Interethnic differences of CYP2C9 alleles in healthy Hungarian and Roma population samples: Relationship to worldwide allelic frequencies

CYP2C9 gene polymorphisms are widely studied in several ethnic groups, however they are less known in the Roma population. The aim of this work was to study the ethnic differences of the CYP2C9 allele distribution in a healthy Roma population in order to compare them with a healthy Hungarian population. A total of 535 Hungarian and 465 Roma volunteers were genotyped for the CYP2C92 (Arg144Cys) and CYP2C93 (Ile359Leu) allelic variants by PCR-RFLP assay. The frequencies of the CYP2C91, 2 and 3 alleles in the Hungarian population were 0.787, 0.125, and 0.088 and in Roma 0.727, 0.118, and 0.155, respectively. We found a significant difference in CYP2C93 prevalence between the Hungarian and Roma populations, which have therapeutic consequences (p < 0.005). The distribution of 1/1, 1/2, 1/3, 2/2, 2/3, and 3/3 genotypes in Hungarians were 0.620, 0.195, 0.139, 0.021, 0.015, and 0.011, while in Roma were 0.533, 0.168, 0.219, 0.011, 0.047, and 0.022, respectively. A significant difference was found between the Hungarian and Roma populations regarding the 1/1, 1/3 and the 2/3 (p < 0.005) genotypes. This is the first study to investigate the polymorphisms of CYP2C9 gene in the two largest populations in Hungary, healthy Hungarians and Roma. The prevalence of variant CYP2C9 alleles in the Hungarian population is similar to that observed in other European populations. In contrast, the Roma population differs from Hungarians, from most of other Caucasian groups, and from Indians in the incidence of CYP2C9 common variants. The difference in allele distribution patterns between the two populations studied has therapeutic implications as it influences the optimization of therapies.     

Netrin-1 prevents ischemia/reperfusion-induced myocardial infarction via a DCC/ERK1/2/eNOSs1177/NO/DCC feed-forward mechanism

We have recently shown that a novel endothelial mitogen netrin-1 potently stimulates nitric oxide (NO) production via a DCC-ERK1/2 dependent mechanism. In view of the well-established cardioprotective role of NO, the present study investigated whether netrin-1 is cardioprotective via NO signaling in the heart. Netrin-1 receptor DCC was abundantly expressed in the C57BL/6J mouse hearts. Perfusion of heart with netrin-1 (100 ng/mL) using a Langendorff system significantly increased NO production. Under ischemia/reperfusion (I/R), netrin-1 induced a substantial reduction in infarct size (21.8 ± 4.9% from 42.5 ± 3.6% in the controls), which was accompanied by an augmented production of NO. Pre-perfusion with DCC-antibody, U0126 (MEK1/2 inhibitor), L-NAME or PTIO (NO scavenger) attenuated protective effects of netrin-1 on infarct size and NO production, indicating upstream roles of DCC and ERK1/2 in NO production, as well as an essential role of NO in cardioprotection. Netrin-1 induced reduction in infarct size was significantly attenuated in DCC+/− mice, confirming an intermediate role of DCC. In additional experiments we found netrin-1 increased ERK1/2 and eNOS_s1177 phosphorylation, and DCC protein expression, which was diminished by I/R. Furthermore, netrin-1-induced DCC upregulation was NO and ERK1/2-dependent, implicating a feed-forward mechanism. DAF-AM staining revealed enhanced NO production in both cardiac endothelial cells (ECs) and myocytes. In primarily isolated cardiomyocytes, netrin-1 also increased NO production, DCC abundance and ERK1/2 phosphorylation. Of note, cardiac apoptosis was significantly attenuated by netrin-1, which was reversed by DCC-antibody, U0126, L-NAME or PTIO. In summary, our data clearly demonstrate that netrin-1 potently protects the heart from I/R injury by stimulating NO production from cardiac ECs and myocytes. This potent effect is mediated by a DCC/ERK1/2/eNOS_s1177/NO/DCC feed-forward mechanism in both cell types.     

Cardioprotective signaling to mitochondria

Mitochondria are central players in the pathophysiology of ischemia–reperfusion. Activation of plasma membrane G-coupled receptors or the Na,K-ATPase triggers cytosolic signaling pathways that result in cardioprotection. Our working hypothesis is that the occupied receptors migrate to caveolae, where signaling enzymes are scaffolded into signalosomes that bud off the plasma membrane and migrate to mitochondria. The signalosome–mitochondria interaction then initiates intramitochondrial signaling by opening the mitochondrial ATP-sensitive K⁺ channel (mitoK_ATP). MitoK_ATP opening causes an increase in ROS production, which activates mitochondrial protein kinase C epsilon (PKC), which inhibits the mitochondrial permeability transition (MPT), thus decreasing cell death. We review the experimental findings that bear on these hypotheses and other modes of protection involving mitochondria.     

Detection of myocardial ischemia by vessel-specific leads derived from the 12-lead electrocardiogram and its subsets

Currently used electrocardiographic criteria for identifying patients with ST-elevation myocardial infarction (STEMI) perform with high specificity but low sensitivity. Our aim was to enhance ischemia-detection ability of conventional STEMI criteria based on 12-lead electrocardiogram (ECG) by adding new criteria using 3 vessel-specific leads (VSLs) derived from 12-lead ECG. Study data consisted of 12-lead ECGs acquired during 99 ischemic episodes caused by balloon inflation in, respectively, left anterior descending coronary artery (LAD; n = 35), right coronary artery (RCA; n = 47), and left circumflex coronary artery (LCx; n = 17). ST deviation was measured at J point in 12 standard leads, and for 3 VSLs, its value was derived from 12-lead ECG by using 8 independent predictor leads or just a pair of precordial leads combined with a pair of limb leads. Mean values of sensitivity (SE) and specificity (SP) of ischemia detection achieved with conventional STEMI vs VSL criteria were then obtained from bootstrap trials. We found that the detection of ischemic state by conventional criteria achieved the mean SE/SP of 60%/96% in the total set of ischemic episodes, 74%/97% in the LAD subgroup, 60%/94% in the RCA subgroup, and 36%/100% in the LCx subgroup. In comparison, the mean SE/SP values of VSLs derived from 8 independent leads of 12-lead ECG were, at 125-μV threshold, 76%/96% in the total set, 91%/97% in the LAD subgroup, 70%/94% in the RCA subgroup, and 71%/100% in the LCx subgroup (with asterisk denoting a statistically significant increase). The mean SE/SP of VSLs derived from some of the 4-predictor lead sets (namely, those including lead V₃) matched or exceeded values achieved by VSLs derived from 8 predictors; for instance, with predictor leads I, II, V₃, V₆ derived VSLs attained at 125-μV threshold the mean SE/SP of 80%/95% in the total set, 91%/97% in the LAD subgroup, 74%/92% in the RCA subgroup, and 71%/100% in the LCx subgroup. Based on these results, we conclude that, in our data set, 3 VSLs derived from the complete standard 12-lead ECG—and even from its subsets—can identify acute ischemia better than existing STEMI criteria.     

Regular moderate exercise reduces advanced glycation and ameliorates early diabetic nephropathy in obese Zucker rats

Advanced glycation end products (AGEs) play a key role in the pathogenesis of diabetes and its complications, including the diabetic nephropathy. The renoprotective effects of exercise are well known; however, the mechanisms remain elusive. Here we examined whether a regular moderate exercise in obese Zucker rats (OZR), a model of diabetes- and obesity-associated nephropathy, will affect the development of early renal injury in OZR possibly via alteration of AGEs formation. The OZR were left without exercise (sedentary) or subjected to 10 weeks intermittent treadmill running of moderate intensity. Compared with sedentary OZR, kidneys of running OZR had significantly less glomerular mesangial expansion and tubulointerstitial fibrosis. Running OZR had significantly lower plasma AGEs-associated fluorescence and N-carboxymethyllysine. Correspondingly, renal AGEs and N-carboxymethyllysine content were lower in running OZR. Systemically, exercise increased aerobic metabolism, as apparent from urinary metabolite profiling. No differences in plasma glucose, insulin, or lipid profile were found between the 2 groups. Apart from lower advanced oxidation protein products (a marker of myeloperoxidase activity), no other marker of inflammation was altered by exercise, either systemically or locally in kidneys. No indication of changed oxidative status was revealed between the groups. Exercise in OZR decreased advanced glycation. This might represent the early event of exercise-induced renoprotection in diabetic nephropathy in OZR. If confirmed in clinical studies, regular moderate exercise could represent an easy and effective nonpharmacologic approach to reduce advanced glycation.     

Effects of mild calorie restriction and high-intensity interval walking in middle-aged and older overweight Japanese

We investigated whether a combination of mild calorie restriction (MCR) and high-intensity interval walking (HIW) improved physical fitness more than HIW alone in middle-aged and older overweight Japanese (40–69 years old, BMI  23.6 kg/m²). Forty-seven women and 16 men were divided into MCR + HIW and HIW groups. All subjects performed HIW: 5 sets of 3-min low-intensity walking (40% peak aerobic capacity for walking, VO_2peak) and 3-min high-intensity walking (70% VO_2peak) per day, 4 days per week, for 16 weeks while energy expenditure was monitored with a tri-axial accelerometer. The MCR + HIW group consumed meal replacement formula (240 kcal): a mixture of low-carbohydrates and -fat and high-protein, for either lunch or dinner everyday and therefore, had 87% of the energy intake of the HIW group during the intervention period. Although the HIW group showed improvements in BMI, blood pressure, and several blood chemicals, the MCR + HIW group had greater improvement. Moreover, the medical expenditure for the 6 months including the intervention period was 59% lower in the MCR + HIW group than in the HIW group. Our strategy of a short-term combination of MCR and HIW may thus prevent lifestyle-associated diseases and improve health in middle-aged and older overweight Japanese.     

Mechanistic insights into folic acid-dependent vascular protection: Dihydrofolate reductase (DHFR)-mediated reduction in oxidant stress in endothelial cells and angiotensin II-infused mice: A novel HPLC-based fluorescent assay for DHFR activity

Folate supplementation improves endothelial function in patients with hyperhomocysteinemia. Mechanistic insights into potential benefits of folate on vascular function in general population however, remain mysterious. Expression of dihydrofolate reductase (DHFR) was markedly increased by folic acid (FA, 50 μmol/L, 24 h) treatment in endothelial cells. Tetrahydrofolate (THF) is formed after incubation of purified DHFR or cellular extracts with 50 μmol/L of substrate dihydrofolic acid. THF could then be detected and quantified by high performance liquid chromatography (HPLC) with a fluorescent detector (295/365 nm). Using this novel and sensitive assay, we found that DHFR activity was significantly increased by FA. Furthermore, FA improved redox status of Ang II treated cells by increasing H₄B and NO bioavailability while decreasing superoxide (O₂⁻) production. It however failed to restore NO levels in DHFR siRNA-transfected or methotrexate pre-treated cells, implicating a specific and intermediate role of DHFR. In mice orally administrated with FA (15 mg/kg/day, 16 days), endothelial upregulation of DHFR expression and activity occurred in correspondence to improved NO and H₄B bioavailability, and this was highly effective in reducing Ang II infusion (0.7 mg/kg/day, 14 days)-stimulated aortic O₂⁻ production. 5′-methyltetrahydrofolate (5′-MTHF) levels, GTPCH1 expression and activity remained unchanged in response to FA or Ang II treatment in vitro and in vivo. FA supplementation improves endothelial NO bioavailability via upregulation of DHFR expression and activity, and protects endothelial cells from Ang II-provoked oxidant stress both in vitro and in vivo. These observations likely represent a novel mechanism (intermediate role of DHFR) whereby FA induces vascular protection.     

Distribution of CYP1A1*2A polymorphism in adult patients with acute lymphoblastic leukemia in a Mexican population

The effects of the CYP1A12A genotype on susceptibility to leukemia have received particular attention in recent years because this enzyme plays a central role in the activation of carcinogens. Several polymorphisms at the CYP1A1 locus have been identified and their genotypes appear to exhibit population frequencies that depend on ethnicity. We evaluated the role of the CYP1A12A genotype in adults with acute lymphoblastic leukemia (ALL) by genotyping 210 patients and 228 healthy controls from the Mexican population. The frequency of the CC genotype was 8% (18/228) in the control group and 42% (88/210) in ALL patients; the frequency of the CT genotype was 39% (89/228) and 29.5% (62/210), respectively; and that of the TT genotype was 53% (121/228) and 28.5% (60/210), respectively. The odds ratio was 8.4 (95% CI, 4.7–15.5; P < 0.001). These data indicate that the CYP1A12A genotype contributes significantly to susceptibility to adult ALL in a sample of the Mexican population.     

Attenuated expression of SECIS binding protein 2 causes loss of telomeric reserve without affecting telomerase

The family of selenoproteins have a broad range of functions, including protection against oxidative damage. Previous studies have shown that elevated levels of oxidative damage can induce accelerated loss of telomeric DNA during proliferation of mammalian cells. The incorporation of selenocysteine (Sec) into proteins in mammalian cells requires the Sec insertion sequence (SECIS) binding protein 2 (SBP2). Thus in the present study we have assessed the effect of knocking down the expression of SBP2 on telomere length. Following knock-down of SBP2 expression in two different human cell lines, the MSTO mesothelioma cell line (5 Kb average telomere length) and SY5Y neuroblastoma cell line (4.2 Kb average telomere length), we observed a significant reduction (−0.6 to −1.1 Kb; P  0.01) in telomere length as compared to control cells. This reduction in telomere length was independent of affects on telomerase, since both telomerase activity levels and Tert mRNA expression levels were not altered by knock-down of SBP2 expression. Furthermore, telomeres were particularly sensitive to S1 nuclease digestion following SBP2 knock-down, indicating an increased frequency of oxidative damage-induced lesions in the telomeric DNA in these cells. Together, these observations imply that selenoproteins may help protect telomeric reserve in mammalian cells.     

Prevalence of iodine deficiency in Europe in 2010

The adverse effects of iodine deficiency (ID)  intellectual impairment, damaged reproduction, goiter and hypo- and hyperthyroidism  are well known and easily corrected with salt iodization, but they continue to impair health and socioeconomic development, with more than two billion people at risk worldwide. During the major global expansion of salt iodization over the past four decades, much of Europe has remained iodine deficient. Although every European country endorsed the goal of eliminating iodine deficiency at the 1992 World Health Assembly, control of iodine deficiency has received low priority in much of Europe. However, there has been recent progress in the region and the number of children with low iodine intakes has decreased by ca. 30% since 2003. This paper presents an estimate of the prevalence of iodine deficiency in Europe in 2010, based on a systematic review to update the WHO Vitamin and Mineral Nutrition Information System (VMNIS) database.     

Kinetic evidence for modulation by glycophorin A of a conformational equilibrium between two states of band 3 (SLC4A1) bound reversibly by the competitive inhibitor DIDS

Recent evidence has suggested that erythrocytes naturally deficient in glycophorin A (GPA) have a reduced V_max for monovalent anion exchange. Unanswered is whether miss-folding of band 3 during biosynthesis, or the absence of GPA modulation of properly folded band 3 is responsible. Here, I determine the effect of selective depletion of GPA on the kinetics of reversible binding of the competitive transport inhibitor DIDS (4,4′-diisothiocyanato-2,2′-stilbenedisulfonate) to properly folded band 3. Reversible binding of DIDS follows biphasic kinetics: a fast phase {DIDS + band 3  (DIDS − band 3), k₁, k_− 1} and a slower phase {(DIDS − band 3)  (DIDS − band 3), k₂, k_− 2}. Selective depletion of GPA was accomplished by pretreating membranes with Triton X-100, over a range where erythrocyte hemolysis is inhibited by the detergent (0% to 0.03%, v/v). Pretreatment with sublytic Triton X-100: (a) virtually completely depleted GPA, (b) did not deplete membrane-bound band 3, and (c) slowed the overall rate of reversible binding of DIDS to band 3. Data analysis and model simulation studies indicated that the decrease in the rate of binding of DIDS was due exclusively to a decrease in k_− 2, with no change in the initial rate of binding. Thus, depletion of GPA does not alter the native conformation of band 3 at the DIDS binding site, but rather modulates a conformational equilibrium between two states of the binary complex formed by the competitive inhibitor DIDS, reversibly bound to properly folded band 3.     

N-RASMutations in Radiation-Induced Murine Leukemic Cells

ABSTRACT:N-rasmutations were examined in DNA samples extracted from the spleen of CBA/Ca mice that developed myeloid leukemia (ML) following exposure to radiations of different qualities. A total of 17 ML cases, i.e. 5 cases of neutron-induced and 12 cases of photon-(3 γ-ray and 9 x-ray) induced ML were included in the study along with 12 DNA samples from the bone marrow cells of control mice. Polymerase chain reaction-single strand conformational polymorphisms (PCR-SSCP) and the direct sequencing of PCR products were used to analyze three regions of theN-rasgene: (i) a 128 base-pair (bp) long portion of exon I (codons 2-37); (ii) a 103 bp long portion of exon II (codons 48-82); and (iii) a 107 bp long portion of exon III (codons 118-150). PCR-SSCP mobility shifts indicated mutations within only exon II of theN-rasgene. Such mutations were more prevalent in samples from mice exposed to fast neutrons. The exact type and location of these mutations were then determined by direct DNA sequencing. Silent point mutations, i.e. base transitions at the third base of codons 57 (GA→GA), 62 (CA→CA), or 70 (CA→CA) were present only in mice that developed ML after exposure to fast neutrons. A base transversion at the third base of codon 61 (CA→CA) was also observed in some ML cases. DNA sequencing demonstrated that ML samples contained normal as well as mutated DNA sequences. The higher frequency ofN-rasmutations in neutron-induced ML suggested that fast neutrons are more effective in inducing genomic instability at theN-rasregion of the genome. More importantly,N-rasmutations are not the initiating event in radiation leukemogenesis. This conclusion was supported by the finding thatN-rasmutations were detected only in mice with an overt leukemic phenotype but not in mice with minimal tissue infiltration of leukemic cells, suggesting that the disease may be present prior to the presence ofN-rasmutations. Alternatively,N-rasmay be present in these mice but a large number of normal spleen cells in these mice interferes with the detection of mutation in a small population of leukemic cells.     

Gallbladder emptying in vivo, bile composition, and nucleation of cholesterol crystals in patients with cholesterol gallstones

Impaired postprandial gallbladder emptying may provide time for progressive bile concentration with formation of instable cholesterol-rich vesicles and fast nucleation of cholesterol crystals. The aim of this study was to assess postprandial gallbladder emptying, bile composition, and nucleation of cholesterol crystals in the same patient. In 30 patients with cholesterol gallstones, postprandial gallbladder emptying was measured ultrasonographically. In each patient, gallbladder bile composition (obtained at cholecystectomy) and nucleation of cholesterol crystals was determined. Patients were divided in 22 strong contractors (>50% postprandial gallbladder emptying) and 8 weak contractors. In weak contractors, bile salt and phospholipid concentrations were much higher than in strong contractors ( and , respectively). Cholesterol concentrations were comparable in strong and weak contractors. Consequently, total lipid concentration was significantly higher (15.5 ± 1.4 and 9.2 ± 0.7 g/dL; P < 0.001) and cholesterol saturation index significantly lower (0.90 ± 0.08 and 1.61 ± 0.17; P < 0.001) in weak contractors. Nucleation time, percentage of cholesterol in vesicles, bile salt species, and molecular species of phosphatidylcholine were not significantly different. Differences in bile composition can be linked to different patterns of post-prandial gallbladder emptying and may point to two different pathways of gallstone formation.     

Histamine inhibits prostaglandin E2-stimulated rabbit duodenal bicarbonate secretion via H2 receptors and enteric nerves

The gastroduodenal epithelium is protected from acid peptic damage by an adherent mucus-bicarbonate layer. Bicarbonate is secreted by the surface epithelial cells into this mucus layer. Patients with duodenal ulcer disease have impaired proximal duodenal bicarbonate secretion. Mast cells, present in large numbers in the duodenal mucosa, release a number of inflammatory mediators, including histamine. Release of such mast cell mediators has been implicated in ulcer disease. In this study, the ability of histamine to regulate bicarbonate secretion was examined. Bicarbonate secretion by rabbit proximal duodenal mucosa was examined in vitro, and the effects of histamine, its agonists, and its antagonists were studied. Histamine essentially eliminated prostaglandin E₂-stimulated duodenal mucosal bicarbonate secretion, an effect reversed both by the neurotoxin, tetrodotoxin, and the histamine H₂-receptor antagonist, cimetidine, as well as reproduced by the H₂-receptor agonist, dimaprit. In addition to the stimulatory action of histamine on gastric acid secretion, histamine expresses an additional antidefensive action by inhibiting prostaglandin E₂-stimulated duodenal epithelial bicarbonate secretion. This effect of histamine is likely mediated via H₂ receptors located on enteric nerves.     

X region deletion mutants associated with surface antigen-positive hepatitis B virus infections

The finding of antibodies against the polymerase of hepatitis B virus in renal dialysis patients before the incubation phase of infection implies underlying virus replication. Hence, the aim of the study was to test for virus during infection. Viremia was assayed in virus-infected and control patients using the polymerase chain reaction and Southern blotting. Six months before the appearance of surface antigen, most patients had detectable core region, but few patients were X region positive. Three months after surface antigen appeared, most carriers had detectable core and X products. Three years after surface antigen appeared, 5 of 8 carriers with persistent hepatitis B e antigen and 1 of 8 carriers with corresponding antibody had these products. Cloning and sequencing showed deletions within the X/precore region of viral DNA. Infection with X region mutants precedes that of wild-type virus, and they reappear after wild-type virus is eliminated in carriers.     

Atrial fibrillation with left bundle branch block and intermittent right axis deviation during acute myocardial infarction

A survey of the litterature was made to precise the characteristics of infectious endocarditis (IE) in the elderly as defined by the diagnostic criteria of Von Reyn. The IE presents at times with a fallacious symptomatology where fever and cardiac bruit are often lacking. With regard to the cardiac disease, there is an increase in the incidence of calcifying degenerative valvulopathy responsible for IE. On the bacteriological point of view, streptococcal infections are still predominent in etiologgy for endocarditis but opportunistic infections dues to other bacterial species are increasing in frequency. Lastly, on the therapeutical side, there is a new trend to reduce the proeminence of curative treatment with aminoglycoside agents in favor of the antibiotic prophylaxis.