Roberto Trevisan

Differential effects of hyperinsulinemia and hyperaminoacidemia on leucine-carbon metabolism in vivo. Evidence for distinct mechanisms in regulation of net amino acid deposition.

The effects of physiologic hyperinsulinemia and hyperaminoacidemia, alone or in combination, on leucine kinetics in vivo were studied in postabsorptive healthy subjects with primed-constant infusions of L-[4,5-3H]leucine and [1-14C]alpha-ketoisocaproate (KIC) under euglycemic conditions. Hyperinsulinemia (approximately 100 microU/ml) decreased (P less than 0.05 vs. baseline) steady state Leucine + KIC rates of appearance (Ra) from proteolysis, KIC (approximately leucine-carbon) oxidation, and nonoxidized leucine-carbon flux (leucine----protein). Hyperaminoacidemia (plasma leucine, 210 mumol/liter), with either basal hormone replacement or combined to hyperinsulinemia, resulted in comparable increases in leucine + KIC Ra, KIC oxidation, and leucine----protein (P less than 0.05 vs. baseline). However, endogenous leucine + KIC Ra was suppressed only with the combined infusion. Therefore, on the basis of leucine kinetic data, hyperinsulinemia and hyperaminoacidemia stimulated net protein anabolism in vivo by different mechanisms. Hyperinsulinemia decreased proteolysis but did not stimulate leucine----protein. Hyperaminoacidemia per se stimulated leucine----protein but did not suppress endogenous proteolysis. When combined, they had a cumulative effect on net leucine deposition into body protein.

Glomerular Hyperfiltration and Renal Disease Progression in Type 2 Diabetes

OBJECTIVE To describe the prevalence and determinants of hyperfiltration (glomerular filtration rate [GFR] ≥120 mL/min/1.73 m2), GFR decline, and nephropathy onset or progression in type 2 diabetic patients with normo- or microalbuminuria. RESEARCH DESIGN AND METHODS We longitudinally studied 600 hypertensive type 2 diabetic patients with albuminuria <200 μg/min and who were retrieved from two randomized trials testing the renal effect of trandolapril and delapril. Target blood pressure (BP) was <120/80 mmHg, and HbA1c was <7%. GFR, albuminuria, and glucose disposal rate (GDR) were centrally measured by iohexol plasma clearance, nephelometry in three consecutive overnight urine collections, and hyperinsulinemic euglycemic clamp, respectively. RESULTS Over a median (range) follow-up of 4.0 (1.7–8.1) years, GFR declined by 3.37 (5.71–1.31) mL/min/1.73 m2 per year. GFR change was bimodal over time: a larger reduction at 6 months significantly predicted slower subsequent decline (coefficient: −0.0054; SE: 0.0009), particularly among hyperfiltering patients. A total of 90 subjects (15%) were hyperfiltering at inclusion, and 11 of 47 (23.4%) patients with persistent hyperfiltration progressed to micro- or macroalbuminuria versus 53 (10.6%) of the 502 who had their hyperfiltration ameliorated at 6 months or were nonhyperfiltering since inclusion (hazard ratio 2.16 [95% CI 1.13–4.14]). Amelioration of hyperfiltration was independent of baseline characteristics or ACE inhibition. It was significantly associated with improved BP and metabolic control, amelioration of GDR, and slower long-term GFR decline on follow-up. CONCLUSIONS Despite intensified treatment, patients with type 2 diabetes have a fast GFR decline. Hyperfiltration affects a subgroup of patients and may contribute to renal function loss and nephropathy onset or progression. Whether amelioration of hyperfiltration is renoprotective is worth investigating.

Insulin Resistance and Microalbuminuria: A Cross-Sectional, Case-Control Study of 158 Patients With Type 2 Diabetes and Different Degrees of Urinary Albumin Excretion

Microalbuminuria is a risk factor for renal and cardiovascular disease. A role for insulin resistance in the pathogenesis of microalbuminuria has been suggested but is still unproven. In this case-control, cross-sectional study, we compared glucose disposal rate (GDR), measured by hyperinsulinemic-euglycemic clamp, in 50 pairs of matched type 2 diabetic patients with micro- or normoalbuminuria (main study) and in 29 matched pairs of diabetic patients with macro- or microalbuminuria (substudy). In the main study, GDR was ∼25% lower in micro- than in normoalbuminuric patients (5.20 ± 1.91 vs. 6.86 ± 2.88 mg · kg−1 · min−1, P < 0.05) and was independently associated with microalbuminuria (P = 0.002), with each 1 mg · kg−1 · min−1 decrease predicting ∼40% increased prevalence (odds ratio 1.37 [95% CI 1.14–1.70]). Microalbuminuria was threefold more frequent in patients with GDR ≤7.50 ± 2.56 mg · kg−1 · min−1 than in those with higher GDR (60% vs. 20%, P < 0.005). In the substudy, GDR in macro- and microalbuminuric patients was comparable (5.52 ± 2.56 vs. 5.16 ± 1.61 mg · kg−1 · min−1) and independent of macroalbuminuria. GDR was significantly correlated with urinary albumin excretion rate in the main study (P = 0.004) but not in the substudy (P = 0.60). In type 2 diabetes, more severe insulin resistance is independently associated with microalbuminuria. Longitudinal studies are needed to clarify the role of insulin resistance in the pathogenesis of microalbuminuria and related complications.

Lipids and Renal Disease

Chronic renal disease is accompanied by characteristic abnormalities of lipid metabolism, which appear as a consequence of nephrotic syndrome or renal insufficiency and are reflected in an altered apolipoprotein profile as well as elevated plasma lipid levels. Experimental and clinical studies have suggested a correlation between the progression of renal disease and dyslipidemia. High cholesterol and triglyceride plasma levels have been demonstrated to be independent risk factors for progression of renal disease in humans. The underlying pathophysiologic mechanisms for the relationship between lipid levels and progression of renal disease are not yet fully understood, although there are data that oxidative stress and insulin resistance may mediate the lipid-induced renal damage. In the animal model, lipid-lowering agents seem to ameliorate glomerular damage, preventing glomerulosclerosis and interstitial fibrosis. Although evidence from clinical studies indicates that statin therapy is associated with significant benefit in individuals with established chronic renal failure, whether lipid reduction can slow the renal functional decline awaits a primary renal outcome lipid-lowering therapy study.

Clinical significance of nonalbuminuric renal impairment in type 2 diabetes.

Objective In type 2 diabetes, prevalence of nonalbuminuric renal impairment is increasing worldwide, though its clinical significance remains unclear. This large-cohort study aimed at evaluating the association of this phenotype with cardiovascular risk factors and other complications. Methods Type 2 diabetic patients from the Renal Insufficiency And Cardiovascular Events (RIACE) Italian Multicenter Study (n = 15 773), visiting consecutively 19 hospital-based Diabetes Clinics in years 2007–2008, were examined. Serum creatinine was assessed by the Jaffe method; albuminuria was measured by immunonephelometry or immunoturbidimetry. Results Of patients with renal impairment, as identified by an estimated glomerular filtration rate (eGFR) less than 60 ml/min per 1.73 m2, 56.6% were normoalbuminuric, 30.8% were microalbuminuric, and 12.6% were macroalbuminuric. Percentages were similar when GFR was estimated using the more accurate Chronic Kidney Disease Epidemiology Collaboration equation instead of the simplified Modification of Diet in Renal Disease formula, and were independent of age, thus indicating that the increasing prevalence of this phenotype does not reflects misclassification of elderly patients. Nonalbuminuric renal impairment was not associated with HbA1c and correlated less strongly with retinopathy and hypertension than albuminuria, either alone or associated with reduced eGFR. It was associated with a higher prevalence of cardiovascular disease (CVD) than albuminuria alone, but lower than albuminuric renal impairment. Female sex correlated with nonalbuminuric renal impairment and male sex with the albuminuric forms. Conclusions These data show that type 2 diabetic patients with nonalbuminuric renal impairment exhibit distinct clinical features, suggesting predominance of macroangiopathy as underlying renal pathology, and that this phenotype is associated with significant CVD burden.

In Type 1 diabetic patients with good glycaemic control, blood glucose variability is lower during continuous subcutaneous insulin infusion than during multiple daily injections with insulin glargine

Aims  The superiority of continuous subcutaneous insulin infusion (CSII) over multiple daily injections (MDI) with glargine is uncertain. In this randomized cross‐over study, we compared CSII and MDI with glargine in patients with Type 1 diabetes well controlled with CSII. The primary end‐point was glucose variability.

Role of Insulin and Atrial Natriuretic Peptide in Sodium Retention in Insulin-Treated IDDM Patients During Isotonic Volume Expansion

Because insulin shows an antinatriuretic effect in healthy humans, insulin therapy resulting in circulating hyperinsulinemia may lead to sodium retention and in turn to hypertension in individuals with insulin-dependent diabetes mellitus (IDDM). Moreover, it has been proved that atrial natriuretic peptide (ANP) plays a major role in modulating natriuresis in humans. This study investigated the relationship between insulin and ANP in modulating sodium metabolism in normotensive and hypertensive IDDM subjects compared with control groups of normotensive and hypertensive nondiabetic subjects. IDDM normotensive and hypertensive subjects had mean ± SE duration of IDDM of 7 ± 2 and 8 ± 2 yr, respectively, and had no clinical features of diabetic nephropathy. All subjects received a saline infusion (2 mmol · kg−1 · 90 min−1) during euglycemia. IDDM normotensive and hypertensive subjects received a subcutaneous insulin infusion (15 mU · kg−1 · h−1), resulting in twofold higher plasma free-insulin levels (16 ± 2 and 19 ± 3 μU/ml, respectively) than in nondiabetic normotensive and hypertensive subjects (7 ± 2 and 8 ± 2 μU/ml, respectively). During saline challenge, sodium excretion increased by 22 ± 4% in normotensive and 49 ± 9% in hypertensive nondiabetic subjects but by only 11 ± 0.4% in normotensive (P < 0.01) and 8 ± 2% in hypertensive (P < 0.01) IDDM subjects. The impaired natriuretic response to saline challenge was mainly due to greater rates of sodium reabsorption by kidney proximal tubules in IDDM than nondiabetic subjects. At baseline, plasma ANP concentrations were significantly higher in both IDDM groups than in control groups (normotensive IDDM and control subjects: 38 ± 4 and 19 ± 2 pg/ml, respectively, P < 0.01 ; hypertensive IDDM and control subjects: 45 ± 6 and 27 ± 4 pg/ml, respectively, P < 0.05). After saline challenge, ANP concentrations rose to 39 ± 4 pg/ml in normotensive and 49 ± 5 pg/ml in hypertensive control subjects, whereas no significant change above baseline value was seen in IDDM subjects. Both IDDM groups showed a 10–12% greater exchangeable Na+pool than control subjects regardless of the presence of hypertension. Subcutaneous insulin infusion, resulting in circulating plasma free-insulin levels in normotensive control subjects comparable to those in IDDM patients, inhibited natriuresis, increased proximal tubule sodium reabsorption at the level of the kidney, and inhibited an adequate ANP stimulation by saline challenge. We conclude that hyperinsulinemia leads to increased proximal tubule sodium reabsorption and impaired ANP response during saline administration. Both mechanisms account for sodium retention in normotensive and hypertensive IDDM patients. Sodium retention seems to characterize insulin-treated diabetes regardless of the presence of hypertension, suggesting that insulin-induced expansion of the Na+ pool alone cannot produce hypertension and requires the concomitant interaction of other pathogenetic factors.

Effect of Metformin on Insulin-Stimulated Glucose Turnover and Insulin Binding to Receptors in Type II Diabetes

Euglycemic insulin glucose-clamp and insulin-binding studies on erythrocytes and monocytes were performed in seven type II (non-insulin-dependent) diabetic subjects before and after 4 wk of metformin treatment (850 mg 3 times/day) and in five obese subjects with normal glucose tolerance. Glucose turnover was also measured at basal insulin concentrations and during hyperinsulinemic euglycemic clamps. During euglycemic insulin-glucose clamps, diabetic subjects showed glucose disposal rates of 3.44 ± 0.42 and 7.34 ± 0.34 mg · kg−1 · min−1 (means ± SD) before metformin at insulin infusion rates of 0.80 and 15.37 mU · kg−1 · min−1, respectively. With the same insulin infusion rates, glucose disposal was 4.94 ± 0.55 (P < .01) and 8.99 ± 0.66 (P < .01), respectively, after metformin treatment. Glucose disposal rates in normal obese subjects were 5.76 ± 0.63 (P < .01) and 10.92 ± 1.11 (P < .01) at 0.80 and 15.37 mU · kg−1 · min−1, respectively. Insulin maximum binding to erythrocytes in diabetics was 9.6 ± 4.2 and 5.8 ± 2.6 × 109 cells (means ± SD) before and after metformin treatment, respectively (NS). Insulin maximum binding to monocytes in diabetics was 6.2 ± 2.3 × 107 cells before and 5.0 ± 1.6% after metformin. Hepatic glucose production was higher in the diabetic patients at basal insulin levels, but not at higher insulin concentrations, and was not significantly changed by drug treatment. Basal glucose and insulin concentrations decreased with metformin. Thus, metformin treatment improved glucose disposal rate without significant effect on insulin-binding capacity on circulating cells. Basal hepatic glucose output was slightly lower after metformin treatment in view of lower (9 vs. 15 μU/ml) insulin levels, potentially indicating increased sensitivity of the liver to insulin.

The GFR and GFR decline cannot be accurately estimated in type 2 diabetics

There are no adequate studies that have formally tested the performance of different estimating formulas in patients with type 2 diabetes both with and without overt nephropathy. Here we evaluated the agreement between baseline GFRs, GFR changes at month 6, and long-term GFR decline measured by iohexol plasma clearance or estimated by 15 creatinine-based formulas in 600 type 2 diabetics followed for a median of 4.0 years. Ninety patients were hyperfiltering. The number of those identified by estimation formulas ranged from 0 to 24:58 were not identified by any formula. Baseline GFR was significantly underestimated and a 6-month GFR reduction was missed in hyperfiltering patients. Long-term GFR decline was also underestimated by all formulas in the whole study group and in hyper-, normo-, and hypofiltering patients considered separately. Five formulas generated positive slopes in hyperfiltering patients. Baseline concordance correlation coefficients and total deviation indexes ranged from 32.1% to 92.6% and from 0.21 to 0.53, respectively. Concordance correlation coefficients between estimated and measured long-term GFR decline ranged from -0.21 to 0.35. The agreement between estimated and measured values was also poor within each subgroup considered separately. Thus, our study questions the use of any estimation formula to identify hyperfiltering patients and monitor renal disease progression and response to treatment in type 2 diabetics without overt nephropathy.

Na+/H+ Antiport Activity and Cell Growth in Cultured Skin Fibroblasts of IDDM Patients With Nephropathy

IDDM patients with incipient and overt nephropathy have been found to exhibit an overactivity of RBC sodium-lithium countertransport. To explore the physiological relevance of this finding, we measured the activity of Na+/H+ antiport in serially passaged cultured skin fibroblasts from IDDM patients with and without nephropathy and from normal, nondiabetic control subjects. Na+/H+ antiport activity (measured as the rate of amiloride-sensitive Na+ influx at pH1 = 6.4, extracellular pH = 8.0, and [Na+] = 1 mM) was elevated significantly in IDDM patients with nephropathy compared with IDDM patients without nephropathy and nondiabetic control subjects (13.35 ± 3.8 vs. 8.54 ± 2.0 vs. 7.33 ± 2.3 nmol Na+ · mg protein−1 · min−1; P < 0.006 and P < 0.001, respectively). A kinetic analysis of Na+/H+ antiport activity showed that the raised activity in IDDM patients with nephropathy was caused by an increased Vmax for extracellular Na+. Km values were similar in the three groups. pH-stimulated amiloride-sensitive Na+ influx also was higher under baseline conditions and after serum stimulation in cells from IDDM patients with nephropathy. pHI values were significantly higher, both during active proliferation and after 10-min exposure to serum, in cells from IDDM patients with nephropathy, compared with IDDM patients without nephropathy and nondiabetic control subjects. Serum-stimulated incorporation of [3H]thymidine into DNA was greater in IDDM patients with nephropathy than in the other two groups (35.7 ± 18.9- vs. 17.4 ± 7.5- vs. 11.9 ± 8.7-fold stimulation above baseline; P < 0.01 for both). These data from cultured skin fibroblasts suggest that intrinsic abnormalities in cell function, independent of the metabolic disturbances of diabetes, are characteristic of IDDM patients who develop nephropathy.