Bjarne Myrup

Effect of low-dose heparin on urinary albumin excretion in insulin-dependent diabetes mellitus

We investigated the effect of heparin on urinary albumin excretion in patients with insulin-dependent diabetes mellitus. 39 patients with persistent urinary albumin excretion of 30-300 mg/24 h were randomly treated for 3 months with subcutaneous injections twice daily of isotonic saline, 5000 IU unfractionated heparin, or 2000 anti-Xa IU low-molecular-weight heparin. Unfractionated and low-molecular-weight heparin induced a significant reduction in urinary albumin excretion (p = 0.04 and p = 0.004). The mechanism and clinical relevance is unknown but deserve further attention.

Elevated fibrinogen and the relation to acute phase response in diabetic nephropathy

Insulin-dependent diabetic patients with nephropathy have a high risk of cardiovascular disease. Chronic inflammation is a part of the pathogenesis of atherosclerosis, and presently we have studied the relation between the inflammatory state, measured as levels of interleukin-6 and C-reactive protein and fibrinogen in diabetic nephropathy. Thirty-three insulin-dependent diabetic patients with diabetic nephropathy (urinary albumin excretion rate (AER) > 300 mg/24-h) and 22 patients with incipient diabetic nephropathy (AER 30-300 mg/24-h) were compared with 14 non-diabetic controls and 17 diabetic patients with normal AER (<30 mg/24-h). Fibrinogen was significantly higher in diabetic nephropathy than in non-diabetic controls and diabetic patients with normal AER (median 8.1, range (5.4-15.6) mu mol/l vs. 6.6 (5.0-12.1) mu mol/l, p < 0.05, and 6.2 (5.0-9.0) mu mol/l, p < 0.005, respectively), while C-reactive protein did not deviate between groups. Interleukin-6 was significantly elevated in all insulin-dependent diabetic patients (diabetic nephropathy (3.2 (1.0-14.5) pg/ml, p < 0.005), incipient nephropathy (3.7 (1.0-22.9) pg/ml, p < 0.005) and diabetic patients with normal AER (2.7 (1.0-9.0) pg/ml, p < 0.05) compared with nondiabetic controls (1.2 (1.0-6.2) pg/ml)). When fibrinogen was adjusted for interleukin-6, C-reactive protein or both, the level of fibrinogen was still higher in patients with diabetic nephropathy than in patients without nephropathy (p < 0.05), which suggests that inflammation is not the only mechanism that increases fibrinogen levels in patients with diabetic nephropathy.

Fish Oil in Diabetic Nephropathy

OBJECTIVE Recent studies in nondiabetic kidney diseases suggest that dietary supplementation with n-3 polyunsaturated fatty acids (fish oil) may have beneficial effects on albuminuria, kidney function, arterial blood pressure, and dyslipidemia. Therefore, we evaluated the long-term effect of fish oil in diabetic nephropathy. RESEARCH DESIGN AND METHODS A 1-year double-blind randomized controlled study comparing fish oil (4.6 g n-3 fatty acids/day) with placebo (olive oil) was performed in an outpatient clinic in a tertiary referral center. Thirty-six normotensive IDDM patients with diabetic nephropathy were included; 18 were treated with fish oil. Seven patients dropped out (four received fish oil), and results for the remaining 29 are presented. Albuminuria (enzyme immunoassay), glomerular filtration rate (51Cr-labeled EDTA plasma clearance), 24-h ambulatory blood pressure, and lipid profile were determined every 6 months. RESULTS Albuminuria increased by 22% (1–46%) (mean [95% CI]) in the fish oil group vs. 15% (−11–49%) in the placebo group (NS). Glomerular filtration rate decreased from 116 ± 7 to 105 ± 7 ml · min−1 · 1.73 m−2 (mean ± SE) vs. from 108 ± 6 to 103 ± 7, fish oil and placebo, respectively (NS). No significant changes occurred in 24-h ambulatory blood pressure: from 141 ± 4/82 ± 2 mmHg to 142 ± 5/83 ± 2 vs. from 140 ± 4/78 ± 2 to 144 ± 4/80 ± 3, fish oil and placebo, respectively (NS). In the fish oil group, serum triglycerides (median [range]) decreased from 0.97 (0.5–4.0) mmol/l to 0.8 (0.4–3.0) vs. from 1.01 (0.4–2.0) to 1.09 (0.4–2.0) in the placebo group (P < 0.05) and VLDL cholesterol decreased from 0.45 (0.23–1.88) to 0.37 (0.21–1.43) mmol/l vs. from 0.44 (0.21–0.94) to 0.41 (0.17–1.94) (P < 0.05), but total and LDL cholesterol rose in the fish oil compared with the placebo group. CONCLUSIONS Our study does not suggest that fish oil has beneficial effects on albuminuria, kidney function, blood pressure, and dyslipidemia in normotensive IDDM patients suffering from diabetic nephropathy.

Raised Serum Sialic Acid Concentration Precedes Onset of Microalbuminuria in IDDM: A 10-year follow-up study

OBJECTIVE Elevated concentrations of serum sialic acid, a potent cardiovascular risk factor in the general population, have been found in patients with IDDM and microalbuminuria. We investigated whether a coincidence exists between the increase of sialic acid concentrations and albuminuria in the transition from normoalbuminuria to microalbuminuria. Furthermore, the predictability of increased sialic acid as well as von Willebrand factor (vWF) and total and HDL cholesterol concentrations in development of persistent microalbuminuria in IDDM was investigated. RESEARCH DESIGN AND METHODS This 10-year prospective study was carried out in a cohort of 209 IDDM patients with normoalbuminuria at baseline. RESULTS Of the cohort, 198 patients completed the follow-up period and 27 developed persistent microalbuminuria (urinary albumin excretion rate [UAER] ≥ 30 mg/24 h). A coincident increase of UAER and serum sialic acid concentration was seen before persistent microalbuminuria was diagnosed. Elevation of serum sialic acid concentrations in those who later developed microalbuminuria occurred 3 years before the diagnosis of persistent microalbuminuria. Baseline serum sialic acid concentrations were significantly higher in the group of patients who later developed microalbuminuria than in the group who remained normoalbuminuric (2.02 ± 0.41 vs. 1.85 ± 0.31 mmol/l [means ± SD], P < 0.05). Baseline serum sialic acid concentration correlated significantly with HbA1c, UAER, blood pressure, total cholesterol, HDL cholesterol, and vWF and was significantly predictive for development of microalbuminuria (hazards ratio [95% CI], 3.1 [1.2–8.1]; P = 0.02) after adjustments for sex, duration of diabetes, smoking, blood pressure, vWF, total cholesterol, and HDL cholesterol. Adjustment for the effects of HbA1c and UAER, however, canceled out the predictive effect of serum sialic acid. CONCLUSIONS UAER and serum sialic acid concentration increase coincidentally before the onset of persistent microalbuminuria. An increased serum sialic acid concentration is predictive for the onset of microalbuminuria independent of age, sex, diabetes duration, smoking, blood pressure, vWF, and total HDL cholesterol.

Effects of heparin and aminoguanidine on glomerular basement membrane thickening in diabetic rats

The effects of heparin and aminoguanidine on glomerular basement membrane thickening were studied in streptozotocin diabetic Sprague‐Dawley rats. A placebo‐treated group and a non‐diabetic group served as controls. All diabetic rats remained severely hyperglycaemic (23 mmol/l) throughout the 8‐month study period. At the end of this time relative kidney weight was significantly increased in diabetic control rats (4.9±0.5 g/kg b.w.) compared with non‐diabetic rats (3.3±0.3 g/kg). This increase was not affected by the intervention treatments. Glomerular basement membrane thickness increased 32% in diabetic control rats (240±24 nm) compared with non‐diabetic rats (182±20 nm). This increase was prevented by s.c. treatment with both unfractionated and low molecular weight heparins, while basement membrane thickness was the same in animals treated with oral heparins and aminoguanidine and untreated diabetic rats. Macroscopic malignant kidney tumours were seen in three aminoguanidine‐treated rats. In conclusion, subcutaneously administered heparin prevents diabetes‐induced glomerular basement membrane thickening.

Increased Tissue Factor Pathway Inhibitor Activity in IDDM Patients With Nephropathy

OBJECTIVE Tissue factor pathway inhibitor (TFPI) is bound to vascular endothelium (presumably to heparan sulfate) and circulates in complex with plasma lipoproteins. It directly binds and inhibits factor Xa. The purpose of the study is to investigate whether plasma TFPI activity is altered in IDDM and nephropathy and to evaluate the possible determinants of the alteration. RESEARCH DESIGN AND METHODS We assessed plasma concentration of TFPI (total, truncated, and domain 3 TFPI) and plasma activity of factor Xa inhibition in nondiabetic control subjects (n = 22) and in IDDM patients with normoalbuminuria (urinary albumin excretion rate [UAE] < 30 mg/24h, n = 17), incipient nephropathy (UAE 30-300 mg/24 h, n = 17), clinical nephropathy (UAE > 300 mg/24h, n = 25). RESULTS Total, truncated, and domain 3 TFPI concentrations were increased in IDDm patients compared with those in control subjects and were more pronounced in IDDM patients with nephropathy. Plasma activity of factor Xa inhibition measured by HEPTEST (Haemachem, St. Louis, MO) assay was increased in IDDM patients, especially in those with nephropathy. TFPI-dependent factor Xa inhibition, obtained as the difference in clotting time with and without adding activity-neutralizing anti-TFPI antibody to samples, was increased in IDDm patients with nephropathy. This was, however, not sufficient to inhibit the biological activity of factor Xa as demonstrated by increased levels of prothrombin fragment 1 + 2. LDL cholesterol and HbA1c were independently correlated to plasma TFPI. CONCLUSIONS Inhibition of factor Xa activity is increased in IDDM patients with nephropathy, mainly because of increased plasma TFPI activity. The increased plasma TFPI activity in these patients may be associated with and regulated by LDL in plasma and metabolic control. The anticoagulant activity of TFPI may attenuate the hypercoagulable state in diabetes but does not seem to be able to normalize hemostasis.

Lack of effect of fish oil supplementation on coagulation and transcapillary escape rate of albumin in insulin-dependent diabetic patients with diabetic nephropathy

OBJECTIVE We studied the effect of a diet supplementation with fish oil in insulin-dependent diabetic patients with nephropathy in order to evaluate whether abnormal transcapillary escape rate of albumin and procoagulant activity in these patients could be modified. METHODS A double-blind, randomized, controlled study was carried out at a tertiary referral centre. The subjects were 29 insulin-dependent diabetic patients with nephropathy. One year of fish oil supplementation (4.6 g n-3 fatty acids/day) was compared with placebo (olive oil). The main outcome measures were N-3 fatty acid proportions of platelet lipids, transcapillary escape rate of albumin, prothrombin fragment 1 + 2, thrombin-antithrombin complexes, markers of fibrinolysis, fibrinogen, factor VII antigen and activity, thrombomodulin, von Willebrand factor, platelet factor 4 and beta-thromboglobulin. These were measured every 6 months. RESULTS Neither transcapillary escape rate of albumin (7.4 (median) (5.0-9.8) (range) % vs. 7.0 (4.6-10.6) %) nor prothrombin fragment 1 + 2 (0.97 (0.72-2.40) nmol/L vs. 1.01 (0.59-3.11) nmol/L) changed after 12 months of fish oil supplementation. CONCLUSION Increased transcapillary escape rate of albumin and activity could not be modified during diet supplementation with fish oil in insulin-dependent diabetic patients with nephropathy.

Aspects of haemostatic function in healthy subjects with microalbuminuria--a potential atherosclerotic risk factor

Microalbuminuria, i.e., slightly elevated urinary albumin excretion rate (UAER), notifies increased risk for atherosclerotic disease and may reflect an early generalized vascular abnormality in healthy subjects. This study was designed in order to examine whether such abnormality is associated with a shift of the haemostatic balance in prothrombotic direction. The following haemostatic factors were measured in two representative groups of clinically healthy subjects, 28 with microalbuminuria (UAER of 6.6-150 micrograms/min) and 60 age- and sex-matched controls with normoalbuminuria (UAER < 6.6 micrograms/min): Coagulation factors: blood platelet count and mean volume, plasma Factor VII antigen concentration and coagulant activity, and plasma concentrations of prothrombin fragment 1 + 2, thrombin-antithrombin III complexes, fibrinogen, and fibrinopeptide A; fibrinolytic and endothelial factors: plasma concentrations of tissue plasminogen activator antigen and plasminogen activator inhibitor type 1 antigen; and endothelial factor: plasma von Willebrand factor antigen concentration. The fibrinolytic and endothelial factors were measured both before and after 10 minutes of venous occlusion of the arm. None of the haemostatic factors were significantly altered in the microalbuminuric group. Plasma fibrinogen concentration tended to be elevated but not statistically significant ((mean (95% C.I.) 7.8 (7.2-8.3) vs. 7.2 (6.9-7.5) mumol/l; p < 0.1). Neither did any of the haemostatic factors correlate with UAER in regression analyses. It is concluded that the haemostatic balance is unaltered in healthy subjects with microalbuminuria. It is unlikely that a prothrombotic state is present as an intermedial factor early in a causal chain between microalbuminuria and atherosclerotic vascular disease.

Procoagulant activity and intimal dysfunction in IDDM.

SummaryThe biological activity of thrombin and coagulation factor Xa was assessed in 62 insulin-dependent diabetic patients. A group of non-diabetic subjects of comparable age and urinary albumin excretion rate (<30 mg/24 h) served as control subjects (group 1,n=14). The patients were divided into three groups according to urinary albumin excretion rate. In group 2, albumin excretion rate was less than 30 mg/24 h (n=17), in group 3 albumin excretion rate was in the range 30–300 mg/24 h (n=20) and in group 4 albumin excretion rate was greater than 300 mg/24 h (n=25). Compared to non-diabetic control subjects an increase in the biological activity of factor Xa was observed in all groups of diabetic patients (prothrombin fragment 1+2 levels were 1.14±0.38 nmol/l in group 2,p<0.005; 1.06±0.45 nmol/l in group 3,p<0.05 and 1.03±0.31 nmol/l in group 4,p<0.05 vs 0.75±0.34 nmol/l in group 1). No difference in the level of antithrombin III was seen between the groups. We reconfirmed the presence of intimal dysfunction in diabetic nephropathy demonstrated by elevated transcapillary escape rate of albumin in group 4 compared with group 2 (8.9±2.0% vs 7.0±1.9%,p<0.05). An overall positive correlation between transcapillary escape rate and prothrombin fragment 1+2 was found (r=0.36,p<0.005). However, in the groups with elevated albumin excretion rate such a correlation was not significant (group 3:r=0.15,p=0.54; group 4:r=0.03,p=0.86) while it was sustained in the groups with albumin excretion rate of less than 300 mg/24 h (group 1:r=0.61,p<0.05; group 2:r=0.64,p<0.05). In conclusion, IDDM patients had elevated biological activity of factor Xa, demonstrated by elevated levels of prothrombin fragment 1+2. This increment could not be explained by a deficiency of antithrombin III. [Diabetologia (1995) 38: 73–78]

Release of endothelium-associated proteins into blood by injection of heparin in normal subjects and in patients with Type 1 diabetes

Aims  Disturbances in heparan sulphate proteoglycans in patients with diabetic nephropathy might contribute to the pathogenesis of vascular disease in these patients. To investigate this possible link, we measured the heparin‐induced, immediate release of eight proteins with heparan sulphate binding properties in patients with nephropathy.