Maria Sambataro

Patterns of renal injury in NIDDM patients with microalbuminuria.

Summary Microalbuminuria predicts overt nephropathy in non-insulin-dependent diabetic (NIDDM) patients; however, the structural basis for this functional abnormality is unknown. In this study we evaluated renal structure and function in a cohort of 34 unselected microalbuminuric NIDDM patients (26 male/8 female, age: 58 ± 7 years, known diabetes duration: 11 ± 6 years, HbA1 c: 8.5 ± 1.6 %). Systemic hypertension was present in all but 3. Glomerular filtration rate (GFR) was 101 ± 27 ml · min–1· 1.73 m–2 and albumin excretion rate (AER) 44 (20–199) μg/min. Light microscopic slides were categorized as: C I) normal or near normal renal structure; C II) changes “typical” of diabetic nephropathology in insulin-dependent diabetes (IDDM) (glomerular, tubulo-interstitial and arteriolar changes occurring in parallel); C III) “atypical” patterns of injury, with absent or only mild diabetic glomerular changes associated with disproportionately severe renal structural changes including: important tubulo-interstitial with or without arteriolar hyalinosis with or without global glomerular sclerosis. Ten patients (29.4 %) were classified as C I, 10 as C II (29.4 %) and 14 as C III (41.2 %); none of these patients had any definable non-diabetic renal disease. GFR, AER and blood pressure were similar in the three groups, while HbA1 c was higher in C II and C III than in C I patients. Diabetic retinopathy was present in all C II patients (background in 50 % and proliferative in 50 %). None of the patients in C I and C III had proliferative retinopathy, while background retinopathy was observed in 50 % of C I and 57 % of C III patients. In summary, microalbuminuric NIDDM patients are structurally heterogeneous with less than one third having “typical” diabetic nephropathology. The presence of both “typical” and “atypical” patterns of renal pathology was associated with worse metabolic control, suggesting that hyperglycaemia may cause different patterns of renal injury in older NIDDM compared to younger IDDM patients. [Diabetologia (1996) 39: 1569–1576]

Inherited Apolipoprotein A-V Deficiency in Severe Hypertriglyceridemia

Objective— Mutations in LPL or APOC2 genes are recognized causes of inherited forms of severe hypertriglyceridemia. However, some hypertrigliceridemic patients do not have mutations in either of these genes. Because inactivation or hyperexpression of APOA5 gene, encoding apolipoprotein A-V (apoA-V), causes a marked increase or decrease of plasma triglycerides in mice, and because some common polymorphisms of this gene affect plasma triglycerides in humans, we have hypothesized that loss of function mutations in APOA5 gene might cause hypertriglyceridemia. Methods and Results— We sequenced APOA5 gene in 10 hypertriglyceridemic patients in whom mutations in LPL and APOC2 genes had been excluded. One of them was found to be homozygous for a mutation in APOA5 gene (c.433 C>T, Q145X), predicted to generate a truncated apoA-V devoid of key functional domains. The plasma of this patient was found to activate LPL in vitro less efficiently than control plasma, thus suggesting that apoA-V might be an activator of LPL. Ten carriers of Q145X mutation were found in the patient’s family; 5 of them had mild hypertriglyceridemia. Conclusions— As predicted from animal studies, apoA-V deficiency is associated with severe hypertriglyceridemia in humans. This observation suggests that apoA-V regulates the secretion and/or catabolism of triglyceride-rich lipoproteins.

Insulin sensitivity and glucose effectiveness: minimal model analysis of regular and insulin-modified FSIGT

The minimal model is widely used to evaluate insulin action on glucose disappearance from frequently sampled intravenous glucose tolerance tests (FSIGT). The common protocols are a regular (rFSIGT, single injection of 0.3 g/kg of glucose) and an insulin-modified test (mFSIGT, with an additional insulin administration at 20 min). This study compared the insulin sensitivity index (SI) and glucose effectiveness (SG) obtained in the same individual (16 normal subjects) with the two tests. SI was 7.11 ± 0.80 10-4 ⋅ min-1 ⋅ μU-1 ⋅ ml in rFSIGT and 6.96 ± 0.83 in mFSIGT ( P = 0.656), regression r = 0.92, P < 0.0001; SG was 0.0260 ± 0.0028 min-1 and 0.0357 ± 0.0052, respectively, statistically different ( P = 0.013) but still with a good regression ( r = 0.66, P = 0.0051). SG and insulin amount during the early period correlated ( r = 0.6, P = 0.015 in rFSIGT and r = 0.76, P = 0.0006 in mFSIGT). In summary, both FSIGTs with minimal model analysis provide the same SI, which is a very robust index. SG was different by 28% due probably to the relationship between SG and the amount of circulating insulin. In studies comparing groups, the simpler rFSIGT can still be used with the advantage of accounting for endogenous insulin, thus offering the possibility of direct inferences on the β-cell activity.The minimal model is widely used to evaluate insulin action on glucose disappearance from frequently sampled intravenous glucose tolerance tests (FSIGT). The common protocols are a regular (rFSIGT, single injection of 0.3 g/kg of glucose) and an insulin-modified test (mFSIGT, with an additional insulin administration at 20 min). This study compared the insulin sensitivity index (SI) and glucose effectiveness (SG) obtained in the same individual (16 normal subjects) with the two tests. SI was 7.11 +/- 0.80 10(-4).min-1.microU-1.ml in rFSIGT and 6.96 +/- 0.83 in mFSIGT (P = 0.656), regression r = 0.92, P < 0.0001; SG was 0.0260 +/- 0.0028 min-1 and 0.0357 +/- 0.0052, respectively, statistically different (P = 0.013) but still with a good regression (r = 0.66, P = 0.0051). SG and insulin amount during the early period correlated (r = 0.6, P = 0.015 in rFSIGT and r = 0.76, P = 0.0006 in mFSIGT). In summary, both FSIGTs with minimal model analysis provide the same SI, which is a very robust index. SG was different by 28% due probably to the relationship between SG and the amount of circulating insulin. In studies comparing groups, the simpler rFSIGT can still be used with the advantage of accounting for endogenous insulin, thus offering the possibility of direct inferences on the beta-cell activity.

Peripheral, rather than hepatic, insulin resistance and atherogenic lipoprotein phenotype predict cardiovascular complications in NIDDM

Abstract Microalbuminuria, hypertension and hyper‐insulinaemia are three independent risk factors for cardiac disease in non insulin‐dependent diabetes (NIDDM). However, it is unknown to what extent hyperinsulinaemia reflects resistance to insulin action at hepatic, extrahepatic or at both sites. A cross‐sectional study from our Department showed that peripheral insulin resistance, hypertension, microalbuminuria and lipid abnormalities are associated in NIDDM. Non diabetic individuals with the so‐called ‘atherogenic lipoprotein phenotype’, characterized by small dense low density lipoproteins (LDL subclass pattern B) have up to 3‐fold higher risk of myocardial infarction. The aim of the present study was to investigate whether impaired peripheral insulin sensitivity, during euglycaemic‐hyperinsulinaemic clamp, as well as abnormalities in lipid concentrations and LDL size, predict abnormalities in albumin excretion rate, blood pressure and cardiac function in 73 consecutive normotensive (<85 mmHg diastolic level) and nor‐moalbuminuric (<15 μg min‐1 daily albumin excretion rate) NIDDM patients. These patients showed a bimodal distribution of whole body glucose utilization rate, a parameter of peripheral insulin sensitivity. The cut‐off point between the two modes of distribution was located close to the mean value minus one standard deviation in a population of 24 control subjects. Therefore, this latter value was used to identify two subgroups inside the overall population of NIDDM patients, i.e. 28 patients (group 1), with whole body glucose utilization rate, above, and 45 patients (group 2), below, the mean value minus 1 SD in the 24 controls. Both groups 1 and 2 had impaired insulin sensitivity at hepatic site, as assessed by the degree of inhibition of hepatic glucose output during insulin administration (controls vs. group 1 vs. group 2: 925±235 vs. 952±166 vs. 506±121; P>0·001). The two groups displayed similar patterns of age, gender, body weight, diabetes duration, HbA1c and cardiac ischaemic events. During 6‐year follow‐up the rate of occurrence of microalbuminuria (>20 μg min‐1) (7% vs. 15%, P<0·01) and diastolic hypertension (>90 mmHg) (14 vs. 30%, P<0·05) was significantly higher in group 2 than in group 1 NIDDM patients. Events of cardiac ischaemic disease were more frequently found among group 2 rather than group 1 patients, during the follow‐up (angina pectoris: 3/28 subjects in group 1 vs. 9/45 subjects in group 2, P<0·05; positive resting ECG: 3/28 subjects in group 1 vs. 9/45 subjects in group 2; positive exercise ECG 4/28 in group 1 vs. 11/45 in group 2). Group 2 patients were also characterized by higher triglyceride and lower high density lipoprotein cholesterol levels and by LDL subclass pattern B. Peripheral, rather than hepatic, resistance and atherogenic lipoprotein phenotype are the clinical hallmark of NIDDM patients who are prone to develop microalbuminuria, hypertension and cardiac ischaemic disease.

Role of Atrial Natriuretic Peptide in the Pathogenesis of Sodium Retention in IDDM With and Without Glomerular Hyperfiltration

The pathogenetic determinants of sodium retention in IDDM are not fully understood. The aim of this study was to elucidate the action of ANP in 11 IDDM patients with high GFR (≥ 135 ml · min−1 · 1.73 m−2), referred to here as HF patients; in 10 IDDM patients with normal GFR (>90 and <135 ml·min−1 · 1.73 m−2), referred to here as NF patients; and 12 control subjects, here called C subjects, at baseline and during saline infusion administered on the basis of either body weight (2 mmol × kg−1 · 60 min−1; Saline 1) or of ECV (12 mM.ECVL−1 · 90 min−1; Saline 2) during euglycemic insulin-glucose clamp. C subjects and both HF and NF IDDM patients received a second Saline 1 infusion accompanied by ANP infusion (0.02 μg · kg−1 · min−1) at euglycemic levels. HF and NF patients were studied again after 3 mo of treatment with (10 mg/day). Quinapril (CI 906, Malesci, Florence, Italy), an ACE inhibitor without sulfhydryl group. At baseline, both HF and NF IDDM patients had higher plasma ANP concentrations than C subjects (HF, 36 ± 4, P < 0.01 and NF, 34 ± 3, P < 0.01 vs. C, 19 ± 3 pg/ml). Plasma ANP and natriuretic response to isotonic volume expansion was impaired both in HF (44 ± 8 pg/ml, NS vs. base) and NF (40 ± 7 pg/ml, NS vs. base) compared with C (41 ± 4 pg/ml, P < 0.01 vs. base) during Saline 1. On the contrary, plasma ANP response to Saline 2 was similar in HF and NF patients and C subjects, but IDDM patients had still lower urinary sodium excretion rates. The simultaneous administration of ANP and Saline 1 resulted in comparable plasma ANP plateaus in C subjects and HF and NF patients. However, urinary sodium excretion rate was significantly lower in HF and NF patients than in C subjects: HF, 267 ± 64, P < 0.01 and NF, 281 ± 42, P < 0.01 vs. C, 424 ± 39 μmol · min−1 · 1.73 m−2. During simultaneous administration of ANP and Saline 1, GFR and FF increased in C subjects, but not in HF and NF patients. HF and NF patients had higher urinary vasodilatory prostanoid excretion rates than C subjects at baseline. Saline infusion did not change urinary excretion rate of prostanoids either in C subjects or IDDM patients (both NF and HF). Quinapril significantly decreased baseline plasma ANP concentrations in both HF and NF, near normalized ANP response to Saline 1, and increased sodium excretion rate both during Saline 1 and 2, without any change in GFR and urinary prostanoid excretion rates. The results of this study suggest that in IDDM patients with high or normal GFR, 1) natriuresis is impaired during saline infusion, even when ANP levels are normal, suggesting the presence of resistance to ANP action, 2) ANP release is normal if volume expansion is proportionate to ECV magnitude, 3) urinary excretion rate of vasodilatory prostanoids is high, and 4) ACE inhibitors ameliorate natriuretic and ANP response to saline infusion. These findings suggest that a resistance to the natriuretic action of ANP, and possibly of vasodilatory prostanoids, contribute to sodium retention in IDDM, irrespective of glomerular hyperfiltration.

Lipoprotein abnormalities in non-insulin-dependent diabetic patients with impaired extrahepatic insulin sensitivity, hypertension, and microalbuminuria.

We investigated whether specific lipoprotein abnormalities are present in non-insulin-dependent diabetes mellitus (NIDDM) patients with hypertension and/or microalbuminuria. Fifteen normotensive normoalbuminuric (H-M-), 32 hypertensive normoalbuminuric (H+M-), and 22 hypertensive microalbuminuric (H+M+) NIDDM patients and 20 sex-, age-, and weight-matched nondiabetic control subjects were studied. Lipoprotein size was measured by nondenaturing polyacrylamide gradient gel electrophoresis; insulin sensitivity was assessed by using a euglycemic hyperinsulinemic clamp and [6,6(2)H]glucose tracer infusion for simultaneous measurement of hepatic glucose output and whole-body glucose utilization. Total plasma and very-low-density lipoprotein cholesterol were higher in H+M+ than in control subjects (5.84 +/- 0.98 versus 4.97 +/- 0.98 and 0.57 +/- 0.54 versus 0.26 +/- 0.21 mmol/L, mean +/- SD, P < .05). Plasma triglycerides were higher in H+M+ than in either control or H-M- subjects (2.17 +/- 1.32 versus 1.18 +/- 0.67 and 1.30 +/- 0.59 mmol/L, respectively; P < .05). The mean low-density lipoprotein diameter was 27.2 +/- 0.8 in control, 26.7 +/- 0.8 in H-M-, 26.5 +/- 0.8 nm in H+M- (P < .05 versus control subjects), and 26.0 +/- 0.8 nm in H+M+ subjects (P < .05 versus control subjects). The mean cholesterol level of the large high-density lipoprotein particles was lower in H+M- and H+M+ (0.37 +/- 0.14 and 0.36 +/- 0.16 mmol/L) than in control and H-M- (0.54 +/- 0.41 and 0.54 +/- 0.27 mmol/L, P < .05) subjects.(ABSTRACT TRUNCATED AT 250 WORDS)

Impaired response to angiotensin II in Type 1 (insulin-dependent) diabetes mellitus. Role of prostaglandins and sodium-lithium countertransport activity

SummaryThe pathogenesis of diabetic nephropathy remains elusive. A role for renal prostaglandins in antagonizing the hormonal effects of renin-angiotensin II has been postulated as a putative factor leading to hyperfiltration in patients with Type 1 (insulin-dependent) diabetes mellitus. Our aim was to elucidate the effects of angiotensin II on kidney haemodynamics and on blood pressure in eight normal subjects, in nine normotensive, in nine hypertensive with normal sodium-lithium countertransport activity in erythrocytes, in seven hypertensive without and in eight hypertensive Type 1 diabetic patients with microalbuminuria and with high sodium-lithium countertransport activity in erythrocytes. Angiotensin II infusion 4ng·kg−1·min−1 for 60 min) decreased the glomerular filtration rate to a greater extent in normal subjects (−20%), than in normotensive patients (−5% p<0.01), in hypertensive patients with normal sodium-lithium countertransport activity in erythrocytes (−8% p<0.01) in hypertensive patients with high sodium-lithium countertransport (−6% p<0.01) and in hypertensive microalbuminuric patients (−5% p<0.01) with Type 1 diabetes. The urinary excretion rate of vasodilatory prostaglandins was two-three fold higher in all patients than in normal subjects. Acute indomethacin treatment restored a normal response to angiotensin II infusion in normotensive patients, but did not change the renal haemodynamic response in normal subjects. With regard to hypertensive patients with and without microalbuminuria indomethacin treatment restored a normal response to angiotensin II in some but not all patients. An inverse relation was found between angiotensin II-induced decrease in the glomerular filtration rate and the sodium-lithium countertransport activity in erythrocytes during indomethacin treatment. Hypertensive and microalbuminuric patients with a sodium-lithium countertransport activity higher than 0.41 mmol·l erythrocyte−1·h−1 (the upper limit in normal subjects) also had a greater intimal plus medial thickness of the carotid artery using an ultrasonic imaging technique. Chronic indomethacin administration (30 days) significantly decreased the baseline overnight fasting glomerular filtration rate in normotensive and in hypertensive patients with normal but not in hypertensive and microalbuminuric patients with high sodium-lithium countertransport activity.In conclusion these results demonstrate that: (1) excessive synthesis of vasodilatory prostaglandins antagonizes the regulation of renal haemodynamics by angiotensin II, at least partially accounting for hyperfiltration in Type 1 diabetes, (2) elevated sodium-lithium countertransport activity in erythrocytes identifies a subgroup of patients with Type 1 diabetes and hypertension, with and without microalbuminuria, in whom the normalization of urinary excretion rate of prostaglandins does not restore a normal response to angiotensin II.

Sensory neuropathy hampers nociception-mediated bone marrow stem cell release in mice and patients with diabetes

Aims/hypothesisUpon tissue injury, peripheral sensory neurons release nociceptive factors (e.g. substance P [SP]), which exert local and systemic actions including the recruitment of bone marrow (BM)-derived haematopoietic stem and progenitor cells (HSPCs) endowed with paracrine pro-angiogenic properties. We herein explore whether diabetic neuropathy interferes with these phenomena.MethodsWe first investigated the presence of sensory neuropathy in the BM of patients with type 2 diabetes by immunohistochemistry and morphometry analyses of nerve size and density and assessment of SP release by ELISA. We next analysed the association of sensory neuropathy with altered HSPC release under ischaemia or following direct stimulation with granulocyte colony-stimulating factor (G-CSF). BM and circulating HSPCs expressing the neurokinin 1 receptor (NK1R), which is the main SP receptor, were measured by flow cytometry. We finally assessed whether an altered modulation of SP secretion interferes with the mobilisation and homing of NK1R-HSPCs in a mouse model of type 2 diabetes after limb ischaemia (LI).ResultsNociceptive fibres were reduced in the BM of patients and mice with type 2 diabetes. Patients with neuropathy showed a remarkable reduction in NK1R-HSPC mobilisation under ischaemia or upon G-CSF stimulation. Following LI, diabetic mice manifested an altered SP gradient between BM, peripheral blood and limb muscles, accompanied by a depressed recruitment of NK1R-HSPCs to the ischaemic site.Conclusions/interpretationSensory neuropathy translates into defective liberation and homing of reparative HSPCs. Nociceptors may represent a new target for treatment of diabetic complications.

Insulin sensitivity correlates with glycogen synthesis rate, but not with von Willebrand factor in type 2 diabetes

BACKGROUND: Whether insulin resistance in type 2 (non-insulin-dependent) diabetes is due to compromised endothelial insulin migration or to impaired intracellular hormone action or both is unclear. Coexistent microalbuminuria reflects possible endothelial pathogenesis in insulin resistance. METHODS: Insulin sensitivity (S(I)) was calculated from an intravenous glucose tolerance test in 23 type 2 albuminuric (AER+), 11 type 2 normoalbuminuric (AER-), and 17 control subjects. Cultured fibroblasts from skin biopsies from these subjects were used to study intracellular insulin action on glycogen synthesis. Endothelial damage in type 2 diabetes was evaluated by plasma concentrations of von Willebrand factor (vWf). Results: S(I) and glycogen synthesis in fibroblasts were lower in AER+ and AER- than in controls. Glycogen synthesis in vitro was related to S(I) in vivo (r=0.55, P<0.001). vWf was 169+/-12% in AER+ and 140+/-5% in AER-, P<0.051. No correlation was observed between vWf and S(I) or plasma insulin clearance. CONCLUSIONS: This study demonstrates that reduced insulin-mediated glucose removal in type 2 diabetes is strictly associated with a decreased glycogen synthesis of cultured skin fibroblasts in vitro, but not with markers of endothelial damage in vivo.

Poor metabolic control and predisposition to hypertension, rather than hypertension itself, are risk factors for nephropathy in type 2 diabetes

Sodium-lithium countertransport (Na+/ Li+ CT) activity in erythrocytes has been shown to be high in a subset of patients with essential but not secondary hypertension and in type 1 (insulin-dependent) diabetic patients with nephropathy. More recently it has been shown that the presence of a major gene for Na+/Li+ CT, or another closely linked gene, rather than the actual level of Na+/Li+ CT, increases the risk of hypertension onset. The aim of the present study was to investigate whether Na+/Li+ CT activity is associated with hypertension and nephropathy in type 2 (non-insulin-dependent) diabetes. We studied 18 type 2 diabetic patients with normal blood pressure levels (systolic ≤140 and diastolic ≤85 mmHg) and albumin excretion rate (≤15 μg/min), 19 type 2 diabetic patients with hypertension (systolic ≥145 and diastolic ≥90 mmHg) and a normal albumin excretion rate (≤15 μg/min) and 19 type 2 diabetic patients with an increased albumin excretion rate (≤20 μg/min), irrespective of blood pressure levels. Eighteen normal subjects, matched for sex and age, served as controls. Na+/Li+ CT activity in erythrocytes was higher in type 2 diabetics with a high albumin excretion rate (486±148 μmol/l erythrocytes per hour,P<0.01) and in hypertensive diabetics (410±129,P<0.05), but not in normotensive diabetics (340±141), than in controls (282±96) (mean±SD). Body mass index was higher in diabetics with hypertension and in those with an abnormal albumin excretion rate than in normotensive diabetics and controls. Blood pressure levels were higher in diabetic patients with an increased albumin excretion rate than in normotensive diabetics and controls. Of diabetic patients with a high albumin excretion rate 26% had normal diastolic blood pressure levels. Diabetics with a high albumin excretion rate had higher glycated haemoglobin, cholesterol and triglyceride levels and a longer duration of diabetes than hypertensive diabetics with a normal albumin excretion rate. The association of these clinical features in type 2 diabetes closely resembles that previously reported in type 1 diabetes. A novel finding of the present study is that predisposition to hypertension, as indicated by high Na+/Li+ CT, seems to confer a susceptibility to the development of renal damage in type 2 diabetes, partially independent of blood pressure levels per se, and that diabetic patients with high Na+/Li+ CT and hypertension are, to some extent, protected against the development of nephropathy when the metabolic control is tighter and the duration of the disease shorter.