other exclusion criteria were history or clinical evidence of coronary or valvular heart disease , congestive heart failure , hyperlipidemia , peripheral vascular disease , chronic gastrointestinal diseases associated with malabsorption , chronic pancreatitis , history of any malignant disease , history of alcohol or drug abuse , liver or kidney failure , and treatments able to modify glucose metabolism .
after 12-h fasting , a 75-g ogtt was performed with 0- , 30- , 60- , 90- , and 120-min sampling for plasma glucose and insulin .
glucose tolerance status was defined on the basis of ogtt using the world health organization ( who ) criteria .
insulin sensitivity was evaluated using the matsuda index ( insulin sensitivity index [ isi ] ) , calculated as follows : 10,000/square root of [ fasting glucose ( millimoles per liter )  fasting insulin ( milliunits per liter ) ]  [ mean glucose  mean insulin during ogtt ] . the matsuda index is strongly related to euglycemic - hyperinsulinemic clamp , which represents the gold standard test for measuring insulin sensitivity ( 13 ) .
the ethics committee approved the protocol , and informed written consent was obtained from all participants .
all of the investigations were performed in accordance with the principles of the declaration of helsinki .
readings of clinic blood pressure were obtained in the left arm of the supine patients , after 5 min of quiet rest , with a mercury sphygmomanometer .
a minimum of three blood pressure readings were taken on three separate occasions at least 2 weeks apart .
systolic blood pressure ( sbp ) and diastolic blood pressure ( dbp ) were recorded at the first appearance ( phase i ) and the disappearance ( phase v ) of korotkoff sounds .
baseline blood pressure values were the average of the last two of the three consecutive measurements obtained at intervals of 3 min .
patients with a clinic sbp > 140 mmhg and/or dbp > 90 mmhg were defined as hypertensive .
plasma glucose was measured by the glucose oxidation method ( beckman glucose analyzer ii ; beckman instruments , milan , italy ) .
triglyceride and total , ldl , and hdl cholesterol concentrations were measured by enzymatic methods ( roche diagnostics , mannheim , germany ) .
plasma insulin concentration was determined by a chemiluminescence - based assay ( roche diagnostics ) .
tracings were taken with the patient in a partial left decubitus position , using a vivid 7 pro ultrasound machine ( ge technologies , milwaukee , wi ) with an annular phased array 2.5-mhz transducer .
echocardiographic readings were made in random order by the investigator , who had no knowledge of patients blood pressure or other clinical data .
the mean values from at least five measurements of each parameter for each patient were computed .
having the same experienced sonographer perform all studies in a dimly lit and quiet room optimized the reproducibility of measurements .
subjects with a left ventricular ejection fraction < 50% were excluded from this study . in our laboratory ,
the intraobserver coefficients of variation were 3.85% for posterior wall thickness , 3.7% for interventricular septal thickness , 1.5% for left ventricular internal diameter , and 5.1% for left ventricular mass ( lvm ) .
tracings were recorded under two - dimensional guidance , and m - mode measurements were taken at the tip of the mitral valve or just below .
measurements of interventricular septal thickness , posterior wall thickness , and left ventricular internal diameter were made at end diastole and end systole , as recommended by the american society of echocardiography ( 14 ) .
lvm was calculated using the devereux formula and normalized by body surface area ( lvmi ) .
left ventricular diastolic function was evaluated according to diagnostic criteria proposed by the american society of echocardiography ( 15 ) .
evaluation of left atrium volume was obtained using the apical four - chamber and two - chamber views .
pulsed doppler transmitral flow velocity profile was obtained from the apical four - chamber view , and the sample volume was positioned at the tip of the mitral valve leaflets .
the following parameters were evaluated for diastolic function : peak transvalvular flow velocity in early diastole ( e wave ) , peak transvalvular flow velocity in late diastole ( a wave ) , e - to - a ratio , deceleration time ( dt ) ( time measured between peak e velocity and the point where the deceleration slope of the e velocity crosses the zero baseline ) , and isovolumic relaxation time ( ivrt ) ( time elapsed between aortic valve closure and mitral valve opening ) .
pulsed wave tissue doppler imaging ( tdi ) was performed at the junction of the septal and lateral mitral annulus .
early diastolic ( septal e and lateral e ) and late diastolic ( septal a and lateral a ) velocities were recorded ; ratio of e - to - e ( average ) was also calculated .
the intraobserver coefficients of variation for measurement of diastolic parameters ranged from 1.8 to 4.1% .
anova for clinical and biological data was performed to test the differences among groups , and the bonferroni post hoc test for multiple comparisons was further performed .
linear regression analysis was performed to relate parameters of diastolic function with the following covariates : age , bmi , sbp , dbp , fasting and 1- and 2-h postload plasma glucose levels , fasting and 1- and 2-h postload insulin , matsuda index , and lvmi .
subsequently , variables reaching statistical significance and sex , as dichotomic value , were inserted in a stepwise multivariate linear regression model to determine the independent predictors of diastolic dysfunction .
correlational analysis was performed for the whole study population and according to different groups of glucose tolerance .
all comparisons were performed using the statistical package spss 16.0 for windows ( spss , chicago , il ) .
readings of clinic blood pressure were obtained in the left arm of the supine patients , after 5 min of quiet rest , with a mercury sphygmomanometer .
a minimum of three blood pressure readings were taken on three separate occasions at least 2 weeks apart .
systolic blood pressure ( sbp ) and diastolic blood pressure ( dbp ) were recorded at the first appearance ( phase i ) and the disappearance ( phase v ) of korotkoff sounds .
baseline blood pressure values were the average of the last two of the three consecutive measurements obtained at intervals of 3 min .
patients with a clinic sbp > 140 mmhg and/or dbp > 90 mmhg were defined as hypertensive .
plasma glucose was measured by the glucose oxidation method ( beckman glucose analyzer ii ; beckman instruments , milan , italy ) .
triglyceride and total , ldl , and hdl cholesterol concentrations were measured by enzymatic methods ( roche diagnostics , mannheim , germany ) .
plasma insulin concentration was determined by a chemiluminescence - based assay ( roche diagnostics ) .
tracings were taken with the patient in a partial left decubitus position , using a vivid 7 pro ultrasound machine ( ge technologies , milwaukee , wi ) with an annular phased array 2.5-mhz transducer .
echocardiographic readings were made in random order by the investigator , who had no knowledge of patients blood pressure or other clinical data .
the mean values from at least five measurements of each parameter for each patient were computed .
having the same experienced sonographer perform all studies in a dimly lit and quiet room optimized the reproducibility of measurements .
subjects with a left ventricular ejection fraction < 50% were excluded from this study . in our laboratory ,
the intraobserver coefficients of variation were 3.85% for posterior wall thickness , 3.7% for interventricular septal thickness , 1.5% for left ventricular internal diameter , and 5.1% for left ventricular mass ( lvm ) .
tracings were recorded under two - dimensional guidance , and m - mode measurements were taken at the tip of the mitral valve or just below .
measurements of interventricular septal thickness , posterior wall thickness , and left ventricular internal diameter were made at end diastole and end systole , as recommended by the american society of echocardiography ( 14 ) .
lvm was calculated using the devereux formula and normalized by body surface area ( lvmi ) .
left ventricular diastolic function was evaluated according to diagnostic criteria proposed by the american society of echocardiography ( 15 ) .
evaluation of left atrium volume was obtained using the apical four - chamber and two - chamber views .
pulsed doppler transmitral flow velocity profile was obtained from the apical four - chamber view , and the sample volume was positioned at the tip of the mitral valve leaflets .
the following parameters were evaluated for diastolic function : peak transvalvular flow velocity in early diastole ( e wave ) , peak transvalvular flow velocity in late diastole ( a wave ) , e - to - a ratio , deceleration time ( dt ) ( time measured between peak e velocity and the point where the deceleration slope of the e velocity crosses the zero baseline ) , and isovolumic relaxation time ( ivrt ) ( time elapsed between aortic valve closure and mitral valve opening ) .
pulsed wave tissue doppler imaging ( tdi ) was performed at the junction of the septal and lateral mitral annulus .
early diastolic ( septal e and lateral e ) and late diastolic ( septal a and lateral a ) velocities were recorded ; ratio of e - to - e ( average ) was also calculated .
the intraobserver coefficients of variation for measurement of diastolic parameters ranged from 1.8 to 4.1% .
tracings were recorded under two - dimensional guidance , and m - mode measurements were taken at the tip of the mitral valve or just below .
measurements of interventricular septal thickness , posterior wall thickness , and left ventricular internal diameter were made at end diastole and end systole , as recommended by the american society of echocardiography ( 14 ) .
lvm was calculated using the devereux formula and normalized by body surface area ( lvmi ) .
left ventricular diastolic function was evaluated according to diagnostic criteria proposed by the american society of echocardiography ( 15 ) .
evaluation of left atrium volume was obtained using the apical four - chamber and two - chamber views .
pulsed doppler transmitral flow velocity profile was obtained from the apical four - chamber view , and the sample volume was positioned at the tip of the mitral valve leaflets .
the following parameters were evaluated for diastolic function : peak transvalvular flow velocity in early diastole ( e wave ) , peak transvalvular flow velocity in late diastole ( a wave ) , e - to - a ratio , deceleration time ( dt ) ( time measured between peak e velocity and the point where the deceleration slope of the e velocity crosses the zero baseline ) , and isovolumic relaxation time ( ivrt ) ( time elapsed between aortic valve closure and mitral valve opening ) .
e - to - a ratio between 1 and 2 was defined as normal . pulsed wave tissue doppler imaging ( tdi ) was performed at the junction of the septal and lateral mitral annulus .
early diastolic ( septal e and lateral e ) and late diastolic ( septal a and lateral a ) velocities were recorded ; ratio of e - to - e ( average ) was also calculated .
the intraobserver coefficients of variation for measurement of diastolic parameters ranged from 1.8 to 4.1% .
anova for clinical and biological data was performed to test the differences among groups , and the bonferroni post hoc test for multiple comparisons was further performed .
linear regression analysis was performed to relate parameters of diastolic function with the following covariates : age , bmi , sbp , dbp , fasting and 1- and 2-h postload plasma glucose levels , fasting and 1- and 2-h postload insulin , matsuda index , and lvmi .
subsequently , variables reaching statistical significance and sex , as dichotomic value , were inserted in a stepwise multivariate linear regression model to determine the independent predictors of diastolic dysfunction .
correlational analysis was performed for the whole study population and according to different groups of glucose tolerance .
all comparisons were performed using the statistical package spss 16.0 for windows ( spss , chicago , il ) .
of 161 patients examined by ogtt , 120 had normal glucose tolerance ( ngt ) , 26 had igt , and 15 had newly diagnosed t2d .
a 1-h postload plasma glucose cutoff point of 155 mg / dl during ogtt was used to stratify ngt subjects into two groups : 90 patients with 1-h postload plasma glucose < 155 mg / dl ( ngt < 155 ) and 30 individuals with 1-h postload plasma glucose 155 mg / dl ( ngt 155 ) .
table 1 shows the demographic , clinical , and biochemical characteristics of the four study groups .
anthropometric , hemodynamic , and biochemical characteristics of the study population according to glucose tolerance data are means  sd unless otherwise indicated .
there were no significant differences among groups for sex , age , bmi , sbp , dbp , and total cholesterol .
from the first to the fourth group , there was a significant increase of triglyceride ( p = 0.002 ) and a significant reduction of hdl cholesterol ( p = 0.034 ) . obviously , a progressive increase of fasting and 1-h and 2-h postload glucose parallels the worsening of glucose tolerance ( p < 0.0001 ) .
fasting and postload insulin values were higher in the ngt 155 group and igt subjects in comparison with those in the ngt<155 group and diabetic patients , respectively .
echocardiographic parameters for the study population , according to glucose tolerance groups , are reported in table 2 .
t2d patients had the highest lvmi value ( p = 0.020 ) , and clinically relevant , ngt 155 subjects showed an lvmi value not significantly different from igt patients ( p = 0.691 ) but significantly higher than ngt < 155 subjects ( p = 0.042 ) .
lvmi and echocardiographic parameters of left ventricular diastolic function according to glucose tolerance data are means  sd unless otherwise indicated .
it is interesting to note that the left atrium volume and ivrt values significantly increased from the first to the fourth group ( p < 0.0001 ) and that ngt 155 subjects showed both parameters significantly higher than ngt < 155 subjects ( p < 0.0001 for left atrium and p = 0.037 for ivrt ) .
moreover , left atrium dimensions in ngt 155 subjects were similar to those of the igt group and t2d patients ; ngt 155 subjects showed an ivrt value similar to that of igt subjects ( p = 0.614 ) and t2d patients ( p = 0.120 ) .
by contrast , e - to - a ratio significantly decreased from the first to the fourth group ( p < 0.0001 ) , and it was significantly lower in ngt 155 than in ngt<155 subjects .
no significant differences among groups were observed for the dt duration ( p = 0.391 ) .
e - to - e ratio significantly increased from the first to the fourth group ( p < 0.044 ) and in ngt 155 subjects was significantly higher than in ngt < 155 subjects ( p < 0.036 ) and similar to that of diabetic patients .
all remaining tissue doppler parameters were significantly decreased from the first to the fourth group , confirming the progressive impairment of left ventricular diastolic dysfunction from the first to the fourth group of glucose tolerance .
it is important to remark that ngt 155 subjects had the same characteristics observed in igt and t2d patients .
a linear regression analysis was performed to test the correlation between echocardiographic parameters and different covariates ( table 3 ) .
one - hour postload glucose was linearly correlated with ivrt ( r = 0.426 ; p < 0.0001 ) and left atrium volume ( r = 0.366 ; p = 0.0001 ) and inversely correlated with septal e-to - a ratio ( r = 0.502 ; p < 0.0001 ) , lateral e ( r = 0.427 ; p < 0.0001 ) , septal e ( r = 0.389 ; p < 0.0001 ) , lateral e-to - a ratio ( r = 0.384 ; p < 0.0001 ) , e - to - e ratio ( r = 0.161 ; p = 0.020 ) , and e - to - a ratio ( r = 0.238 ; p = 0.001 ) . linear regression analysis
( r / p ) between diastolic function parameters and different covariates bsa , body surface area ; dt , deceleration time .
thus , variables reaching statistical significance and sex , as dichotomic value , were inserted in a stepwise multivariate linear regression model to determine the independent predictors of left ventricular diastolic function parameters ( table 4 ) . in the study population ,
1-h postload glucose was the first predictor of left atrium volume , ivrt , and dt , explaining 25.6 ( p < 0.0001 ) , 18.2 ( p < 0.0001 ) , and 2.9% ( p = 0.031 ) of its variation , respectively .
age , sex , and dbp added another 5.6 ( p = 0.001 ) , 3.1 ( p = 0.013 ) , and 2.8% ( p = 0.032 ) to explaining left atrium volume , ivrt , and dt , respectively .
as for the tissue doppler parameters , we observed that 1-h postload glucose was retained as the first correlated of septal e , septal e-to - aratio , lateral e , and lateral e-to - a ratio , explaining 15.2 , 25.2 , 18.3 , and 14.8% of their variations , respectively .
age and sbp were retained as the independent predictors of e - to - e ratio , explaining 8.3% of its variation .
stepwise multiple regression analysis on diastolic function parameters as dependent variables in the whole study population bsa , body surface area .
echocardiographic parameters for the study population , according to glucose tolerance groups , are reported in table 2 .
t2d patients had the highest lvmi value ( p = 0.020 ) , and clinically relevant , ngt 155 subjects showed an lvmi value not significantly different from igt patients ( p = 0.691 ) but significantly higher than ngt < 155 subjects ( p = 0.042 ) .
lvmi and echocardiographic parameters of left ventricular diastolic function according to glucose tolerance data are means  sd unless otherwise indicated .
it is interesting to note that the left atrium volume and ivrt values significantly increased from the first to the fourth group ( p < 0.0001 ) and that ngt 155 subjects showed both parameters significantly higher than ngt < 155 subjects ( p < 0.0001 for left atrium and p = 0.037 for ivrt ) .
moreover , left atrium dimensions in ngt 155 subjects were similar to those of the igt group and t2d patients ; ngt 155 subjects showed an ivrt value similar to that of igt subjects ( p = 0.614 ) and t2d patients ( p = 0.120 ) .
by contrast , e - to - a ratio significantly decreased from the first to the fourth group ( p < 0.0001 ) , and it was significantly lower in ngt 155 than in ngt<155 subjects .
no significant differences among groups were observed for the dt duration ( p = 0.391 ) .
e - to - e ratio significantly increased from the first to the fourth group ( p < 0.044 ) and in ngt 155 subjects was significantly higher than in ngt < 155 subjects ( p < 0.036 ) and similar to that of diabetic patients .
all remaining tissue doppler parameters were significantly decreased from the first to the fourth group , confirming the progressive impairment of left ventricular diastolic dysfunction from the first to the fourth group of glucose tolerance .
it is important to remark that ngt 155 subjects had the same characteristics observed in igt and t2d patients .
a linear regression analysis was performed to test the correlation between echocardiographic parameters and different covariates ( table 3 ) .
one - hour postload glucose was linearly correlated with ivrt ( r = 0.426 ; p < 0.0001 ) and left atrium volume ( r = 0.366 ; p = 0.0001 ) and inversely correlated with septal e-to - a ratio ( r = 0.502 ; p < 0.0001 ) , lateral e ( r = 0.427 ; p < 0.0001 ) , septal e ( r = 0.389 ; p < 0.0001 ) , lateral e-to - a ratio ( r = 0.384 ; p < 0.0001 ) , e - to - e ratio ( r = 0.161 ; p = 0.020 ) , and e - to - a ratio ( r = 0.238 ; p = 0.001 ) .
linear regression analysis ( r / p ) between diastolic function parameters and different covariates bsa , body surface area ; dt , deceleration time .
thus , variables reaching statistical significance and sex , as dichotomic value , were inserted in a stepwise multivariate linear regression model to determine the independent predictors of left ventricular diastolic function parameters ( table 4 ) . in the study population ,
1-h postload glucose was the first predictor of left atrium volume , ivrt , and dt , explaining 25.6 ( p < 0.0001 ) , 18.2 ( p < 0.0001 ) , and 2.9% ( p = 0.031 ) of its variation , respectively .
age , sex , and dbp added another 5.6 ( p = 0.001 ) , 3.1 ( p = 0.013 ) , and 2.8% ( p = 0.032 ) to explaining left atrium volume , ivrt , and dt , respectively .
as for the tissue doppler parameters , we observed that 1-h postload glucose was retained as the first correlated of septal e , septal e-to - aratio , lateral e , and lateral e-to - a ratio , explaining 15.2 , 25.2 , 18.3 , and 14.8% of their variations , respectively .
age and sbp were retained as the independent predictors of e - to - e ratio , explaining 8.3% of its variation .
stepwise multiple regression analysis on diastolic function parameters as dependent variables in the whole study population bsa , body surface area .
this study , conducted in a cohort of never - treated and well - characterized hypertensive patients , showed that the worsening of glucose tolerance was associated with an impairment of left ventricular diastolic function .
the main finding of this study is that 1-h postload plasma glucose in ngt subjects is associated with left ventricular diastolic dysfunction .
this result persists after adjustment for all significant covariates reported in table 3 . of interest and clinically relevant , ngt 155 subjects had significantly worse diastolic function compared with ngt<155 subjects and similar compared with igt and t2d patients . to our knowledge , this is the first study that demonstrates this association , confirming the usefulness of early diagnosis in the stratification of overall cardiovascular risk ; in addition , it highlights the links between early alterations of glucose tolerance and early alterations in cardiac function .
there is a consistent body of evidences demonstrating the association between t2d or insulin resistance and diastolic dysfunction , which is recognized as the first stage of diabetic cardiomyopathy that may lead to chronic heart failure , independent of other cardiovascular heart diseases such as hypertension or atherosclerotic ischemic coronary disease ( 16,17 ) .
the development of diabetic cardiomyopathy is likely multifactorial involving several mechanisms including metabolic disturbances , endothelial dysfunction , coronary microvascular impairment , modification in the extracellular matrix , and sympathetic hyperactivity ( 16,17 ) .
all of these factors contribute to the increase of ventricular stiffness , promoting cardiac structure abnormalities such as left ventricular remodeling or hypertrophy .
cardiac fibrosis causes an imbalance between extracellular matrix deposition and degradation within the heart resulting in excessive fibroblast proliferation .
in addition , ventricular fibrosis causes progressive stiffening of the ventricular wall resulting in ventricular dysfunction , increase in end diastolic pressure , and atrial dilatation . some of these effects could be related to chronic hyperglycemia that induces , in diabetic patients , nonenzymatic glycation of circulating and cellular membrane proteins , leading to the formation of advanced glycation end products ( ages ) and , through protein kinase c activation , to reactive oxygen species production with increased oxidative stress ( 18 ) .
ages accumulation , in the myocardium and arterial wall , makes irreversible and stable links with collagen polymers , leading to fibrosis development with reduction of ventricular compliance and increase of lvm , as observed in animal models of igt ( 19 ) .
moreover , under chronic hyperglycemia condition , there is an increased turnover of free fatty acids , with a shift of myocardial metabolism toward the oxidation of the latter , with intracellular accumulation of intermediate products that lead , via increased oxidative stress , to deleterious effects ( 20 ) . in keeping with this ,
our results , obtained in ngt 155 subjects , clearly indicate that these modifications begin early , at a clinically silent phase , and support reconsideration of the notion that ngt subjects are a homogeneous group with a low cardiovascular risk profile .
the activation of both the renin - angiotensin - aldosterone system and sympathetic nervous system is another important mechanism potentially involved in activation of cardiac fibroblasts and collagen production ( 21 ) , leading to fibrosis and likely subsequent to the development of diastolic dysfunction .
finally , we should not ignore the role of coronary microcirculation abnormalities that may lead to myocardial cell injury and reactive fibrosis / hypertrophy . of interest
, the impairment of coronary microcirculation , occurring without obstructive atherosclerotic lesions on epicardial coronary arteries , induces a reduction of coronary flow reserve , as demonstrated in type 1 diabetic patients ( 22 ) and hypertensive subjects ( 23 ) .
the reduction of coronary flow reserve seems to be a direct consequence of elevated glycemia ( 24 ) .
( 25 ) showing that postprandial hyperglycemia induces myocardial perfusion defects in t2d patients , secondary to deterioration in microvascular function causing a decrease in myocardial blood flow .
the most clinically relevant information from this study , is that there is a statistically significant and direct correlation between 1-h postload plasma glucose and diastolic dysfunction in ngt hypertensive patients .
our data have allowed us to identify a new early predictor of subclinical organ damage and emphasize the importance of performing an ogtt in all subjects affected by essential hypertension , paying attention not only to 2- h but also to 1-h postload plasma glucose values , which are more strongly associated with diastolic dysfunction , in order to better stratify the global cardiovascular risk in hypertensive patients .
 objectiveto address whether glucose tolerance status , and in particular 1-h postload plasma glucose levels , may affect diastolic function in 161 never - treated hypertensive white subjects . 
 impaired left ventricular relaxation , an early sign of diastolic dysfunction , represents the first manifestation of myocardial involvement in diabetic cardiomyopathy . 
 a plasma glucose value 155 mg / dl for the 1-h postload plasma glucose during an oral glucose tolerance test ( ogtt ) is able to identify subjects with normal glucose tolerance ( ngt ) at high risk for type 2 diabetes and with subclinical organ damage.research design and methodssubjects underwent ogtt and standard echocardiography . 
 diastolic function was assessed by pulsed doppler transmitral flow velocity and tissue doppler imaging . 
 insulin sensitivity was assessed by matsuda index.resultsamong the participants , 120 had ngt , 26 had impaired glucose tolerance ( igt ) , and 15 had type 2 diabetes . according to the 1-h postload plasma glucose cutoff point of 155 mg / dl , we divided ngt subjects as follows : ngt < 155 mg / dl ( n = 90 ) and ngt 155 mg / dl ( n = 30 ) . those with ngt 155 mg / dl 
 had higher left atrium dimensions ( p < 0.0001 ) and isovolumetric relaxation time ( ivrt ) ( p = 0.037 ) than those with ngt < 155 mg / dl . by contrast , early / late transmitral flow velocity and all tissue doppler parameters were significantly lower in those with ngt 155 mg / dl than in those with ngt<155 mg / dl . at multiple regression analysis , 
 1-h glucose was the major determinant of left atrium area , ivrt , septal e , septal e-to - a ratio , lateral e , and lateral e-to - a ratio.conclusionsthe main finding of this study is that 1-h postload plasma glucose is associated with left ventricular diastolic dysfunction . 
 subjects with ngt 155 mg / dl had significantly worse diastolic function than those with ngt<155 mg / dl . 