[Show abstract][Hide abstract] ABSTRACT: To assess candidate genes for association with osteoarthritis (OA) and identify promising genetic factors and, secondarily, to assess the candidate gene approach in OA.
A total of 199 candidate genes for association with OA were identified using Human Genome Epidemiology (HuGE) Navigator. All of their single-nucleotide polymorphisms (SNPs) with an allele frequency of >5% were assessed by fixed-effects meta-analysis of 9 genome-wide association studies (GWAS) that included 5,636 patients with knee OA and 16,972 control subjects and 4,349 patients with hip OA and 17,836 control subjects of European ancestry. An additional 5,921 individuals were genotyped for significantly associated SNPs in the meta-analysis. After correction for the number of independent tests, P values less than 1.58 × 10(-5) were considered significant.
SNPs at only 2 of the 199 candidate genes (COL11A1 and VEGF) were associated with OA in the meta-analysis. Two SNPs in COL11A1 showed association with hip OA in the combined analysis: rs4907986 (P = 1.29 × 10(-5) , odds ratio [OR] 1.12, 95% confidence interval [95% CI] 1.06-1.17) and rs1241164 (P = 1.47 × 10(-5) , OR 0.82, 95% CI 0.74-0.89). The sex-stratified analysis also showed association of COL11A1 SNP rs4908291 in women (P = 1.29 × 10(-5) , OR 0.87, 95% CI 0.82-0.92); this SNP showed linkage disequilibrium with rs4907986. A single SNP of VEGF, rs833058, showed association with hip OA in men (P = 1.35 × 10(-5) , OR 0.85, 95% CI 0.79-0.91). After additional samples were genotyped, association at one of the COL11A1 signals was reinforced, whereas association at VEGF was slightly weakened.
Two candidate genes, COL11A1 and VEGF, were significantly associated with OA in this focused meta-analysis. The remaining candidate genes were not associated.
[Show abstract][Hide abstract] ABSTRACT: The widespread use of pesticides substances nowadays largely guarantees the protection of crops and people from undesired pests. However, exposure to pesticides was related to a variety of human health effects. The present study was conducted in the region of Thessaly which is characterized by intensive agricultural activities and wide use of pesticides. The study aimed at estimating the oxidative damage to DNA in different subpopulations in Thessaly region (Greece) and investigating its correlation with exposure to pesticides and other potential risk factors. In total, the study involved 80 pesticide sprayers, 85 rural residents and 121 individuals, inhabitants of the city of Larissa. Demographic characteristics, habits, medical history and exposure history of the participants to pesticides were recorded by personal interviews. Blood and urine samples were collected from all participants. For the measurement of exposure to organophosphorus insecticides, dialkylphosphate (DAP) metabolites were quantified in urine, by gas chromatography–mass spectrometry. Genomic DNA was extracted from peripheral blood samples and the oxidation by-product 8-hydroxydeoxyguanosine (8-OHdG) was determined by Enzyme Immuno-Assay. Urinary metabolite concentrations were not associated with 8-OHdG levels but it was found that pesticide sprayers had significantly higher levels of 8-OHdG (p = 0.007) in comparison to the control group. Last season's exposure to insecticides and fungicides, expressed as total area treated multiplied by the number of applications, showed a statistically significant association with the risk of having high 8-OHdG levels [RR: 2.19 (95%CI:1.09–4.38) and RR: 2.32 (95% CI:1.16–4.64) respectively]. Additionally, from the subgroups of pesticides examined, seasonal exposure to neonicotinoid insecticides [RR: 2.22 (95% CI:1.07–4.63)] and glufosinate ammonium [RR: 3.26 (95% CI:1.38–7.69)] was found to have the greater impact on 8-OHdG levels.
This study produced findings that support the hypothesis that pesticide exposure is involved in the induction of oxidative damage to DNA and identified chemical groups of pesticides which should be given greater attention in future investigations.
Science of The Total Environment 01/2014; 496:358–364. · 3.26 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Glucuronidation, mediated by the UDP-glucuronosyltransferase 1A1 (UGT1A1) enzyme, is an important metabolic process during which steroids are converted to more easily excreted compounds in steroid target tissues, such as the prostate. The aim of our study was to investigate the possible correlation between UGT1A1 promoter gene polymorphism and benign prostatic hyperplasia. 421 blood samples were obtained from 138 consecutive patients diagnosed with benign prostatic hypeplasia (BPH group) and 283 healthy volunteers (control group). A(TA)6TAA promoter polymorphism of UGT1A1 gene was studied using the Fragment Analysis Software of an automated DNA sequencer and three genotypes (homozygous 7/7, heterozygous 6/7 and normal homozygous 6/6) were identified. No significant differences were observed between the BPH group and controls regarding the genotyping distribution of the three UGT1A1 promoter genotypes (P = 0.39). Also, no association was found between overall disease risk and the presence of the polymorphic homozygous genotype (TA(7)/TA)7) vs. TA(6)/TA(7) + TA(6)/TA(6)) (P = 0.31). Our data suggest that the TA repeat polymorphism of UGT1A1 is not associated with increased BPH risk susceptibility in Caucasian men.
[Show abstract][Hide abstract] ABSTRACT: Osteoarthritis (OA) is the most common form of arthritis with a clear genetic component. To identify novel loci associated with hip OA we performed a meta-analysis of genome-wide association studies (GWAS) on European subjects.
We performed a two-stage meta-analysis on more than 78 000 participants. In stage 1, we synthesised data from eight GWAS whereas data from 10 centres were used for 'in silico' or 'de novo' replication. Besides the main analysis, a stratified by sex analysis was performed to detect possible sex-specific signals. Meta-analysis was performed using inverse-variance fixed effects models. A random effects approach was also used.
We accumulated 11 277 cases of radiographic and symptomatic hip OA. We prioritised eight single nucleotide polymorphism (SNPs) for follow-up in the discovery stage (4349 OA cases); five from the combined analysis, two male specific and one female specific. One locus, at 20q13, represented by rs6094710 (minor allele frequency (MAF) 4%) near the NCOA3 (nuclear receptor coactivator 3) gene, reached genome-wide significance level with p=7.9×10(-9) and OR=1.28 (95% CI 1.18 to 1.39) in the combined analysis of discovery (p=5.6×10(-8)) and follow-up studies (p=7.3×10(-4)). We showed that this gene is expressed in articular cartilage and its expression was significantly reduced in OA-affected cartilage. Moreover, two loci remained suggestive associated; rs5009270 at 7q31 (MAF 30%, p=9.9×10(-7), OR=1.10) and rs3757837 at 7p13 (MAF 6%, p=2.2×10(-6), OR=1.27 in male specific analysis).
Novel genetic loci for hip OA were found in this meta-analysis of GWAS.
Annals of the rheumatic diseases 08/2013; · 8.11 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cell adhesion proteins that connect each cell to neighboring cells and the extracellular matrix play a fundamental role in metastasis. Mitogen-inducible gene-2 (MIG2), is a cell-matrix adhesion protein, which through migfilin, interacts with filamin-A, being linked to actin cytoskeleton. Aim: Recent studies have implicated both MIG2 and migfilin in cancer, but little is known regarding their expression in breast cancer. In this study, we investigated this topic.
mRNA and protein expression was examined in 30 breast cancer samples and compared to that of normal adjacent tissue using real time-polymerase chain reaction (PCR) and western blotting.
Our results showed that expression of MIG2 and migfilin was significantly reduced in the majority of the breast cancer tissues compared to normal tissues regardless of metastatic status and disease stage.
Both MIG2 and migfilin are down-regulated in breast cancer.
Anticancer research 05/2013; 33(5):1977-81. · 1.71 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Fibroblast growth factor 23 (FGF-23) is a bone-derived circulating phosphaturic factor that decreases serum concentration of phosphate and vitamin D, suggested to actively participate in a complex renal-gastrointestinal-skeletal axis. Serum FGF-23 concentrations, as well as various other laboratory parameters involved in bone homeostasis, were measured and analyzed with regard to various diseases and patients' characteristics in 44 patients with Crohn disease (CD) and 20 healthy controls (HCs) included in this cross-sectional study. Serum FGF-23 levels were significantly lower in patients with CD (900.42 ± 815.85pg/mL) compared with HC (1410.94 ± 1000.53pg/mL), p = 0.037. Further analyses suggested FGF-23 as a factor independent from various parameters including age (r = -0.218), body mass index (r = -0.115), 25-hydroxy vitamin D (r = 0.126), parathyroid hormone (r = 0.084), and bone mineral density (BMD) of hip and lumbar (r = 0.205 and r = 0.149, respectively). This observation remained even after multivariate analyses, exhibiting that BMD was not affected by FGF-23, although parameters such as age (p = 0.026), cumulative prednisolone dose (p < 0.0001), and smoking status (p = 0.024) were strong determinants of BMD regarding hip. Lower FGF-23 levels in patients with bowel inflammation are accompanied but not directly correlated with lower vitamin D levels, showing no impact on BMD determination of young adults with CD. The downregulation of serum FGF-23 levels in CD appears as a secondary compensatory effect on the bone and mineral metabolism induced by chronic intestinal inflammation.
Journal of Clinical Densitometry 04/2013; · 1.71 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Osteoarthritis (OA) is a debilitating disease of the joints characterized by cartilage degradation but to date there is no available pharmacological treatment to inhibit disease progression neither is there any available biomarker to predict its development. In the present study, we examined the expression level and possible involvement of novel cell-ECM adhesion-related molecules such as Iintegrin Linked Kinase (ILK), PINCH, parvin, Mig-2 and Migfilin in OA pathogenesis using primary human articular chondrocytes from healthy individuals and OA patients. Our findings show that only ILK and Migfilin were upregulated in OA compared to the normal chondrocytes. Interestingly, Migfilin silencing in OA chondrocytes rather exacerbated than ameliorated the osteoarthritic phenotype, as it resulted in even higher levels of catabolic and hypertrophic markers while at the same time induced reduction in ECM molecules such as aggrecan. Furthermore, we also provide a link between Migfilin and β-catenin activation in OA chondrocytes, showing Migfilin to be inversely correlated with β-catenin. Thus, the present study emphasizes for the first time to our knowledge the role of Migfilin in OA and highlights the importance of cell-ECM adhesion proteins in OA pathogenesis.
Biochemical and Biophysical Research Communications 12/2012; · 2.41 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: INTRODUCTION: We aimed to explore the involvement of a multiallelic functional polymorphism in knee osteoarthritis (OA) susceptibility as a prototype of possible genetic factors escaping GWAS detection. METHODS: OA patients and controls from three European populations (Greece, Spain and the UK) adding up to 1003 patients (716 women, 287 men) that had undergone total knee joint replacement (TKR) due to severe primary OA and 1543 controls (758 women, 785 men) lacking clinical signs or symptoms of OA were genotyped for the D6S1276 microsatellite in intron 1 of BMP5. Genotype and mutiallelic trend tests were used to compare cases and controls. RESULTS: Significant association was found between the microsatellite and knee OA in women (P from 3.1 x10-4 to 4.1 x10-4 depending on the test), but not in men. Three of the alleles showed significant differences between patients and controls, one of them of increased risk and two of protection. The gender association and the allele direction of change were very concordant with the previously reported for hip OA. CONCLUSIONS: We have found association of knee OA in women with the D6S1276 functional microsatellite that modifies in cis the expression of BMP5 making this a sounder OA genetic factor and extending its involvement to other joint. This result also shows the interest of analysing other multiallelic polymorphisms.
Arthritis research & therapy 11/2012; 14(6):R257. · 4.27 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: BACKROUND: A case control study to evaluate whether a single serum measurement of Angiopoietin-1 (ANG-1) and Angiopoietin-2 (ANG-2) at 6-8weeks gestation can differentiate failed pregnancies, whether ectopic pregnancies (EP) or missed abortions (MA), from healthy intrauterine pregnancies (IUP). INTERVENTION(S): Serum and tissue mRNA determination of ANG-1 and ANG-2 levels by ELISA and RTPCR,from 60 (30 EP, 30 MA) patients with failed early pregnancy and 33 IUPs. RESULTS: ANG-1 and ANG-2 concentrations and their ratio are lower in EP (median, 689 and 302pg/ml, respectively) and MA cases (median, 810 and 402pg/ml, respectively) compared to IUP (median, 963 and 1477pg/ml, respectively) (p<0.05, for all). Unlike ANG-2, serum ANG- 1 discriminates an EP from a MA (p=0.011). Trophoblastic ANG-1 mRNA expression levels are lower in EP compared to MA and IUP (p<0.05), while ANG-2 mRNA is higher in EP and MA than in IUP (p<0.05). CONCLUSIONS: A single measurement of serum ANG-1 and ANG-2 at 6-8weeks of gestation designate the outcome of a pregnancy, as their levels are significantly decreased in failed than normal pregnancies. Serum ANG-1 showed potential to discriminate MA from EP.
Clinica chimica acta; international journal of clinical chemistry 10/2012; · 2.54 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Clinical and molecular description of a fetus in prenatal diagnosis with a rare de novo ring 10 and deletions of 12.59 Mb in 10p15.3ep14 and 4.22 Mb in 10q26.3 a b s t r a c t Ring chromosomes are rare cytogenetic findings and are mostly associated with an abnormal phenotype. We report on the prenatal diagnosis of a ring chromosome 10 in a fetus in which talipes equinovarus was incidentally found during routine obstetric ultrasound at 22 weeks of gestation. Amniocentesis was undertaken and cytogenetic analysis revealed a de novo non-mosaic apparently stable ring chromosome 10 replacing one of the two homologs. Multiplex Ligation-dependent Probe Amplification (MLPA) revealed subtelomeric deletions in both the short and long arm of chromosome 10. Analysis with high resolution micro-array based comparative genomic hybridization (array-CGH), defined the ring chro-mosome as del 10p15.3ep14 (12.59 Mb in size) and del 10q26.3 (4.22 Mb in size) and revealed the genes that are deleted. After elected termination of the pregnancy at 27th week of gestation a detailed autopsy of the fetus allowed for genotypeephenotype correlations. To our knowledge, this is the first case of a de novo ring chromosome 10 which is reported during prenatal diagnosis and is thoroughly investigated with array CGH and autopsy study. Ó 2011 Elsevier Masson SAS. All rights reserved. 1. Methods of detection 1.1. Cytogenetics Chromosomal analyses were performed on amniotic fluid, parental peripheral blood, placenta and fetal skin using conven-tional GTG-banding techniques at the 550 band level. 1.2. Array-CGH Agilent Human Genome CGH 244K microarrays with an average spatial resolution of 12 kb were used in the study (Agilent Tech-nologies, Santa Clara, CA). Genomic DNA from the proband and pooled normal male reference DNA (Promega Corporation, Madi-son, WI) were digested with AluI and RsaI (Promega Corporation) and labeled with Agilent Genomic DNA labeling Kit according to manufacturer's instructions. Patient and reference DNA were labeled with Cy3 and Cy5, respectively and were co-hybridized to arrays for 40 h at 65 C in a rotating oven (Agilent Technologies) at 20 rpm. The arrays were then washed and scanned with an Agilent Microarray Scanner. Data were extracted using Feature Extraction 9.1 software (Agilent Technologies) and analyzed using CGH Analytics 3.4 software (Agilent Technologies). Genomic copy number changes were identified with the assistance of the Aber-ration Detection Method 1 algorithm with a threshold of 6. Centralization and fuzzy zero corrections were applied to remove putative variant intervals with small average log 2 ratios. For the location of genes in the deleted/duplicated genomic segments the) and the Ensemble Genome Browser (http://www.ensembl.org) were used. 1.3. Chromosomal anomalies The fetus presented with talipes equinovarus at 22 weeks of gestation during a routine scan and an amniocentesis was per-formed. Rapid QF-PCR (quantitative fluorescence PCR) results were consistent with euploidy for the chromosomes investigated (13, 18, 21, X and Y). Conventional cytogenetic analysis revealed a 46,XY,r(10) abnormal non-mosaic fetal karyotype with a ring
[Show abstract][Hide abstract] ABSTRACT: Events normally taking place in the terminal chondrocyte differentiation in the growth plate are also observed during osteoarthritis (OA) development, suggesting that molecules, such as Wnts and bone morphogenetic proteins (BMPs) regulating chondrocyte activity in the growth plate, may play a key role in osteoarthritis pathogenesis. The aim of the study was to investigate the possible cross-talk between BMP-2 and Wnt/β-catenin pathways in OA progression.
Low-density-lipoprotein receptor-related protein 5 (LRP-5) and 6, BMP-2, -4, and -7, bone morphogenetic protein receptor-IA and IB (BMPR-IA and BMPR-IA), lymphoid enhancer factor-1 (LEF-1), and transcription factor 4 (TCF-4) expression levels were investigated in normal and osteoarthritic chondrocytes. LRP-5, β-catenin (phospho and active form), matrix metalloproteinases (MMPs) 7, 9, 13, 14, ADAMTS-4, 5, as well as collagen X (COL10A1) expression levels were evaluated after LRP-5 silencing in BMP-2-treated chondrocytes. The investigation of Smad1/5/8 binding to LRP-5 promoter was assessed with chromatin immunoprecipitation (ChIP). Furthermore, we evaluated the effect of experimental activation of the Wnt/β-catenin pathway with LiCl and LEF-1 silencing, in LiCl-treated chondrocytes, on matrix metalloproteinases (MMPs) 7, 9, 13, 14, ADAMTS-4, 5, and collagen X (COL10A1) expression, as well as possible interactions between LEF-1 and MMPs and COL10A1 promoters by using a ChIP assay.
LRP-5, BMP-2, BMP-4, BMPR-IA, and LEF-1 mRNA and protein expression levels were found to be significantly upregulated in osteoarthritic chondrocytes compared with normal. We showed that treatment of cultured chondrocytes with BMP-2 resulted in increased β-catenin nuclear translocation and LRP-5 expression and that the BMP-2-induced LRP-5 upregulation is mediated through Smad1/5/8 binding on LRP-5 promoter. LRP-5 silencing reduced nuclear β-catenin protein levels, MMPs and collagen X expression, whereas increased phospho-β-catenin protein levels in BMP-2-treated chondrocyte. Furthermore, we demonstrated that activation of the Wnt/β-catenin signaling pathway by LiCl and LEF-1 downregulation by using siRNA regulates MMP-9, 13, 14, ADAMTS-5, and COL10A1 expression, evidenced by the observed strong binding of LEF-1 to MMP-9, 13, 14, ADAMTS-5 and COL10A promoters.
Our findings suggest, for the first time to our knowledge, that BMP-2-induced Wnt/β-catenin signaling activation through LRP-5 may contribute to chondrocyte hypertrophy and cartilage degradation in osteoarthritis.
Arthritis research & therapy 04/2012; 14(2):R82. · 4.27 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Hypertrophy and impaired mineralization are two processes closely associated with osteoarthritis (OA). 1,25-dihydroxyvitamin D(3) (1a,25(OH)(2)D(3)) and inorganic phosphate (Pi) are two important factors that are implicated in calcium and phosphate homeostasis of bone metabolism and both can be regulated by the circulating phosphaturic factor fibroblast growth factor 23 (FGF23). The objective of this study was to investigate the role of 1a,25(OH)(2)D(3) and Pi and the molecular mechanism through which they contribute to hypertrophy and mineralization in human osteoarthritic chondrocytes. For this purpose, primary human chondrocytes were obtained from articular cartilage which was collected after total knee replacement surgery in OA patients. FGF23, fibroblast growth factor receptor 1c (FGFR1c), vitamin D(3) receptor (VDR), and phosphate inorganic transporter-1 and -2 (PiT-1 and PiT-2) expression levels were evaluated and found to be significantly higher in OA chondrocytes compared with normal. In addition, we observed that the binding of FGF23 to FGFR1c was stronger in OA chondrocytes compared with normal. Chromatin immunoprecipitation (ChIP) assay revealed, for the first time, the presence of two vitamin D response elements (VDREs) in the FGF23 promoter. Treatment of normal chondrocytes with 1a,25(OH)(2)D(3) or Pi resulted in significant up-regulation of VDR, FGF23, PiT-1, PiT-2 mRNA and protein levels, extracellular signal-regulated kinases 1/2 (ERK1/2) phosphorylation and induction of hypertrophy markers collagen type X (COL10A1), osteopontin (OPN), osteocalcin (OC), catabolic markers metalloproteinase-13 (MMP-13) and the apoptotic marker caspase-9. Furthermore, VDR silencing in OA chondrocytes negatively regulated FGF23, COL10A1, OPN, OC, MMP-13 and caspase-9 expressions and ERK1/2 phosphorylation. Finally, combined VDR silencing and PiT-1, PiT-2 inhibition in OA chondrocytes resulted in additive down-regulation of FGF23 expression, ERK1/2 activation and COL10A1, OPN, OC, MMP-13 and caspase-9 expression levels. We propose that 1a,25(OH)(2)D(3) and Pi act synergistically through FGF23 signaling and ERK1/2 phosphorylation contributing to late hypertrophic events and impaired mineralization in osteoarthritic chondrocytes.
Experimental Biology and Medicine 03/2012; 237(3):241-53. · 2.80 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Osteoarthritis (OA) is a common disease with a genetic component for its etiology. Recently, a genetic association of a single nucleotide polymorphism (SNP), rs17039192 in HIF-2α with knee OA has been reported in a Japanese population; however, controversy exits for its replication and a role of HIF-2α in OA. This study aimed to evaluate the association of the SNP by a large-scale replication study. A total of 8,457 subjects (3,129 OA cases and 5,328 controls) from seven independent cohorts from six countries (Japan, China, Taiwan, Korea, Greece, and Australia) were recruited and genotyped. The association of rs17039192 with knee OA was evaluated by meta-analyses. The association of the HIF-2α SNP was not replicated in any of the populations. Contrary to the previous report, the odds ratios (ORs) of the risk allele frequency were all less than 1. A combined analysis for the seven populations also showed no replication of the association (OR = 0.91, 95% confidence interval = 0.81-1.03). Our large-scale meta-analysis showed that the association of rs17039192 in HIF-2α with knee OA is negative. The significance of HIF-2α in human OA (idiopathic OA as a common disease) should be further evaluated carefully.
Journal of Orthopaedic Research 01/2012; 30(8):1244-8. · 2.88 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Recent studies have implied that osteoarthritis (OA) is a metabolic disease linked to deregulation of genes involved in lipid metabolism and cholesterol efflux. Sterol Regulatory Element Binding Proteins (SREBPs) are transcription factors regulating lipid metabolism with so far no association with OA. Our aim was to test the hypothesis that SREBP-2, a gene that plays a key role in cholesterol homeostasis, is crucially involved in OA pathogenesis and to identify possible mechanisms of action.
We performed a genetic association analysis using a cohort of 1,410 Greek OA patients and healthy controls and found significant association between single nucleotide polymorphism (SNP) 1784G>C in SREBP-2 gene and OA development. Moreover, the above SNP was functionally active, as normal chondrocytes' transfection with SREBP-2-G/C plasmid resulted in interleukin-1β and metalloproteinase-13 (MMP-13) upregulation. We also evaluated SREBP-2, its target gene 3-hydroxy-3-methylglutaryl-coenzymeA reductase (HMGCR), phospho-phosphoinositide3-kinase (PI3K), phospho-Akt, integrin-alphaV (ITGAV) and transforming growth factor-β (TGF-β) mRNA and protein expression levels in osteoarthritic and normal chondrocytes and found that they were all significantly elevated in OA chondrocytes. To test whether TGF-β alone can induce SREBP-2, we treated normal chondrocytes with TGF-β and found significant upregulation of SREBP-2, HMGCR, phospho-PI3K and MMP-13. We also showed that TGF-β activated aggrecan (ACAN) in chondrocytes only through Smad3, which interacts with SREBP-2. Finally, we examined the effect of an integrin inhibitor, cyclo-RGDFV peptide, on osteoarthritic chondrocytes, and found that it resulted in significant upregulation of ACAN and downregulation of SREBP-2, HMGCR, phospho-PI3K and MMP-13 expression levels.
We demonstrated, for the first time, the association of SREBP-2 with OA pathogenesis and provided evidence on the molecular mechanism involved. We suggest that TGF-β induces SREBP-2 pathway activation through ITGAV and PI3K playing a key role in OA and that integrin blockage may be a potential molecular target for OA treatment.
PLoS ONE 01/2012; 7(5):e35753. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Ring chromosomes are rare cytogenetic findings and are mostly associated with an abnormal phenotype. We report on the prenatal diagnosis of a ring chromosome 10 in a fetus in which talipes equinovarus was incidentally found during routine obstetric ultrasound at 22 weeks of gestation. Amniocentesis was undertaken and cytogenetic analysis revealed a de novo non-mosaic apparently stable ring chromosome 10 replacing one of the two homologs. Multiplex Ligation-dependent Probe Amplification (MLPA) revealed subtelomeric deletions in both the short and long arm of chromosome 10. Analysis with high resolution micro-array based comparative genomic hybridization (array-CGH), defined the ring chromosome as del 10p15.3-p14 (12.59 Mb in size) and del 10q26.3 (4.22 Mb in size) and revealed the genes that are deleted. After elected termination of the pregnancy at 27th week of gestation a detailed autopsy of the fetus allowed for genotype-phenotype correlations. To our knowledge, this is the first case of a de novo ring chromosome 10 which is reported during prenatal diagnosis and is thoroughly investigated with array CGH and autopsy study.
European journal of medical genetics 09/2011; 55(1):75-9. · 1.57 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Radiotherapy is an important treatment modality against cancer resulting in apoptosis and inhibition of cell growth. Survivin is an important cancer biomarker conferring to tumour cells increased survival potential by inhibiting apoptosis. In the present study, we investigated the implication of breast cancer cells features, as hormone receptors and p53 status, in the radio-resistance of breast cancer cells and in the regulation of survivin's expression by nuclear factor (NF)-κB and c-myc. Six breast cancer cell lines Michigan Cancer Foundation (MCF-7), MCF-7/Human Epidermal Growth Factor Receptor (HER)2, M. D. Anderson - Metastatic Breast (MDA-MB-231), SK-BR-3, BT-474 and Human Breast Lactating (HBL-100) were irradiated and cell viability as well as cell cycle distribution were evaluated by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry, respectively. Survivin mRNA and protein levels were evaluated by real time PCR and Western blot analysis. Survivin and HER2 gene knockdown was performed with siRNA technology and investigation of transcription factors binding to survivin and c-myc gene promoters was assessed by chromatin immunoprecipitation. Student's t-test and F-statistics were used for statistical evaluation. Our results demonstrated that only HER2(+) breast cancer cells up-regulated survivin upon irradiation, whereas HER2 knockdown in HER2(+) cells led to survivin's down-regulation. Survivin and especially HER2 knockdown abolished the observed G2/M cell cycle checkpoint and reduced the radio-resistance of HER2 overexpressing breast cancer cells. Additionally, HER2 was found to regulate survivin's expression through NF-κB and c-myc transcription factors. This study revealed the significance of HER2 in the radio-resistance of HER2(+) breast cancer cells through induction of transcription factors NF-κB and c-myc, leading to activation of survivin, a downstream target oncogene preventing apoptosis.
Journal of Cellular and Molecular Medicine 07/2011; 15(7):1542-50. · 4.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Diagnosis of early pregnancy failure is hampered by the lack of reliable serological markers.
We assessed whether a single serum measurement of placental growth factor (PlGF) and the soluble Flt-1 (sFlt-1) receptor of vascular endothelial growth factor at 6-8 wk gestation could differentiate failed pregnancies, whether ectopic pregnancies (EP) or missed abortions (MA), from healthy intrauterine pregnancies (IUP).
We conducted a prospective clinical study at a tertiary university hospital between January 2009 and February 2011.
A total of 78 consecutive patients (38 EP, 40 MA) with failed early pregnancy and 50 IUP (control group) participated in the study. Intervention(s): Determination of serum PlGF and sFlt-1 has been carried out by ELISA. Gene expression of PlGF and Flt-1 in trophoblasts was performed by RT-PCR.
We investigated whether a single, combined serum measurement of the above markers could contribute to the differential diagnosis.
PlGF and sFlt-1 concentration was lower in both EP (mean, 14.60 ± 3.42/178.16 ± 76.03 pg/ml) and MA (mean, 16.25 ± 4.73/399.42 ± 337.54 pg/ml) compared to IUP (mean, 21.64 ± 5.68/1390.32 ± 655.37 pg/ml). sFlt-1 (P = 0.033) and sFlt-1/PlGF ratio (P = 0.029) but not PlGF had the ability to discriminate MA from EP. Compared to women with viable IUP, mRNA gene expression levels of PlGF and Flt-1 were considerably lower in women with MA and in women with EP.
Combined measurement of sFlt-1 and PlGF levels can differentiate normal from failed pregnancies, whereas sFlt-1 as well as sFlt-1/PlGF ratio can also discriminate EP from MA. PlGF and Flt-1 gene expression in trophoblasts from women with EP and MA appears impaired.
The Journal of clinical endocrinology and metabolism 06/2011; 96(9):E1444-51. · 6.50 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Telomerase activity (TA), frequently observed in cancer, compensates for telomere shortening thus preventing cell senescence and conferring resistance to therapy. In the present study, we investigated the expression of human telomerase reverse transcriptase (hTERT) and TA and their regulation, as well as apoptotic rates and correlation with the presence of human epidermal growth factor receptor 2 (HER2), in irradiated tumour-derived breast cancer cells.
In 50 breast cancer tissue samples hTERT mRNA expression and TA were correlated with cell features (HER2, Estrogen and Progesterone Receptor status). Cells from six samples were then irradiated with 10 and 20 Gy; apoptotic rates were measured by flow cytometry, hTERT mRNA expression by real-time polymerase chain reaction and TA by telomeric repeat amplification protocol assay, at 24-144 h post-irradiation. Chromatin immunoprecipitation was performed to investigate hTERT and cellular-myelocytomatosis (c-myc) promoters' activity. HER2 gene knockdown was performed using small interfering RNA technology.
hTERT/TA were found increased only in irradiated HER2-positive cells, which were found to be more radioresistant, while HER2 knockdown led to hTERT/TA downregulation. HER2 was found to mediate hTERT expression through activation of Nuclear Factor-kappa B (NF-κB) and c-myc.
The present study suggests that following irradiation, HER2 receptor activates hTERT/telomerase, increasing the breast cancer cells' survival potential, through sequential induction of transcription factors NF-κΒ and c-myc.
International Journal of Radiation Biology 06/2011; 87(6):609-21. · 1.84 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Toll-like receptors (TLRs) are involved in mediating cell activation on stimulation with microbial components. Our objective was to investigate the role of TLR-2 mediated by the NF-κB pathway in septic arthritic chondrocytes. TLR-1, -2, and -6 mRNA expression levels were investigated in septic and normal chondrocytes using real-time reverse transcription-PCR. TLR-2 and MMP-13 mRNA and protein levels were measured using real-time PCR and Western blot analysis, respectively. Blocking TLR-2 mRNA expression was performed using small interfering RNA (siRNA) against TLR-2 and subsequently MMP-3, MMP-13, IL-1β, and IL-6 mRNA levels, as well as p65 NF-κB, IkBα, and MMP-13 protein levels were evaluated using real-time PCR and Western blot analysis. IL-6 protein levels were measured using ELISA assay. We observed that TLR-1, -2, and -6 mRNA expression levels were significantly higher in septic compared to normal chondrocytes. MMP-13 mRNA and protein expressions were also significantly upregulated in septic arthritic cartilage. Blocking TLR-2 mRNA expression in septic chondrocytes resulted in significant increase of inactivated nonphosphorylated p65 NF-κB and IkBα protein levels and reduction in MMP-13, IL-1β, and IL-6 expression. Our findings suggest the pro-inflammatory and catabolic role of TLR-2 mediated by the NF-κB pathway in septic arthritis. Modulation of TLR-mediated signaling may be a potential therapeutic strategy for the prevention of postinfectious cartilage degradation in articular joints.
Journal of Orthopaedic Research 02/2011; 29(2):247-51. · 2.88 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To study whether common genetic variants of the genes involved in the complex regulatory mechanism determining the intracellular bio-availability of T3 influence osteoarthritis onset.
In total 17 genetic variants within the genes encoding WD40-repeat/SOCS-box protein 1, ubiquitin specific protease 33, thyroid hormone receptor α, deiodinase, iodothyronine, type III (DIO3) and Indian hedgehog were measured and associated with osteoarthritis in a meta-analyses in European populations from the UK, The Netherlands, Greece and Spain containing a total of 3252 osteoarthritis cases and 2132 controls.
The minor allele of the DIO3 variant rs945006 showed suggestive evidence for protective association in the overall meta-analyses, which was supported by individual osteoarthritis studies and osteoarthritis subtypes. The association appeared most significant in cases with knee and/or hip with an allelic OR of 0.81 (95% CI 0.70 to 0.930) with a nominal p value of 0.004 and a permutation-based corrected p value for multiple testing of 0.039.
The findings suggest that the DIO3 gene modulates osteoarthritis disease risk; however, additional studies are necessary to replicate our findings. To elucidate the molecular mechanisms focus should be on the local adaptation to T3 availability either during the endochondral ossification process or during ageing of the articular cartilage.
Annals of the rheumatic diseases 01/2011; 70(1):164-7. · 8.11 Impact Factor