Research On Serum Visfatin In Metabolic Syndrome And The Association With Tribble 3 Gene Polymorphisms

Abstract of dissertationâ… RESEARCH ON SERUM VISFATIN IN METABOLIC SYNDROME AND THE ASSOCIATION WITH TRIBBLE 3 GENE POLYMORPHISMSBackgroundMetabolic syndrome(MS) is a clustering of some clinical syndroms,with the common pathophysiological basis including obesity,hypertension,insulin resistence, hyperglycemia and dyslipidaemia,and the clinical character of combination of several metabolic disorders.Each of the components is the risk factor of cardiovascular diseases at the same time.The more components cause the more serious conditions. Epidemiological data showed that MS correlated closely with cardiovascular diseases and atherosclerosis.The morbidity and mortality of cardiovascular disease in MS are both higher than that in the normal people.So studying the relationship of MS and cardiovascular diseases at the level of pathology is of strategic meanings to make effective prevention and treatment,and to decrease the morbidity and mortality of cardiovascular disease in MS.The pathogenesis of MS is very complex,and the genetic and inflammatory factors attract more attention recently.Many adipocytokines which at the same time are also inflammatory factors were found to participate in the pathogenesis of MS. Vistatin,a newly recognized adipocytokine who has the insulin mimic effect was found in the human atherosclerotic plaque.Studies have found it had important effect in the inflammatory course.But till now,the changes of serum visfatin concentration and its correlation with clinical indexes and IMT in MS and atherosclerotic patients were steel open questions.Besides the environmental factors the genetic factors also participate in the pathogenesis of MS.Insulin resistance one of the most important etiological factors of MS was found to be affected by genetic reasons.The gene TRB3 who was the mammalian homologue of Drosophila tribbles,was proved to be a candidate gene of insulin resistance and DM.Studies have found that Q84R(+251A/G)polymorphism of TRB3 correlated closely with insulin resistance and cardiovascular risk.Based on these results,we bring forward some problems as follows:what is the serum visfatin concentration of MS? And what is the further change if MS is combining with atherosclerosis? Is the visfatin associated with each component of MS? And is TRB3 associated with visfatin in MS? Nowadays,there are few studies referring to these questions.Therefore,we carried out the present study and tried to find satisfying answers to all the above problems.Objectives1.To investigate the serum concentration of visfatin of MS by EIA method and determine the association of visfatin and each component of MS.2.To explore the relationship between visfatin and IMT,inner diameter of carotid artery by use of a high resolution ultrasound system3.Genotyping the +251A/G polymorphisms of TRB3 gene by PCR-RFLP, exploring the relationship between the +251A/G polymorphisms of TRB3 and visfatin in MS.Subjects and methodsIn our study,we recruited 139 consecutive subjects with MS(78 females,54±8 yr of age) and 114 age- and sex-matched normal subjects(70 females,54±8 yr of age) without cardiovascular diseases or diabetes mellitus.The MS patients were further divided in two groups:with and without carotid atherosclerotic plaque.All subjects completed a questionnaire on present condition,family and medical histories of cardiovascular risk factors and complications.Height,weight,body mass index(BMI), waist circumference,hip circumference and blood pressure(BP) were measured. Furthermore,fasting blood sample was collected from each subjects after 12-14 hours fast to determine the fasting blood glucose(FBG),insulin,total cholesterol(TC), low-density lipoprotein cholesterol(LDL-C),high-density lipoprotein cholesterol (HDL-C),triglycerides(TG) HBA1C and uric acid(UA).visfatin was measured by EIA method.DNA was obtained from blood samples using Phenol/chloroform method and saved at -80℃until the assay.Genotyping of the TRB3 polymorphism was carried out by restriction fragment length polymorphism-polymerase chain reaction(RFLP-PCR) methods.B-mode ultrasonography of the carotid arteries was performed and Ultrasound images were acquired using a 5～10MHz linear array transducer and a commercially available ultrasound machine(Vivid 7 dimension; General Electric Medical Systems,Horten,Norway).The association of visfatin and each component of MS,the relationship between visfatin and IMT,inner diameter of carotid artery as well as the relationship between the +251A/G polymorphisms of TRB3 and visfatin were determined.Results(1)Clinical characteristics of the control group and MS group:There were no significant differences in age and gender between MS group and control group. Compared with controls,BMI,waist circumference,waist-to-hip ratio,systolic blood pressure(SBP),diastolic blood pressure(DBP),TG,TC,LDL-C,FBG,fasting insulin (FINS),HOMA-insulin resistance(HOMA-IR),HbA1c,UA were significantly higher in the patients with MS(all P＜0.001),whereas HDL-C was significantly lower in MS group(P＜0.001).(2)The changes of serum visfatin,IMT and carotid inner diameter:compare with the control group the serum concentration of visfatin,Max-IMT,Mean IMT and carotid inner diameter increased significantly(P＜0.001).(3)Clinical characteristics of the non-carotid atherosclerotic plaque group and carotid atherosclerotic plaque group:Compared with the control group,BMI,waist circumference,waist-to-hip ratio,SBP,DBP,TG,TC,LDL-C,FBG,FINS,HbA1c and HOMA-IR were significantly higher in carotid atherosclerotic plaque group, whereas HDL-C was significantly lower(P＜0.001).Compared with the non-carotid atherosclerotic plaque group FBG and HbA1c were significantly higher in carotid atherosclerotic plaque group(P＜0.01).whereas other clinical characteristics had no significant differences.(4)The changes of serum visfatin,IMT and inner diameter of carotid artery of carotid atherosclerotic plaque group:compared with the non-carotid atherosclerotic plaque group,the serum concentration of visfatin,Max-IMT(P＜0.001) and Mean IMT (P＜0.01) increased significantly,the carotid inner diameter increased but did not reach the statistical criteria.(5)The result of Pearson linear correlation of log visfatin and each index and the result of Multiple regression analysis with log visfatin as the dependent variable was performed by using stepwise linear regression analysis:Log visfatin was associated with total cholesterol(R=0.192,P=0.015),LDL(R=0.219,P=0.007) and HOMA (R=0.163,P=0.010) in simple regression analysis in the patients with MS,whereas in multiple regression analysis,only LDL-cholesterol remains positively correlated with log visfatin(R=0.219,P=0.013).(6)The result of Pearson linear correlation of max-IMT and each index and the result of Multiple regression analysis with log visfatin as the dependent variable was performed by using stepwise linear regression analysis:max-IMT was associated with age(R=0.215,P=0.012),DBP(R=-0.162,P=0.044)and log visfatin(R=0.282, P=0.001) in simple regression analysis in the patients with MS,whereas in multiple regression analysis,age(R=0.230,P=0.012) and log visfatin(R=0.293, P=0.001) remain positively correlated with log visfatin(R=0.219,P=0.013).(7)The ability of log visfatin to detect patients with MetS and carotid plaque:The area under the curve for the ROC curve of Ts-max was 0.740(95%confidential interval, 0.654-0.827).A log visfatin value of＞1.08 ng/ml had 70%sensitivity and 67% specificity for detecting patients with MetS and carotid plaque.(8)+251A/G genotyping of TRBs3:the PCR product of TRBs3 is 593 bp,after Restriction Enzyme MspI cutting,QQ genotype is 593bp,QR genotype are 593bp and 297bp,RR genotype is 297bp.(9)The log visfatin between the different +251A/G(Q84R) gene of TRIB3: the log visfatin have no difference between the three different genotype in control group;while in MS group the log visfatin of RR84 genotype is significantly higher thanQQ84(P＜0.01) and RR84(P＜0.05) genotype.(10)The correlation of TRIB3 +251A/G genotype and log visfatin in MS:the QQ genotype has no relation with log visfatin(P=0.227) in MS;while the RR genotype correlate significantly with log visfatin in MS(P=0.009).Conclusions(1)The concentration of serum visfatin was significantly elevated in MS,and was even higher in the MS patients with carotid atherosclerosis,which indicated that visfatin participated not only in the pathology of MS,but also in the pathology of atherosclerosis,and maybe a bridge of MS and atherosclerosis.(2)The concentration of serum visfatin was independently correlated with LDL concentration,and was an independent risk factor of carotid max-IMT,which indicated visfatin correlated closely with cholesterol metabolism,and may participate in the pathology of atherosclerosis by modulating lipid metabolism.(3)Among the three genotypes of TRIB3 +251A/G in MS,the concentration of serum visfatin of RR84 genotype was significantly higher than that of QQ84 and QR84 genotype,which indicated TRB3 polymorphism may have modulating effect on visfatin.(4)The RR84 genotype of MS was significantly correlated with the concentration of serum visfatin,the TRIB3 RR84 genotype of MS may cause the elevation of concentration of serum visfatin. BackgroundThe correlation of metabolic syndrome and many other clinical diseases such as diabetic mellitus and cardiovascular diseases has been recognized and studied by many researchers, While researchers have reached consensus on the metabolic syndrome as the independent risk factor of kidney disease in recent years. The mechanism how MS caused or aggravated kidney disease is still unclear, hypertension and diabetic mellitus were thought to be the most important reasons, while microinflammation and lipids toxicity have caused more and more attention recently.Researchers have found that adipose tissue may secrete many cytokines which may participate in the renal lesion as inflammation cytokines. Recently a newly recognized adipocytokine, Visfatin was found to have close relationship with obesity, glucose and lipid metabolism and atherosclerosis. Our research showed that the concentration of serum Visfatin of MS was significantly higher than that of control, and was even higher in MS with carotid atherosclerosis than that without carotid atherosclerosis. The result implied the close relationship of Visfatin and MS, atherosclerosis. As an important target organ of MS, the kidney and the relationship with Visfatin also have attracted more and more attention.It has been reported that the serum concentration of Visfatin in hemodialysis patients were significantly elevated, also in peritoneal dialysis patients. But there were till no reporting of Visfatin and renal lesion in MS patients. Based on these results, we bring forward some problems as follows: How does the GFR change in the early stage of MS? Is there any relation between the Visfatin and GFR in MS? Nowadays, thereare few studies referring to these questions. Therefore, we carried out the presentstudy and tried to find satisfying answers to the above problems.Objectives1.To calculate the GFR of MS patients using MDRD formula and find out the changesof GFR of MS patients2. To investigate the serum concentration of Visfatin of MS by EIA method and determine the association of Visfatin and GFR of MS Subjects and methodsThe subjects were the same as those of dissertation I, among whom 85 MS patients were tested of serum Cr, (51 females, 55±8 yr of age) and 88 age- and sex-matched normal subjects (51 females, 53±9 yr of age). The MS patients were further divided in two groups: with GFR higher than 90 ml/min·1.73m and lower than 90 ml/min·1.73m². All subjects completed a questionnaire on present condition, family and medical histories of cardiovascular risk factors and complications. Height, weight, body mass index (BMI), waist circumference, hip circumference and blood pressure (BP) were measured. Furthermore, fasting blood sample was collected from each subjects after 12-14 hours fast to determine the fasting blood glucose (FBG), insulin, total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C) ,triglycerides (TG) HBA1C , uric acid (UA) and cretinine(Cr).Visfatin was measured by EIA method. GFR was calculated by MDRD formula: GFR (ml/min·1.73m²) =186x [Cr(mg/ml] ^-1.154×[Age (year)] ^0.203×[Female×0.742. Then the data were analyzed with SPSS 16.0.Results1. Clinical characteristics of the control group and MS group: There were nosignificant differences in age and gender between MS group and control group. Compared with controls, BMI, waist circumference, waist-to-hip ratio, systolic blood pressure (SBP), diastolic blood pressure (DBP), TG, TC, LDL-C, FBG, fasting insulin (FINS), HOMA-insulin resistance (HOMA-IR), HbA1c, UA were significantly higher in the patients with MS (all P<0.001), whereas HDL-C was significantly lower in MS group (P<0.001).2. The changes of Visfatin, Cr and GFR of MS patients group: the log Visfatin (p <0.05 ) and serum Cr concentration (p <0.01) were significantly higher than those of control group, the GFR of MS group was significantly lower than that of control group (p <0.01) .3. The relationship of serum Cr and each component of MS3.1 The result of Pearson linear correlation of serum Cr and clinical, laboratory indexes: serum Cr was associated positively with age (r=-0.190, p <0.05) , waist circumference (r=0.249, p <0.001), waist-to-hip ratio (r=0.379, p<0.001), systolic blood pressure (SBP) (r=0.233 ,p <0.01), diastolic blood pressure (DBP) (r=0.240, p<0.01), TG (r=0.194, p <0.01), UA (r=-0.375, p <0.001) and HOMA (r=0.206, p<0.01) , negatively with HDL-c (r=-0.233, p<0.01) in simple regression analysis in the patients with MS.3.2 The result of partial correlation of serum Cr and clinical, laboratory indexes: controlling other correlating factors, serum Cr still correlated with waist-to-hip ratio(r=0.232, p<0.01) and UA (r=0.196, p<0.05)3.3 The result of multiple regression analysis: in multiple regression analysis when the Cr was dependent variable, age, BMI, waist circumference, waist-to-hip ratio, systolic blood pressure (SBP), diastolic blood pressure (DBP), TG, TC, LDL-C, FBG, fasting insulin (FINS), HOMA-insulin resistance (HOMA-IR), HbA1c, UA and log Visfatin were independent variables, waist-to-hip ratio (β=0.276,β<0.01) and UA (β=0.188? p <0.05) remain positively correlated with Cr.4. The relationship of GFR and each component of MS4. 1 The result of Pearson linear correlation of GFR and clinical, laboratory indexes: serum Cr was associated negatively with age (r=-0.25, p <0.01) , systolic blood pressure (SBP) (r=-0.26, p<0.01) , diastolic blood pressure (DBP) (r=-0.18, p <0.05) , TC (r=-0.24, p <0.01) ,TG (r=-0.18, p <0.05) , UA (r=-0.375, p <0.001) ,LDL-c (r=-0.24, p<0.01) ,HOMA (p=-0.19, p<0.05) and log Visfatin (r=-0.19, p<0.05) in simple regression analysis in the patients with MS.4. 2 The result of partial correlation of GFR and clinical, laboratory indexes: controlling other correlating factors, GFR still correlated positively with age(r=-0.271, p<0.01) , SBP (r=-0.187, p<0.05) ,TC (r=-0.202, p<0.05) ,TG(r=-0. 158, p<0.05) and log Visfatin (r=-0.209, p<0.05) .4. 3 The result of multiple regression analysis: in multiple regression analysis when the GFR was dependent variable, age, BMI, waist circumference, waist-to-hip ratio, systolic blood pressure (SBP), diastolic blood pressure (DBP), TG, TC, LDL-C, FBG, fasting insulin (FINS), HOMA-insulin resistance (HOMA-IR), HbA1c, UA and log Visfatin were independent variables, age (β=-0.28, p <0.01) , SBP (β=-0.23, p <0.01) , TC (β=-0.16, p <0.05), TG (β=-0.16, p <0.05) and log Visfatin (β=-0.17, p <0.01) remain negatively correlated with Cr, which suggesting age, SBP, TG, TC and serum Visfatin were independent risk factors of GFR in MS patients.5. The changes of each index in those MS patients whose GFR were decreased: age, TC, TG, LDL-c, UA, FBG and HOMA-IRI of those MS patients whose GFR were decreased were higher than those of MS patients whose GFR were normal. Among those indexes, age(p=0.05) and log Visfatin (p<0.05) meet the statistical standard.6. The changes of serum Visfatin concentration of the MS patients whose GFR were decreased: the log Visfatin of MS patients whose GFR was lower than 90 ml/min·1.73m² was significantly higher than that of MS patients whose GFR was more than GFR>90ml/min·1.73m².7. Divided by GFR , the result of multiple regression analysis between Log Visfatin and other indexes of MS patients: no statistically significant result.Conclusions(1)The GFR decreased before clinical CKD in MS patients, which indicated that the renal malfunction emerged before DM and hypertension, and there were probably other mechanisms besides DM and hypertension that caused renal malfunction in MS.(2)The serum concentration of Visfatin of those MS patients whose GFR decreased significantly elevated. The serum concentration of Visfatin correlated with GFR in MS patients and was an independent risk factor of GFR in MS which suggested that Visfatin can be taken as a marker and also a risk factor of early renal injury of MS patients. Visfatin maybe an important factor involving in the kidney disease of MS.