Letter to the Editor
Insulin Resistance and Increased Intimal Medial Thickness in Glucose Tolerant Offspring of Type 2 Diabetic Subjects Carrying the D298D Genotype of Endothelial Nitric Oxide Synthase
To the Editor:
Endothelial nitric oxide synthase (eNOS) deficiency was found to affect insulin sensitivity and vascular homeostasis in animal models.1,2 Blood flow regulation by endothelial NO was suggested to modulate glucose uptake in the skeletal muscle.3 Therefore, it is possible that also in human subjects defects in eNOS activation or levels could affect insulin sensitivity.3 The D298D eNOS variant reduces eNOS protein half-life under stress conditions4 and it has been associated to increased cardiovascular disease (CVD) risk.5 The aim of present study was to assess whether among healthy offspring of patients with type 2 diabetes mellitus (T2DM) carriers of the D298D genotype are characterized by insulin resistance and early signs of atherosclerosis.
We recruited 142 young white offspring of T2DM with these inclusion criteria: age <45 years, 1 parent affected by T2DM, normal glucose tolerance (fasting plasma glucose <110 mg/dL; 2-hour plasma glucose <140 mg/dL), body mass index (BMI) less than 30 kg/m2, and absence of diseases or drugs able to modify glucose metabolism.
The phenotypization protocol, performed in all patients, was previously described and included blood sampling, blood pressure evaluation, oral glucose tolerance test and euglycemic hyperinsulinemic clamp technique.6 Moreover, high resolution B-mode ultrasound (HDI 3000 ultrasound system) was utilized to measure intima-media thickness (IMT). Briefly, the anterior, lateral, and posterolateral projections were used to image longitudinally the right and left common carotid arteries. At each projection, 3 determinations of IMT were made at 2 cm proximal to the bulb, at the site of greatest thickness. The values at each site were averaged, and the greatest value of the averaged IMT was used for each individual.7
E298D polymorphism was studied with polymerase chain reaction (PCR) amplification followed by enzymatic digestion with MBOI. All statistical analyses were performed using the SPSS software program (version 12.0 for Windows, SPSS, Inc., Chicago, Illinois).
Among subjects, 38.7% (n=51) were homozygous for the wild-type E298 eNOS, 46.2% (n=61) were heterozygous and 15.1% (n=20) homozygous for D298 eNOS allele, similarly to other studies in white population.6
No significant differences were observed among carriers of the 3 genotypes for age, sex, BMI and body composition. Carriers of D298D eNOS genotype have higher levels of triglycerides (P<0.05) compared with both E298D and E298E genotypes, and Non Esterified Fatty Acids (NEFA; P<0.05), compared with E298E subjects (Table 1). Fasting insulin (P<0.05) was lower in E298E genotype compared with D298D homozygous subjects. Consistently, homozygous subjects for the D298D allele showed a significantly lower insulin sensitivity, evaluated by glucose infusion rate (mg/kg FFM−1 min−1), than wild-type for E298 variant (P<0.05). IMT was higher in D298D genotype subjects compared with both D298E and E298E genotypes subjects (P<0.01). Insulin resistance, evaluated by clamp, and IMT remained statistically different after multivariate analysis (P<0.05 for both). Accordingly, a statistically significant inverse correlation between IMT and insulin sensitivity was observed (r=−0,37, P<0.01).
In the present study we show that the eNOS D298D genotype affects insulin sensitivity in glucose tolerant adult offspring of T2DM patients. In fact, carriers of the D298D eNOS genotype are characterized by insulin-resistance, hyperinsulinemia, hypertriglyceridemia, increased NEFA and IMT, compared with E298E and E298D carriers. Increased blood NEFA are a marker of adipose tissue insulin resistance and they are known to affect whole-body insulin sensitivity inhibiting insulin-stimulated glucose uptake and insulin regulated lipid metabolism.7 No differences in terms of blood pressure came out from genotyping distribution of subjects, probably due to the young mean age of the participants (35 years). Altogether these defects suggest that the D298D eNOS genotype is linked to impaired insulin sensitivity. The observation that only the D298D homozygous subjects show significant clinical alterations is in keeping with previous works observing increased cardiovascular risk and features of metabolic syndrome in T2DM subjects only in carriers of D298D genotype.5,7 Taken together our results suggest that the D298D eNOS genotype could represent one of the common variants contributing to the common soil linking insulin resistance and cardiovascular diseases in humans.
This work was supported in part by grants from PRIN-COFIN 2003 2003067733-006, RFS 2002 Farmacogenetica (to M.F.) and European Community “EuroDiabetesGene” Grants QLG1-CT-1999-00674 (to G.S.).
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