Maria Cristina Sanguedolce

Association of a Polymorphism in a Gene Encoding a Urate Transporter with CKD Progression

BACKGROUND AND OBJECTIVES Hyperuricemia predicts a high risk for CKD progression but there is no large clinical trial in humans indicating that this relationship is causal in nature. The rs734553 single-nucleotide polymorphism (SNP) of the GLUT9 urate transporter gene was strongly associated with uric acid (UA) levels in a large meta-analysis. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS This prospective study adopted the Mendelian randomization approach. The rs734553 SNP was used as an instrumental variable to investigate the relationship between UA and renal outcomes in a cohort of 755 patients with CKD who were enrolled between October 18, 2005, and October 2, 2008. The association between the polymorphism and UA was preliminary confirmed in a series of 211 healthy volunteers enrolled between January 1, 2001, and July 12, 2011, from the same geographic area as the patients with CKD. The study end point was a composite renal-end point (i.e., >30% decrease in the GFR, dialysis, or transplantation). Patients were followed up for a median of 36 months. RESULTS In healthy individuals, serum UA levels were highest in homozygotes for the T allele (risk allele), intermediate in heterozygotes for the same allele, and lowest in those without the risk allele (P<0.001), but no such relationship was found in patients with CKD. In the CKD cohort, homozygotes (TT) and heterozygotes (GT) for the risk allele had a 2.35 times higher risk (hazard ratio, 2.35; 95% confidence interval, 1.25 to 4.42; P=0.008) of CKD progression. The risk for CKD progression by rs734553 remained unmodified in analyses adjusting for proteinuria, GFR, and other classical and CKD-peculiar risk factors. CONCLUSIONS A GLUT9 polymorphism, which is strongly associated with serum UA levels in healthy individuals of the general population with normal renal function, holds a strong predictive power for CKD progression. These findings are compatible with the hypothesis that the link between UA and CKD progression is causal in nature.

A polymorphism in the major gene regulating serum uric acid associates with clinic SBP and the white-coat effect in a family-based study.

Objectives: Hyperuricemia associates with hypertension, but it is uncertain whether this relationship is causal in nature. Glucose transporter 9 (GLUT9) gene is a major genetic determinant of plasma uric acid levels in humans. Since polymorphisms are randomly distributed at mating (Mendelian randomization), studies based on GLUT9 polymorphisms may provide unconfounded assessment of the nature of the link between uric acid and hypertension. Methods: We tested the association between uric acid, the rs734555 polymorphism of the GLUT9 gene and arterial pressure in a family-based study including 449 individuals in a genetically homogenous population in Southern Italy. Results: Serum uric acid levels were strongly associated (P < 0.001) with all components of clinic and 24-h ambulatory blood pressures (BPs). However, only clinic SBP and the white-coat effect (the difference in clinic systolic and daytime systolic ambulatory blood pressure monitoring) associations remained significant after adjustment for classical risk factor and the estimated glomerular filtration rate. Serum uric acid was strongly associated with the risk allele (T) of the rs734555 polymorphism (P < 0.001). Furthermore, TT individuals showed higher clinic SBP (129 + SEM 1 mmHg) than GT (125 + 1 mmHg) and GG individuals (122 + 3 mmHg), as well as a higher white-coat effect (P = 0.02), confirming that the association between uric acid and these BP components is unconfounded by environmental risk factors. Conclusion: Results in this family-based study are compatible with the hypothesis that uric acid is a causal risk factor for hypertension. Trials testing uric acid-lowering interventions are needed to definitively establish the causal implication of hyperuricemia in human hypertension.

A genetic marker of uric acid level, carotid atherosclerosis, and arterial stiffness: a family-based study.

BACKGROUND Hyperuricemia associates with atherosclerosis complications, but it is uncertain whether this relationship is causal in nature. The urate transporter GLUT9 (encoded by the SLC2A9 gene) is a major genetic determinant of serum uric acid level in humans. Because polymorphisms are distributed randomly at mating (Mendelian randomization), studies based on GLUT9 polymorphisms may provide unconfounded assessment of the nature of the link between uric acid and atherosclerosis. STUDY DESIGN Cross-sectional study. SETTING & PARTICIPANTS Family-based study including 449 individuals in 107 families in a genetically homogeneous population in Southern Italy. FACTOR Serum uric acid level, rs734553 allele, and age. OUTCOME Ultrasound biomarkers of atherosclerosis (intima-media thickness [IMT] and internal diameter) and pulse wave velocity (PWV). RESULTS Serum uric acid level was dose-dependently associated with the T allele of rs734553, a polymorphism in SLC2A9 (P=8×10(-6)). Serum uric acid level was a strong modifier of the relationship between age and IMT in fully adjusted analyses (β=0.33; P=0.01), whereas no such relationship was found for internal diameter (β=-0.15; P=0.3) or PWV (β=0.10; P=0.6). The T allele coherently associated with carotid IMT, internal diameter, and PWV and emerged as an even stronger modifier of the age-IMT and age-internal diameter relationships in both crude and fully adjusted (β=0.40 [P<0.001] and β=0.48 [P=0.003], respectively) analyses. LIMITATIONS This is a hypothesis-generating study. CONCLUSIONS Results in this family-based study implicate uric acid as an important modifier of the age-dependent risk for atherosclerosis. Trials testing uric acid-lowering interventions are needed to prove this hypothesis.

eNOS and Caveolin-1 Gene Polymorphisms Interaction and Intima Media Thickness: A Proof of Concept Study in ESRD Patients

BACKGROUND Caveolae are a prominent microdomain in endothelial cells and appropriate localization in caveolae is fundamental for endothelial nitric oxide synthase (eNOS) activity. Since the Glu298Asp variant in the eNOS gene alters caveolar localization of the corresponding enzyme, we tested the interaction between this variant and the rs4730751 polymorphism of the caveolin-1 (CAV-1) gene as related to arterial remodeling in end-stage renal disease (ESRD) patients. METHODS One hundred and thirty-three ethnically homogeneous ESRD patients underwent carotid ultrasonographic studies to measure intima-media thickness (IMT) and carotid cross-sectional area (CSA). Genotyping was performed by high-throughput allelic discrimination assays on real-time PCR. RESULTS Arterial remodeling was associated to the number of G alleles of CAV-1 polymorphism, GG homozygotes displaying an IMT and a CSA that were, respectively, 16% and 21% higher than those in patients without the risk allele (P < 0.012). In multiple linear regression analyses including the CAV-1 and the eNOS polymorphisms and adjusting for classical risk factors and risk factors peculiar to ESRD both polymorphisms were independent correlates of IMT (CAV-1: β = 0.20, P = 0.01; eNOS β = 0.25, P = 0.001) and CSA (CAV-1: β = 0.20, P = 0.01: eNOS β = 0.13, P = 0.09). Furthermore, strong interactions emerged between the two polymorphisms for explaining the variability in IMT (P = 0.001) and in CSA (P = 0.038) in these patients. CONCLUSION Overall these findings form preliminary evidence that disturbed interaction between CAV-1 and eNOS may be of relevance for arterial disease in ESRD and perhaps in other human diseases.

The fat-mass and obesity-associated gene (FTO) predicts mortality in chronic kidney disease of various severity

BACKGROUND Polymorphisms in the FTO (fat-mass and obesity-associated) gene have been associated with the body mass index, cancer, type 2 diabetes and hypertension. METHODS We investigated the relationship between 17 tag single-nucleotide polymorphisms (SNPs) and all-cause mortality in three cohorts of dialysis patients (CREED-1, North Apulian and CREED-2 cohorts; n = 783) and in one cohort of stage 2-5 CKD patients (n = 757). RESULTS We first explored the association between the 17 tag SNPs and all-cause mortality in the CREED-1 cohort and found that patients with the A allele of the FTO rs708259 polymorphism had an elevated risk of mortality (hazard ratio, HR: 1.52, 95% confidence interval (CI) 1.11-2.08; P = 0.008). Similarly, the A allele was associated with an increased risk of death also in the other two dialysis cohorts (North Apulian cohort, risk: +23%; CREED-2 cohort, risk: +21%). The elevated risk portended by this allele was even higher in the stage 2-5 CKD cohort (+97%). However, the risk of mortality associated with the A allele in the three confirmatory cohorts failed to achieve formal statistical significance. In a meta-analysis including the four cohorts (n = 1540; total deaths, n = 381), individuals with the A allele had a 42% excess risk of death (HR: 1.42, 95% CI 1.14-1.76, P = 0.002). CONCLUSION The A allele of the FTO rs708259 polymorphism is an independent predictor of all-cause mortality in patients with CKD of various severity. These data support our hypothesis that the FTO gene may be a relevant genetic risk factor for mortality in this population.

A polymorphism in a major antioxidant gene (Kelch-like ECH-associated protein 1) predicts incident cardiovascular events in chronic kidney disease patients: an exploratory study.

Objective: Oxidative stress is considered a major pathway conducive to cardiovascular disease in chronic kidney disease (CKD) patients. However, observational studies and clinical trials testing this relationship are controversial. The Nuclear factor-erythroid-2-related factor 2 (Nrf2)-Kelch-like ECH-associated protein 1 (Keap1) system is a major system regulating antioxidant mechanisms in living organisms. Owing to the fact that genes are transmitted randomly (Mendelian randomization), genetic variants may provide unconfounded assessment of putative causal risk factors. Methods: We have therefore explored the association of eight polymorphisms in the Nrf2 gene and three polymorphisms in the Keap1 gene (capturing over 80% of the genetic variance in the same genes) with cardiovascular events in a multicenter cohort study of 758 CKD patients. Results: During the follow-up period, 117 patients had fatal and nonfatal cardiovascular events and 42 died. The hazard rate of fatal and nonfatal cardiovascular outcomes was about twice higher in patients with the AA or the CA genotype (dominant model) in the rs110857735 polymorphism of the Keap1 gene (hazard rate: 1.85, 95% CI: 1.20–2.84, P = 0.005) than in those with the CC genotype. Further analyses adjusting for Framingham risk factors and CKD-specific risk factors and a bootstrapping validation analysis did not modify the strength of this association. No association was registered between other Keap1 and Nrf2 polymorphisms and cardiovascular disease in the same cohort. Conclusion: In this exploratory study a gene-variant in Keap1, a major gene regulating the antioxidant response, predicts incident cardiovascular events in CKD patients. This finding is in keeping with the hypothesis implicating oxidative stress in cardiovascular disease in this population.

SLEEP DISORDERED BREATHING IN RENAL TRANSPLANT PATIENTS: A LONGITUDINAL STUDY

Objective: Sleep disordered breathing (SDB) is a pervasive problem in stage 5 CKD patients maintained on regular dialysis treatment. Renal transplantation improves SDB but the longitudinal, long term evolution of SDB after kidney grafting has not been investigated. The issue is relevant because renal transplantation abolishes uremic toxicity but introduces classical risk factors for SDB, like overweight and obesity. Design and method: We investigated the long term evolution of polysomnographic recordings in 221 stable renal transplant patients. Overall 404 recordings over a median time of 3 years (interquartile range 2 – 4) were performed. Longitudinal analysis was performed by the linear mixed model (LMM). Results: The apnea-hypopnea index (AHI) in renal transplant patients was 5.44 ± SD 0.75 and the corresponding number of O2 desaturation (Des O2) episodes was 4.40 ± 0.60. The number of patients with AHI > 5 (the threshold for identifying mild SDB) rose from 25% at baseline to 43% at the end of the follow up. Over follow-up the AHI rose to 9.94 ± 2.37 (P < 0.001) and the number of desaturation episodes to 5.55 ± 2.1 (P = 0.010) which went along with a decline in the mean O2-saturation overnight (Mean SatO2: from 95.08 ± 0.14 to 94.64 ± 0.32,P = 0.001). LMM showed that age, sex, BMI, ESA treatment but neither eGFR nor the presence of proteinuria associated with the AHI. On multivariate analyses only male gender (3.42; 95%CI: 0.35 to 6.49; P = 0.003) and BMI (0.87; 95% CI 0.46 to 1.27; P < 0.001) associated with the AHI evolution over follow-up. Similar associations were registered also for the other parameters of SDB (Des O2, Mean Sat O2). Conclusions: Even though renal transplantation produces an early improvement in SDB in renal transplant patients, this treatment does not stabilize the disturbance which gradually re-emerges over longitudinal observation. Among risk factors underlying the risk of SDB re-emergence after transplantation the rise in BMI, i.e. a modifiable risk factor, appears to be of paramount importance. Tackling the surge of SDB after transplantation is of obvious relevance because this disturbance is a potent driver of sympathetic over-activity and of the attendant risk of cardiovascular events in this population.