Clinical Research On Association Of Serum AAP And LAP Activity With Metabolic Syndrome

BackgroundMetabolic syndrome is a constellation of cardiovascular risk factors,including obesity,hypertension,dyslipidemia and hyperglycemia,which are correlated with cardiovascular disease and diabetes.These risk factors interact and interrelate with each other,amplyfing the adverse effect,directly promoting the development of target organ damage.Because of the epidemic of overweight and ageing world-wide,the metabolic syndrome is becoming increasingly commonThe syndrome is characterized by insulin resistance and is also known as the insulin resistance syndrome.In the twentieth century,available data and clinical observations which suggest that there is a pathogenetic relationship between hypertension,diabetes mellitus,and atherosclerosis have provided a concept of the X syndrome,by which hypertensive patients,mainly males,have impaired insulin tolerance along with hyperinsulinemia and concurrent atherogenic disorders of lipid metabolism.Then they saw insulin resistance is fundemental factor in the pathogenesis.So the concept "insulin resistant syndrome"come into being.With regard to the risk of CVD and type 2 diabetes mellitues,we give it a new concept "the metabolic syndrome".In the pathogensis,insulin resistance is the key facter,maybe the cause of the metabolic symdrome.Aminopeptidases are metallopeptidases that cleave N-terminal residues from proteins and peptides.Aminopeptidases constitute a diverse set of peptidases with important roles in cell maintenance,growth and development,and defense.Historically,aminopeptidases were regarded as a ubiquitous set of enzymes with mundane roles in cell homeostasis. Aminopeptidases are often thought to solely provide the final events in the journey of a polypeptide through a cell,converting the peptides released by endoproteases or the proteasome to their amino acid constituents.In recent years,aminopeptidases have been identified as true renaissance proteins harboring a kaleidoscope of functions,such as recepter of cellular signals,the compannion molecular of GLUT 4.And it take part in the regulation of RAAS system,and have relation with blood pressure.It is clear that the functional roles of this very versatile set of peptidases may still be underestimated.Bsaed on such background,we investigated the aminopeptidases activty in MS.Objectives(1) To investigate the serum AAP and LAP activity in patients with MS,and to probe into the potential mechanism underlying these changes.(2) To investigate the alterations in structure and function of the carotid artery,as well as the association of serum AAP and LAP activity levels with these changes.(3) To investigate the alterations in structure and function of the heart,as well as the association of serun AAP and LAP activity levels with these changes.Subjects and MethodsAccording to IDF 2005 criteria of MS,we enrolled 103 patients with MS(49 females and 54 males,51.1±7.0 yr of age) and 97 people matched with age and sex(P＞0.05) without MS(51 females and 46 males,48.8±9.7 yr of age).Height(H),weight(W), systolic blood pressure(SBP),diastolic blood pressure(DBP),waist circumference (WC) and hip circumference(HC).Pulse pressure(PP),body mass index(BMI) and waist-hip ratio(WHR) were calculated.Furthermore,fasting blood sample was collected from each subjects after 12-14 hours fast to determine the total cholesterol(TC), triglycerides(TG),high-density lipoprotein cholesterol(HDL-C),low-density lipoprotein cholesterol(LDL-C),and uric acid(UA),fasting blood glucose(FBG),insulin(FINS) and HOMA-IR.Chromogenic substrates were used to measure AAP and LAP activities.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).ECG data were recorded concurrently.carotid ultrasonic parameters, including lumen diameter(D),intima-media thickness(IMT),plaque index,flow velocity (V),pressure -strain elastic modulus(E_p),normalized E_p,arterial stiffness(β), arterial compliance(AC),resistance index(RI) and pulse index(PI),were measured.Echocardiograms were obtained with a commercially available ultrasound machine(Vivid 7;GE Vingemed Ultrasound,Horten,Norway) with a 1- to 3-MHZ phased array transducer.The M-mode echocardiography,two-dimensional echocardiography, pulsed-Doppler echocardiography and tissue Doppler echocardiography were performed respectively.Cardiac ultrasonic parameters,including ascending aorta diameter(AO), left atrium(LA) diameter,left ventricle(LV) diameter,interventricular septum(IVS) thickness,LV posterior wall thickness,left ventricle mass(LVM),left ventricle mass index (LVMI),left atrium index,left ventricular ejection fraction(LVEF),early diastolic mitral inflow velocity(E),late diastolic mitral inflow velocity(A),E/A,systolic mitral annular velocity(V_s),early diastolic mitral annular velocity(V_e),late diastolic mitral annular velocity(V_a),V_e/V_a and E/V_e,were measured.All statistical analyses were performed using the SPSS 13.0 statistical program. Normally distributed data were expressed as mean±SD or±SEM as indicated,and non-normally distributed data are presented as medians(quartile range).Continuous variables were compared between groups by unpaired Student's t test.The associations between categorical ones was analyzed using the chi-square test.Co- rrelation coefficients between the variables were determined using the Pearson's or Spearman's method, meanwhile multiple regression analysis was performed to determine relationships between variables of interest.A P value＜0.05 wasconsidered statistically significant. Results(1) Comparison of clinical and biochemical characteristics of the subjectsThere were no significant differences in age and gender between MS group and control group.Compared with the controls,the MS group showed significantly higher systolic blood pressure(SBP),diastolic blood pressure(DBP),pulse pressure(PP), W,BMI,WC,HC,WHR,TC,TG and LDL-C,UA,FBG,FINS,HOMA-IR, but lower HDL-C(P＜0.001,respectively).(2) Comparison of carotid ultrasonic parametersThe mean IMT(P＜0.001),maximum IMT(P=-0.044),D_d(P=0.011),plaque index (P=-0.003) of the MS group were significantly higher than those of the control group. Furthermore the MS group showed significantly decreases in V_s(P=0.002),V_d(P=-0.005), V_m(P=-0.004),RI(P＜0.001) and AC(P＜0.001),with significantly increases in PI (P＜0.001),E_p(P＜0.001),normalized E_p(P＜0.001) andβ(P＜0.001).(3) Comparison of cardiac ultrasonic parametersThe IVSd,LVPWd,LVM,LVMI,LA,AO of the MS group were significantly higher than those of the control group.Furthermore the MS group showed significantly decreases in E/A,TVI-E,TVI-E/A,TVI-S,with significantly increases in mitral-A.(4) Comparison of serum enzymatic activity parametersThe serum AAP activity levels(51.84±14.01vs.46.79±9.49,P=0.003) and AAP activity levels(41.27±10.17 vs.37.12±7.07,P=0.001)of the MS group were significantly higher than those of the control group.(5) Correlation of serun AAP activity levels with clinical and biochemical characteristicsSerun AAP activity levels was significantly correlated positively with DBP,Wt,S,BMI,WC,WHR,TC,LDL-c,FINS,HOMA-IR..(6) Effects of the MS components on serum AAP activity levelsStepwise multiple linear regression model for serum AAP activity expression level was as follows:serum AAP activity=0.200HOMA-IR,P=0.008.R2=0.040.Only HOMA-IR were enrolled,suggesting the association of serum AAP activity level with insulin resistance. (7) Correlation of serum AAP activity levels with carotid ultra-sonic parametersserum AAP activity levels was independently significantly correlated negatively with IR(P=0.017).(8) Effects of the serum AAP activity level on RI.Stepwise multiple linear regression model for RI was as follows: RI=-0.261SBP-0.285TG-0.228BMI-0.530sex-0.217UA-0.229smoking-0.135 Serum AAP activitySerum AAP activity was enrolled(P=0.038),suggesting the association of RI with serum AAP activity level.(9) Correlation of serum AAP activity levels with carotid ultra-sonic parametersSerum AAP activity levels was independently significantly correlated negatively with LV(P=0.026).(10) Effects of the serum AAP activity level on LVStepwise multiple linear regression model for RI was as follows: LV=0.252S+0.188smoking.LV was not enrolled.(11) Correlation of serun AAP activity levels with clinical and biochemical characteristicsSerun AAP activity levels was significantly correlated positively with Wt,BMI,S,WC,WHR,DBP,TC,LDL,TG,FBG,FINS,HOMA-IR(P＜0.05),significantly correlated negatively with HDL(P＞0.05).(12) Effects of the MS components on serum LAP activity levelsStepwise multiple linear regression model for serum AAP activity expression level was as follows:serum LAP activity=0.223HOMA-IR,P=0.003.R2=0.050.Only HOMA-IR were enrolled,suggesting the association of serum LAP activity level with insulin resistance.(13) Correlation of serum LAP activity levels with carotid ultra-sonic parametersSerum AAP activity levels was significantly correlated negatively with RI(P=0.011).Partial correlation show serum AAP activity levels was significantly independently correlated negatively with RI(P=0.027),was significantly independently correlated positively with Vd(P=0.030). (14) Effects of the serum LAP activity level on RIStepwise multiple linear regression model for serum AAP activity expression level was as follows:RI=-0.277SBP-0.276TG-0.233BMI-0.347sex-0.201UA-0.153 serum LAP activity.Serum LAP activity was enrolled(P=0.021),suggesting the association of serum LAP activity level with perphral resistance.(15) Effects of the serum LAP activity level on VdStepwise multiple linear regression model for serum AAP activity expression level was as follows:Vd=-0.401WHR-0.330age+0.145 serum LAP activity.Serum LAP activity was enrolled(P=0.036),suggesting the association of serum LAP activity level with Vd.(16) Effects of the serum LAP activity level on AOStepwise multiple linear regression model for serum AAP activity expression level was as follows:AO=0.330WHR+0.246SBP+0.185UA-0.177 serum LAP activity.Serum LAP activity was enrolled(P=0.021),suggesting the association of serum LAP activity level with AO.(17) Correlation of serum LAP activity levels with cardiac ultra-sonic parametersSerum AAP activity levels was significantly correlated positively with RI(P=0.011).Partial correlation show no relation.Conclusions(1) The serum AAP and LAP activity levels of the MS group were obviously increased.Elevated AAP and LAP levels activity were biomarks of insulin resistance,they both were independent risk factors for insulin resistance,even the cause of it.(2) Significant alterations to carotid arteries of the MS patients include thickened IMT, increased plaque index,distended vessel diameter,reduced elasticity and augmented stiffness.(3) Left ventricular structure and function were impaired in patients with metabolic syndrome,which were characterized as LV remolding and fragmental and global diastolic dysfunction.The left ventricular global systolic function was demonstrated normal,while tissue Doppler imaging revealed a reduced myocardial velocity,a process possibly happened earlier than the development of global systolic dysfunction in MS.(4) Serum AAP and LAP activity levels was significantly correlated negatively with RI, Serum LAP activity levels was significantly correlated positively with Vd,negatively with AO,suggesting that elevated serum AAP and LAP activity levels can decrease periphral resistance as an protective mechanisem. BackgroundMetabolic syndrome is a constellation of cardiovascular risk factors,including obesity,hypertension,dyslipidemia and hyperglycemia,which are correlated with cardiovascular disease and diabetes.These risk factors interact and interrelate with each other,amplify the risk of CVD and type 2 diabetes mellitus.Because of the epidemic of over-weight and sedentary lifestyle world-wide,the metabolic syndrome is becoming increasingly common.The mechanism of MS is complex.Recently the role of chronic systematic inflammation in the pathogenesis of MS has received more and more attention.So researchers pay much attention to expression of inflammation factors in the MS.Aminopeptidase N(APN)/CD13 is a membrane-bound surface ectopeptidase with a ubiquitous distribution.As an ectopeptidase,its spectrum encompasses some inflammatory peptides,exerting an anti-inflammation function.And it has some other functions related to the regulation of blood pressure and lipid metabolism.Until now,the CD13 expression in MS has not been investigated.Doppler ultrasound as the noninvasive diagnostic method has been widely applicated in clinical practice.The high-resolution sonography,providing a direct and dynamic evaluation of artery structure and function,improves the prediction of cardiovascular events greatly.Tissue Doppler imaging,a relatively new echocardiographic technique,offers a more sensitive method for the detection of the left ventricular long-axis myocardial velocity changes,which reflect the alterations in cardiac structure more accurately,and thus for the assessment of cardiac systolic and diastolic function.Based on these knowledge,we investigated the expression levels of CD13 in patients with MS, observed alterations in structure and function of the heart and carotid arteries,and discussed the relationship between expression of these genes and MS components,as well as its effect on the target organ damage.Objectives(1) To investigate the mRNA expression levels of CD13 in peripheral blood mononuclear cells(PBMCs)in patients with MS,and to probe into the potential mechanism underlying these changes.(2) To investigate the alterations in structure and function of the carotid artery,as well as the association of CD13 mRNA expression levels with these changes.(3) To investigate the alterations in structure and function of the heart,as well as the association of CD13 mRNA expression levels with these changes.Subjects and MethodsAccording to IDF 2005 criteria of MS,we enrolled 48 patients with MS(25 females and 23 males,53.50±9.73 yr of age) and 49 people matched with age and sex(P＞0.05) without MS(35 females and 14 males,50.86±9.24 yr of age).Height(H),weight(W), systolic blood pressure(SBP),diastolic blood pressure(DBP),waist circumference (WC) and hip circumference(HC).Pulse pressure(PP),body mass index(BMI) and waist-hip ratio(WHR) were calculated.Furthermore,fasting blood sample was collected from each subjects after 12-14 hours fast to determine the total cholesterol(TC), triglycerides(TG),high-density lipoprotein cholesterol(HDL-C),low-density lipoprotein cholesterol(LDL-C),and uric acid(UA),fasting blood glucose(FBG),insulin(FINS) and HOMA-IR.Briefly,PBMCs were isolated.Total RNA was extracted using Trizol one-step method.The RT-PCR technique was applied to detect the mRNA expressions of Ob-R and ghrelin in PBMCs.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).ECG data were recorded concurrently,carotid ultrasonic parameters,including lumen diameter(D),intima-media thickness(IMT),plaque index, flow velocity(V),pressure -strain elastic modulus(E_p),normalized E_p,arterial stiffness (β),arterial compliance(AC),resistance index(RI) and pulse index(PI),were measured.Echocardiograms were obtained with a commercially available ultrasound machine(Vivid 7;GE Vingemed Ultrasound,Horten,Norway) with a 1- to 3- MHZ phased array transducer.The M-mode echocardiography,two-dimensional echocardiography, pulsed-Doppler echocardiography and tissue Doppler echocardiography were performed respectively.Cardiac ultrasonic parameters,including ascending aorta diameter(AO), left atrium(LA) diameter,left ventricle(LV) diameter,interventricular septum(IVS) thickness,LV posterior wall thickness,left ventricle mass(LVM),left ventricle mass index (LVMI),left atrium index,left ventricular ejection fraction(LVEF),early diastolic mitral inflow velocity(E),late diastolic mitral inflow velocity(A),E/A,systolic mitral annular velocity(V_s),early diastolic mitral annular velocity(V_e),late diastolic mitral annular velocity(V_a),V_e/V_a and E/V_e,were measured.All statistical analyses were performed using the SPSS 13.0 statistical program. Normally distributed data were expressed as means±SD or±SEM as indicated,and non-normally distributed data are presented as medians(quartile range).Continuous variables were compared between groups by unpaired Student's t test.The associations between categorical ones was analyzed using the chi-square test.Co- rrelation coefficients between the variables were determined using the Pearson's or Spearman's method, meanwhile multiple regression analysis was performed to determine relationships between variables of interest.A P value＜0.05 wasconsidered statistically significant.Results(1) Comparison of clinical and biochemical characteristics of the subjectsThere were no significant differences in age and gender between MS group and control group.Compared with the controls,the MS group showed significantly higher systolic blood pressure(SBP),diastolic blood pressure(DBP),pulse pressure(PP),W, BMI,WC,HC,WHR,TC,TG and LDL-C,UA,FBG,FINS,HOMA-IR,but lower HDL-C(P＜0.001,respectively).(2) Comparison of carotid ultrasonic parametersThe mean IMT(P＜0.001),maximum IMT(P=0.044),D_d(P=0.011),plaque index (P=0.003) of the MS group were significantly higher than those of the control group. Furthermore the MS group showed significantly decreases in V_s(P=0.002),V_d(P=0.005), V_m(P=0.004),RI(P＜0.001) and AC(P＜0.001),with significantly increases in PI (P＜0.001),E_p(P＜0.001),normalized E_p(P＜0.001) andβ(P＜0.001).(3) Comparison of cardiac ultrasonic parametersThe IVSd(P＜0.001),LVPWd(P＜0.001),LVM(P＜0.001),LVMI(P＜0.001), LA(P＜0.001),AO(P＜0.001) of the MS group were significantly higher than those of the control group.Furthermore the MS group showed significantly decreases in E/A (P＜0.001),TVI-E(P=0.004),TVI-E/A(P＜0.001),TVI-S(P=0.001),with significantly increases in mitral-A(P=0.002).(4) Comparison of gene expression parametersThe mRNA expression levels of CD13(-6.833±1.224 vs.-6.777±1.270,P=0.827) in PBMCs of the MS group were not changed significantly.(5) Correlation of CD13 mRNA expression levels with clinical and biochemical characteristicsCD13 mRNA expression level was significantly correlated negatively with age(P=0.002),SBP(P=0.014),PP(P=0.013).(6) Effects of the MS components on CD13 mRNA levelsStepwise multiple linear regression model for CD13 mRNA expression level was as follows:CD13 mRNA=-0.345WHR+0.380SBP.WHe(P=0.002)and SBP(P=0.001) was enrolled,sugesting the association of CD13 mRNA with WHR and SBP.(7) Correlation of CD13 mRNA expression levels with carotid ultra-sonic parametersCD13 mRNA expression level was significantly correlated negatively with IMTthickness(P=0.024).Patial corrlation show no relation.(8) Correlation of CD13 mRNA and ghrelin expression levels with cardiac ultra-sonic parametersCD13 mRNA expression level was significantly correlated negatively with LVEF (P=0.030),and positively with TVIA(P=0.038).Patial corrlation show CD13 mRNA expression level was significantly correlated negatively with TVIE(P=0.014).(9) Effects of the CD13 mRNA expression on cardiac functionStepwise multiple linear regression model for TVIE was as follows: TVIE=-0.360age-0.389BMI-0.263CD13mRNA.CD13mRNA was enrolled,suggesting the association of TVIE with CD13 mRNA expression level.Conclusions1.The mRNA expression levels of CD13 in PBMCs of the MS group were not changed;2.The mRNA expression levels of CD13 in PBMCs was negatively correlated with WHR,suggesting abdominal obesity can inhibit the mRNA expression levels of CD13,maybe via the inflammation factors.And this chang can be seen as indicater of inflammation state.3.The mRNA expression levels of CD13 in PBMCs was positively correlated with ABP,.And it is a manifestion of the inflammation state.4.The mRNA expression levels of CD13 in PBMCs was negatively correlated with TVIE, suggesting CD13 can inhibit the cardiac relaxation,through influencing the fibrosis of the cardiac muscle.