compounds of cereal selenized onion biscuits showed potential to attenuate the adverse impact of high fat diet on antioxidative status.

Vitamin D Levels Decline with Rising Number of Cardiometabolic Risk Factors in Healthy Adults: Association with Adipokines, Inflammation, Oxidative Stress and Advanced Glycation Markers

Introduction Hypovitaminosis D associates with obesity, insulin resistance, hypertension, and dyslipoproteinemia. We asked whether the presence of multiple cardiometabolic risk factors, and which particular combination, exerts additive negative effects on 25(OH)D3 levels; and whether 25(OH)D3 levels associate with markers of inflammation and oxidative stress. Subjects and Methods In non-diabetic medication-free adults central obesity (waist-to-height ratio > 0.5); elevated blood pressure (systolic BP≥130 mm Hg and/or diastolic BP ≥85 mm Hg); increased atherogenic risk (log(TAG/HDL) ≥ 0.11); and insulin resistance (QUICKI < 0.322) were considered as cardiometabolic risk factors. 25(OH)D3 status was classified as deficiency (25(OH)D3 ≤20 ng/ml); insufficiency (levels between 20-to-30 ng/ml), or as satisfactory (>30 ng/ml). Plasma adipokines, inflammatory and oxidative stress markers, advanced glycation end-products, and their soluble receptor were determined. Results 162 subjects were cardiometabolic risk factors-free, 162 presented increased (i.e. 1 or 2), and 87 high number (i.e. 3 or 4) of cardiometabolic risk factors. Mean 25(OH)D3 decreased with rising number of manifested risk factors (36 ± 14 ng/ml, 33 ± 14 ng/ml, and 31 ± 15 ng/ml, respectively; pANOVA: 0.010), while prevalence of hypovitaminosis D did not differ significantly. Elevated blood pressure and insulin resistance appeared as significant determinants of hypovitaminosis D. Subjects presenting these risk factors concurrently displayed the lowest 25(OH)D3 levels (29 ± 15 ng/ml). Plasma adipokines, inflammatory and oxidative stress markers, advanced glycation end-products, and their soluble receptor generally differed significantly between the groups, but only advanced oxidation protein products and advanced glycation end-products associated fluorescence of plasma showed significant independent association with 25(OH)D3 levels. Conclusion In apparently healthy adults increasing number of cardiometabolic risk factors associates with poorer 25(OH)D3 status, while the association between 25(OH)D3 status and inflammatory or oxidative stress markers remains equivocal.

Association of sVAP-1, sRAGE, and CML with lactation-induced insulin sensitivity in young non-diabetic healthy women

BACKGROUND In comparison with non-lactating women breast-feeding mothers display higher insulin sensitivity. Recent data suggest that advanced glycation end products, soluble receptor for advanced glycation end products (sRAGE) and soluble vascular adhesion protein-1 (sVAP-1) may play a role in insulin resistance even in healthy subjects. AIM We studied whether breast-feeding induced insulin sensitivity associates with changes in concentrations of circulating sVAP-1, sRAGE and N(ε)-(carboxymethyl)lysine (CML) - chemically defined advanced glycation end product and RAGE ligand. METHODS In 74 lactating non-diabetic mothers, 45 weaned non-diabetic mothers and 50 age-matched non-parous women insulin sensitivity was assessed using Quantitative insulin-sensitivity check index (QUICKI). sVAP-1, sRAGE and CML levels were determined. RESULTS Lactating mothers were more insulin sensitive than their weaned and non-parous counterparts. Lactating mothers displayed the highest concentrations of sRAGE, and higher sVAP-1 levels if compared to weaned mothers. Both groups of mothers presented with lower CML levels than the non-parous women. CONCLUSION Lactation-induced insulin sensitivity is associated with higher sVAP-1 and a tendency towards higher sRAGE levels. Lactation-associated rise in sVAP-1 may promote effective glucose utilization in the mother. Lactation-induced insulin sensitivity vanishes shortly after weaning. In young healthy women CML levels are of no clinical relevance to insulin sensitivity.

Effects of high fat diet, ovariectomy, and physical activity on leptin receptor expression in rat brain and white fat tissue.

Aim To evaluate in a rat animal model whether ovariectomy, high fat diet (HFD), and physical activity in the form of running affect leptin receptor (Ob-R) distribution in the brain and white fat tissue compared to sham (Sh) surgery, standard diet (StD), and sedentary conditions. Methods The study included 48 female laboratory Wistar rats (4 weeks old). Following eight weeks of feeding with standard or HFD, rats were subjected to either OVX or Sh surgery. After surgery, all animals continued StD or HFD for the next 10 weeks. During these 10 weeks, ovariectomy and Sh groups were subjected to physical activity or sedentary conditions. Free-floating immunohistochemistry and Western blot methods were carried out to detect Ob-R in the brain and adipose tissue. Results StD-ovariectomy-sedentary group had a greater number of Ob-R positive neurons in lateral hypothalamic nuclei than StD-Sh-sedentary group. There was no difference in Ob-R positive neurons in arcuatus nuclei between all groups. Ob-R distribution in the barrel cortex was higher in HFD group than in StD group. Ob-R presence in perirenal and subcutaneous fat was decreased in StD-ovariectomy group. Conclusion HFD and ovariectomy increased Ob-R distribution in lateral hypothalamic nuclei, but there was no effect on arcuatus nuclei. Our results are first to suggest that HFD, ovariectomy, and physical activity affect Ob-R distribution in the barrel cortex, which might be correlated with the role of Ob-R in election of food in rats.

The Cell Surface Markers Expression in Postmenopausal Women and Relation to Obesity and Bone Status

The age-related changes and hormonal deprivation in postmenopausal women are associated with the immune response alteration. The excessive fat accumulation, local and systemic inflammation may lead to dysregulation in immune function and relevant health problems, including obesity and osteoporosis. We analyzed the expression of cell surface markers in the venous blood specimens, stained with fluorophores-conjugated monoclonal antibodies and analysed by multicolour flow cytometry. The significant changes of cytotoxic, naive, and memory T-lymphocytes, plasmacytoid dendritic cells (DCs) were in postmenopausal women versus fertile women. Body mass index (BMI) affected markedly the cell surface expression of CD265/RANK. Osteoporosis is linked to reduced percentage of plasmacytoid DCs, and elevated natural Treg cells (p < 0.05). The confounding factors such as women age, BMI, bone mineral density (BMD), waist size and tissue fat affect the expression of RANK on myeloid DCs and CD40L on T-lymphocytes that might be the immunophenotypic modulators after menopause.

A pilot study of a genetic CJD risk factor (E200K) in the general Slovak population

Creutzfeldt-Jakob disease (CJD) is a fatal, incurable human transmissible spongiform encephalopathy, or prion disease. Classical CJD has been recognised since the 1920s. CJD occurs in three forms : sporadic, of unknown origin, genetic, related to disease specific point mutations, insertions or deletions of the prion protein gene (PRNP) and iatrogenic (iCJD), caused by accidental transmission of the disease in the course of medical or surgical treatment. A new variant of CJD, which is causally related to bovine spongiform encephalopathy (BSE) was first described by Will et al. in 1996. The risk of horizontal transmission of CJD has been confirmed in classical [1] as well as in variant CJD. Precautions and measures to prevent iCJD are focused mainly on clinical CJD patients, but a potential risk from preclinical cases, for example asymptomatic carriers of CJDspecific mutations of the PRNP, cannot be excluded. These carriers represent a ‘‘genetic CJD-risk group’’ in the general population. More than 37 disease-specific mutations have been described in the PRNP coding sequence. The most common point mutation associated with genetic CJD is at codon 200 (E200K) [2]. While in the majority of countries the genetic form represents about 10–15 % of all CJD patients, in Slovakia it has never been \65 % since 1975. [3]. Except for one case with the R208H mutation, all 202 genetic CJD patients in Slovakia carry the E200K mutation (gCJD). Most of these cases are concentrated in two clusters in central Slovakia [4]. Voluntary genetic testing of healthy blood relatives has revealed that 35 % carry the E200K mutation [3]. Besides the risk of gCJD (59 % of carriers will develop the disease) (3), ‘‘healthy’’ carriers could act as a source of iatrogenic infection. To our knowledge, the prevalence of the mutation E200K has not been studied. The aim of this pilot study is to investigate the occurrence and frequency of asymptomatic carries of E200K in the Slovak general population, comparing the areas of focal accumulation of gCJD with extrafocal areas, i.e. comparison of regions with contrasting incidence rates of gCJD. In the present study the occurrence of the mutation E200K and the distribution of the M129 V polymorphism of PRNP were analysed in 2,662 subjects with no known link to patients with gCJD. Tested were DNA from two different sources: 1. anonymised DNA samples of newborns provided by Slovak Newborn Screening Center were from regions with different incidence of gCJD: 1a) 893 samples from two focal accumulations (clusters) of gCJD in central Slovakia (mean incidence of gCJD 3.55/million) and 1b) 757 samples from extrafocal regions in western and E. Mitrova D. Kosorinova M. Gajdos I. Tomeckova Medical Faculty, Slovak Medical University, Bratislava, Slovakia

Enalapril Inhibits Growth and Proliferation of Various Tissues in Rat Normotensive Four–Sixths Kidney Ablation Nephropathy

Most experimental studies on kidney proliferation and its attenuation by angiotensin–converting enzyme inhibitors were performed in the rat hypertensive remnant–kidney model with a five–sixths kidney ablation. The developing hypertension rose the objections on the hypertension and its treatment in control rats. A normotensive four–sixths remnant–kidney model (Nx) was elaborated, compared with sham–operated (S) animals, and a subantihypertensive dosage of enalapril (E) was administered for 4 weeks of intensive kidney tissue proliferation (NxE). The pair–fed groups increased their body weight and blood pressure comparably. Moderately increased plasma creatinine and urea concentrations were found in the Nx group; markedly increased levels in the NxE group. Nx increased proteinuria, and E attenuated its increase. The remnant–kidney weight (Nx 912±31 vs. S 1,111±36 mg, p<0.001) was still lower, but collagen (Col; Nx 164±2 vs. S 148±5 mg/100 g, p<0.05) and tubular protein/DNA ratio (Nx 26.2±10.8 vs. S 9.8±1.0, p<0.05) increased markedly in the Nx group; E attenuated the kidney growth (NxE 719±31 vs. Nx 912±31 mg, p<0.01) and decreased the tubular protein/DNA ratio remarkably (NxE 15.3±10.5 vs. Nx 26.2 ±10.8), but E did not inhibit the Col accumulation. Nx decreased the heart (Nx 1,002±28 vs. S 1,130±41 mg, p<0.05), but not liver weights and did not influence Col concentrations or protein/DNA ratios either in heart or liver. E potentiated the weight decrease of heart (NxE 862±20 vs. Nx 1,002±28 mg, p<0.01) and liver (NxE 8.3±0.44 vs. Nx 10.3±0.51 g, p<0.001) and Col accumulation (heart: NxE 113±6 vs. Nx 92±5 mg/100 g, p<0.01; liver: NxE 134±8 vs. Nx 101±9 mg/100 g, p<0.01). Nx did not influence either the soleus muscle weight or its Col accumulation, but it increased its protein/DNA ratio (Nx 66.3±4.7 vs. S 35.5±2.8 mg/100 g, p<0.01). E increased the Col concentration in muscle (NxE 141±3 vs. Nx 110±5 mg/100 g, p<0.01), while it attenuated the increase in protein/DNA ratio (NxE 36.6±2.1 vs. Nx 66.3±4.7, p<0.01). In conclusion, kidney ablation nephropathy stimulating kidney proliferation evokes only minor changes in heart, liver and striated muscle. E inhibits markedly the kidney proliferation and functional recovery, but does not prevent the Col accumulation. E evokes antiproliferative changes also in the heart and surprisingly even in the liver. Alterations in soleus muscle are only borderline.

A different effect of obesity on ECG in premenopausal and postmenopausal women

Both obesity and menopause are significant cardiovascular risk factors. In postmenopausal women the protective effect of estrogens is reduced and menopause is frequently associated with occurrence of other significant cardiovascular factors including obesity. This study was focused on evaluating the effect of obesity on the QRS complex in pre- and postmenopausal women. We present results of analysis of 199 electrocardiograms of pre- and postmenopausal women analyzed in relation to the body mass index within normal limits (BMI 20 to 24.9 kg/m2) and obesity (BMI > 30 kg/m2), respectively. Obesity in premenopausal women and menopause significantly affected both the electrical axis (EA) and maximum QRS spatial vector magnitude (QRSmax). The highest QRSmax and electrical axis values were observed in premenopausal lean women, and they were significantly higher as than in the premenopausal obese women, postmenopausal lean and obese women (QRSmax: 1.66 ± 0.4 mV, 1.17 ± 0.35 mV, 1.4 ± 0.46 mV, and 1.35 ± 0.39 mV, resp.). (EA: 56.4 ± 18.0°, 38.22 ± 18.38°, 45.82 ± 18.63°, and 36.75 ± 17.51°). The differences between obese premenopausal women, lean and obese postmenopausal women were not statistically significant. These differences were reflected in 12-lead ECG amplitude. The presence of additional cardiovascular risk factors did not affect the ECG parameters. Obesity significantly affected QRS complex in premenopausal women. This effect was comparable with the effect of menopause. Because all QRS complex changes were within normal limits, these results suggest that ECG evaluation in women should go beyond traditional diagnostic categories and consider the relationship between ECG changes and two cardiovascular risk factors - obesity and menopause.