CFTR gene analysis in patient with atypical cystic fibrosis

U ovom radu je prikazan slučaj atipične cistične fibroze sa graničnom vrednošću znojnog testa. U većini slučajeva znojni test je glavni dijagnostički parametar za dijagnostikovanje cistične fibroze, ali se dijagnoza može potvrditi samo na osnovu rezultata molekularno-genetičkog testiranja. Utvrđeno je da je pacijent složeni heterozigot za dve CFTR mutacije, F508del i D1152H. Prisustvo mutacije F508del detektovano je PSM metodom, dok je za analizu prisustva druge mutacije korišćena DGGE metoda. Strategija detekcije mutacija kod pacijenata sa cističnom fibrozom, naročito onih sa atipičnim prezentacijama bolesti koji nose ređe mutacije, trebalo bi da uključuje i direktne i indirektne metode molekularne dijagnostike.This paper reports a case of a patient presenting with atypical cystic fibrosis whose sweat test shows borderline values. In vast majority of cases the sweat test is essential diagnostic tool for establishing the diagnosis of cystic fibrosis, but only after the molecular genetic testing the diagnosis can be confirmed. The patient was found to be compound heterozygote for two CFTR mutations, F508del and D1152H. The presence of F508del mutation was analyzed by PSM method, while the screening for the second mutation was performed using DGGE. The strategy of mutation detection in cystic fibrosis patients, especially those with atypical presentations who carry less frequent mutations, should include both direct and indirect methods of molecular diagnostics

Association of functional variants of phase I and II genes with chronic obstructive pulmonary disease in a Serbian population

Uvod: Hronična opstruktivna bolest pluća (HOBP) jeste složeno oboljenje koje karakteriše povišen oksidativni stres. Funkcionalne varijante gena faze I i II ksenobiotičkog metabolizma mogu uticati na ravnotežu oksidanti-antioksidanti i mogu dovesti do razvoja HOBP. Cilj ove studije je bio ispitivanje uloge funkcionalnih genskih varijanti u genima za citohrom P450 (CYP), glutation S-transferazu (GST) i mikrozomalnu epoksidnu hidrolazu (mEH) u patogenezi HOBP u srpskoj populaciji. Metode: U ovoj studiji analizirane su genske varijante CYP1A1 *1A/*2A, CYP2E1 *1A/*5B, GSTM1 null, GSTT1 null, GSTP1 Ile105Val, mEH Tyr113His i mEH His139Arg u grupi obolelih od HOBP koja je obuhvatala 122 ispitanika i kontrolnoj grupi koja je obuhvatala 100 ispitanika sa normalnom funkcijom pluća. Rezultati: Dobijeni rezultati su pokazali da je GSTM1 null varijanta statistički značajno povišena u grupi obolelih od HOBP u poređenju sa kontrolnom grupom (61,5% i 47,0%; or = 1,80; p = 0,042). Takođe, uočena je značajna razlika u zastupljenosti kombinacije genotipova GSTM1 null i GSTP1 105Val/(Val) (38,5% i 24,0%; or = 1,98; p = 0,029), kao i kombinacije CYP1A1 *1A/*2A, GSTM1 null i mEH 113H is/(H is) (7,4% i 1,0% ; or = 7 ,88; p = 0,025). Zaključak: Ovo su prvi podaci o ulozi genskih varijanti gena faze I i II u patogenezi HOBP u srpskoj populaciji. Rezultati dobijeni u ovoj studiji otvaraju mogućnost za detaljniju analizu uloge genetičkih faktora u HOBP na većim grupama ispitanika. Pored toga, podaci dobijeni u našoj studiji potvrđuju važnost genetičkih determinanti povezanih sa HOBP u prethodnim studijama, ali takođe otkrivaju nove genetičke faktore, koji nisu objavljeni do sada.Background: Chronic obstructive pulmonary disease (COPD) is a complex disorder characterized by increased oxidative stress. Functional genetic variants of phase I and II genes are implicated in oxidants-antioxidants imbalance and may be involved in COPD development. In this study, we aimed to investigate the role of cytochrome P450 (CYP), glutathione S-transferase (GST) and microsomal epoxide hydrolase (mEH) functional variants in the pathogenesis of COPD in a Serbian population. Methods: The genotypes of l2 2 COPD patients and 100 controls with normal lung function were determined for CYP1A1 *1A /*2A , CYP2E1 *1 A /*5B , GSTM1 null, GSTT1 null GSTP1 Ile105Val, mEH Tyr113His and mEH His139Arg gene variants. Results: Results obtained showed that GSTM1 null variant was significantly more represented in COPD patients than in controls (61.5% vs. 47.0%; or = 1.80; p = 0.042). Also, a significant difference was observed for combinations of GSTM1 null and GSTP1 105Val/(Val) (38.5% vs. 24.0%; or = 1.98; p = 0.029), as well as for CYP1A1 *1A/*2A, GSTM1 null and mEH 113His/(His) genotypes (7.4% vs. 1.0%; or = 7.88; p = 0.025). Conclusions: These are the first data concerning the analysis of the variants of phase I and II genes in the pathogenesis of COPD in a Serbian population. Results obtained in this study open up the possibility for thorough analyses of the role of genetic factors in COPD on larger cohorts. Also, they implicate the importance of previously described genetic associations with COPD in our population, as well as reveal a new one, not reported so far

Koncentracija magnezijuma u serumu i urinu kod bolesnika sa akutnim i hroničnim plućnim bolestima

In this study we determined magnesium concentration in serum and in 24-hour urine, at the start (To) and at the end of treatment (T1), in 56 patients with acute pulmonary disease (B1) and in 58 patients with chronic obstructive pulmonary disease - COPD (B2). In group B1 there was disbalance of Mg in serum in 14-25% patients at the start of treatment (To) which decreased significantly at the end of treatment (T1) and persisted in 4-7.1% patients (p lt 0.05). In group B2 distribution of normal, decreased and increased values of Mg in serum was similar in patients in period To and T1 (p > 0.05). In group B1, 9 (16.1%) patients had hypomagnesemia at the start of treatment (To), which was accompanied by increased concentration of Mg in 24-hour urine of only 4 (7.2%) patients. There is a possibility that there was extrarenal elimination of Mg in patients with acute pulmonary disease or there was some kind of transcellular distribution. In group B2 in period To, there was proportional ratio between hypomagnesemia (12-20.7% patients) and increased concentration of Mg in 24-hour urine (20 -34.5% patients). This could be because of renal loss. Simultaneous determination and follow up of magnesium in serum and in 24-hour urine can give us reliable information about homeostasis of this electrolyte in acute and chronic pulmonary diseases.U ovom radu određivana je koncentracija magnezijuma u serumu i u 24-urinu, na početku (To) i na kraju hospitalnog lečenja (T1) kod 56 bolesnika sa akutnim plućnim bolestima (B1) i kod 58 sa hroničnim plućnim bolestima (B2). U grupi B1 postojao je disbalans Mg u serumu kod 14-25% bolesnika na početku lečenja (To) koji se značajno smanjio na kraju lečenja (T1) i postojao je kod 4-7,1% bolesnika (p lt 0,05). U grupi B2 distribucija normalnih, snizenih i povišenih vrednosti Mg u serumu bila je slična u periodu To i T1 (p> 0,05). Hipomagnezemiju u grupi B1 imalo je 9 (16,1%) bolesnika na početku lečenja (To) što je bilo praćeno povećanom koncentracijom Mg u 24-časovnom urinu samo kod 4 (7,2%) bolesnika. Ovo je bilo moguće zbog ekstrarenalnog gubitka elektrolita ili je došlo do transcelularne preraspodele. U grupi B2 u periodu To postojao je proporcionalni odnos hipomagnezemije (12,0-20,7% bolesnika) sa povećanom koncentracijom Mg u 24-časovnom urinu (20,0-34,5% bolesnika). Ovo je bilo moguće zbog renalne eliminacije elektrolita. Istovremeno određivanje i praćenje magnezijuma u serumu i 24-časovnom urinu daje pouzdane informacije o homeostazi ovog elektrolita kod akutnih i hroničnih plućnih bolesti

Alpha-1-Antitrypsin Deficiency in Serbian Adults with Lung Diseases

Aim: Alpha-1-antitrypsin (A1AT) is the main inhibitor of neutrophil elastase, and severe alpha-1-antitrypsin deficiency (A1ATD) is a genetic risk factor for early-onset emphysema. Despite the relatively high prevalence of A1ATD, this condition is frequently underdiagnosed. Our aim was to determine the distribution of the A1ATD phenotypes/alleles in patients with lung diseases as well as in the Serbian population. Methods: The study included the adults with chronic obstructive pulmonary disease (COPD) (n = 348), asthma (n = 71), and bronchiectasis (n = 35); the control was 1435 healthy blood donors. The A1ATD variants were identified by isoelectric focusing or polymerase chain reaction-mediated site-directed mutagenesis. Results: PiMZ heterozygotes, PiZZ homozygotes, and Z allele carriers are associated with significantly higher risk of developing COPD than healthy individuals (odds ratios 3.43, 42.42, and 5.49 respectively). The calculated prevalence of PiZZ, PiMZ, and PiSZ was higher in patients with COPD (1:202, 1:8, and 1:1243) than in the Serbian population (1: 5519, 1: 38, and 1:5519). Conclusion: The high prevalence of A1ATD phenotypes/allele in our population has confirmed the necessity of screening for A1ATD in patients with COPD. On the other hand, on the basis of the estimated number of those with A1ATD among the COPD patients, it is possible to assess the diagnostic efficiency of A1ATD in the Serbian population

Alpha-1-Antitrypsin Deficiency in Serbian Adults with Lung Diseases

Aim: Alpha-1-antitrypsin (A1AT) is the main inhibitor of neutrophil elastase, and severe alpha-1-antitrypsin deficiency (A1ATD) is a genetic risk factor for early-onset emphysema. Despite the relatively high prevalence of A1ATD, this condition is frequently underdiagnosed. Our aim was to determine the distribution of the A1ATD phenotypes/alleles in patients with lung diseases as well as in the Serbian population. Methods: The study included the adults with chronic obstructive pulmonary disease (COPD) (n = 348), asthma (n = 71), and bronchiectasis (n = 35); the control was 1435 healthy blood donors. The A1ATD variants were identified by isoelectric focusing or polymerase chain reaction-mediated site-directed mutagenesis. Results: PiMZ heterozygotes, PiZZ homozygotes, and Z allele carriers are associated with significantly higher risk of developing COPD than healthy individuals (odds ratios 3.43, 42.42, and 5.49 respectively). The calculated prevalence of PiZZ, PiMZ, and PiSZ was higher in patients with COPD (1:202, 1:8, and 1:1243) than in the Serbian population (1: 5519, 1: 38, and 1:5519). Conclusion: The high prevalence of A1ATD phenotypes/allele in our population has confirmed the necessity of screening for A1ATD in patients with COPD. On the other hand, on the basis of the estimated number of those with A1ATD among the COPD patients, it is possible to assess the diagnostic efficiency of A1ATD in the Serbian population

Evaluation of magnesium in serum and urine in patients with pulmonary diseases

We evaluated magnesium (Mg) in serum and 24-hour urine in patients with acute and chronic pulmonary diseases. Mg was determined in 114 patients, 56 with acute pulmonary diseases (group I) and 58 patients with acute exacerbation of chronic obstructive pulmonary disease (group II), at the start (To) and at the end of hospital treatment (T1). In group I, in period To, there were disturbances of Mg in serum in 14 patients (25%) which decreased in period T1 and persisted in 4 patients (7.1%) (p lt 0.05). In group II the distribution of normal, decreased and increased Mg in serum was similar in periods To and T1 (p > 0.05). Hypomagnesemia was found in 9 patients (16.1%) in group I at the start of treatment (To), with accompanying increased Mg in 24-hour urine in only 4 patients (7.2%). Extrarenal elimination of Mg or transcellular distribution was a possibility. In group II in period To there was a proportional ratio between hypomagnesemia (12-20.7% patients) and increased concentration of Mg in 24-hour urine (20-34.5% patients) probably due to renal loss. Simultaneous determination and follow-up of Mg in serum and in 24-hour urine can give information about electrolyte disturbances in acute and chronic pulmonary diseases

Association of Functional Variants of Phase I and II Genes with Chronic Obstructive Pulmonary Disease in a Serbian Population / Udruženost Funkcionalnih Varijanti Gena Faze I I Ii Sa Hroničnom Opstruktivnom Bolešću Pluća U Srpskoj Populaciji

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is a complex disorder characterized by increased oxidative stress. Functional genetic variants of phase I and II genes are implicated in oxidants–antioxidants imbalance and may be involved in COPD development. In this study, we aimed to investigate the role of cytochrome P450 (CYP), glutathione S-transferase (GST) and microsomal epoxide hydrolase (mEH) functional variants in the pathogenesis of COPD in a Serbian population. METHODS: The genotypes of 122 COPD patients and 100 controls with normal lung function were determined for CYP1A1 *1A/*2A, CYP2E1 *1A/*5B, GSTM1 null, GSTT1 null GSTP1 Ile105Val, mEH Tyr113His and mEH His139Arg gene variants. RESULTS: Results obtained showed that GSTM1 null variant was significantly more represented in COPD patients than in controls (61.5% vs. 47.0%; OR=1.80; p=0.042). Also, a significant difference was observed for combinations of GSTM1 null and GSTP1 105Val/(Val) (38.5% vs. 24.0%; OR=1.98; p=0.029), as well as for CYP1A1 *1A/*2A, GSTM1 null and mEH 113His/(His) genotypes (7.4% vs. 1.0%; OR=7.88; p=0.025). CONCLUSIONS: These are the first data concerning the analysis of the variants of phase I and II genes in the pathogenesis of COPD in a Serbian population. Results obtained in this study open up the possibility for thorough analyses of the role of genetic factors in COPD on larger cohorts. Also, they implicate the importance of previously described genetic associations with COPD in our population, as well as reveal a new one, not reported so far

Gene-environment interaction between the MMP9 C-1562T promoter variant and cigarette smoke in the pathogenesis of chronic obstructive pulmonary disease

The aetiology of chronic obstructive pulmonary disease (COPD) is complex. While cigarette smoking is a well-established cause of COPD, a myriad of assessed genetic factors has given conflicting data. Since gene-environment interactions are thought to be implicated in aetiopathogenesis of COPD, we aimed to examine the matrix metalloproteinase (MMP) 9 C-1562T (rs3918242) functional variant and cigarette smoke in the pathogenesis of this disease. The distribution of the MMP9 C-1562T variant was analyzed in COPD patients and controls with normal pulmonary function from Serbia. Interaction between the C-1562T genetic variant and cigarette smoking was assessed using a case-control model. The response of the C-1562T promoter variant to cigarette smoke condensate (CSC) exposure was examined using a dual luciferase reporter assay. The frequency of T allele carriers was higher in the COPD group than in smoker controls (38.4% vs. 20%; OR=2.7, P=0.027). Interaction between the T allele and cigarette smoking was identified in COPD occurrence (OR=4.38, P=0.005) and severity (P=0.001). A functional analysis of the C-1562T variant demonstrated a dose-dependent and allele-specific response (P lt 0.01) to CSC. Significantly higher MMP9 promoter activity following CSC exposure was found for the promoter harboring the T allele compared to the promoter harboring the C allele (P lt 0.05). Our study is the first to reveal an interaction between the MMP9-1562T allele and cigarette smoke in COPD, emphasising gene-environment interactions as a possible cause of lung damage in the pathogenesis of COPD. Environ. Mol. Mutagen. 57:447-454, 2016

Alpha-1-antitrypsin Phenotypes and Neutrophil Elastase Gene Promoter Polymorphisms in Lung Cancer

Imbalance between neutrophil elastase and alpha-1-antitrypsin (AAT) leads to emphysema in smokers as well as in patients with inherited alpha-1-antitrypsin deficiency. AAT as a proven inhibitor of apoptosis may play role in lung cancer (LC) progression. The aim was to analyse AAT protein variants and polymorphism in promoter region of the neutrophil elastase gene (ELA2) in patients with primary lung cancer. AAT phenotypisation by isoelectric focusing method and ELA2 gene promoter characterization by DNA sequencing were performed in 66 patients with primary lung cancer. Results showed that the frequency of M1 allele and PiM1 homozygotes in LC patients was significantly higher when compared to the healthy subjects (f = 0.6360 and 0.7424 respectively). The most frequent ELA2 promoter region genotypes in LC patients were -903TT and -741GG. There were significantly more patients with intermediate and high ELA2 genotype activity, compared to those with low activity (91% vs. 9%, respectively). In conclusion, we found that PiM1 homozygosity could be associated with the lung cancer, probably due to increased synthesis of this antiapoptotic protein. Non-MM variants of AAT and ELA2 genotypes with predicted intermediate or high activity could also represent a risk factor for aggressive form of lung cancer associated with extrathoracic metastases

Matrix Metalloproteinases Gene Variants in Idiopathic Disseminated Bronchiectasis

Background: The excess of matrix metalloproteinases (MMPs) might be associated with the airways destruction or dilatation in bronchiectasis. The functional promoter polymorphisms of MMPI and MMP9 genes, involved in the extracellular matrix remodeling, might increase the expression of MMPs leading to the development of bronchiectasis. Methods: Detection of MMP1 G-1607GG and MMP9 C-1562T gene variants was performed on 37 patients with idiopathic disseminated bronchiectasis and 102 control subjects. We also described a novel method for simple and rapid detection of MMP1 G-1607GG polymorphism. Results: The frequency of -1607GG allele was significantly higher in the group of patients than in control subjects (P = 0.014). The heterozygote genotype showed association with bronchiectasis (odds ratio, 5.3; 95% confidence intervals, 1.4-20.0). The association was even stronger in homozygotes for -1607GG allele (odds ration, 8.7; 95% confidence intervals, 1.9-41.0). The allelic and genotype frequencies of MMP9 C-1562T variant did not show significant differences between the groups. Conclusions: This is the first report concerning a role of MMP1 G-1607GG and MMP9 C-1562T variants in pathogenesis of idiopathic disseminated bronchiectasis. The results of our study revealed the association of -1607GG allele and the lack of association of MMP9 C-1562T variant with the disease