Dissertations / Theses on the topic 'Gene polymorphisms'

Systemic lupus erythematosus (SLE) is an autoimmune disease, with a strong genetic component. It is characterised by hyperactive T and B cells, chronic inflammation and the production of antinuclear autoantibodies. SLE affects mostly women of child baring age, with a 9:1 ratio, women to men and has been reported to be more prevalent in people of non-European ancestry. In the era of genome-wide association studies (GWAS), elucidating the genetic factors present in SLE has been very successful, with over 28 confirmed disease susceptibility loci mapped and a number of candidate genes identified. During this thesis I fine mapped IL18 as it had previously been reported to be associated with SLE, SNP rs360719. After fine mapping and subphenotype analysis in UK and African American cohorts, I was unable to replicate the published association. Although genetic data did not confirm IL18 to be associated with SLE, I demonstrated increased IL-18 serum levels in SLE renal patients compared to SLE patients. I further analysed IL10, another previously associated SLE candidate loci in our current SLE GWAS cohort (4000 cases and 9000 controls) of European ancestry. I again was unable to replicate the previous association, however using other SLE GWAS data showed SNP rs3024505 to be associated in Northern European samples. Further analysing our SLE GWAS, I located IKZF3 as a candidate loci. I identified an associated block of 56 SNPS and located the association to a single SNP rs2941509, p=1.46xlQ-8. Furthermore, I demonstrated an allelic imbalance in this SNP, with the protective G allele being expressed 1.5 times greater than the risk A allele, in controls. These data here demonstrated in this thesis indicates the importance of fine mapping candidate loci and verification of previously associated loci. This thesis contributes to the current knowledge of SLE by demonstrating discrepancies in published association data and showing the importance of larger studies.

Experimental evidence has implicated a function for TGF-β1 in many pathological processes, including atherosclerosis, cancer, fibrotic diseases and osteoporosis. In this study 1115 bp of the TGF-β1 promoter, the 5' untranslated region and exons 1, 2 and 3 were screened by single stranded conformational polymorphism analysis. Four polymorphisms were identified in the TGF-β1 gene at -800 and -509 of the promoter and at +869 (codon 10) and +914 (codon 25) of exon 1. A simple non-radioactive assay was developed for each of these polymorphisms and also for a polymorphism located at +1628 (exon 5, codon 263) which was reported in the literature. These polymorphisms were analysed in a study of postmenopausal female twins and the -800, -509 and +869 polymorphisms were associated with the concentration of TGF-β1 in the serum detected by ELISA. The TGF-β1 promoter polymorphisms (-509 and -800) were associated with increased and decreased serum TGF-β1 respectively in the twin study. These promoter isoforms were assayed in vitro to establish whether there was a functional effect of the polymorphisms on the basal activity of the TGF-β1 promoter. However, the activity of the TGF-β1 promoter was weak, and in the cell line studied no difference in activity between the isoforms of the TGF-β1 promoter was observed. The five TGF-β1 polymorphisms were analysed in 457 subjects recruited to the St. George's Heart Disease study. None of the polymorphisms showed any association with either coronary artery disease or hypertension in this study. However, strong linkage disequilibrium was observed at the TGFB1 locus and haplotypes of the locus were also considered in this study. The haplotype G(-800)C(-509)T(+869)G(+914)C(+1628) arose more frequently in the control subjects than the coronary artery disease cases (p=0.024), suggesting that it may be cardioprotective. The -509 and -800 TGF-β1 promoter polymorphisms were analysed in subjects of a breast cancer study. Subjects homozygous for the -800A allele appeared to have an increased risk for breast cancer in this study (relative risk 3.9, 95% confidence interval 0.90-17), but few homozygotes were observed and this risk was not insignificant.

Beta-2-Adrenergic receptors (ADRB2) participate in the physiologic responses of the lung, including bronchodilation and bronchoprotection, through mechanisms such as mucociliary clearance, fluid accumulation and mediator release from mast cells and basophils. Thus, these receptors may also play an important role in the pathophysiology of asthma. The gene encoding ADRB2 is extremely polymorphic, and studies of this gene improves our understanding of asthma and possibly lead to new methods to prevent, diagnose and treat it. When you are citing the document, use the following link http://essuir.sumdu.edu.ua/handle/123456789/33568

Thesis (Ph.D. in Epidemiology) -- University of Colorado at Denver and Health Sciences Center, 2005.
Typescript. Includes bibliographical references (leaves 95-118). Free to UCDHSC affiliates. Online version available via ProQuest Digital Dissertations;

Abstract Cytochrome P450 2A6 (CYP2A6) is involved in the 7-hydroxylation of coumarin, C-oxidation of nicotine, and the metabolism of tobacco specific nitrosamines. Initially in 1995 Fernandez-Salguero et al. reported a genotyping method for three alleles: CYP2A6*1 (wild-type), CYP2A6*2 (variant 1), and CYP2A6*3 (variant 2). Later studies presented in this thesis indicated that the original genotyping method produces erroneous results for the CYP2A6*3 allele due to unspecific PCR conditions and previously unknown CYP2A6*1B allele. Furthermore, the CYP2A6*2 allele genotyping caused erroneous genotypes (CYP2A6*2/*2 was misclassified as CYP2A6*1/*2). In this work, new PCR based genotyping methods were developed for CYP2A6*2 and for several new alleles (CYP2A6*1B, CYP2A6*4A/*4D and CYP2A6*5). In population-based studies, the deletion alleles (pooled as CYP2A6*4) turned out to be more prevalent among Asians (15.1%) than Caucasians (0.5%). The frequencies of the other inactive alleles varied within 0–3% in both populations. Asians totally lacked the CYP2A6*2 allele, whereas Caucasians lacked the CYP2A6*5 allele. The frequencies of two wild-type alleles, CYP2A6*1A and CYP2A6*1B alleles were 66.5% and 30.0% in Caucasians, and 43.2% and 40.6% in Asians, respectively. Correlation studies between the phenotype, as tested by the administration of coumarin, and the genotype demonstrated that individuals with the CYP2A6*2/*2 genotype were totally defective, while CYP2A6*1/*2 subjects exhibited intermediate and CYP2A6*1/*1 subjects full capablility of producing 7-hydroxycoumarin. Upon phenotyping with nicotine, individuals with the CYP2A6*1/*2 or CYP2A6*1/*4 genotype were shown to have a lower enzyme activity (one fourth of the normal activity), compared to those with the CYP2A6*1/*1 genotype. Defective CYP2A6 activity has been hypothesised to reduce the risk of environmentally (especially tobacco smoke) induced diseases either by decreasing production of genotoxic metabolites or by preventing addiction to tobacco smoking. However, in our case-control studies on Spanish patients with liver cirrhosis (n = 83) and liver cancer (n = 90) and their controls (n = 237) no significant association between the CYP2A6 genotypes and disease proneness was found. The odds ratio (OR) for developing liver cancer was was 1.4 (95% confidence interval [CI] 0.5–3.7) for genotypes containing at least one CYP2A6*2 allele. For liver cancer the respective OR was 1.3 (95% CI 0.4–4.5). Similarly, no statistically association between CYP2A6 alleles and the risk of lung cancer was observed in our Finnish study population cinsisting of 177 cases and 1089 controls; the OR for combined CYP2A6 variant allele containing genotypes (CYP2A6*1/*2 and CYP2A6*1/*4) was 1.19 (95% CI 0.56–2.45). Our studies therefore do not indicate any major modifying role for the CYP2A6 genotypes in individual susceptibility to environmentally induced diseases.

Calpastatin is widely known as an endogenous specific inhibitor to the ubiquitously expressed calpain an enzyme responsible for proteolysis of myofibrillar proteins during post-mortem degradation of muscle. The presence of the calpastatin polypeptide in muscle indicates that the activity of calpain can be potentially down regulated which could result in meat toughness. Asssement of calpastatin activity in meat could be a predictive marker to meat tenderness and variation in the gene has the potential to become a candidate genetic marker which is associated with meat tenderness. The variability and inconsistency produced in meat tenderness post-mortem could be reduced if animals could be selected based on this potential genetic marker prior to slaughter which in turn will reduce the cost in meat processing and ultimately achieve the main objective of producing consistently tender meat. Previous studies have successfully sequenced bovine calpastatin cDNA and found that a series of promoters in the 5’ region are responsible for transcribing Type I, II and III mRNA for calpastatin. The presence and length of CA tandem repeat sequence 5’ to the transcription start site of Type I calpastatin mRNA is believed to play a significant role in regulating the transcriptional activity of this promoter. This thesis investigated the hypothesis that there was a relationship between length polymorphisms of CA repeat located 5’ to the promoter region of Type I bovine calpastatin which altered the level of calpastatin transcripts and ultimately influenced meat shear force value due to the variation in calpain inhibition. Apart from this, transcriptional activity of promoter for Type I, II, and Type III calpastatin were also assessed as well as their response towards agents involved in signalling cascade associated with the agents that stimulate hypertrophic growth. In order to investigate the CA tandem repeat polymorphisms, a PCR based cloning strategy was developed in this study which allowed amplification of this region. Cattle (n=6) of different breed and meat tenderness had their CA tandem repeat sequences amplified which were then cloned into a ZsGreen based reporter construct and transcriptional activity of the promoter were measured using fluorescence imager (Typhoon Trio). From the results, there was no direct correlation (R=0.28) found between the CA tandem repeat length and the shear force value of the meat. However, transcriptional activity for Type I promoter was significantly affected (P<0.05) by changing the length of CA tandem repeat (40-60bp). In general, the calpastatin promoters displayed negative response towards treatment with cAMP(P<0.05) and there were no significant changes to the promoter activity when it was treated with forskolin. Furthermore, a significant reduction in promoter activity (P<0.05) was observed from all calpastatin promoters with calcimycin treatment. The research shows that the type I calpastatin promoter has transcriptional activity and is regulated by secondary messengers which activate cAMP dependent kinases. Although altering the CA tandem repeat length alters promoter activity, there appears to be no simple relationship between its length and toughness, as determined by shear force. However the differential activity of the three calpastatin promoters indicates that there are potentially multiple mechanisms by which its activity can be regulated.

Background Essential hypertension remains a major risk factor for coronary heart disease (CHO) and stroke, and its prevalence is greater, more severe, occurs earlier, and is less well controlled among black individuals than among white individuals, at all ages after young adult hood (Comoni-Huntley et al, 1989). In Caucasians, studies have shown that β2-adrenoceptor polymorphism accounts for the variability in the vascular responsiveness to the agonist isoprenaline (Cockcroft et al, 1994 and Lang et al, 1995). Individuals homozygous for Gln 27 β2-adrenoceptor showed reduced responses due to chronic down regulation of β2-adrenoceptor in the vasculature. Therefore, variability in response to isoprenaline was determined by β2-adrenoceptor gene polymorphism. Aim and Objectives This study investigated whether there is a relationship between the ArglGly16 and Gln/Glu 27 β2-adrenoceptor polymorphisms by examining whether the incidence of occurrence is prevalent in African Trinidadians. In addition, comparison data of vascular responses with arterial compliance using pulse wave analysis (PWA) was correlated. The study aimed to give evidence if these polymorphisms contributed fully or in part, to determine the disease severity, or response to therapy in hypertensive individuals. It also aimed to prove that PWA is a reliable and therapeutic tool, in diagnosing and treating blood pressure, as reliance on brachial artery recording of blood pressure, alone, is becoming a poor indicator and predictor of risk. Methods The study genotyped 408 African Trinidadian subjects for the β2-adrenoceptor polymorphism and used the technique of applanation tonometry to analyse the central pulse wave, generating information on arterial compliance, left ventricular function and coronary perfusion. Blood pressure was measured in triplicate using a semiautomatic blood pressure meter after 15 minutes of supine rest and bloods lipids assessed using a validated portable lipid cartridge. This was achieved by subjects attending a nurse-led cardiovascular risk clinic. Results There is no significant association between the Arg-Glyl6 polymorphism and the Gln-Glu27 polymorphism and hypertension in African Trinidadians. Interestingly, the appearance of the Glu27 polymorphism was very uncommon in African Trinidadians and this is constant with findings by Candy et al, 2000. Conclusion There is no difference in the frequency of β2-polymorphisms between normotensive and hypertensive African Trinidadians, and are unlikely to be a contributing factor for essential hypertension. Therefore, hypertension would indicate that it is polygenic with complex gene to gene and gene environmental interactions, through multiple, indirect and intermediate phenotypes and interactions.

Pharmacogenetics is the study of variability in drug response attributed to genetic variation. Olanzapine (OLA) is a widely used antipsychotic drug for schizophrenia treatment. The pharmacokinetics of OLA display large inter-individual variation leading to multiple-fold differences in drug exposure between patients at a given dose. This variation in turn gives rise to the need of individualized dosing in order to avoid concentration-dependent adverse effects and therapeutic failure. The observed variability has been partially explained by environmental and physiological factors. Genetically determined differences in drug metabolism represent a less studied source of variability. Precluded contribution by cytochrome P450 (CYP) 2D6 calls for evaluation of the other major OLA metabolizing enzymes. The objective of this thesis was to study pharmacogenetic influence of flavin-containing monooxygenase (FMO) 1 and 3, CYP1A2 and uridine diphosphate-glucuronosyltransferase (UGT) 1A4 on therapeutic OLA exposure. We conducted genetic association studies applying gene re-sequencing and genotyping of candidate and tagging SNPs. Patients carrying the FMO1*6 allele displayed increased dose-adjusted concentrations (C/Ds) of OLA, in serum as well as cerebrospinal fluid. Patients who were homozygous for the FMO3 K158-G308 compound variant showed reduced C/Ds of OLA N-oxide metabolite, but no alteration in OLA exposure. This compound variant is expected to have clinical relevance primarily for non-African populations, since low frequencies were detected among native Africans. Deviation in OLA exposure was observed in carrier of a rare FMO3 mutation, predicted in silico to affect gene splicing. Reduced OLA exposure was observed in UGT1A4*3 carriers. The CYP1A2 -163(A) (CYP1A2*1F) variant was not associated with increase in CYP1A2-catalyzed OLA metabolism or reduction in OLA exposure. Correlations were detected for two cis-acting variants within the inter-genetic region of the CYP1A cluster and a trans-acting variant located upstream the locus encoding aryl hydrocarbon receptor. The inconsistent data reported for CYP1A2*1F could be explained by presence of ethnic specific haplotype structures incorporating the -163(A) variant. A continuously improved understanding of the wide range of factors that can influence pharmacokinetics and pharmacodynamics will increase the likelihood of achieving optimal treatment response for individual patients.

Background and Aims Acute Pancreatitis is an inflammatory disorder of varied aetiology and outcome. Tumour necrosis factor (TNF) and interleukin-10 are important mediators of disease pathogenesis. To investigate if the TNF and IL-10 gene loci influence susceptibility to and severity of acute pancreatitis, 135 patients with acute pancreatitis, ethnically matched normal controls, and alcoholics without pancreatic disease were studied. Methods Aetiology was classified as being secondary to alcohol, gallstones, or idiopathic. Patients were stratified into groups according to disease severity by assigning an organ failure score. Three TNF microsatellite loci (TNFa, TNFb, and TNFc), the -308 polymorphism within the TNF gene, the IL-10.G microsatellite locus, and 3 hi-allelic polymorphisms in the 5' flanking region of the IL-l 0 gene were typed using the polymerase chain reaction. Results There was no difference in allelic frequency of any of the cytokine gene loci between groups stratified according to disease severity. When patients were stratified according to aetiology of disease there was a decrease in the frequency of the TNFa2 allele in those patients with alcoholic acute pancreatitis compared to controls (14.3 vs. 35.5%, χ²=7.24, p=0.007). There was also a reduction in the frequency of the IL-10.Gl3 allele in patients with alcoholic pancreatitis compared to controls (4.8 vs. 21.3%, χ² =6.46, p=0.011). Data is also presented showing that a number of haplotypes exist as well as linkage disequilibrium across all 4 loci of the IL-10 gene, which contrasts with findings from previous work. The 3 locus haplotypes GCC and ATA are in strongest linkage disequilibrium, as is the microsatellite allele G9 and -1117.A and G9 with the 3-locus haplotype ATA. Conclusions This work has identified an allele within the TNF gene locus, and an allele within the IL-1 0.G locus which have different frequencies in patients with alcohol induced acute pancreatitis compared to other aetiologies. This finding may in part explain individuals' differing susceptibility to the development of acute pancreatitis after excessive alcohol consumption. Haplotypes not previously described exist across the IL-10 locus.

Folic acid is an essential vitamin utilized in the one-carbon metabolism pathway for the synthesis of purine and thymidine nucleotides, which are necessary for cell growth and proliferation. As a result, the enzymes that participate in the metabolism of folic acid have been good targets for cancer chemotherapy. Folylpoly-γ-glutamate synthetase (FPGS) is an enzyme in the folate metabolism pathway that catalyzes the addition of glutamic acid to the naturally occurring folates, thereby allowing the retention of folate cofactors in cells. Similarly, in the case of cancer chemotherapy, antifolates, such as Lometrexol and Tomudex are retained in cells through the activity of FPGS. Consequently, any single nucleotide polymorphisms (SNPs) that exist in the fpgs gene may decrease or increase the cytotoxicity of antifolates and, ultimately, the clinical response rate to antifolate therapy. The goal of this project is to define the position and frequency of single nucleotide polymorphisms (SNPs) in the mRNA made from the fpgs gene from peripheral blood of one hundred normal individuals. Six Polymerase Chain Reaction (PCR) primers were designed to amplify the gene as three overlapping pieces and four primers were designed for sequencing of the three PCR products. In this study, we found polymorphic sites at nucleotides 64, 123, 253, 423, 1334 and 1781. The majority of the samples (49/88) expressed rnRNA with point mutations on at least one allele at base 64, while 8 samples had a SNP at base 123. At nucleotide 123, 6 samples expressed the heterozygote GIA genotype, and one sample expressed the homozygote A/A allele at this site. At nucleotide 423, two samples expressed a G allele and also the common C allele. While the SNPs at nucleotide 64, 123, and 423 caused a silent or conservative mutation in the gene, in sample 82, a mutation C253T produced an amino acid change from an arginine to tryptophan, which may cause a functional change in the fpgs protein, due to the significant change in size and charge of the wild type amino acid. Similarly, sample 26 expessed a homozygote T/T allele at nucleotide 1334 instead of the common C/C allele expressed in the remaining samples. This point mutation caused a valine to alanine amino acid change. We also detected a SNP that is expressed after the stop codon in sample 40.

Orientador: Carmen Silvia Bertuzzo
Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas
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Resumo: A Asma é uma doença inflamatória crônica de vias aéreas, na qual várias células têm papel fundamental, tais como os eosinófilos, mastócitos e linfócitos T. O processo inflamatório está associado à hiper-responsividade brônquica a uma variedade de estímulos. A asma é uma doença genética complexa e multifatorial que não acompanha os padrões monogênicos de hereditariedade. Estudos de ligação sugerem que múltiplos genes estão envolvidos na patogenia dessa doença. O objetivo desse trabalho é verificar em uma amostra de pacientes com asma leve, moderada e grave, uma possível associação entre os polimorfismos dos genes TGF-'beta'1, CD14, IL-4, IL-4Ra, ADAM33 e a gravidade da asma. A análise do polimorfismo T869C do gene TGF-'beta'1 foi realizada pela técnica de PCR + ARMS. Os outros polimorfismos C-509T do gene TGF-'beta'1, C-159T do gene CD14, C-590T da IL-4, Ile50Val da IL-4Ra , S2 do gene ADAM33 foram detectados por PCR + enzima de restrição. Em relação ao polimorfismo T869C (TGF-?1), em nossa amostra observou-se uma associação entre o genótipo CC e os pacientes portadores de asma grave. Quanto ao polimorfismo C-509T(TGF-'beta'1), nenhuma associação foi encontrada. Quando se comparou a distribuição da frequência genotípica do polimorfismo C-159T(CD14) na asma grave com o grupo controle foi observado um resultado significativo com genótipos TT e CT. Não encontramos associação entre C-590T (IL4) e asma. Houve significância entre o genótipo Val/Val (IL-4Ra) e a asma leve. O heterozigoto GC do polimorfismo S_2 (ADAM33) com a asma leve e grave. O homozigoto CC com a asma leve. Nossos resultados indicam que os polimorfismos T869C(TGF-?1) e C-159T(CD14) podem estar envolvidos na modulação da gravidade da asma
Abstract: Asthma is a chronic inflammatory disease affecting air pathways, in which several cells, such as eosinophils, mastocytes and T lymphocytes, play fundamental roles. The inflammatory process is associated with bronchial hyperresponsiveness to a variety of stimulus types. Asthma is a complex and multifactorial genetic disease that does not follow the monogenic line of hereditary patterns. Linkage studies suggest that multiple genes are involved in its pathogenesis. The objective of this work was to verify, in a sample of patients with mild, moderate and acute asthma, a possible association between TGF-'beta'1, CD14, IL-4, IL-4Ra, ADAM33 gene polymorphisms and asthma severity. The T869C polymorphism of the TGF-'beta'1 gene was analyzed using the ARMS-PCR technique. The other polymorphisms C-509T of the TGF-'beta'1 gene, C-159T of the CD14 gene, C-590T of the IL-4 gene, Ile50Val of the IL-4Ra gene , S2 of the ADAM33 gene were be detected by PCR + restriction analysis. The T869C polymorphism (TGF-'beta'1) in our sample showed an association between the CC genotype and the patients who presented acute asthma. No association was found concerning the C-509T (TGF-'beta'1) polymorphism. Comparing the genotype distribution of the C-159T (CD14) polymorphism on the control group of acute asthma, a significant result with TT and CT genotypes was observed. There was no association between C-590T (IL4) and asthma. There was a significant association between Val/Val (IL-4Ra) genotype and mild asthma. The GC heterozygote of the S_2 polymorphism (ADAM33) with acute and mild asthma. The CC homozygote and mild asthma. The results indicated that T869C (TGF-'beta'1) and C-159T (CD14) polymorphisms can modulate asthma severity
Doutorado
Genetica Medica
Mestre em Ciências Médicas

Thesis (M. Phil.)--Hong Kong University of Science and Technology, 2003.
Includes bibliographical references (leaves 95-105). Also available in electronic version. Access restricted to campus users.

The incidences of endogenous viral genes (ev-genes) and myb-gene polymorphisms were determined in substrains of strains R, M and G which had been divergently selected or susceptibility to tumour formation induced by Rous Sarcoma virus (RSV) of type A and B. Frequencies of myb gene polymorphisms were also determined in two replicates of strains selected for high and low multiple immune response to challenge with Pasteurella multocida and Mycoplasma gallisepticum. Strains R, M and G were found to contain different sets of ev-genes reflecting their distinct genetic origins. Among eleven ev-genes identified, two showed a significantly increased frequency in the susceptible substrains. One was ev-6, which expresses the viral envelope protein of the endogenous avian leucosis, while the other was a new endogenous viral gene New-E, whose phenotype is unknown. A significant increased incidence of ev-4, reported to be a silent ev-gene, was observed in resistant rather than susceptible substrain. Myb gene polymorphisms were assessed using a cDNA probe and a genomic probe yielding 2 and 3 restriction fragment length polymorphisms (RFLPs) respectively. In strains M and G, only one polymorphism (PM5$ sp+$) observed at an Msp I site located downstream of the last of the c-myb exons was found to be significantly co-selected for susceptibility. Analysis of RFLPs of myb-gene in strains selected for high or low multiple immune response did not reveal any significant response to selection. Rather, polymorphisms seemed to reflect a founder effect as revealed by opposite frequencies obtained in the two replicates. DNA methylation, a possible epigenetic mechanism regulating gene expression, was also investigated in the myb-gene. DNA from semen, blood, spleen, liver and thymus was extracted from organs obtained from chickens at different ages.

Breast cancer is a world health problem and is a leading cause of cancer-related death among women in the United States. However, breast cancer risks were reported to be reduced through exposure to Vitamin D through its Receptors identified as the p53 target gene. The purpose of this study was to assess the associations between VDR gene polymorphisms knowledge/awareness and decisions to reduce breast cancer risks and likelihood of mammogram screening among women in Texas. Data from survey were used. Roy adaptation model was the theoretical framework that guided this quasi- experimental, quantitative research. The dependent variables were decisions to reduce breast cancer risks and likelihood of mammogram screening. The independent variables were knowledge about VDR gene polymorphisms and exposure to vitamin D. The covariates were level of education, awareness, lifestyle, breast self-exams, mammograms, age, early menarche, late menopause, and family history of breast cancer. The chi-square test and regression analysis were used to test the stated research hypotheses and to answer the research questions. Knowledge of VDR gene polymorphisms and exposure to vitamin D were not significantly associated with breast cancer risk, ï?£2 (3, N= 250) =3.84, p > 0.05. Also, awareness of the risk factors for breast cancer was not significantly associated with decisions to go for mammogram screenings or to enroll in breast cancer risk-reduction programs, ï?£2 (3, N= 250) =1.58, p > 0.05. To advocate for the promotion of awareness of the importance of pharmacogenetic testing for VDR gene polymorphisms for early detection of breast cancer, which would help to undertake appropriate therapeutic measures in a timely manner to prevent cancer metastasis, further research is warranted.

The paraoxonase gene family consists of three members (PON1, PON2, and PON3) with both distinct and overlapping roles in human health. These enzymes influence oxidative stress, inflammation, and bacterial infections, along with a large number of diseases and disorders, such as atherosclerosis. The wide-reaching effects of the PON gene family make them an important and highly advantageous subject of study. Their ability to be modified by diet, lifestyle, and environmental exposures, as well as various polymorphisms and genetic influences, provide for a complex, highly modifiable internal form of individual protection. The overall goal of this project was to determine what factors affect individual variations in paraoxonase activity, as well as the influence of individual PON members on health and exposure outcomes. The initial study in this project provided the first data about intra-individual PON1 variations over a time of about 15 years, showing levels remain relatively stable in an agricultural population. This study also contributed data regarding the polymorphic distributions of influential PON SNPs and the influence of lifestyle factors on PON1 activity. The use of a twin population for the next study allowed for examination of the heritability of PON1 activity and antioxidant capacity, and provided novel data regarding the influence of genetic variations on PON1 activity. To further attempt to eliminate the complexity of influences on these genes and individual polymorphisms, the third study in this project characterized an innovative transgenic Drosophila melanogaster model with the goal of analyzing the influence of individual PON family members on exposure and disease outcomes without the effects of compensation from other PONs. By further elucidating the effects of the PONs at the individual level, human populations will be able to be advised regarding the most at-risk individuals and modifiable changes to improve PON levels, and therefore overall health.

Background: Vascular remodeling generated by reactive oxygen species contributes to aneurysm formation. The NADPH oxidase system is a major source of superoxide anion not only in phagocytes, but also in endothelial and vascular smooth muscle cells. Polymorphisms of p22phox, an essential component of the NADPH oxidase system, are found to be associated with atherosclerosis, while a recent study found a significant association between the 214C>T polymorphism and the occurrence of ischemic cerebrovascular disease. We conducted a case-control study to investigate the relationship of five polymorphisms of the p22phox gene and the occurrence of cerebral aneurysms. Methods: The study population consisted of 113 patients with intracranial aneurysms and 53 control subjects. The 214C>T polymorphism was investigated by restriction fragment length polymorphism analysis, while polymorphisms 381T>C, 480G>A, 521C>T, and *24A>G were analyzed by direct sequencing of exon 6 and adjacent intronic sequences. Results: The analysis of a primary study sample comprising 35 cases and 28 controls failed to show a significant association between any of the five polymorphisms and the occurrence of intracranial aneurysms using both allele frequencies and genotypes (all nominal p > 0.05). Although there was a deviation from Hardy-Weinberg equilibrium in cases at the 521C>T locus (nominal p < 0.05), this could not be confirmed in a second study sample of 78 patients. Haplotypes were constructed regarding three frequent polymorphisms (214C>T, 521C>T, and *24A>G); haplotype frequencies in cases and controls were not significantly different. Conclusion: Although polymorphisms of the p22phox gene located in the coding region and the 3′-untranslated region were reported to be associated with atherosclerosis and cerebrovascular disease, our data provide evidence that there is no association between these polymorphisms and the occurrence of cerebral aneurysms in Caucasians
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v It was found that most of the classical scrapie genotypes belong to R3 risk group, whereas atypical scrapie genotypes belonging to zero (0) and one (1) risk groups were frequently seen in sheep analyzed. In other words, Turkish sheep is found to have intermediate risk of classical scrapie and low atypical scrapie risk, in general. The data from the current study may help to establish a breeding program for classical scrapie control in Turkey and will be beneficial for both the animal and public health in the country. In addition, the outcomes of the study will fill the gap which is present in the geographic distribution data of PrP gene polymorphisms in Eurasia.Scrapie is an infectious fatal disease of sheep and goats which affects the central nervous system. In the present study, samples of 14 native Turkish sheep breeds (n=655) were analyzed with respect to their polymorphisms of PrP gene (at codons: 136, 141, 154 and 171) and their classical and atypical scrapie risk levels were identified. Turkish sheep are found to have the highest PrP genetic variability with 13 classical scrapie alleles and 14 atypical scrapie alleles compared to all previous studies. Classical scrapie-susceptible and wild-type ARQ allele was found as the most frequent allele in Turkish sheep examined. The most classical scrapie-susceptible allele, VRQ was detected at low frequencies in 5 of the breeds (Ç
ine Ç
apari, Dagliç
, Kivircik, Karayaka and Gö
kç
eada). One novel allele (TL141HQ) was observed in Sakiz breed for the first time in this study.

Abstract The cholesteryl ester transfer protein (CETP) is a plasma protein that transfers cholesteryl esters and triglycerides between plasma lipoproteins. Humans with a genetic CETP deficiency have high plasma high density lipopoprotein cholesterol (HDL-C) levels, whereas the CETP transgene lowers plasma HDL-C levels in mice. The role of CETP in the development of atherosclerosis is unclear due to the controversial results of many human and animal studies. The present research was designed to investigate the CETP gene as a candidate gene in the regulation of plasma HDL-C levels and the development of atherosclerosis in humans. The CETP gene was screened for mutations and polymorphisms associated with these traits in a well-characterized, homogenous population sample of 515 men and women and in a sample of 115 men with low HDL-C levels and coronary heart disease (CHD). Using polymerase chain reaction and single-strand conformation polymorphism analysis (PCR-SSCP), three polymorphic sites were found (A373P, I405V, R451Q) in the exons of the CETP gene, one in intron 9 and one in the 3'untranslated region of the CETP gene. In addition, the genotypes of a functional promoter polymorphism were determined. The V405 allele was associated with lower plasma CETP activity in the whole population sample, and the Q451 allele and the P373 allele were associated with higher plasma CETP activity in men, whereas the genotypes of the promoter polymorphism were not significantly associated with plasma CETP activity. The genotypes of the CETP gene explained about 20 % of the variation of plasma CETP activity in men. The CETP gene polymorphisms were found to be a minor regulator of plasma HDL-C levels, and these associations interacted with alcohol consumption, sex and triglyceride levels. The strongest association was detected between the promoter polymorphism and HDL-C levels in women. The variation at the CETP gene locus explained about 8 % of the variation in plasma HDL-C levels in women, but less than 1 % in men. CETP gene polymorphisms (A373P, I405V and R451Q) were associated with carotid intima-media thickness, explaining about 6 % of the variation in men and 4 % in women. However, none of the polymorphisms were associated significantly with the CHD risk. In conclusion, the CETP gene was found to be polymorphic and a minor regulator of plasma HDL-C levels and the development of atherosclerosis.

Abstract Pulmonary surfactant is a lipid-protein mixture that lines the inner surface of the lung. The main function of surfactant is to reduce surface tension at the air-liquid interface, thus preventing alveolar collapse at the end of expiration. Lack of surfactant is the main cause of respiratory distress syndrome (RDS) in preterm infants. Very preterm babies are at risk of developing a lung disease called bronchopulmonary dysplasia (BPD). The surfactant proteins SP-A, -B, -C and -D have important functions in surfactant structure, homeostasis and innate immunity of the lung. The genes of these proteins have been studied as candidates for several multifactorial lung diseases both in adults and in children. The aim of the present study was to examine the genetic variation in SP genes and to evaluate the role of SP gene polymorphism in the etiology of severe pulmonary infantile diseases, including RDS, BPD and severe respiratory syncytial virus (RSV) infection among the Finnish population. Conventional allelic association methods in combination with multiparameter analysis and family-based transmission disequilibrium test (TDT) were used. The SP-D Met11 allele was associated with a risk for severe RSV bronchiolitis in a matched case-control setting of 84 infants with severe RSV infection and 93 control infants. The variants of the SP-C gene had no detectable association with BPD. However, a modest association of SP-C Asn138 and Asn186 alleles with RDS was found. A length variation in the SP-B gene was associated with BPD among very preterm infants born before 32 weeks of gestation. The SP-B intron 4 deletion variant allele increased the risk for BPD especially in very low birth weight infants. The association was confounded by birth order, being evident only among presenting infants, who are more prone to ascending infections during a preterm birth process. The present study provides new evidence about the significance of SP gene polymorphisms in the etiology of complex infantile pulmonary diseases, including RDS, BPD and severe RSV bronchiolitis. The results help us to understand the molecular mechanisms underlying these diseases and may, in the long run, enable better treatment of these life-threatening diseases
Tiivistelmä Keuhkosurfaktantti on keuhkon sisäpintaa peittävä kalvomainen rasva-proteiinikompleksi, jonka tärkein ominaisuus on pintajännityksen vähentäminen keuhkorakkuloissa. Surfaktantin puutos ennenaikaisesti syntyneillä lapsilla aiheuttaa hengitysvaikeusoireyhtymän, RDS-taudin (respiratory distress syndrome). Alle 30 raskausviikon iässä syntyneistä, useimmiten RDS-taudin saaneista keskosista n. 30 % sairastuu vakavaan krooniseen keuhkotautiin, BPD-tautiin (bronchopulmonary dysplasia). Surfaktanttiproteiineilla SP-A, -B, -C ja -D on osoitettu olevan tärkeä tehtävä surfaktantin toiminnassa ja keuhkon synnynnäisessä immuniteetissa. Tämän tutkimuksen tavoitteena oli selvittää surfaktanttiproteiineissa esiintyvän geneettisen muuntelun määrää ja merkitystä keskosten RDS- ja BPD-taudeissa sekä pienten lasten vakavassa respiratory syncytial -viruksen (RSV) aiheuttamassa keuhkotulehduksessa. Tutkimuksen laajin osa keskittyi tutkimaan keskosten BPD-tautia ja surfaktanttiproteiinien geenien osuutta siinä. Geneettisen muuntelun merkitystä tarkasteltiin populaatiogeneettisin keinoin tapaus-verrokkiasetelmissa ja perheaineistojen avulla. Yhteensä analysoitiin noin tuhannen lapsen ja yli kahdensadan vanhemman DNA-näytteet. Tutkimuksessa havaittiin SP-D-geenissä olevan metioniini11-geenimuodon liittyvän pienten lasten vakavaan RSV-infektioon. Lisäksi saatiin uutta tietoa SP-C-geenin populaatiotason yleisestä muuntelusta ja todettiin SP-C:n asparagiini138 ja asparagiini186 -geenimuotojen yhteys keskosten RDS-taudin esiintymiseen. Merkittävin löydös oli SP-B-geenissä olevan deleetiovariantin kytkeytyminen alle 32-viikkoisina syntyneiden keskosten BPD-tautiin. Geneettisen altistuksen lisäksi BPD-tautiin sairastumiseen vaikuttivat lukuisat keskosuudelle ominaiset seikat, kuten alhainen syntymäpaino, RDS-tauti ja syntymähetkellä todettu hapenpuute. Geneettisen tekijän vaikutus oli voimakkain erittäin pienipainoisilla keskosilla. Tutkimuksen tulokset ovat tuoneet arvokasta lisätietoa surfaktanttiproteiinien geenien osuudesta keskosten RDS- ja BPD-taudeissa sekä pienten lasten vakavassa RSV-infektiossa. Ne auttavat ymmärtämään näiden molekyylibiologisia syntymekanismeja ja voivat ajan mittaan olla edistämässä uusien hoitomuotojen kehittämistä

Thyroid cancer is the most common endocrine malignancy and a complex disease with a largely unknown aetiology. Thyroid carcinomas are derived from thyroid follicular cells and parafollicular cells. The majority of thyroid cancer cases comprise both papillary (PTC) and follicular carcinomas (FTC). The evaluation of genetic susceptibility could give valuable information regarding the risk of thyroid cancer development. There are many genes associated with the thyroid function that modulates the risk of tumour development. Among them, is the vitamin D receptor gene (VDR), located on chromosome 12q12-q14, and includes eight protein coding exons (exons 2-9) and one untranslated exon (exons 1a-1f). The most common VDR polymorphisms investigated are FokI (rs10735810 C>T), located in exon 2 of VDR, BsmI (rs1544410 G>A) and ApaI (rs7975232 G>T), located in intron 8, and TaqI (rs731236 T>C), located in exon 9 of VDR. The importance of vitamin D and its receptor VDR, in many signalling pathways is well known. Therefore we aim to verify in which way VDR polymorphisms influence the predisposition for thyroid cancer development. The contribution of four well-known VDR polymorphisms (FokI, BsmI, ApaI and TaqI) for the genetic susceptibility of thyroid cancer in the Portuguese population was analysed, including haplotypes comparisons. The following parameters were studied: thyroid cancer type differences (PTC vs. FTC), age (≤45 vs. >45 years), gender (male vs. female), carcinoma size (≤10mm vs. >10mm), lymph node metastasis and distant metastasis multicentricity, and stage of cancer (I-II vs. III-IV). All the participants in the study were Caucasian Portuguese inhabitants, being subdivided into two groups: patients with thyroid cancer (N = 208) and a control group (N = 248). In conclusion, there were some statistically significant differences in some parameters assessed. However, the results considered were only those with a statistical significant p-value < 0.005, according to Bonferroni’s correction. Therefore, the results suggest that BsmI polymorphism genotype AA (p = 0.004) may influence lymph node metastasis or distant metastasis in patients with DTC. Moreover, the TT genotype (p = 0.004) of ApaI polymorphism may increase the predisposition for more aggressive phenotypes of DTC, since it is overrepresented in patients with more advanced cancer stages (III-IV).
O cancro da tiróide é, de todas as neoplasias endócrinas, a mais comum, revelando ser uma patologia complexa e com uma etiologia desconhecida em parte. Tem uma incidência mundial que tem tendência a aumentar, contabilizando cerca de 1.7% dos cancros diagnosticados. Adicionalmente, o cancro da tiróide é mais prevalente em pacientes de meia idade e idosos, onde mais de metade dos indivíduos diagnosticados têm uma idade superior aos 45 anos. Ademais, esta neoplasia endócrina é mais comum nas mulheres, com uma incidência de 3 a 5 vezes maior. Os nódulos que surgem na tiróide são diagnosticados em cerca de 5% da população adulta mundial, e podem ser adenomas ou lesões malignas. Os carcinomas da tiróide derivam quer das células foliculares da tiróide, bem como das células C, porém, a grande maioria deles tem origem nas células foliculares. De todas as variantes de carcinomas da tiróide, os carcinomas papilar e folicular da tiróide são os mais predominantes, sendo a variante papilar a mais comum de entre todos, seguida da variante folicular. Apesar da elevada incidência mundial de cancro da tiróide, a taxa de mortalidade associada permanece estável. O tratamento do cancro da tiróide é um processo multifatorial, envolvendo a combinação de terapias cirúrgicas, hormonais ou de medicina nuclear. Sabe-se que o cancro da tiróide, em especial os tumores diferenciados da tiróide, estão a aumentar de incidência em alguns países desenvolvidos. Existem muitos fatores de risco que aumentam a predisposição para este tipo de cancro, incluindo fatores genéticos com risco associado a esta patologia. A título de exemplo, o cancro diferenciado da tiróide está associado a uma forte hereditariedade, aumentando a suscetibilidade genética do indivíduo em desenvolver cancro de acordo com o seu historial familiar. Para além disso, a presença de polimorfismos genéticos podem determinar a suscetibilidade individual do indivíduo para o desenvolvimento de cancro da tiróide. Atualmente são conhecidos vários genes associados com a função tiroideia e que modulam o risco para a tumorigénese. O VDR é um membro da superfamília de recetores nucleares, sendo a única proteína com afinidade para a 1α,25-dihidroxivitamina D, também conhecida como calcitriol. Nos mamíferos, a expressão do VDR encontra-se aumentada em tecidos metabólicos tais como o intestino, rins, pele e glândula da tiróide. O impacto biológico do VDR surge quando este se liga aos seus elementos localizados nas regiões promotores dos genes alvo, interferindo assim em muitas ações celulares e moleculares que vão desde a regulação do metabolismo de cálcio até à regulação de péptidos antimicrobiais. Desta forma, a ação molecular da vitamina D/ VDR está envolvida na regulação mineral e homeostase óssea, modulação do crescimento, eventos cardiovasculares, prevenção de cancro e regulação de respostas imunes. Uma disfunção do VDR ou défice de vitamina D podem levar a consequências no desenvolvimento e saúde óssea assim como aumentar a predisposição do indivíduo para o desenvolvimento de algumas doenças crónicas, incluindo o cancro. Os polimorfismos associados ao gene VDR já provaram estar implicados como um fator principal de risco em vários tipos de cancro, tais como o cancro da próstata, mama ou cólon. Ao longo do tempo, estudos de associação têm sido feitos de modo a se poder correlacionar os polimorfismos genéticos e o seu impacto na saúde do indivíduo. Assim, no presente trabalho pretende-se estudar a suscetibilidade genética do cancro da tiróide associada aos polimorfismos do gene VDR. Neste trabalho, foram estudados quatro polimorfismos diferentes gene do VDR. Para tal, através do uso de enzimas de restrição, foi possível analisar áreas restritas do gene VDR, localizado no cromossoma 12q12-q14 de forma a se poder observar variações da sequência de DNA. Os quatro polimorfismos estudados no âmbito deste projeto foram o FokI (rs10735810 C>T), localizado no exão 2 do VDR, BsmI (rs1544410 G>A) e ApaI (rs7975232 G>T), localizados no intrão 8, e TaqI (rs731236 T>C), localizado no exão 9 do VDR. Estes quatro polimorfismos foram analisados com o objetivo de verificar de que forma influenciam a predisposição de um indivíduo para o desenvolvimento de cancro da tiróide. Desta forma, este estudo realizado na população Portuguesa, fez a análise destas variantes do VDR, e o seu impacto no desenvolvimento de cancro da tiróide de acordo com os seguintes parâmetros: tipo de cancro, idade de diagnóstico, sexo, dimensões do carcinoma, metástases ganglionares e à distância, multicentricidade tumoral, e estádios de cancro. Todos os participantes deste estudo foram indivíduos caucasianos de origem Portuguesa. Estes indivíduos foram divididos em dois grupos distintos. Um dos grupos foi composto por indivíduos com cancro diferenciado da tiróide (N = 208), provenientes do Instituto Português de Oncologia de Coimbra. O grupo de indivíduos saudáveis (N = 248), que constituíam o grupo controlo, consistiram em dadores voluntários de sangue Portugueses caucasianos, que não possuíam um historial clínico de cancro da tiróide. Após o recrutamento dos indivíduos e obtenção das amostras de sangue dos mesmos, procedeu-se a uma série de metodologias práticas que visaram como objetivo final genotipar as amostras recolhidas. A cada indivíduo, doente ou controlo, foi atribuído um número de código único, de forma a poder identificar e diferenciar a amostra em estudo. O processo clínico dos doentes foi registado com todos os dados necessários para este estudo. Quanto aos indivíduos saudáveis, estes permaneceram no anonimato, sendo apenas registado a idade, sexo, peso, altura e naturalidade. Após estes procedimentos de registo, o DNA genómico foi extraído das amostras de sangue recolhidas através do método de “salting-out”. De seguida o DNA extraído foi quantificado e armazenado. Para efeitos de genotipagem, o DNA de cada indivíduo participante no estudo foi submetido à técnica “polymerase chain reaction”, mais conhecida por PCR. Com este procedimento pretende-se amplificar o fragmento do gene VDR onde se encontra cada polimorfismo. Após a amplificação do fragmento do VDR que se pretendeu estudar, conforme o polimorfismo, procedeu-se à digestão enzimática utilizando a respetiva enzima. Desta forma, conseguimos determinar o genótipo do indivíduo, através da visualização desses produtos digeridos num gel de agarose de 3%. Para além deste método, a genotipagem foi também confirmada através da sequenciação de DNA, sendo utilizada uma amostra representativa de cada genótipo para cada polimorfismo. Terminada a genotipagem de todos os indivíduos participantes deste estudo, para os quatro polimorfismos do VDR, procedeu-se ao tratamento estatístico dos dados analisando os parâmetros acima referidos. Como resultados, verificaram-se, em alguns parâmetros, algumas diferenças de frequências dos polimorfismos. De entre esses resultados, nas comparações entre pacientes de sexo diferente, o genótipo GA do polimorfismo BsmI foi mais frequente no sexo masculino (p = 0.044). Na análise de metástases ganglionares e à distância, o genótipo AA (p = 0.004) do BsmI e o alelo A (p = 0.014) e o genótipo CC (p = 0.024) do TaqI foram mais frequentes no grupo de doentes com metástases. No estudo da multicentricidade tumoral, o alelo C (p = 0.041) do FokI, o genótipo AA (p = 0.013) do BsmI, e o genótipo CC (p = 0.017) do TaqI foram mais frequentes nos doentes com multicentricidade. No estudo dos estádios de cancro, os genótipos GT (p = 0.012) e TT (p = 0.004) do ApaI, e seu respetivo alelo T (p = 0.031) foram mais frequentes em doentes com estádios mais avançados. A correção estatística de Bonferroni para comparações múltiplas revelou que os resultados foram estatisticamente significativos apenas para o genótipo AA do polimorfismo BsmI, que parece estar envolvido na presença de metástases ganglionares em indivíduos com cancro diferenciado da tiróide. Para além disso, também o genótipo TT do polimorfismo ApaI revelou diferenças estatisticamente significativas, podendo estar associado a um estádio mais avançado de cancro da tiróide. Desta forma, os polimorfismos do gene do VDR podem servir como marcadores de risco úteis para pacientes com cancro diferenciado da tiróide, uma vez que estes já forma associados em outros tipos de cancro. No entanto, não é possível retirar conclusões a partir destes resultados uma vez que são necessários mais estudos que permitam compreender as ações celulares e moleculares do VDR. Para tal, estudos funcionais genómicos serão necessários, para que se possa clarificar de que forma os polimorfismos deste gene podem influenciar a suscetibilidade genética para o cancro da tiróide.