Polymorphisms in the osteopontin promoter affect its transcriptional activity
ABSTRACT Understanding the molecular mechanisms that underlie regulation of transcription of the human osteopontin encoding gene (OPN) may help to clarify several processes, such as fibrotic evolution of organ damage, tumorigenesis and metastasis, and immune response, in which OPN overexpression is observed. With the aim to evaluate variants with functional effect on transcription, we have analyzed the promoter region and focused our investigation on three common variants present in the first 500 bp upstream of the transcription start site. Transfection of constructs carrying the four most frequent haplotypes relative to variants at -66, -156, and -443 fused to the luciferase reporter gene in a panel of different cell lines showed that one haplotype conferred a significantly reduced level of reporter gene expression in all tested cell lines. We describe that the -66 polymorphism modifies the binding affinity for the SP1/SP3 transcription factors, the -156 polymorphism is included in a yet uncharacterized RUNX2 binding site, and the -443 polymorphism causes differential binding of an unknown factor. The finding of differential effects of various combination of variants in haplotypes may contribute to explain data of association studies reported in several already published articles. Future association studies using haplotypes instead of single OPN variants will allow to achieve more accurate results referable to differential expression of OPN in several common diseases, in which OPN is considered a candidate susceptibility gene.
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ABSTRACT: Duchenne muscular dystrophy (DMD) is an X-linked human disorder in which absence of the protein dystrophin causes degeneration of skeletal and cardiac muscle. For the sake of treatment development, over and above definitive genetic and cell-based therapies, there is considerable interest in drugs that target downstream disease mechanisms. Drug candidates have typically been chosen based on the nature of pathologic lesions and presumed underlying mechanisms and then tested in animal models. Mammalian dystrophinopathies have been characterized in mice (mdx mouse) and dogs (golden retriever muscular dystrophy [GRMD]). Despite promising results in the mdx mouse, some therapies have not shown efficacy in DMD. Although the GRMD model offers a higher hurdle for translation, dogs have primarily been used to test genetic and cellular therapies where there is greater risk. Failed translation of animal studies to DMD raises questions about the propriety of methods and models used to identify drug targets and test efficacy of pharmacologic intervention. The mdx mouse and GRMD dog are genetically homologous to DMD but not necessarily analogous. Subcellular species differences are undoubtedly magnified at the whole-body level in clinical trials. This problem is compounded by disparate cultures in clinical trials and preclinical studies, pointing to a need for greater rigor and transparency in animal experiments. Molecular assays such as mRNA arrays and genome-wide association studies allow identification of genetic drug targets more closely tied to disease pathogenesis. Genes in which polymorphisms have been directly linked to DMD disease progression, as with osteopontin, are particularly attractive targets.ILAR journal / National Research Council, Institute of Laboratory Animal Resources 06/2014; 55(1):119-49. DOI:10.1093/ilar/ilu011 · 1.05 Impact Factor
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ABSTRACT: Carotid intima media thickness (CIMT) is clearly associated with atherosclerosis. Studies in ischemic stroke patients reveal that there is a significant association between CIMT with monocyte chemoattractant protein-1 (MCP-1) and osteopontin (OPN) promoter polymorphism. This research aims to explain the effect of MCP-1 and OPN promoter polymorphism toward CIMT changes identified in Javanese Indonesian children. Subjects were 54 children: 27 were from parents with ischemic stroke (cases), and 27 were from healthy parents (controlled). The CIMT was examined by utilizing high resolution B-mode ultrasound. Physical examination and genotyping analysis of MCP-1 promoter were conducted by employing PCR method. Research results indicate that two polymorphisms were obtained, that is, A-2138T and G-2464A, respectively. A-2138T polymorphism was found in 5% of case children and in 14.3% of controlled children. G-2464A polymorphism was found in 5% of case children. CIMT of case children was significantly different from that of controlled children (0.61 ± 0.012 mm versus, 0.52 ± 0.015 mm, P = 0.021). Subjects with MCP-1 promoter polymorphism have 1.471 times higher tendency to have thicker CIMT than subjects with no polymorphism in MCP1 promoter. OPN promoter T-66G was also studied but it did not indicate occurrence of polymorphism in samples.04/2014; 2014:176535. DOI:10.1155/2014/176535
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ABSTRACT: BACKGROUND: In chronic heart failure (CHF), it remains unclear whether the minimal dose of beta-blockade is related to survival benefits and which parameter predicts morbidity and mortality. We sought to determine the minimal dose related to survival benefits by comparing the efficacy and safety of three doses of carvedilol and the best predictive parameter for effective outcomes in Japanese patients with CHF. METHODS: In this prospective, randomized, stratified trial, 364 patients with mild to moderate CHF were assigned to a daily carvedilol dose of 2.5, 5, or 20mg, plus optimal standard therapy. FINDINGS: During the mean 3-year follow-up, resting heart rate (HR) and BNP were significantly reduced with dose-response relations in the early period but without dose-response relations in the late period. The LVEF and the LVDd were increased and decreased, respectively, without a dose-response relation. No significant difference was seen in the composite primary endpoint of all-cause mortality and hospitalization for cardiovascular diseases and heart failure. Multivariate analysis indicated early decreases in HR and BNP predicted long-term outcomes. However, adverse events increased dose-dependently. Among 237 polymorphisms in 87 heart failure-related genes, the osteopontin G-156 del genotype was associated with an event-free survival rate (Wilcoxon test, P=0.030). CONCLUSIONS: A low carvedilol dose is effective if the HR and/or plasma BNP has been reduced. Carvedilol therapy should be guided by reductions in HR and/or BNP, especially by initial HR reduction, but not only by its dose. OPN might be a surrogate genetic marker for long-term event-free survival.International journal of cardiology 12/2012; 164(2). DOI:10.1016/j.ijcard.2012.11.051 · 6.18 Impact Factor