Sequence variation in gene promoters is often associated with disease risk. In this study, a 1,418 bp sequence of the 5'-flanking region of the APOH (encoding for β₂-glycoprotein I) has been functionally characterized by in vitro analysis. Associations of APOH promoter SNPs with systemic lupus erythematosus (SLE) risk and related phenotypes along with their effect on human plasma β₂GPI levels were examined. Transient transfections, dual-luciferase reporter gene assays were performed in COS-1 and HepG2 cells. HepG2 nuclear extracts was used for electrophoretic mobility shift assay (EMSA). A case-control design and genotyped 345 SLE women and 454 healthy control women for 12 APOH promoter single nucleotide polymorphisms (SNPs) (-1284C>G, -1219G>A, -1190G>C, -1076G>A, -1055T>G, -759A>G, - 700C>A, -643T>C, -627A>C, -581A>C, -363C>T -38G>A, and -32C>A). Haplotype analyses were performed using EH (Estimate Haplotype-frequencies) and Haploview programs. Deletion analysis localized the core promoter of APOH ∼160 bp upstream of ATG codon with the presence of critical cis-acting elements between -166 and -65. Functional relevance for three SNPs (-1219G>A, -643T>C and -32C>A) that resulted in lower promoter activity (51%, 40% and 37%, respectively) as compared to the wild-type alleles in COS-1 cells. EMSA demonstrated HepG2 nuclear protein(s) bind to the elements located in the regions of the three SNPs. Overall haplotype distribution of the APOH promoter SNPs was significantly different between cases and controls (P = 0.009). The -643C allele was found to be protective against carotid plaque formation (adjusted OR = 0.37, P = 0.013) among SLE patients. Three-site haplotype analysis revealed one haplotype carrying -32A to be significantly associated with decreased plasma β₂GPI levels (P < 0.001) and another haplotype harboring the minor allele for -1219A showed a significant albeit less pronounced association (P = 0.046). Our data indicate that APOH promoter variants may be involved in the etiology of SLE, especially the risk for autoimmune-mediated cardiovascular disease. Public Health Significance: Cardiovascular disease is the leading cause of death in the U.S. and other developed nations. Understanding its pathogenesis will help in formulating newer therapeutic strategies and treatment that may decrease the adverse consequences of accelerated atherosclerosis in SLE patients.
| Identifer | oai:union.ndltd.org:PITT/oai:PITTETD:etd-12032008-175014 |
| Date | 29 January 2009 |
| Creators | Suresh, Sangita |
| Contributors | F. Yesim Demirci, MD., Iliya Lefterov, MD, Ph.D., Robert E. Ferrell, Ph.D., Candace M. Kammerer, Ph.D., M. Ilyas Kamboh, Ph.D. (Advisor) |
| Publisher | University of Pittsburgh |
| Source Sets | University of Pittsburgh |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.library.pitt.edu/ETD/available/etd-12032008-175014/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to University of Pittsburgh or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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