Association Of Polymorphisms In The RAGE Gene With Clinical Characteristics And Prognosis Of Cardiovascular Diseases In The Chinese Han Population

BackgroundThe receptor for advanced glycation end products (RAGE), a member of the immunoglobulin superfamily, is a transmembrane protein on the cellular surface that recognizes tridimensional molecules, instead of aminoacid sequences. Besides advanced glycation end products (AGE), a large body of evidence suggests that RAGE has other pro-inflammatory ligands such as:high mobility group box1protein (HMGB1), the group of calcium binding cellular factors S100(also termed calgranulin), amiloid beta peptides and Mac-1, a beta-2integrin (CD11b/CD18). The engagement of RAGE by these ligands triggers a series of RAGE-dependent signaling events, including the activation of the nucleus of transcription factor NF-kB, cAMP response element binding protein(CREB) and the rapid generation of reactive oxygen species(ROS), amplifies immune/inflammatory reaction, perturbs endothelium properties, upregulate irreversible formation of cross-linked collagen and induces cardiac myocytes apoptosis and the like, thereby imparting a potential impact in cardiovascular system. In addition, subsequent evidence has been presented that RAGE expression is highly upregulated in human atherosclerotic plaques, particularly in macrophages located at the vulnerable regions of atherosclerotic lesions that contain inflammatory mediators. These considerations highlight RAGE as the vital receptors linked to chronic cardiovascular perturbation, especially to coronary artery disease(CAD), the degree of coronary artery stenosis and left ventricular hypertrophy(LVH), as well as their prognosis.Clearly, the upregulation and pathogenic effects of RAGE in vascular disease highlight RAGE gene variants as likely candidates involved in the pathogenesis of cardiovascular diseases. The gene encoding for RAGE (LocusID177; chromosome6p21.3) is in the major histocompatibility complex (MHC). Polymorphic differences within key domains of the RAGE gene may alter gene expression and influence proinflammatory mechanisms, thereby inducing varied effects of RAGE in inflammatory settings. Over50SNPs are reported for RAGE, including functional polymorphisms-429T/C (dbSNP rs1800625) in the promoter region that related to increased RAGE expression,1704G/T (dbSNP rs184003) in intron7and G82S (dbSNP rs2070600) in exon3within the V-type immunoglobulin domain in the RAGE extracellular region, all have been reported in the development of various vascular diseases. For example, Kankova K et al. have reported that the RAGE haplotype bearing minor alleles in positions-429and2184(CTGGGG) was associated with DN in type2diabetics and with earlier DN onset and, thus, could be regarded as a marker for DN. A recent study suggested the HLA8.1ancestral haplotype was strongly linked to the C allele of the-429T>C polymorphism of the RAGE gene. Furthermore, an association of the G82S polymorphism with diabetic retinopathy (DR) has been reported recently and the G-A haplotype containing1704G and82S alleles was suggested to be a risk marker of DR in Chinese type2diabetic patients. Regarding the G82S mutation, its location within the V-type immunoglobulin domain in the RAGE extracellular region has been illustrated to exhibit increasing receptor-binding affinity for RAGE ligands. Experimental studies elucidated that cells expressing the RAGE G82S polymorphism displayed enhanced ligand-binding affinity and led to increased ligand-stimulated activation of proinflammatory mediators compared to the common RAGE G82isoform. Park SJ et al. have reported that the G82S polymorphism promotes N-linked glycosylation of Asn(81), which might explain the enhanced function associated with the G82S polymorphic RAGE variant. Interestingly, it is noteworthy that nondiabetic/nonobese persons with homozygosity for the minor S allele (S/S) of the G82S polymorphism had elevated risk factors for cardiovascular disease, including low soluble form of RAGE (sRAGE) levels, inflammation, oxidative stress, and insulin resistance (IR) compared with those bearing at least one G allele. All of these might indicate a possible genetic contribution of functional variants in the RAGE gene to the development of clinical features and prognosis of cardiovascular diseases.C-reactive protein (CRP), first described as an inflammatory biomarker, is known to be a strong independent predictor of future coronary events in healthy subjects. Recent in vitro and in vivo studies have shown that CRP stimulates cell adhesion molecules (CAMs), chemokines, endothelin-1release from endothelial cells, upregulation of RAGE and E-selectin expression, and enchancement of monocyte-endothelial cell adhesion. In the vessel wall, CRP promotes the migration and proliferation of vascular smooth muscle cells (VSMCs) and amplifies the production of ROS. These observations argue strongly in favor of a specific proatherosclerotic effect of CRP, in that CRP functions not only as a biomarker for the risk of cardiovascular disease, but also as an active mediator of atherosclerosis. Therefore, functional variants of the RAGE gene may be associated with elevated CRP levels during the progression of atherosclerotic events.However, to date, few gene association studies have been conducted to assess the role of common RAGE gene polymorphisms (-429T/C,1704G/T and G82S) on the pathogenesis of clinical features and prognosis of cardiovascular diseases in the Chinese Han population. Our previous studies found that RAGE gene polymorphisms (-429T/C,1704G/T and G82S) were not significantly associated with an increased risk of DM and HP in Chinese Han population, but RAGE gene G82S polymorphism might be associated with CAD. We anticipate our investigation will lead to a better understanding of the influence of these genetic variants to CAD, the degree of coronary artery stenosis and LVH, as well as the prognosis of cardiovascular diseases.Part oneAssociation of polymorphisms in the RAGE gene with serum CRP levels and coronary artery disease in the Chinese Han populationObjective The role of an advanced glycation end product/receptor for advanced glycation end product (AGE/RAGE) system in the pathogenesis of coronary artery disease (CAD) is not fully understood. To clarify whether polymorphisms of the RAGE gene were related to CAD, we performed a case-control study in Chinese Han patients.Methods The allele frequencies and genotype distribution combinations of the-429T/C,1704G/T and G82S polymorphisms of the RAGE gene were compared in200cases of HP,155cases of CAD combined with hypertension (CAD&HP),175cases of CAD, and170control subjects. PCR-RFLP was used for detection of genotypic variants.Results The S allele frequency of the G82S polymorphism was higher in the CAD (odds ratio (OR),2.303,95%CI1.553～3.416; P<0.001, Pcorr<0.003) and the CAD&HP (OR,1.842,95%CI1.219～2.785; P<0.003, Pcorr<0.009) groups respectively when compared with the control group. However, the S allele frequency was not significantly different between the CAD and the CAD&HP patient groups (P =0.223), while no statistically significant difference of genotype or allele frequency distributions was observed in the HP group (P>0.05). Meanwhile, serum CRP was significantly associated with the G82S variant. Haplotype-based logistic regression analysis revealed that haplotype G-Ser-T (OR,1.670,95%CI,1.017～2.740, P=0.043), compared to the reference haplotype T-Gly-T, was associated with an increased risk of CAD after adjusting for other risk factors. Further analysis limited to non-diabetic participants exhibited similar significant findings.Conclusion The haplotype carrying the G82S variant of the RAGE gene was significantly associated with an increased risk of CAD. Moreover, a remarkable association of the G82S variant with serum CRP levels implied that the prevalence of RAGE82S allelic variation might influence susceptibility to CAD by affecting vascular inflammation. Part twoAssociation of polymorphisms in the RAGE gene with the degree of coronary artery stenosis in the Chinese Han populationObjective To investigate whether polymorphisms of the RAGE gene were associated with the degree of coronary artery stenosis, we performed a case-control study in Chinese Han patients.Methods The allele frequencies and genotype distribution combinations of the-429T/C,1704G/T and G82S polymorphisms of the RAGE gene were compared in109cases of mild/moderate coronary stenosis (MCS),134cases of serious coronary stenosis (SCS) and170control subjects. PCR-RFLP was used for detection of genotypic variants. Results The S allele frequency of the G82S polymorphism was higher in the SCS (OR,2.086,95%CI1,494～2.914; P<0.001, Pcorr<0.003) group when compared with the control group. However, the S allele frequency was not significantly different between the MCS and the SCS patient groups (P=0.023, Pcorr=0.069), while no statistically significant difference of allele frequency distributions was observed in the MCS group (P=0.075). Haplotype-based logistic regression analysis revealed that haplotype G-Ser-T (OR,1.961,95%CI,1.150-3.343, P=0.013), compared to the reference haplotype T-Gly-T, was associated with an increased risk of SCS after adjusting for other risk factors.Conclusion The haplotype carrying the G82S variant of the RAGE gene was significantly associated with an increased risk of SCS. Part threeAssociation of polymorphisms in the RAGE gene with Left Ventricular Hypertrophy in the Chinese Han populationObjective To assess whether polymorphisms of the RAGE gene were associated with left ventricular hypertrophy (LVH) in the Chinese Han population.Methods The study was conducted from July2004to December2005and enrolled531(282/249, male/female) subjects from the city of Guangzhou in the GuangDong province.The allele frequencies and genotype distribution combinations of the-429T/C,1704G/T and G82S polymorphisms of the RAGE gene were compared in157cases of LVH,204cases of hypertension and170control subjects. PCR-RFLP was used for detection of genotypic variants.Results The S allele frequency of the G82S polymorphism was higher in the LVH (OR,1.881,95%CI1.247～2.837; P=0.002, Pcorr=0.006) group when compared with the control group. However, the S allele frequency was not significantly different between the hypertension and the LVH patient groups (P=0.046, Pcorr=0.138), while no statistically significant difference of allele frequency distributions was observed in the hypertension group (P=0.224). Logistic regression analysis revealed that G82S polymorphism of the RAGE gene was associated with an increased risk of LVH after adjusting for other risk factors (P=0.003, Pcorr=0.009).Conclusion The G82S variant of the RAGE gene was significantly associated with an increased risk of LVH in Chinese Han population. Part fourRelation between polymorphisms in the RAGE gene and prognosis of cardiovascular diseases in the Chinese Han populationObjective To investigate whether polymorphisms of the RAGE gene were associated with the prognosis of cardiovascular diseases in the Chinese Han population.Methods The study was conducted from July2004to December2005and enrolled425subjects from the city of Guangzhou in the GuangDong province. Genotyping of the three polymorphisms (-429T/C,1704G/T and G82S) in the RAGE gene was done by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Patients were followed for6.5years for the development of cardiovascular events and mortality.Results Subjects with S mutation of the G82S polymorphism had significantly higher risk of all-cause mortality (GS&SS vs. GG,26.1%vs.15.8%, OR,1.883,95%CI (1.163～3.048), χ2=6.742, P=0.009) and myocardial infarction (MI)(GS&SS vs. GG, 22.4%vs.13.8%, OR,1.799,95%CI (1.083～2.988),χ2=5.221, P=0.022) than those with wild-type homozygote. Logistic regression analysis revealed that G82S polymorphism of the RAGE gene was associated with significantly increased risk of all-cause mortality and MI after adjusting for other risk factors (P=0.017and P=0.045, respectively). Further analysis limited to those with cardiovascular diseases (HP or CAD) revealed that G82S polymorphism of the RAGE gene was associated with an increased risk of all-cause mortality after adjusting for other risk factors (P=0.019). However, the-429T/C and1704G/T polymorphism had no effect on the prognosis of the subjects.Conclusion The G82S variant of the RAGE gene was significantly associated with an increased risk of all-cause mortality and MI in Chinese Han population.