Sergej Nadalin

BanI polymorphism of cytosolic phospholipase A2 gene is associated with age at onset in male patients with schizophrenia and schizoaffective disorder

The enzymes phospholipases A2 are believed to be involved in the pathology of schizophrenia. We investigated allelic and genotype frequencies of PLA2G4A BanI polymorphism and the rs4375 in PLA2G6A in Croatian schizophrenic patients (n=81) and controls (n=182), using PCR/RFLP. Genotype and allelic frequencies of both loci, alone or in combination did not show significant difference (chi2-test). Allele-wise and genotype-wise meta-analyses of BanI polymorphism in case-control and family-based studies also revealed no significant association with schizophrenia. Multiple logistic regression analyses revealed statistically significant association between several items from PANSS general psychopathology scale and BanI polymorphism in PLA2G4A. BanI polymorphism further showed a significant impact on mean age of the onset of disease in males (betaA1=0.351, P=0.021; Spearmans rA1=0.391, P=0.010) indicating lower mean age at admission in homozygous A2A2 males.

The impact of PLA2G4A and PTGS2 gene polymorphisms, and red blood cell PUFAs deficit on niacin skin flush response in schizophrenia patients

We investigated the etiology of the attenuated niacin skin flush response in schizophrenia patients. Skin response to topical niacin of 0.1M, 0.01 M, 0.001 M, and 0.0001 M concentrations was rated using method of volumetric niacin response (VNR) and correlated to two functional A/G polymorphisms in genes: phospholipase A2 group IVA (BanI of the PLA2G4A), and rs689466 of the prostaglandin synthase-2 (PTGS2). We further tested the possible correlation between niacin response and fatty acid (FA) content of red blood cells (RBCs). We detected statistically significant but weak impact of both polymorphisms on niacin flush response in schizophrenia patients. The dosage of the G alleles of both polymorphisms was associated with higher VNR values, although each polymorphic variant accounted for only 1% of the overall flush response variability. Regarding FA content, both n-3 and n-6 polyunsaturated FAs (PUFAs) were significantly reduced in the patient group, but an association with niacin sensitivity was not detected.

The impact of hemochromatosis mutations and transferrin genotype on gonadotropin serum levels in infertile men

OBJECTIVE To address the possibility that HFE mutations and TF gene polymorphism cause dysfunction of spermatogenesis and/or the hypothalamic-pituitary-gonadal axis via contribution to long-term iron overload in the testes and brain. DESIGN Case-control and association study. SETTING Clinic of obstetrics and gynecology and university-based research laboratory. PATIENT(S) 127 infertile men (including 97 with idiopathic infertility) and 188 controls of proven fertility. INTERVENTION(S) Polymerase chain reaction/restriction fragment length polymorphism (PCR-RFLP). MAIN OUTCOME MEASURE(S) HFE mutations and transferrin allelic polymorphism, and testosterone, prolactin, and gonadotropin serum levels. RESULT(S) The frequencies of the analyzed alleles and genotypes showed no statistically significant difference between infertile men and controls. Sperm count and progressive sperm motility did not correlate with HFE or TF genotype, or their combination. After excluding patients with clinical hypogonadism or varicocele from further analysis, a statistically significant correlation between serum follicle-stimulating hormone and luteinizing hormone levels and the combined HFE H63D/TFC2 genotype was found in 97 men with idiopathic infertility. CONCLUSION(S) The combined HFE H63D/TF-C2 genotype contributed to 4.1% and 10.6% of follicle-stimulating hormone and luteinizing hormone variation, respectively, in infertile men, raising mean hormonal values above the normal physiologic range. Therefore, HFE and TF genes together may influence the hypothalamic-pituitary-gonadal axis, functioning at the pituitary or testes level.

Angiotensin-converting enzyme gene insertion/deletion polymorphism is not associated with schizophrenia in a Croatian population.

Because angiotensin converting enzyme (ACE) plays an important role in dopamine system functioning in the brain and the insertion/deletion (I/D) polymorphism of a 287 nucleotide fragment of the ACE gene correlates with enzyme activity, several studies have investigated the role of ACE I/D polymorphism in psychiatric diseases (Segman et al., 2002 ; Crescenti et al., 2009). Two recent studies yielded contradictory results: the D allele was identified as a protective factor in a Spanish population, while a protective effect toward schizophrenia and bipolar disorder was attributed to the I allele in a Turkish population (Crescenti et al., 2009 ; Kucukali et al., 2010). We tested whether schizophrenia risk was associated with ACE I/D polymorphism in a Croatian population, and examined its possible impact on schizophrenia symptom severity. Our study group consisted of 211 Croatian patients (115 male, 96 female) who met DSM-IV criteria for schizophrenia (n = 187) and schizoaffective disorder (n = 24), and 270 healthy blood donors (135 male, 135 female). All participants gave informed consent for the analysis. The study was approved by the Ethics Committee of the School of Medicine, University of Rijeka, Croatia. Genotyping was performed by polymerase chain reaction (PCR) as previously described (Rigat et al., 1990). To exclude mistyping of the heterozygotes as DD homozygotes, all DD genotype samples were confirmed with insertion-specific PCR (Shanmugan et al., 1993). The significance of differences in genotype and allele frequencies between patients and controls was determined using the χ2 test. Potential correlation between Positive and Negative Symptom Scale (PANSS) scores and I/D polymorphism was tested using linear regression analysis (stepwise selection), adjusted for age at PANSS assessment and sex, in patients with schizophrenia. PANSS evaluation was performed during the last hospitalization due to a psychotic episode. P-values < 0.01 were considered statistically significant. Allelic and genotypic frequencies of I/D polymorphisms were consistent with Hardy-Weinberg equilibrium and were not significantly different between groups. Allele frequencies were 230 (D) and 192 (I) in the patient group, and 285 (D) and 255 (I) in the control group. Genotype frequencies were 0.299 (DD = 63), 0.493 (ID = 104) and 0.208 (II = 44) in the patient group, and 0.274 (DD = 74), 0.507 (ID = 137) and 0.219 (II = 59) in the control group. Therefore, our data did not support results from Spanish or Turkish samples. However, after adjusting for sex and age at PANSS assessment we observed a significant correlation between ACE genotype and psychopathology evaluated by PANSS. Increased negative and total PANSS scores were significantly correlated with the number of D alleles (b = 0.26, F = 7.803 ; P = 0.006 and b = 0.29, F = 7.557 ; P = 0.002, respectively). Increased symptom severity of the general psychopathology scale in males with schizophrenia could also be attributed to the number of D alleles (b = 0.37, F = 9.910 ; P = 0.008). This is the first study reporting significant correlation between clinical expression of illness and I/D polymorphic variants. I/D polymorphism may affect symptom severity by modulating ACE expression/activity. Further studies in other populations/ethnic groups may help clarify the relationship between ACE activity and schizophrenia.

The impact of ACE gene I/D polymorphism on plasma glucose and lipid concentrations in schizophrenia patients.

A functional 287 nucleotide fragment insertion/deletion (I/D) polymorphism of the angiotensin converting enzyme (ACE) gene, by affecting ACE levels, may influence various pleiotropic effects of ACE. The I/D polymorphism has been extensively associated with the development of the metabolic syndrome and its separate components such as diabetes mellitus and dyslipidemia in the general population (Mittal et al., 2011). Since the ACE enzyme plays an important role in the functioning of the dopamine system in the brain, the I/D polymorphism has also been investigated in schizophrenia, yet with conflicting results (Crescenti et al., 2009; Nadalin et al., 2012; Hui et al., 2014; Zhang et al., 2014). Schizophrenia patients have a significantly higher risk of developing diabetes, dyslipidemia and obesity and it has been determined that treatment with atypical antipsychotic medications particularly contributes to abnormalities in the glucose and lipid metabolism in those patients (Kapur and Remington, 2001). In our recent study we found no association between the I/D polymorphism and schizophrenia risk in the Croatian population, but we revealed a significant polymorphisms impact on the clinical expression of the illness (Nadalin et al., 2012). We undertook the current study to determine whether and to what extent plasma glucose and lipid concentrations in schizophrenia patients may be influenced by ACE-I/D polymorphism. Our study group consisted of 211 chronically ill schizophrenia patients (115 males, 96 females) from the Department of Psychiatry of the Clinical Medical Centre in Rijeka, Croatia (N1⁄4110) and the Psychiatric Hospital in Rab, Croatia (N1⁄4101), aged 43.1710.8 years, all Croatian citizens. Diagnoses were assessed according to the Diagnostic and Statistical Manual of Mental Disorders (DSM-IV) criteria using the structural clinical interview. The investigation was carried out in accordance with the latest version of the Declaration of Helsinki. The study was approved by the Ethics Committee of the School of Medicine, University of Rijeka, Croatia. All patients gave informed consent to the analysis. Allele/genotype frequencies were determined by polymerase chain reaction/restriction fragment length polymorphism in the Laboratory for Molecular Genetics (Department of Biology and Medical Genetics, School of Medicine, Rijeka). Biochemical measurements (determination of fasting plasma glucose, total cholesterol, HDL cholesterol, LDL cholesterol and triglyceride levels) were carried out in the Department of Clinical Laboratory Diagnostics of the Clinical Medical Centre Rijeka. The difference between means of plasma glucose and lipid levels according to the ACE-I/D polymorphism was revealed by the one-way ANOVA analysis of variance (ANOVA). The association between glucose and lipid levels and ACE-I/D allele/genotype variants was tested using multiple stepwise regression analysis, adjusted for the patients age. Considering the possible interaction of the biochemical measurements with the patients gender, we performed separate analyses among males and females. Statistical analyses were conducted using Statistica for Windows, version 9 (StatSoft, Inc., Tulsa, OK, USA). The P values less than 0.05 (Po0.05) were considered statistically significant. The frequencies of the genotypes DD, ID and II were 0.299 (N1⁄463), 0.493 (N1⁄4104) and 0.208 (N1⁄444), and the frequencies of the D and I alleles were 0.545 (N1⁄4230) and 0.455 (N1⁄4192), respectively. We did not find significant gender differences neither in the frequencies of the genotypes DD, ID and II (males vs. females: 0.278, 0.470 and 0.252 vs. 0.323, 0.521 and 0.156) nor in the frequencies of the D and I alleles (males vs. females: 0.513 and 0.487 vs. 0.583 and 0.417) (P40.05). The multiple stepwise regression analysis revealed a significant association between the ACE genotype andmean plasma glucose levels in females (β1⁄4 0.60, F1⁄48.40, P1⁄40.005). Females heterozygous for the ACE genotype (ID; N1⁄450) had significantly higher glucose levels when compared to those DD homozygous (N1⁄431) and II homozygous (N1⁄415) (6.171.2 vs. 5.370.7 and 5.470.9; one-way ANOVA test: F1⁄43.84, Po0.026). The ACE genotype accounted for approximately 9% of plasma glucose levels variability (R change1⁄40.09). Furthermore, the ACE-ID polymorphism affected plasma triglyceride levels in males, although its impact only approached statistical significance (multiple stepwise regression analysis: β1⁄40.31, F1⁄43.92, P1⁄40.055). Males carrying the D allele in the ACE genotype (DD homozygous or ID heterozygous; N1⁄486) trended toward higher triglyceride levels than those II homozygous (N1⁄429) (2.371.4 vs. 1.570.8; one-way ANOVA test: F1⁄43.92, P1⁄40.055). This is the first report investigating the possible influence of ACE-I/ D polymorphism on plasma glucose and lipid levels in schizophrenia. Our results suggest gender dependent effect of the ACE-I/D polymorphism on glucose and lipid metabolism. Thus, males carrying the D allele in their ACE genotype (DD homozygous or ID heterozygous) trended toward an elevated risk for developing dyslipidemia. Furthermore, female patients heterozygous for the ACE genotype (ID) had significantly greater risk for diabetes mellitus than those DD homozygous and II homozygous. The strength of the observed association (P1⁄40.005) between glucose levels in female patients and ACE genotype is strong, and, to our opinion, the ACE genotype satisfactorily described plasma glucose levels (multiple stepwise regression: R change1⁄40.09). Intriguingly, this is also the first study reporting highest plasma glucose concentrations in ACE-ID heterozygous individuals while investigating the association between ACE-I/D polymorphism and glucose levels. Previous studies revealed highest plasma glucose levels in the ACE homozygous (DD) genotype as well as in the ACE homozygous (II) genotype, in patients with the metabolic syndrome (Alvarez-Aguilar et al., 2007; Mittal et al., 2011). Moreover, Zingone et al. (1994) determined the highest glucose levels in the ACE homozygous (DD) genotype among a random sample of male subjects from the general population. Thus, our results indicate that the mechanism by which the ACE may maintain glucose homoeostasis possibly differs in schizophrenia compared to

PPARα-L162V polymorphism is not associated with schizophrenia risk in a Croatian population

Disturbances of lipid and glucose metabolism have been repeatedly reported in schizophrenia. A functional L162V polymorphism in peroxisome proliferator-activated receptor alpha (PPARα) gene has been extensively investigated in etiology of abnormal lipid and glucose metabolism, yet not in schizophrenia. We determined whether the schizophrenia risk was associated with L162V polymorphism and we examined the impact of L162V variant on age of onset, and data of psychopathology scores. We also hypothesized that plasma glucose and lipid concentrations in patients may be influenced by L162V polymorphism. Genotype and allele frequencies between 203 patients and 191 controls did not differ significantly. Females heterozygous for the PPARα genotype (L162V) manifested significantly lower negative symptom scores, tended toward an earlier onset, and had significantly greater triglyceride levels. The PPARα-L162V polymorphism is not associated with schizophrenia risk in Croatian population, but it impacts clinical expression of the illness and plasma lipid concentrations in female patients.

An association between the PPARα-L162V polymorphism and nicotine dependency among patients with schizophrenia

OBJECTIVE Patients with schizophrenia are more likely to be smokers than the general population, which makes them an interesting group with which to study the etiology of nicotine dependency. We studied the prevalence of a gene variant of peroxisome proliferator-activated receptor alpha (PPARα) in schizophrenia, together with nicotine dependency, to investigate whether the PPARα-L162V polymorphism (rs1800206) influences nicotine dependency in schizophrenia. Given evidence suggesting that smoking influences the severity of schizophrenia, together with our recent data linking the PPARα-L162V polymorphism to clinical manifestations of schizophrenia (in the Croatian population), we hypothesized that interactions between the two (smoking and the PPARα-L162V polymorphism) might contribute to disease onset and scores for the Positive and Negative Syndrome Scale. To the best of our knowledge, this is the first study to investigate the possible associations between the PPARα gene and nicotine dependency. PATIENTS AND METHODS Genotyping was performed for 267 chronically ill schizophrenia patients (males/females: 140/127) by polymerase chain reaction. RESULTS A significant excess of PPARα-L162V genotypes and PPARα-162V alleles were detected among female smokers in comparison to female nonsmokers (18.2% vs. 2.0%, and 9.1% vs. 1.0%, p<0.01, respectively). We also revealed a significant PPARα genotype-smoking interaction that predicted positive symptom severity among male patients (F=4.43, p<0.05). These data indicated that the PPARα-L162V heterozygous genotype, depending on smoking status, might be of relevance as either protective, or a risk factor, for the severity of positive symptoms. No interaction between the PPARα-L162V polymorphism and smoking for the time of onset of schizophrenia was detected (p>0.05, respectively). CONCLUSION We demonstrated two significant yet weak effects. The first showed an effect of the PPARα-L162V polymorphism on the risk of nicotine dependency. The second linked the PPARα genotype-smoking interaction to positive symptoms severity among schizophrenia patients; both effects manifested in a gender-specific fashion.

The advantages of humeral anteromedial plate osteosynthesis in the middle third shaft fractures

ZusammenfassungHINTERGRUND: 1,2 % aller Frakturen entfallen auf Humerusschaftfrakturen, die unter etwas jüngeren Altersgruppen verzeichnet werden. Stolpern, Abstürze oder Verkehrsunfälle zählen zu den Ursachen, obwohl diese Frakturen auch pathologisch bedingt sein können. Um die Frage zu klären, welche von den zwei chirurgischen Behandlungen, die in unserer Abteilung für Behandlung von Humerusschaftfrakturen durchgeführt werden mehr Vorteile aufweist (anterolaterale oder anteromediale Platte durch anterolaterale Methode), haben wir die Inzidenz der postoperativen iatrogenischen Radialislähmungen und die jeweilige durchschnittliche Operationsdauer analysiert. METHODEN: Im Zeitraum von Januar 1992 bis Dezember 2009 wurden an der Traumatologie der chirurgischen Abteilung des Klinikzentrums Rijeka (KBC Rijeka) Frakturen im mittleren Drittel des Humerusschaftes von 420 Patienten (340 männliche und 80 weibliche Patienten im Durchschnittsalter von 38,11 ± 9,29 Jahren) behandelt. Dabei wurde die anterolaterale Methode und die interne Fixation mit Hilfe von AO/DCP oder LCP-Platten angewandt, die bei 141 Patienten (33,57 %) auf die anteromediale und bei 279 Patienten (66,43 %) auf die anterolaterale Humerusfläche gelegt wurde. ERGEBNISSE: Kein Patient, bei dem die Platte in der Osteosynthese auf der anteromedialen Humerusfläche angewandt wurde, hatte Läsion des Radialnervs. Demzufolge zeigte der χ2 Test eine erheblich höhere Häufigkeit von postoperativen Verletzungen des Radialnervs bei denjenigen Patienten, bei den die anterolaterale Platte angewandt worden ist als bei denjenigen, bei denen die anteromediale Platte gelegt worden ist (χ2 = 17,51; p < 0,05). Die Anwendung der anterolateralen Platte benötigte eine längere durchschnittliche Operationszeit als die der anteromedialen Platte. Die unterschiedliche Dauer, bestimmt durch den t-Test für unabhängige Muster, zeigte einen statistisch bedeutenden Unterschied. (t= 14,57; p< 0,05). SCHLUSSFOLGERUNG: Die Anwendung der anteromedialen Platte bei Humerusschaftfrakturen durch anterolaterale Methode hat sich als einfache, sichere, wirksame und als schnelle chirurgische Behandlung erwiesen. Demzufolge wird unsererseits diese Operationstechnik zur Behandlung von Humerusschaftfrakturen nachdrücklich empfohlen.SummaryBACKGROUND: Humeral shaft fractures account for 1.2% of all fractures and occur in a slightly younger population. Their causes include a fall from standing or from height, motor vechicle accident, but can be also pathological. In order to clarify which of both surgeries we performed in our Department for treating humeral shaft fractures had more advantages (anterolateral or anteromedial plating through anterolateral approach) we analyzed incidence of postoperative iatrogenic radial palsies and mean operation time required to complete each surgery. METHODS: During January 1992 to December 2009 on Department of Surgery, Division for Traumatology of Clinical Hospital Center Rijeka, 420 patients (340 males and 80 females with mean age of 38.11 ± 9.29 years) were treated for middle third humeral shaft fracture by anterolateral approach and internal fixation using AO/DCP or LCP plates that was positioned on anteromedial humeral surface in 141 patients (33.57%) and on anterolateral humeral surface in 279 patients (66.43%). RESULTS: None of the patients who had osteosynthesis by using plate on anteromedial humeral sufrace had lesions of the radial nerve. Therefore, χ2 test revealed significantly higher frequency of postsurgical radial nerve injuries in patients who were treated by anterolateral plating than in patients where anteromedial plating was performed (χ2 = 17.51; p< 0.05). Anterolateral plating required longer mean operation time than anteromedial plating and the difference in its duration determined by t-test for independent samples showed statistically significant difference (t= 14.57; p< 0.05). CONCLUSION: An anteromedial plating of humeral shaft fractures through anterolateral approach was determinated to be a simple, safe, effective and also fast surgical treatment and we highly recommend it as operative technique for treating humeral shaft fractures.

Polymorphisms in PLA2G6 and PLA2G4C genes for calcium-independent phospholipase A2 do not contribute to attenuated niacin skin flush response in schizophrenia patients

We hypothesized that attenuated niacin skin flushing in schizophrenia patients might be associated with polymorphic variants in PLA2G6 and PLA2G4C genes (rs4375 and rs1549637 variations) which encode calcium-independent phospholipase A2 beta (iPLA2β) and cytosolic phospholipase A2 gamma (cPLA2γ) enzymes. The iPLA2β and cPLA2γ may play an important role in niacin-mediated signaling; in addition to their major role - mediating phospholipids remodeling, which alters membrane receptors and signal transduction, they regulate the reservoir of arachidonic acid for prostaglandins synthesis. Skin response to topical niacin of 0.1M, 0.01M, 0.001M and 0.0001M concentrations in 75 schizophrenia patients was rated using the method of volumetric niacin response (VNR). Neither PLA2G6 nor PLA2G4C gene polymorphisms were significantly associated with VNR values. Furthermore, polymorphisms׳ synergy on niacin skin flushing was also not detected.

An association between the BanI polymorphism of the PLA2G4A gene for calcium-dependent phospholipase A2 and plasma glucose levels among females with schizophrenia

Abnormal glucose and lipid metabolism may be associated with altered cytosolic Ca2+-dependent phospholipase A2 (cPLA2) signaling in patients with schizophrenia. The relationship between schizophrenia and the functional BanI polymorphism (rs10798059 variant, A/G polymorphism) of the PLA2G4A gene for cPLA2 has been extensively investigated. We previously reported that it can influence several clinical features of schizophrenia, and it was shown to contribute to schizophrenia risk in several population studies. We performed PCR/RFLP genotyping of 263 Croatian patients (males/females: 139/124) to investigate the relationship between the BanI polymorphism and fasting plasma glucose and lipid levels in patients with schizophrenia. Our results indicate that the BanI polymorphic variant contributes significantly to plasma glucose levels in female patients. Females carrying the PLA2G4A-G allele (PLA2G4A-GG homozygous and PLA2G4A-AG heterozygous) presented with lower glucose levels than PLA2G4A-AA homozygous carriers, and the PLA2G4A genotype contributed approximately 6% of plasma glucose level variability in this group of patients.