Accumulation of advanced glycation end products is associated with macrovascular events and glycaemic control with microvascular complications in Type 2 diabetes mellitus.

AIM: The United Kingdom Prospective Diabetes Study (UKPDS) study showed that glycaemic control (HbA1c ) can predict vascular complications in Type 2 diabetes mellitus. The Diabetes Control and Complications Trial (DCCT) study showed that accumulation of advanced glycation end products (AGEs) from skin biopsies predicts vascular complications in Type 1 diabetes. Previously, we showed that tissue AGEs can be measured non-invasively using skin autofluorescence (SAF). The aim of this study was to compare the predictive value of HbA1c and SAF for new macrovascular events and microvascular complications in people with Type 2 diabetes. METHODS: A prospective cohort study of 563 participants, median age 64 years [interquartile range (IQR) 57-72], diabetes duration of 13 years, from five Dutch hospitals was performed. RESULTS: After a median follow-up of 5.1 (IQR 4.3-5.9) years, 79 (15%) participants had died and 49 (9%) were lost to follow-up. Some 133 (26%) developed a microvascular complication and 189 (37%) a macrovascular event. Tertiles of HbA1c were significantly associated with development of microvascular complications (log rank P = 0.022), but not with macrovascular events. Tertiles of SAF were significantly associated with macrovascular events (log rank P = 0.003). Cox regression analysis showed SAF was associated with macrovascular events: crude hazard ratio (HR) 1.53 (P < 0.001) per unit increase, HR 1.28 (P = 0.03) after correction for UKPDS score. HbA1c was predictive for microvascular complications: crude HR 1.20 (P = 0.004), HR 1.20 (P = 0.004) after correction for UKPDS score. CONCLUSION: This study shows that tissue accumulation of AGEs, assessed by SAF, is associated with development of macrovascular events in people with Type 2 diabetes, whereas HbA1c is associated with the development of microvascular complications.

Skin autofluorescence and risk of micro- and macrovascular complications in patients with Type 2 diabetes mellitus-a multi-centre study.

AIMS: Skin autofluorescence is a non-invasive marker of advanced glycation end product accumulation. In a previous study, skin autofluorescence correlated with and predicted micro- and macrovascular complications in Type 2 diabetes in a primary care setting. The present cross-sectional study aims to confirm the association between skin autofluorescence and diabetic complications in patients with Type 2 diabetes in a multi-centre secondary care setting. METHODS: We analysed 563 subjects with Type 2 diabetes mellitus from five Dutch hospitals. RESULTS: Median age was 64 years, median duration of diabetes 13 years and median HbA(1c) 58 mmol/mol (7.5%). Sixty-one per cent of patients had microvascular complications (38% nephropathy, 36% retinopathy, 35% neuropathy) and 42% had macrovascular complications. Median UK Prospective Diabetes Study 10-year risk for coronary events was 19%. Median skin autofluorescence was elevated compared with age-matched healthy control subjects: 2.77 (interquartile range 2.39-3.28) vs. 2.46 (2.08-2.84) arbitrary units. Skin autofluorescence was particularly increased in patients with complications: no complications, median 2.56 (2.26-2.90); microvascular complications, 2.79 (2.38-3.29); macrovascular complications, 2.85 (2.41-3.41); both micro- and macrovascular complications, 2.96 (2.56-3.60) arbitrary units, P < 0.001. Logistic regression analysis showed that age, duration of diabetes, renal function, gender, atrial fibrillation and skin autofluorescence were independently associated with macrovascular complications. Multiple regression analysis identified age, smoking, renal function, macrovascular complications and the number of microvascular complications as the determinants of skin autofluorescence. CONCLUSIONS: This study confirms that skin autofluorescence is increased in patients with Type 2 diabetes in a secondary care setting. Skin autofluorescence was associated with macrovascular complications in patients with diabetes and this association was independent of classical risk factors.

Human plasma phospholipid transfer protein activity is decreased by acute hyperglycaemia: studies without and with hyperinsulinaemia in Type 1 diabetes mellitus.

AIMS: Little is known about the regulation of phospholipid transfer protein (PLTP), that plays a key role in lipoprotein metabolism. PLTP secretion may be up-regulated by glucose in vitro, whereas plasma PLTP activity is decreased by exogenous hyperinsulinaemia and glucose-induced hyperinsulinaemia in vivo. In the present study, we evaluated the separate effects of hyperglycaemia and hyperinsulinaemia in C-peptide-negative Type 1 diabetic patients. METHODS: The protocol was carried out in 16 patients (eight females). In each individual, plasma PLTP mass and activity (measured by enzyme-linked immuno-sorbent assay and liposome-high density lipoprotein system, respectively) as well as plasma cholesteryl ester transfer protein (CETP) activity, lipids and apolipoprotein levels were determined at the end of four different glucose clamps, each lasting 210 min: standard insulin (30 mU/kg/h) and standard glucose (glucose 5.0 mmol/l) (SI-SG), standard insulin and high glucose (glucose 12 mmol/l) (SI-HG), high insulin (150 mU/kg/h) and standard glucose (HI-SG), and high insulin and high glucose (HI-HG). RESULTS: Plasma lipids and (apo)lipoproteins, measured at the end of the SI-HG, HI-SG and HI-HG clamps, were not significantly different compared with the levels obtained at the end of the SI-SG clamp. Median plasma PLTP mass and activity at the end of the SI-SG clamp were 12.8 mg/l and 13.2 micromol/ml/h, respectively. Median plasma PLTP mass decreased by 9.1% at the end of the HI-HG clamp (P < 0.01), whereas the changes at the end of the SI-HG and HI-SG clamps were not significant. Median plasma PLTP activity decreased by 5.7, 4.6 and 8.6% at the end of the SI-HG, HI-SG and HI-HG clamps, respectively (all P < 0.05). Median plasma CETP activity was 177 nmol/ml/h at the end of the SI-SG clamp, and decreased by 4.9% (P < 0.05) and by 8.3% (P < 0.05) at the end of the HI-SG and the HI-HG clamps, respectively. Plasma CETP activity did not change significantly at the end of the SI-HG clamp. CONCLUSIONS: The present study demonstrates that plasma PLTP activity is independently decreased by acute hyperglycaemia and hyperinsulinaemia in humans in vivo. These data do not support a direct role of short-term hyperglycaemia in up-regulating plasma PLTP levels.

Simple non-invasive assessment of advanced glycation endproduct accumulation.

AIMS/HYPOTHESIS: The accumulation of AGE is thought to play a role in the pathogenesis of chronic complications of diabetes mellitus and renal failure. All current measurements of AGE accumulation require invasive sampling. We exploited the fact that several AGE exhibit autofluorescence to develop a non-invasive tool for measuring skin AGE accumulation, the Autofluorescence Reader (AFR). We validated its use by comparing the values obtained using the AFR with the AGE content measured in extracts from skin biopsies of diabetic and control subjects. METHODS: Using the AFR with an excitation light source of 300-420 nm, fluorescence of the skin was measured at the arm and lower leg in 46 patients with diabetes (Type 1 and 2) and in 46 age- and sex-matched control subjects, the majority of whom were Caucasian. Autofluorescence was defined as the average fluorescence per nm over the entire emission spectrum (420-600 nm) as ratio of the average fluorescence per nm over the 300-420-nm range. Skin biopsies were obtained from the same site of the arm, and analysed for collagen-linked fluorescence (CLF) and specific AGE: pentosidine, N(epsilon)-(carboxymethyl)lysine (CML) and N(epsilon)-(carboxyethyl)lysine (CEL). RESULTS: Autofluorescence correlated with CLF, pentosidine, CML, and CEL ( r=0.47-0.62, p

No effects of acute hyperglycaemia and hyperinsulinaemia on skin microcirculation and endothelial markers in Type II diabetes mellitus.

BACKGROUND: Increased microvascular permeability is a hallmark of microangiopathy in Type I diabetes mellitus and is associated with endothelial dysfunction and haemodynamic alterations. Type II diabetes mellitus is characterized by insulin resistance and hyperinsulinaemia. The purpose of this study was to determine whether acute hyperinsulinaemia, under both normoglycaemic and hyperglycaemic conditions, increases skin capillary permeability through its effect on skin haemodynamics, capillary recruitment or circulating markers of endothelial dysfunction in Type II diabetes. METHODS: Nine Type II diabetic patients without microalbuminuria, (pre-) proliferative retinopathy or clinical neuropathy underwent three glucose clamps of 210 min., in random order, on separate days. A "standard" clamp (insulin-infusion rate 30 mU kg(-1) h(-1), glucose-target 5.0 mmol/L) was compared with a hyperinsulinaemic (insulin-infusion rate 150 mU kg(-1) h(-1), glucose-target 5.0 mmol/L) and a hyperinsulinaemic, hyperglycaemic (insulin-infusion rate 150 mU kg(-1) h(-1), glucose-target 12.0 mmol/L) clamp. Skin capillary permeability and density were measured using large-window sodium fluorescein videodensitometry, and skin blood flow by laser Doppler flowmetry. Endothelial dysfunction was estimated from increases in soluble intercellular adhesion molecule-1 (sICAM-1) and von Willebrand factor antigen (vWF). RESULTS: No differences were found in skin capillary permeability, skin haemodynamics and capillary density at the end of the three glucose clamp periods. sICAM-1 and vWF did not increase as compared to the standard glucose clamp. sICAM-1 (r=-0.76, p<0.05) and vWF (r=-0.71, p<0.05) correlated negatively with insulin sensitivity, but not with skin microcirculatory parameters. CONCLUSIONS: Acute hyperinsulinaemia, both with and without concomitant hyperglycaemia, does not increase skin microvascular permeability, haemodynamics or parameters of endothelial dysfunction in Type II diabetic patients. Furthermore, these data suggest that the coexistence of hyperinsulinaemia and endothelial dysfunction in Type II diabetes does not indicate a causal relationship, but may rather indicate decreased insulin sensitivity as a common underlying cause.

Sympathetic mediated vasomotion and skin capillary permeability in diabetic patients with peripheral neuropathy.

AIMS/HYPOTHESIS: A loss of sympathetic function could lead to changes in capillary fluid filtration in diabetic patients. We investigated whether a decreased sympathetically mediated vasomotion in the skin in diabetic patients with peripheral neuropathy is associated with an abnormal capillary leakage. METHODS: Three matched groups were studied: 18 diabetic patients with documented peripheral neuropathy (DN), 18 diabetic patients without peripheral neuropathy (D), and 18 healthy control subjects (C). Sensory and motor nerve function of the distal extremities were assessed by standard neurography, and expressed in a sensory-motor nerve function score. Sympathetic vasomotion of the skin microcirculation was assessed by determining the power of blood flow variability in the low-frequency (0.02-0.14 Hz) band by spectral analysis of laser Doppler flowmetry at the median ankle. Skin capillary leakage was evaluated by sodium fluorescein videodensitometry at the same site of the foot. RESULTS: Sympathetically mediated vasomotion of the foot skin microcirculation was lower in diabetic patients with documented peripheral neuropathy compared with diabetic patients without peripheral neuropathy and control subjects (p<0.001). Capillary sodium fluorescein leakage was larger in 18 diabetic patients with documented peripheral neuropathy than in diabetic patients without peripheral neuropathy (p<0.02) and C (p<0.005). Multiple regression analysis disclosed that a reduced sympathetically mediated vasomotion, together with a lower sensory-motor nerve function score, independently contributed to the variance in sodium fluorescein leakage, for 30% (p<0.001) and 17% (p<0.01), respectively. CONCLUSIONS: A loss of sympathetic tone, apart from sensory-motor nerve dysfunction, seems to be a major determinant of an increased capillary permeability in diabetic patients with neuropathy.

Growth hormone replacement does not elevate albuminuria in GH-deficient adults.

Minor elevations in urinary albumin excretion rate (Ualb.V) are likely to be associated with renal function loss and increased cardiovascular risk. Since urinary albumin excretion is affected by the growth hormone (GH)-insulin-like growth factor-1 (IGF-1) axis, we evaluated the effect of 6 months GH replacement (1U/day, n=8 and 2U/day, n=16) on urinary protein handling in 24 non-diabetic, normotensive GH-deficient adults (12 men and 12 women), of whom 8 patients received placebo during 6 months, followed by active GH treatment. Plasma IGF-1 increased from 11.4 (8.1-15.8) nmol/L (median, interquartile range) to 35.4 (22.3-44.1) nmol/L (p<0.001 versus change after placebo; median Z-score reached +0.2) after 6 months GH. and remained unaltered after placebo. Overnight Ualb.V was 4.0 (3.1-4.9) microg/min before and 4.6 (2.4-8.2) microg/min after GH (p>0.10). Likewise, urinary IgG excretion rate did not significantly change after GH (0.71 (0.52-1.20) microg/min before GH versus 0.80 (0.51-1.56) microg/min after GH; p>0.10). No significant changes were observed in creatinine clearance (p>0.10) and mean arterial pressure (p>0.10). No changes in any of these parameters were observed after placebo. Individual changes in Ualb.V were positively correlated with changes in plasma IGF-1 (r(s)=0.41, p=0.05) and with changes in mean arterial pressure (r(s)=0.49, p<0.02). The present study shows that 6 months GH replacement, increasing plasma IGF-1 within the physiological range, does not result in a clinically relevant increase in urinary protein excretion in GH-deficient adults. The correlation between changes in plasma IGF-1 and in albuminuria supports the rationale to avoid supraphysiological IGF-1 levels.

Reduced capillary permeability and capillary density in the skin of GH-deficient adults: improvement after 12 months GH replacement.

OBJECTIVE: Several lines of evidence suggest that the GH-IGF-1 axis affects capillary permeability and angiogenesis. We evaluated skin capillary permeability and capillary density in GH-deficient adults, before and after GH replacement therapy. PATIENTS: Seven normotensive, nondiabetic GH-deficient adults (two women) were matched with 14 control subjects. MEASUREMENTS: Large-window videodensitometry with sodium fluorescein was performed in all subjects. Capillary permeability was expressed as the average relative light intensity over the first 7 min after the appearance of fluorescein in the skin capillaries; Iav(7). Skin capillary density was determined by counting the visualized capillaries and was expressed as n/mm2. The GH-deficient patients were restudied after 12 months of GH replacement therapy (2 U/day). RESULTS: Both capillary permeability and capillary density were lower in untreated GH-deficient patients than in control subjects (median, interquartile range): Iav(7) in GH-deficient patients 47.1 (45.1-52.2)% vs. 57.5 (50.5-64.8)% in controls, P < 0.05; capillary density in GH-deficient patients 18 (12-24)/mm2 vs. 32 (26-36)/mm2 in controls, P < 0.05. GH treatment normalized plasma IGF-1 from 4.3 (1.0-13.4) to 22.2 (19.8-48.2) nmol/l (P < 0.05). Furthermore, both capillary permeability [Iav(7) 53.1 (48.8-58.4)%, P < 0.05] and capillary density [26 (17-34)/mm2, P < 0.05] increased to a level that was not different from that in control subjects. CONCLUSIONS: The present study demonstrates that the growth hormone deficiency syndrome is associated with microvascular alterations, which are responsive to growth hormone replacement therapy.

Acute hyperglycemia and hyperinsulinemia enhance vasodilatation in Type 1 diabetes mellitus without increasing capillary permeability and inducing endothelial dysfunction.

Uncomplicated Type 1 (insulin-dependent) diabetes mellitus is characterized by generalized vasodilatation. Its possible correlates, increased microvascular permeability and endothelial dysfunction, have been associated with long-term complications. The objective was to study the effects of acute hyperglycemia and hyperinsulinemia, both separately and in combination, on skin microvascular flow, capillary permeability, capillary recruitment, and endothelial dysfunction in Type 1 diabetes mellitus. Sixteen Type 1 diabetic patients (all normoalbuminuric, no (pre-)proliferative retinopathy) underwent a euglycemic (glucose target 5.0 mmol/L, insulin infused at 30 mU x kg(-1) x h(-1)), a hyperglycemic (glucose target 12.0 mmol/L, insulin 30 mU x kg(-1) x h(-1)), a hyperinsulinemic (glucose target 5.0 mmol/L, insulin 150 mU x kg(-1) x h(-1)), and a hyperglycemic-hyperinsulinemic (glucose target 12.0 mmol/L, insulin 150 mU x kg(-1) x h(-1)) clamp on separate days, in random order. Skin microvascular flow was measured by laser Doppler flowmetry. Capillary permeability and density were determined by large-window sodium-fluorescein videodensitometry. Increases in serum soluble intercellular adhesion molecule-1 (sICAM-1) and plasma von Willebrand factor antigen (vWF-Ag) were considered to represent abnormal endothelial function. Hyperglycemia (P < 0.01) and hyperinsulinemia (P < 0.05) as well as both interventions combined (P < 0.001) induced an increase in laser Doppler flow, without capillary recruitment. Transcapillary leakage of sodium-fluorescein and sICAM-1 and vWF-Ag levels were unaffected by hyperglycemia or hyperinsulinemia. Microvascular permeability appears to be determined primarily by properties of the capillary wall and not by acute changes in local hemodynamics. The acute hyperglycemia- and hyperinsulinemia-induced vasodilatation is not accompanied by changes in microvascular permeability or endothelial markers.