kidney transplant recipients ( ktrs ) are known to be at increased risk for cardiovascular disease and by three years after transplant about 40% of ktrs have experienced a cardiovascular disease - related event . the incidence of disease is occurring in the context of best medical practice for treatment of established cardiovascular risk factors . thus there is a need to identify novel risk factors with the objective of improving treatment and reduction of the incidence of cardiovascular disease . end - stage renal disease , the precursor to ktr , is increasing in individuals of south asian ethnicity relative to subjects of caucasian ethnicity . there is a need to characterize these subjects and determine whether novel risk factors for cardiovascular disease are present in an ethnicity - dependent manner . the current study was designed to compare two novel risk factors , adiponectin and paraoxonase 1 ( pon1 ) , as candidates for differing risk between south asian and caucasian ktr . stage 3 chronic renal disease , defined as cystatin c - based estimated glomerular filtration rate ( egfr ) less than 60 , was associated with lower pon1 mass in subjects with diabetes . in this paper we present the analysis for pon1 and report that pon1 gln192arg ( q192r ) phenotype is different by ethnicity . subjects were recruited with age ( 10 years ) and gender matched in a 1  :  2 , south asian  :  caucasian ratio . inclusion criteria were ktr of stated ethnicity with stable renal function ( less than 20% fluctuation of creatinine in the 2 months preceding recruitment ) , within 3 to 60 months of transplant , and able to provide informed consent . subjects were excluded if ethnicity was uncertain , if they were multiorgan transplant recipients , if they had unstable renal function , or if they were unable to understand the consent process . subjects who consented provided a blood sample at the time of clinic visit in the nonfasting state . a total of 60 caucasian males , 30 south asian males , 26 caucasian females , and 13 south asian females were studied , slightly exceeding the study design target sample of 120 subjects . a complete description of the subjects can be found in prasad et al . . routine clinical laboratory measurements were performed for standard blood clinical chemistry analytes , including creatinine , glucose , and hemoglobin a1c . clinical records were used for determination of the presence of diabetes according to the 2008 canadian diabetes association guidelines . high - density lipoprotein cholesterol was determined by a homogeneous assay on the beckman lx-20 instrument ( beckman , mississauga , on ) . cystatin c , apolipoprotein b , and apolipoprotein ai were measured using the dade - behring bn prospec ( dade - behring , mississauga , on ) . pon1 mass was measured on serum samples by western blot with a recombinant standard and calibrated with a normal serum sample . pon1 arylesterase activity was measured using 3  mm phenyl acetate to determine the arylesterase activity and expressed in u / ml . pon1 phenotype was determined using the two - substrate assay of richter et al . data for the hydrolysis of 4-(chloromethyl)phenyl acetate ( cmpa ) was plotted on the x - axis and data for hydrolysis of phenyl acetate in the presence of 2  m nacl ( high salt conditions ) was plotted on the y - axis . data was transformed using the arctangent of the ratio of cmpa / arylesterase activity to derive the radians from the origin . the activity for purified pon1 q192 and pon1 r192 was used to confirm identity of the qq and rr samples . the pon1 phenotypes were assigned as follows : qq , arctangent range from 11.8 to 17.08 degrees ; qr , arctangent range from 20.7 to 34.9 degrees ; and rr , arctangent range from 51.3 to 57.4 degrees . subjects were well matched by age at transplant ( 50 versus 52 years for south asian versus caucasian ) and time after transplant ( 65 versus 71 months for south asian versus caucasian ) . none of the established risk factors were significantly different between south asian and caucasian subjects by univariate analysis . pon1 mass and arylesterase activity were similar between the two groups and had a similar distribution as seen by comparison of the 25th and 75th percentiles . univariate spearman correlation was determined for pon1 mass and arylesterase activity by gender . among women , arylesterase activity correlated with pon1 mass ( r = 0.71 , p < 0.0001 ) , hdl cholesterol ( r = 0.45 , p = 0.004 ) , and apoai ( r = 0.35 , p = 0.03 ) . among men , arylesterase activity correlated with pon1 mass ( r = 0.78 , p < 0.0001 ) , but not with hdl cholesterol ( r = 0.14 , p = ns ) and weakly with apoai ( r = 0.21 , p = 0.05 ) . pon1 mass was not significantly correlated with either hdl cholesterol or apoai in either gender . pon1 phenotype was determined as described by richter et al .   using the two - substrate method . we observed clear separation of the results into three phenotypes representing the qq , qr , and rr phenotypes ( figure 1 ) . the frequency of qq , qr , and rr was significantly different (  = 7.72 , p = 0.02 ) between south asian and caucasian subjects with the rr phenotype being 3 times more prevalent in south asian subjects ( table 2 ) . the determinants of pon1 mass were determined with ethnicity and the covariates pon1 phenotype , lipoprotein concentrations , diabetes status , and renal status as covariates . in spite of the difference in pon1 phenotype , in contrast , pon1 phenotype , diabetes status , hdl cholesterol , and ldl cholesterol were all significantly associated with pon1 mass ( table 3 ) .   next the determinants of serum arylesterase were studied ( table 4 ) . again , ethnicity was not significantly associated with arylesterase activity . among the variables tested , only hdl cholesterol was significantly associated with arylesterase activity ( p = 0.003 ) . when regression analysis was performed including pon1 mass as an independent variable , pon1 mass had a partial r of 0.67 ( p < 0.001 ) , pon1 phenotype had a partial r of 0.11 ( p < 0.001 ) , and hdl cholesterol had a partial r of 0.007 ( p = 0.05 ) . thus 77% of the variation on arylesterase activity was accounted for by pon1 mass and phenotype and less than 1% was accounted for by hdl cholesterol . neither creatinine - based ( mdrd ) nor cystatin c - based ( arnal - dade ) estimates of glomerular filtration rate ( egfr ) showed a relationship with pon1 mass or arylesterase activity ( data not shown ) . pon1 has been extensively studied as a risk factor for cardiovascular disease independent of lipoprotein concentrations . differences in pon1 between subjects have been shown to be due to genetic polymorphisms [ 7 , 8 ] , presence of diabetes , presence of hepatic disease , and presence of renal disease [ 3 , 1115 ] . the complexity of the genetic polymorphism at the pon1 locus may confound studies of pon1 and thus it has been suggested that determination of the pon1 phenotype , mass , and activity be measured [ 5 , 16 ] . we report that the phenotypic prevalence of the qr and rr phenotype is higher in south asian compared with caucasian ktr subjects . the frequency of the phenotypes in caucasians is similar to that reported for renal transplant patients in debrecen , hungary [ 17 , 18 ] . the frequency of the phenotypes in south asians is consistent with recently published data on the frequency of the of gln192arg polymorphism in north - west indian punjabis . it is also similar to that reported for patients with cad or t2 dm from hyderabad , india . interestingly , the frequency of the rr was 11.7% in control subjects from hyderabad , similar to the frequency we observed for caucasians . in contrast to the difference in phenotype by ethnicity , there was no difference in the mass or arylesterase activity of pon1 by ethnicity in ktr . further , there was a difference in pon1 mass , but not arylesterase activity by diabetes status . analysis of indices of diabetes ( glucose , hemoglobin a1c ) did not show significant associations with pon1 mass or arylesterase activity ( data not shown ) . post hoc analyses for an interaction between ethnicity and diabetes status were also negative . thus , in this group of ktr subjects , diabetes was a limited determinant of pon1 status . similarly , an a priori hypothesis was that renal function would be a determinant of pon1 . however , neither creatinine nor cystatin c - based estimates of glomerular filtration rate were associated with pon1 . it should also be noted that the study design was to restrict subjects to a range of post renal transplant from 3 to 60 months . thus although most of the subjects with diabetes had pre - transplant diabetes , the duration of posttransplant factors was limited to the relatively early time points . most previous studies of non - ktr have been of subjects with an average duration of diabetes greater than 15 years [ 20 , 21 ] . thus one explanation for the absence of associations with diabetes and egfr could be the shorter duration after transplant in this study . pon1 mass , but not arylesterase activity , has been reported to be inversely correlated with mortality in patients on dialysis . in contrast , pon1 genotypes have not been associated with increased cardiovascular risk in renal transplant recipients . future studies will require measurement of pon1 phenotype by enzymatic activity and pon1 concentration in order to fully evaluate the predictive value of pon1 as a cardiovascular risk factor in kidney transplant recipients .
south asian renal transplant recipients have a higher incidence of cardiovascular disease compared with caucasian renal transplant recipients . 
 we carried out a study to determine whether paraoxonase 1 , a novel biomarker for cardiovascular risk , was decreased in south asian compared with caucasian renal transplant recipients . 
 subjects were matched two to one on the basis of age and sex for a total of 129 subjects . 
 paraoxonase 1 was measured by mass , arylesterase activity , and two - substrate phenotype assay . 
 comparisons were made by using a matched design . 
 the frequency of pon1 qq , qr and rr phenotype was 56% , 37% , and 7% for caucasian subjects versus 35% , 44% , and 21% for south asian subjects 
 ( 2 = 7.72 , p = 0.02 ) . 
 pon1 mass and arylesterase activity were not significantly different between south asian and caucasian subjects . 
 pon1 mass was significantly associated with pon1 phenotype ( p = 0.0001 ) , hdl cholesterol ( p = 0.009 ) , ldl cholesterol ( p = 0.02 ) , and diabetes status ( p < 0.05 ) . 
 arylesterase activity was only associated with hdl cholesterol ( p = 0.003 ) . 
 thus the frequency of the pon1 rr phenotype was higher and that of the qq phenotype was lower in south asian versus caucasian renal transplant recipients . 
 however , ethnicity was not a significant factor as a determinant of pon1 mass or arylesterase activity , with or without analysis including pon1 phenotype . 
 the two - substrate method for determining pon1 phenotype may be of value for future studies of cardiovascular complications in renal transplant recipients .