RESUMO
We performed a molecular analysis of formalin-fixed paraffin embedded and decalcified bone marrow trephine biopsies of 41 patients with a B-cell disorder with lymphoplasmacytic differentiation to enable a more precise diagnosis and to describe potentially prognostic and therapeutic relevant mutations. Analysis was performed with a commercially available next-generation sequencing (NGS) lymphoma panel (Lymphoma Solution, SophiaGenetics). Results were correlated with clinical and pathological parameters. Our group covered a spectrum of B-cell disorders with plasmacytic differentiation ranging from Waldenstroem's macroglobulinemia (WM), to small-B-cell lymphomas with plasmacytic differentiation (SBCL-PC) to IgM myeloma (MM). The most helpful diagnostic criteria included morphology and immuno-phenotype as a prerequisite for the interpretation of molecular analysis. MYD88 mutation was present in nearly all WM, but also in 50% of SBCL-PCs, while MM were consistently negative. Driver mutations, such as TP53, were already detectable early in the course of the respective diseases indicating a higher risk of progression, transformation, and reduced progression-free survival. In addition, we report on a novel BIRC3 frameshift mutation in one case of a progressive WM. Our data indicate that patients with LPL/WM might benefit from thorough pathological work-up and detailed molecular analysis in terms of precise diagnosis and targeted treatment allocation.
Assuntos
Leucemia Linfocítica Crônica de Células B , Linfoma , Macroglobulinemia de Waldenstrom , Humanos , Linfoma/patologia , Macroglobulinemia de Waldenstrom/diagnóstico , Macroglobulinemia de Waldenstrom/genética , Macroglobulinemia de Waldenstrom/tratamento farmacológico , Leucemia Linfocítica Crônica de Células B/patologia , Medula Óssea/patologia , Mutação , Fator 88 de Diferenciação Mieloide/genéticaRESUMO
Lipoprotein (a) [Lp(a)] concentrations are among the strongest genetic risk factors for cardiovascular disease and present pronounced interethnic and interindividual differences. Approximately 90% of Lp(a) variance is controlled by the LPA gene, which contains a 5.6-kb-large copy number variation [kringle IV type 2 (KIV-2) repeat] that generates >40 protein isoforms. Variants within the KIV-2 region are not called in common sequencing projects, leaving up to 70% of the LPA coding region currently unaddressed. To completely assess the variability in LPA, we developed a sequencing strategy for this region and report here the first map of genetic variation in the KIV-2 region, a comprehensively evaluated ultradeep sequencing protocol, and an easy-to-use variant analysis pipeline. We sequenced 123 Central-European individuals and reanalyzed public data of 2,504 individuals from 26 populations. We found 14 different loss-of-function and splice-site mutations, as well as >100, partially even common, missense variants. Some coding variants were frequent in one population but absent in others. This provides novel candidates to explain the large ethnic and individual differences in Lp(a) concentrations. Importantly, our approach and pipeline are also applicable to other similar copy number variable regions, allowing access to regions that are not captured by common genome sequencing.
Assuntos
Variações do Número de Cópias de DNA , Genômica , Kringles/genética , Lipoproteína(a)/química , Lipoproteína(a)/genética , Polimorfismo de Nucleotídeo Único , Humanos , MutaçãoRESUMO
AIMS: Lp(a) concentrations represent a major cardiovascular risk factor and are almost entirely controlled by one single locus (LPA). However, many genetic factors in LPA governing the enormous variance of Lp(a) levels are still unknown. Since up to 70% of the LPA coding sequence are located in a difficult to access hypervariable copy number variation named KIV-2, we hypothesized that it may contain novel functional variants with pronounced effects on Lp(a) concentrations. We performed a large scale mutation analysis in the KIV-2 using an extreme phenotype approach. METHODS AND RESULTS: We compiled an discovery set of 123 samples showing discordance between LPA isoform phenotype and Lp(a) concentrations and controls. Using ultra-deep sequencing, we identified a splice site variant (G4925A) in preferential association with the smaller LPA isoforms. Follow-up in a European general population (n = 2892) revealed an exceptionally high carrier frequency of 22.1% in the general population. The variant explains 20.6% of the Lp(a) variance in carriers of low molecular weight (LMW) apo(a) isoforms (P = 5.75e-38) and reduces Lp(a) concentrations by 31.3 mg/dL. Accordingly the odds ratio for cardiovascular disease was reduced from 1.39 [95% confidence interval (CI): 1.17-1.66, P = 1.89e-04] for wildtype LMW individuals to 1.19 [95%CI: 0.92; 1.56, P = 0.19] in LMW individuals who were additionally positive for G4925A. Functional studies point towards a reduction of splicing efficiency by this novel variant. CONCLUSION: A highly frequent but until now undetected variant in the LPA KIV-2 region is strongly associated with reduced Lp(a) concentrations and reduced cardiovascular risk in LMW individuals.
Assuntos
Doenças Cardiovasculares/genética , Kringles/genética , Lipoproteína(a)/genética , Adulto , Idoso , Variações do Número de Cópias de DNA/genética , Feminino , Genótipo , Humanos , Desequilíbrio de Ligação/genética , Lipoproteína(a)/metabolismo , Masculino , Pessoa de Meia-Idade , Fenótipo , Polimorfismo de Nucleotídeo Único/genética , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Fatores de RiscoRESUMO
OBJECTIVE: The enzyme heme oxygenase-1 (HO-1) exerts cytoprotective effects in response to various cellular stressors. A variable number tandem repeat polymorphism in the HO-1 gene promoter region has previously been linked to cardiovascular disease. We examined this association prospectively in the general population. APPROACH AND RESULTS: Incidence of stroke, myocardial infarction, or vascular death was registered between 1995 and 2010 in 812 participants of the Bruneck Study aged 45 to 84 years (49.4% males). Carotid atherosclerosis progression was quantified by high-resolution ultrasound. HO-1 variable number tandem repeat length was determined by polymerase chain reaction. Subjects with ≥32 tandem repeats on both HO-1 alleles compared with the rest of the population (recessive trait) featured substantially increased cardiovascular disease risk (hazard ratio [95% confidence interval], 5.45 [2.39, 12.42]; P<0.0001), enhanced atherosclerosis progression (median difference in atherosclerosis score [interquartile range], 2.1 [0.8, 5.6] versus 0.0 [0.0, 2.2] mm; P=0.0012), and a trend toward higher levels of oxidized phospholipids on apolipoprotein B-100 (median oxidized phospholipids/apolipoprotein B level [interquartile range], 11364 [4160, 18330] versus 4844 [3174, 12284] relative light units; P=0.0554). Increased cardiovascular disease risk in those homozygous for ≥32 repeats was also detected in a pooled analysis of 7848 participants of the Bruneck, SAPHIR, and KORA prospective studies (hazard ratio [95% confidence interval], 3.26 [1.50, 7.33]; P=0.0043). CONCLUSIONS: This study found a strong association between the HO-1 variable number tandem repeat polymorphism and cardiovascular disease risk confined to subjects with a high number of repeats on both HO-1 alleles and provides evidence for accelerated atherogenesis and decreased antioxidant defense in this vascular high-risk group.
Assuntos
Aterosclerose/enzimologia , Aterosclerose/genética , Doenças Cardiovasculares/enzimologia , Doenças Cardiovasculares/genética , Heme Oxigenase-1/genética , Repetições Minissatélites , Polimorfismo Genético , Regiões Promotoras Genéticas , Idoso , Idoso de 80 Anos ou mais , Apolipoproteína B-100/sangue , Aterosclerose/sangue , Aterosclerose/diagnóstico , Aterosclerose/mortalidade , Áustria/epidemiologia , Doenças Cardiovasculares/sangue , Doenças Cardiovasculares/diagnóstico , Doenças Cardiovasculares/mortalidade , Progressão da Doença , Feminino , Marcadores Genéticos , Predisposição Genética para Doença , Heterozigoto , Homozigoto , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , Infarto do Miocárdio/enzimologia , Infarto do Miocárdio/genética , Infarto do Miocárdio/mortalidade , Oxirredução , Fenótipo , Fosfolipídeos/sangue , Modelos de Riscos Proporcionais , Estudos Prospectivos , Fatores de Risco , Acidente Vascular Cerebral/enzimologia , Acidente Vascular Cerebral/genética , Acidente Vascular Cerebral/mortalidadeRESUMO
Several infrequent genetic polymorphisms in the SERPINA1 gene are known to substantially reduce concentration of alpha1-antitrypsin (AAT) in the blood. Since low AAT serum levels fail to protect pulmonary tissue from enzymatic degradation, these polymorphisms also increase the risk for early onset chronic obstructive pulmonary disease (COPD). The role of more common SERPINA1 single nucleotide polymorphisms (SNPs) in respiratory health remains poorly understood. We present here an agnostic investigation of genetic determinants of circulating AAT levels in a general population sample by performing a genome-wide association study (GWAS) in 1392 individuals of the SAPALDIA cohort. Five common SNPs, defined by showing minor allele frequencies (MAFs) >5%, reached genome-wide significance, all located in the SERPINA gene cluster at 14q32.13. The top-ranking genotyped SNP rs4905179 was associated with an estimated effect of ß = -0.068 g/L per minor allele (P = 1.20*10(-12)). But denser SERPINA1 locus genotyping in 5569 participants with subsequent stepwise conditional analysis, as well as exon-sequencing in a subsample (N = 410), suggested that AAT serum level is causally determined at this locus by rare (MAF<1%) and low-frequent (MAF 1-5%) variants only, in particular by the well-documented protein inhibitor S and Z (PI S, PI Z) variants. Replication of the association of rs4905179 with AAT serum levels in the Copenhagen City Heart Study (N = 8273) was successful (P<0.0001), as was the replication of its synthetic nature (the effect disappeared after adjusting for PI S and Z, P = 0.57). Extending the analysis to lung function revealed a more complex situation. Only in individuals with severely compromised pulmonary health (N = 397), associations of common SNPs at this locus with lung function were driven by rarer PI S or Z variants. Overall, our meta-analysis of lung function in ever-smokers does not support a functional role of common SNPs in the SERPINA gene cluster in the general population.
Assuntos
Estudo de Associação Genômica Ampla , Doença Pulmonar Obstrutiva Crônica/genética , alfa 1-Antitripsina/sangue , Dinamarca , Frequência do Gene , Predisposição Genética para Doença , Genótipo , Humanos , Pulmão/patologia , Família Multigênica , Polimorfismo de Nucleotídeo Único/genética , Doença Pulmonar Obstrutiva Crônica/sangue , Doença Pulmonar Obstrutiva Crônica/patologia , alfa 1-Antitripsina/genéticaAssuntos
Medula Óssea/patologia , Fator 88 de Diferenciação Mieloide/análise , Plasmócitos/patologia , Reação em Cadeia da Polimerase/métodos , Macroglobulinemia de Waldenstrom/patologia , Idoso , Biópsia , Diferenciação Celular , Feminino , Humanos , Linfoma/química , Linfoma/diagnóstico , Linfoma/patologia , Masculino , Pessoa de Meia-Idade , Fator 88 de Diferenciação Mieloide/genética , Macroglobulinemia de Waldenstrom/diagnósticoRESUMO
Oxidative stress and inflammation are major contributors to accelerated age-related relative telomere length (RTL) shortening. Both conditions are strongly linked to leptin and adiponectin, the most prominent adipocyte-derived protein hormones. As high leptin levels and low levels of adiponectin have been implicated in inflammation, one expects adiponectin to be positively associated with RTL while leptin should be negatively associated. Within the ENGAGE consortium, we investigated the association of RTL with adiponectin and leptin in seven independent cohorts with a total of 11,448 participants. We performed partial correlation analysis on Z-transformed RTL and LN-transformed leptin/adiponectin, adjusting for age and sex. In extended models we adjusted for body mass index (BMI) and C-reactive protein (CRP). Adiponectin showed a borderline significant association with RTL. This appeared to be determined by a single study and when the outlier study was removed, this association disappeared. The association between RTL and leptin was highly significant (r = -0.05; p = 1.81 × 10(-7)). Additional adjustment for BMI or CRP did not change the results. Sex-stratified analysis revealed no difference between men and women. Our study suggests that high leptin levels are associated with short RTL.
Assuntos
Adiponectina/genética , Proteína C-Reativa/metabolismo , Leptina/genética , Telômero/genética , Adiponectina/sangue , Adulto , Biomarcadores/sangue , Índice de Massa Corporal , Estudos de Coortes , Estudos Transversais , Feminino , Humanos , Inflamação , Leptina/sangue , Masculino , Pessoa de Meia-Idade , Estresse OxidativoRESUMO
Family studies suggest a genetic component to the etiology of chronic kidney disease (CKD) and end stage renal disease (ESRD). Previously, we identified 16 loci for eGFR in genome-wide association studies, but the associations of these single nucleotide polymorphisms (SNPs) for incident CKD or ESRD are unknown. We thus investigated the association of these loci with incident CKD in 26,308 individuals of European ancestry free of CKD at baseline drawn from eight population-based cohorts followed for a median of 7.2 years (including 2,122 incident CKD cases defined as eGFR <60ml/min/1.73m(2) at follow-up) and with ESRD in four case-control studies in subjects of European ancestry (3,775 cases, 4,577 controls). SNPs at 11 of the 16 loci (UMOD, PRKAG2, ANXA9, DAB2, SHROOM3, DACH1, STC1, SLC34A1, ALMS1/NAT8, UBE2Q2, and GCKR) were associated with incident CKD; p-values ranged from pâ=â4.1e-9 in UMOD to pâ=â0.03 in GCKR. After adjusting for baseline eGFR, six of these loci remained significantly associated with incident CKD (UMOD, PRKAG2, ANXA9, DAB2, DACH1, and STC1). SNPs in UMOD (ORâ=â0.92, pâ=â0.04) and GCKR (ORâ=â0.93, pâ=â0.03) were nominally associated with ESRD. In summary, the majority of eGFR-related loci are either associated or show a strong trend towards association with incident CKD, but have modest associations with ESRD in individuals of European descent. Additional work is required to characterize the association of genetic determinants of CKD and ESRD at different stages of disease progression.
Assuntos
Receptores ErbB/genética , Nefropatias/genética , Falência Renal Crônica/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Adulto , Idoso , Doença Crônica , Creatinina/sangue , Feminino , Seguimentos , Estudos de Associação Genética , Humanos , Nefropatias/etiologia , Falência Renal Crônica/etiologia , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Uromodulina/genética , População Branca/genéticaRESUMO
Recent studies demonstrated a strong influence of rare genetic variants on several lipid-related traits. However, their impact on free fatty acid (FFA) plasma concentrations, as well as the role of rare variants in a general population, has not yet been thoroughly addressed. The adipose triglyceride lipase (ATGL) is encoded by the PNPLA2 gene and catalyzes the rate-limiting step of lipolysis. It represents a prominent candidate gene affecting FFA concentrations. We therefore screened the full genomic region of ATGL for mutations in 1,473 randomly selected individuals from the SAPHIR (Salzburg Atherosclerosis Prevention program in subjects at High Individual Risk) Study using a combined Ecotilling and sequencing approach and functionally investigated all detected protein variants by in-vitro studies. We observed 55 novel mostly rare genetic variants in this general population sample. Biochemical evaluation of all non-synonymous variants demonstrated the presence of several mutated but mostly still functional ATGL alleles with largely varying residual lipolytic activity. About one-quarter (3 out of 13) of the investigated variants presented a marked decrease or total loss of catalytic function. Genetic association studies using both continuous and dichotomous approaches showed a shift towards lower plasma FFA concentrations for rare variant carriers and an accumulation of variants in the lower 10%-quantile of the FFA distribution. However, the generally rather small effects suggest either only a secondary role of rare ATGL variants on the FFA levels in the SAPHIR population or a recessive action of ATGL variants. In contrast to these rather small effects, we describe here also the first patient with "neutral lipid storage disease with myopathy" (NLSDM) with a point mutation in the catalytic dyad, but otherwise intact protein.
Assuntos
Variação Genética , Lipase/genética , Lipase/metabolismo , Adulto , Idoso , Áustria , Ácidos Graxos não Esterificados/sangue , Feminino , Nível de Saúde , Humanos , Lipase/química , Masculino , Pessoa de Meia-Idade , Mutação , Polimorfismo de Nucleotídeo Único , Estrutura Terciária de Proteína , População Branca/genéticaRESUMO
Osteogenesis imperfecta (OI) is a genetically and clinically heterogeneous disorder characterized by bone fragility and reduced bone mass generally caused by defects in type I collagen structure or defects in proteins interacting with collagen processing. We identified a homozygous missense mutation in SEC16B in a child with vertebral fractures, leg bowing, short stature, muscular hypotonia, and bone densitometric and histomorphometric features in keeping with OI with distinct ultrastructural features. In line with the putative function of SEC16B as a regulator of trafficking between the ER and the Golgi complex, we showed that patient fibroblasts accumulated type I procollagen in the ER and exhibited a general trafficking defect at the level of the ER. Consequently, patient fibroblasts exhibited ER stress, enhanced autophagosome formation, and higher levels of apoptosis. Transfection of wild-type SEC16B into patient cells rescued the collagen trafficking. Mechanistically, we show that the defect is a consequence of reduced SEC16B expression, rather than due to alterations in protein function. These data suggest SEC16B as a recessive candidate gene for OI.
Assuntos
Colágeno Tipo I , Osteogênese Imperfeita , Criança , Humanos , Colágeno/genética , Colágeno Tipo I/genética , Colágeno Tipo I/química , Colágeno Tipo I/metabolismo , Mutação , Osteogênese Imperfeita/genética , Osteogênese Imperfeita/metabolismo , Estresse do Retículo EndoplasmáticoRESUMO
Lipoprotein(a) [Lp(a)] is a major cardiovascular risk factor, which is largely genetically determined by one major gene locus, the LPA gene. Many aspects of the transcriptional regulation of LPA are poorly understood and the role of epigenetics has not been addressed yet. Therefore, we conducted an epigenome-wide analysis of DNA methylation on Lp(a) levels in two population-based studies (total n = 2208). We identified a CpG site in the LPA promoter which was significantly associated with Lp(a) concentrations. Surprisingly, the identified CpG site was found to overlap the SNP rs76735376. We genotyped this SNP de-novo in three studies (total n = 7512). The minor allele of rs76735376 (1.1% minor allele frequency) was associated with increased Lp(a) values (p = 1.01e-59) and explained 3.5% of the variation of Lp(a). Statistical mediation analysis showed that the effect on Lp(a) is rather originating from the base change itself and is not mediated by DNA methylation levels. This finding is supported by eQTL data from 208 liver tissue samples from the GTEx project, which shows a significant association of the rs76735376 minor allele with increased LPA expression. To evaluate, whether the association signal at rs76735376 may actually be derived from a stronger eQTL signal in LD with this SNP, eQTL association results of all correlated SNPs (r2≥0.1) were integrated with genetic association results. This analysis pinpointed to rs10455872 as the potential trigger of the effect of rs76735376. Furthermore, both SNPs coincide with short apo(a) isoforms. Adjusting for both, rs10455872 and the apo(a) isoforms diminished the effect size of rs76735376 to 5.38 mg/dL (p = 0.0463). This indicates that the effect of rs76735376 can be explained by both an independent effect of the SNP and a strong correlation with rs10455872 and apo(a) isoforms.
Assuntos
Metilação de DNA , Estudo de Associação Genômica Ampla/métodos , Lipoproteína(a)/genética , Lipoproteína(a)/metabolismo , Fígado/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Ilhas de CpG , Epigênese Genética , Feminino , Regulação da Expressão Gênica , Frequência do Gene , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Regiões Promotoras Genéticas , Locos de Características Quantitativas , Sequenciamento Completo do GenomaRESUMO
The armamentarium of anti-leukemic drugs has increased substantially since anti-leukemic activities were recently found for a variety of non-classical cytostatic drugs, among them the histone deacetylase (HDAC) inhibitor valproic acid (VPA). This study investigated the effect of VPA on proliferation and apoptosis of human Philadelphia chromosome-positive (Ph+) acute lymphatic (ALL) and chronic myeloid leukemia (CML) cells and on colony formation of human chronic-phase CML progenitor cells. Strong anti-proliferative and pro-apoptotic effects of VPA were observed on human ALL and CML cell lines at concentrations achievable in vivo. These effects were most pronounced in ALL cell lines as well as in primary ALL cells. Notably, VPA revealed enhanced activity with imatinib mesylate, nilotinib, the farnesyl transferase inhibitor SCH66336, interferon-alpha and cytosine arabinoside. VPA inhibited the growth of colony-forming cells from 12 Ph+ chronic-phase CML patients but also of those from normal healthy controls in a dose-dependent fashion. HDAC-inhibiting activity of VPA was confirmed on ALL and CML cells. In conclusion, VPA, whether alone or in combination with other non-classical anti-leukemic compounds, exerts significant anti-leukemic effects on human ALL and CML cells.
Assuntos
Antineoplásicos/farmacologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Cromossomo Filadélfia , Piperazinas/farmacologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Pirimidinas/farmacologia , Ácido Valproico/farmacologia , Antineoplásicos/administração & dosagem , Apoptose/efeitos dos fármacos , Sequência de Bases , Benzamidas , Estudos de Casos e Controles , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Primers do DNA/genética , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/administração & dosagem , Inibidores Enzimáticos/farmacologia , Genes abl , Células-Tronco Hematopoéticas/efeitos dos fármacos , Células-Tronco Hematopoéticas/patologia , Inibidores de Histona Desacetilases , Humanos , Mesilato de Imatinib , Técnicas In Vitro , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Piperazinas/administração & dosagem , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Pirimidinas/administração & dosagem , Ácido Valproico/administração & dosagemRESUMO
We examined the methylation status of the Ep-CAM promoter region of human breast cancer cell lines and breast cancer tissue using MethyLight technology and bisulfite sequencing. We found the promoter of Ep-CAM-negative breast cancer cell lines Hs 578T to be methylated to a higher degree as compared to positive cell lines MCF-7. Demethylation of cell lines was performed using 5-aza-2'-deoxycytidine. Ep-CAM RNA and protein expression could be partially restored by treating cells with 5-Aza-2'-deoxycytidine. In most primary breast cancer tissue, methylation of the Ep-CAM gene could be detected at a low level and no correlation was found with Ep-CAM protein expression in tumour tissue. Taken together, these data suggest that methylation of the Ep-CAM promoter is not a crucial mechanism for regulation of Ep-CAM expression in breast cancer. Thus, most important regulatory mechanisms have to be supposed in vivo.
Assuntos
Antígenos de Neoplasias/genética , Neoplasias da Mama/genética , Moléculas de Adesão Celular/genética , Metilação de DNA , Regiões Promotoras Genéticas/genética , Antígenos de Neoplasias/metabolismo , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Mama/metabolismo , Neoplasias da Mama/patologia , Moléculas de Adesão Celular/metabolismo , Linhagem Celular Tumoral , Colo/metabolismo , Ilhas de CpG , Decitabina , Molécula de Adesão da Célula Epitelial , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Mucosa Intestinal/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
INTRODUCTION: Calciphylaxis/calcific uremic arteriolopathy affects mainly end-stage kidney disease patients but is also associated with malignant disorders such as myeloma, melanoma and breast cancer. Genetic risk factors of calciphylaxis have never been studied before. METHODS: We investigated 10 target genes using a tagging SNP approach: the genes encoding CD73/ ecto-5'-nucleotidase (purinergic pathway), Matrix Gla protein, Fetuin A, Bone Gla protein, VKORC1 (all related to intrinsic calcification inhibition), calcium-sensing receptor, FGF23, Klotho, vitamin D receptor, stanniocalcin 1 (all related to CKD-MBD). 144 dialysis patients from the German calciphylaxis registry were compared with 370 dialysis patients without history of CUA. Genotyping was performed using iPLEX Gold MassARRAY(Sequenom, San Diego, USA), KASP genotyping chemistry (LGC, Teddington, Middlesex, UK) or sequencing. Statistical analysis comprised logistic regression analysis with adjustment for age and sex. RESULTS: 165 SNPs were finally analyzed and 6 SNPs were associated with higher probability for calciphylaxis (OR>1) in our cohort. Nine SNPs of three genes (CD73, FGF23 and Vitamin D receptor) reached nominal significance (p< 0.05), but did not reach statistical significance after correction for multiple testing. Of the CD73 gene, rs4431401 (OR = 1.71, 95%CI 1.08-2.17, p = 0.023) and rs9444348 (OR = 1.48, 95% CI 1.11-1.97, p = 0.008) were associated with a higher probability for CUA. Of the FGF23 and VDR genes, rs7310492, rs11063118, rs13312747 and rs17882106 were associated with a higher probability for CUA. CONCLUSION: Polymorphisms in the genes encoding CD73, vitamin D receptor and FGF23 may play a role in calciphylaxis development. Although our study is the largest genetic study on calciphylaxis, it is limited by the low sample sizes. It therefore requires replication in other cohorts if available.
Assuntos
5'-Nucleotidase/genética , Calciofilaxia/genética , Fatores de Crescimento de Fibroblastos/genética , Receptores de Calcitriol/genética , Uremia/genética , Idoso , Idoso de 80 Anos ou mais , Calciofilaxia/etiologia , Estudos de Casos e Controles , Feminino , Fator de Crescimento de Fibroblastos 23 , Proteínas Ligadas por GPI/genética , Predisposição Genética para Doença , Alemanha , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Sistema de Registros , Diálise Renal/efeitos adversos , Uremia/etiologiaRESUMO
Measurement of telomere length is widely used in epidemiologic studies. Insufficient standardization of the measurements processes has, however, complicated the comparison of results between studies. We aimed to investigate whether DNA extraction methods have an influence on measured values of relative telomere length (RTL) and whether this has consequences for epidemiological studies. We performed four experiments with RTL measurement in quadruplicate by qPCR using DNA extracted with different methods: 1) a standardized validation experiment including three extraction methods (magnetic-particle-method EZ1, salting-out-method INV, phenol-chloroform-isoamyl-alcohol PCI) each in the same 20 samples demonstrated pronounced differences in RTL with lowest values with EZ1 followed by INV and PCI-isolated DNA; 2) a comparison of 307 samples from an epidemiological study showing EZ1-measurements 40% lower than INV-measurements; 3) a matching-approach of two similar non-diseased control groups including 143 pairs of subjects revealed significantly shorter RTL in EZ1 than INV-extracted DNA (0.844 ± 0.157 vs. 1.357 ± 0.242); 4) an association analysis of RTL with prevalent cardiovascular disease detected a stronger association with INV than with EZ1-extracted DNA. In summary, DNA extraction methods have a pronounced influence on the measured RTL-values. This might result in spurious or lost associations in epidemiological studies under certain circumstances.
Assuntos
DNA/genética , DNA/isolamento & purificação , Homeostase do Telômero/genética , Telômero/genética , Adulto , DNA/química , Impressões Digitais de DNA/métodos , Estudos Epidemiológicos , Feminino , Humanos , Masculino , Telômero/químicaRESUMO
Lung function is an independent predictor of mortality and serves as an aging marker in never smokers. The protein sirtuin-1 of gene SIRT1 has profound anti-inflammatory effects and regulates metabolic pathways. Its suggested longevity effects on lower organisms remain poorly studied in humans. In 1132 never smokers of the population-based SAPALDIA cohort, we investigated associations between single nucleotide polymorphisms (SNPs; rs730821, rs10997868, rs10823116) of SIRT1 and aging-related lung function decline over 11 years in terms of change in forced expiratory volume in the first second (FEV1), forced vital capacity (FVC), FEV1/FVC ratio, and forced expiratory flow between 25 and 75 % of FVC (FEF25-75) using multiple linear regression models. Interactions between the SIRT1 SNPs and adiposity parameters (body mass index (BMI), its change and weight gain) were tested by including multiplicative interaction terms into the models. SIRT1 polymorphisms exhibited no main effects, but modified the association between obesity measures and FEV1/FVC and FEF25-75 decline (p = 0.009-0.046). Per risk allele, FEV1/FVC decline was accelerated up to -0.5 % (95 % CI -1.0 to 0 %) and -0.7 % (-1.3 to -0.2 %) over interquartile range increases in BMI (2.4 kg/m(2)) or weight (6.5 kg), respectively. For FEF25-75 decline, corresponding estimates were -57 mL/s (-117 to 4 mL/s) and -76 mL/s (-1429 to -9 mL/s). Interactions were not present in participants with genetically lowered C-reactive protein concentrations. Genetic variation in SIRT1 might therefore affect lung function and human longevity by modifying subclinical inflammation arising from abdominal adipose tissue.
Assuntos
Gordura Abdominal/metabolismo , Envelhecimento , Pneumopatias/genética , Pulmão/fisiopatologia , Polimorfismo de Nucleotídeo Único , Sirtuína 1/genética , Capacidade Vital/fisiologia , Adolescente , Adulto , Índice de Massa Corporal , DNA/genética , Feminino , Seguimentos , Volume Expiratório Forçado , Genótipo , Humanos , Pneumopatias/metabolismo , Pneumopatias/fisiopatologia , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Sirtuína 1/metabolismo , Fatores de Tempo , Adulto JovemRESUMO
Carotid intima media thickness (cIMT) is a marker for subclinical atherosclerosis. The most recent genome-wide association meta-analysis (GWAMA) from the CHARGE consortium identified four genomic regions showing either significant (ZHX2, APOC1, PINX1) or suggestive evidence (SLC17A4) for an association. Here we assess these four cIMT loci in a pooled analysis of four independent studies including 5446 individuals by providing updated unbiased effect estimates of the cIMT association signals. The pooled estimates of our four independent samples pointed in the same direction and were similar to those of the GWAMA. When updating the independent second stage replication results from the earlier CHARGE GWAMA by our estimates, effect size estimates were closer to those of the original CHARGE discovery. A fine-mapping approach within a ±50 kb region around each lead SNP from CHARGE revealed 27 variants with larger estimated effect sizes than the lead SNPs but only three of them showed a r(2) > 0.40 with these respective lead SNPs from CHARGE. Some variants are located within potential functional loci.
Assuntos
Doenças Cardiovasculares/genética , Espessura Intima-Media Carotídea , Polimorfismo de Nucleotídeo Único , Adulto , Idoso , Idoso de 80 Anos ou mais , Aterosclerose/genética , Mapeamento Cromossômico , Interpretação Estatística de Dados , Saúde da Família , Feminino , Estudo de Associação Genômica Ampla , Genômica , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Observacionais como Assunto , Estudos Prospectivos , Projetos de PesquisaRESUMO
PURPOSE: The identification of malignant cells in effusions by conventional cytology is hampered by its limited sensitivity. The aim of this study was to improve tumor cell detection in effusions by molecular approaches. MATERIALS AND METHODS: A total of 157 effusions from patients with tumors and 72 effusions from patients without a history or evidence of malignancy were included in this study. All effusion specimens were evaluated in parallel by cytology, fluorescence in situ hybridization (FISH) for aneuploidy, and reverse-transcriptase polymerase chain reaction (RT-PCR) for expression of human mammaglobin (hMAM) and mammaglobin B (hMAM-B). RESULTS: In effusions from patients with tumors, the sensitivities of tumor cell detection by cytology, FISH, and hMAM and hMAM-B detection were 46.2%, 53.3%, 36.4%, and 57.7%, respectively. The corresponding specificities were 94.4%, 97.0%, 87.1%, and 88.6%. Notably, a high percentage of effusions containing malignant cells were in fact transudates, indicating the necessity for molecular diagnostic work-up of transudates collected from patients with tumors. Dependent on the tumor type, the use of appropriate marker combinations improved tumor cell detection in effusions significantly. By combining all four diagnostic tests, a positive test result indicating the presence of malignancy was achieved in 81.1%, with a fairly good specificity of 70.1%. CONCLUSION: Molecular techniques are definitely useful to detect malignancy in cytologically negative effusions. Tumor cell detection in effusions can be significantly improved by FISH and PCR techniques applying appropriate molecular markers. This finding should help to improve tumor staging, prognostic assessment, and treatment monitoring.
Assuntos
Aneuploidia , Líquido Ascítico/metabolismo , Proteínas de Neoplasias/genética , Neoplasias/metabolismo , Derrame Pleural Maligno/metabolismo , Uteroglobina/genética , Biomarcadores Tumorais/análise , Técnicas Citológicas , Células Epiteliais/metabolismo , Feminino , Humanos , Hibridização in Situ Fluorescente , Masculino , Mamoglobina A , Neoplasias/patologia , RNA Mensageiro/metabolismo , RNA Neoplásico , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sensibilidade e EspecificidadeRESUMO
Telomere length is considered as a biological marker for aging. It is expected that telomeres shorten with age and with conditions associated with oxidative stress and inflammation. Both are present in patients with chronic kidney disease (CKD) who have a very high cardiovascular risk. We investigated whether CKD duration is associated with relative telomere length (RTL) in 4802 patients from the German Chronic Kidney Disease (GCKD) study. We measured RTL in each sample in quadruplicates using a quantitative polymerase chain reaction (qPCR). We observed a U-shaped association of RTL with CKD duration: the longest RTL was found in those 339 patients who reported the shortest disease duration (<6 months) and shorter RTL in 2108 patients with duration between 6 months and less than 5 years. Most importantly, those 2331 patients who reported a CKD duration of 5 years and more had significantly longer RTL compared to those with intermediate CKD duration (6 months to less than 5 years): mean 0.954, 95%CI 0.946-0.961 versus 0.937, 95%CI 0.929-0.944, p=0.002). Due to the cross-sectional nature of the study these surprising results have to be considered with caution and as hypothesis-generating. Whether the longer RTL in patients with long-lasting disease is caused by an activation of telomerase to counteract the shortening of RTL due to oxidative stress and inflammation or whether they are caused by a survival bias needs to be investigated in longitudinal studies. Our data are in support of a higher plasticity of shortening and elongations of RTL as until recently anticipated.
Assuntos
Envelhecimento/genética , Insuficiência Renal Crônica/genética , Telômero/ultraestrutura , Idoso , Biomarcadores , Estudos Transversais , Feminino , Alemanha , Humanos , Modelos Lineares , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Reação em Cadeia da Polimerase em Tempo Real , Fatores de RiscoRESUMO
BACKGROUND: Chronic kidney disease (CKD) affects 10-15% of the general population and affected individuals are at an increased risk for cardiovascular disease (CVD). Since telomere length is considered to be involved in biological aging, we tested whether relative telomere length (RTL) might be a marker for these two diseases. METHODS: The German Chronic Kidney Disease (GCKD) study is an ongoing prospective cohort study including patients with CKD of moderate severity. RTL was measured by qPCR in 4955 out of 5217 GCKD patients at baseline. RESULTS: RTL was distributed in the cohort with a mean ± SD of 0.95 ± 0.19. CVD was present in 1266 patients. Each decrease of RTL by 0.1 unit was associated with a higher probability for prevalent CVD: OR = 1.06, 95% CI 1.02-1.11, p = 0.007 (adjusted for age, sex, eGFR, BMI, ln-CRP, smoking, hypertension, diabetes, and lipids). Similar findings were observed for history of specific CVD entities, such as coronary artery disease (OR = 1.05, p = 0.025), myocardial infarction (OR = 1.08, p = 0.013) and percutaneous transluminal coronary angioplasty (OR = 1.06, p = 0.032). The strongest associations were found for interventions at the carotid arteries (OR = 1.25, p = 0.001) as well as aortic aneurysms (OR = 1.22, p = 0.001). CONCLUSIONS: In the presence of CKD there is a significant association between shorter RTL and CVD manifestations. RTL appears to be a marker reflecting changes in homeostasis associated with CKD that may contribute to the excess CVD risk.