Study On The Relationship Between Klotho Gene Polymorphism And Protein Level And Arteriosclerosis In Chronic Kidney Disease

Objective: To explore the effect of Klotho gene polymorphisms and serum levels of klotho protein on Chronic Kidney Disease and Arteriosclerosis.Methods: 216 subjects, including 156 CKD patients(The non dialysis group included 56 cases of CKD2-3, 60 cases of CKD4-5, and 40 cases of hemodialysis group)and 60 normal controls were enrolled in the study. The intima-media thickness of carotid artery was measured by Color Doppler ultrasound in patients with CKD. One SNP genotype of Klotho gene was detected by FQ-PCR method and the serum levels of Klotho protein were tested using enzyme linked immuno sorbent assay(ELISA) in all subjects. Meanwhile, other related clinical and biochemical parameters were also evaluated in patients with CKD. The genotypes frequencies, allele frequencies and Klotho protein, clinical indexes were compared among groups.Results:(1)The site of G-395 A had polymorphism and present three kinds of genotypes of which distribution conformed to Hardy-Weinberg Balance.(2)Statistical differences were observed in genotype and allele frequency of G-395 A between CKD and control group(P<0.05).The GA+AA genotype and A allele in non dialysis group were lower than those in control group(P<0.01). Statistical differences were observed in callele frequency of G-395 A between control group and hemodialysis group.(3)The GA+AA genotype and A allele in CKD2-5 group with arteriosclerosis were lower than those in CKD2-5 non arteriosclerosis group(P<0.01). The frequency of GA+AA genotype and A allele frequency in patients hemodialysis with arteriosclerosis were lower than that in non atherosclerosis group(P<0.05).(4)There were significant differences in age, diabetes, P, i PTH, e GFR, FGF23, and klotho between atherosclerosis and non atherosclerosis group in CKD non hemodialysis patients(P<0.05). There were no differences in clinical index and klotho level between GG and GA+AA group. Logistic regression analysis showed: The age,Diabetes mellitus, FGF23,e GFR, klotho protein and G-395 A allele(GA+AA) is associated with atherosclerosis in CKD patients with non hemodialysis, serum levels of klotho protein was associated with occurrence of CKD.(5) There were significant differences in age, Diabetes mellitus,SBP, hs CRP and FGF23 between atherosclerosis and non atherosclerosis group in CKD hemodialysis patients(P<0.05). There were no differences in clinical index and klotho level between GG and GA+AA group. Logistic regression analysis showed: The age,hs CRP,FGF23 and G-395 A allele(GA+AA) is associated with atherosclerosis in CKD hemodialysis patients.Conclusion:(1)The G-395 A site of Klotho gene exists polymorphisms in study population.(2)In CKD patients,with increasing of serum klotho protein level,the risk of CKD is decreased,and it confirmed that serum level of klotho protein may be independent protective factor for CKD.(3)The A allele carriers might be genetic protective factors of CKD complicated by arteriosclerosis.The serum level of klotho protein is independent protective factor for CKD with arteriosclerosis in non hemodialysis patients.(4)Age, Diabetes mellitus,e GFR and serum FGF23 level make contribution to aggravated arteriosclerosis in CKD non hemodialysis patients.(5)Age,hs CRP and serum FGF23 level make contribution to aggravated arteriosclerosis in CKD hemodialysis patients.Objective:The purpose of this study was to test whether serum klotho and fibroblast growth factor 23(FGF23) levels, might be associated with the mineral metabolic disorders and arteriosclerosis(AS) in chronic kidney disease(CKD) patients.Methods: 163 patients with chronic kidney disease according to the e GFR group were divided CKD2 group(23 cases), CKD3 group(32 cases), CKD4 group( 30 cases), CKD5 group[78 cases,including non-hemodialysis(41 cases) and hemodialysis patients(37 cases)]. The carotid artery intima thickness was divided into 54 cases in the AS group and 72 cases in the non arteriosclerosis(N-AS) group by the neck color Doppler ultrasound in the CKD2-5 patients. Another healthy control group of 33 cases. Enzyme-linked assay FGF23(immunosorbent) was used to detect the levels of ELISA and klotho, and the relationship between FGF23, klotho and clinical indexes were analyzed statistically.Result:(1) Klotho levels in non-dialysis patients with CKD stages 2-5 were found to be ignificantly lower compared with normal control group(P<0.01); Klotho levels of atients with CKD stages 4 and 5 were lower than in patients with CKD stages 2 and 3(P <0.01). FGF23 and i PTH levels in patients with CKD stages 3-5 were found to be significantly higher than in the control ones(P <0.01); There were significant differences in the levels of FGF23 and i PTH in CKD2-5 phase group(P<0.01);. Klotho levels were negatively correlated with FGF23(r=-0.823, P<0.01), serum phosphorus(r=-0.785, P<0.01), calcium phosphorus product(r=-0.697, P<0.01) and i PTH(r=-0.692, P<0.01). After adjustment for age, gender and e GFR, Klotho remained independently associated with age and e GFR. FGF23 was positively correlated with serum phosphorus(r=0.838, P<0.01), calcium phosphorus product(r=0.794, P<0.01) and i PTH(r=0.813, P<0.01). FGF23 was negatively correlated serum calcium(r=-0.675,P<0.01).(2)In HD and non-dialysis patients with CKD stages 5: compared with the HD group klotho levels were significantly increased in non-dialysis patients(P<0.01), while FGF23 levels showed no significant difference(P>0.05). PTH level of non-dialysis group was higher than HD(P<0.05),while serum calcium and phosphorus and calcium phosphorus product were lower than HD(P<0.01).(3)To illustrate the relationship of klotho, FGF23, i PTH, phosphorus, calcium versus e GFR with LOESS in non-dialysis patients with CKD stages 2-5, a graph was plotted. Klotho linearly decreased with e GFR. Klotho levels declined by 5.35pg/ml(95%CI5.02-5.68pg/ml,P<0.01) as e GFR declined by 1ml/min/1.73m2. After adjustment for age, we estimated that the mean Klotho change was 5.31pg/m L(95%CI5.01-5.64pg/ml, P<0.01) for each 1ml/min/1.73m2 GFR change. FGF23, i PTH and serum phosphorus showed a baseline at CKD stages 2 for FGF23, stages 2–3 for i PTH and phosphorus, and then a dramatic increase at higher respective CKD stages. FGF23 started to rise at an e GFR of 42 m L/min/1.73m2,whereas i PTH rose from the baseline at an e GFR of 28 m L/min/1.73m2 and serum phosphorus rose at an e GFR of 27ml/min/1.73m2 respectively.(4)In non-dialysis patients with CKD,age,BMI, HGB, i PTH and FGF23 were increased in AS group(P<0.05) compared with the N-AS group,while klotho level was lower than the N-AS group(P<0.05). By means of multivariate analysis with binary logistic regression, we found klotho(OR=0.988, P<0.05), FGF23(OR=1.086,P<0.05) were independent factors for the occurrence of arteriosclerosis.Conclusions:(1) In non-dialysis patients with chronic kidney disease, an increase in FGF23 precedes an increase in serum phosphorus or parathyroid hormone, whereas a decline in klotho levels precedes an increase in FGF23. Klotho may be a sensitive marker for the early detection of the mineral metabolic disorder in chronic kidney disease.(2)In non-dialysis patients with chronic kidney disease, Klotho and may be the influential factors for the occurrence of AS, Klotho is a protective factor, while FGF23 is an independent risk factor.