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The total low-density lipoprotein (LDL) fraction was isolated from 21 patients with type I diabetes and 7 nondiabetic normolipemic subjects. The LDL was separated into two subfractions, one glycated (G-LDL) and one nonglycated (N-LDL), using affinity chromatography. G-LDL comprised 21.1 +/- 3.6 and 5.2 +/- 0.6% of the total LDL in diabetic patients and normal subjects, respectively. G-LDL isolated from both diabetic patients and normal subjects was significantly more glycated than N-LDL isolated from the same subject. G-LDL isolated from both diabetic patients and normal subjects was enriched in triglycerides. The metabolism of N-LDL and G-LDL was investigated in human fibroblasts, which express only the classical LDL receptor, and in human monocyte-derived macrophages, which also express a receptor for G-LDL. In fibroblasts, the rates of receptor-mediated accumulation of N-LDL isolated from normal subjects and diabetic patients were significantly greater (P < 0.01) than those of G-LDL. In contrast, when the same LDL subfractions were incubated with human monocyte-derived macrophages, the rates of receptor-mediated accumulation of G-LDL isolated from both groups were significantly greater (P < 0.01) than those of N-LDL. Rates of degradation of G-LDL by human macrophages were not significantly different from those of N-LDL during short-term incubations but reached statistical significance (P < 0.05) when LDL subfractions were incubated with cells for 24 h.(ABSTRACT TRUNCATED AT 250 WORDS)

Objective: To compare platelet plasminogen activator inhibitor 1 (PAI-1) concentration in type II diabetic patients and healthy control subjects., Research Design and Methods: We studied a group of 12 diabetic patients whose disease was controlled by diet or sulfonylurea therapy and a group of 17 nondiabetic control subjects. All subjects were free of clinically advanced vascular disease. PAI-1 antigen concentrations were measured in 5 x 10(8) isolated platelets, which were lysed by 1% Triton X-100., Results: Mean platelet PAI-1 was significantly higher in diabetic patients (264 +/- 83 ng/5 x 10(8) platelets) compared with control subjects (202 +/- 71 ng/5 x 10(8) platelets) (P < 0.05). A significant independent positive correlation was found between platelet PAI-1 concentrations and fasting plasma specific insulin levels in the diabetic patients (r = 0.63, P = 0.03)., Conclusions: These findings suggest that 1) a higher platelet PAI-1 concentration may contribute to enhanced thrombosis in type II diabetes and 2) megakaryocyte PAI-1 synthesis may be under the control of insulin.

Diabetes mellitus is associated with altered platelet function and endothelial damage, but their relationship remains unclear. We examined the effect of short-term metabolic control with insulin in 14- and 28-day streptozocin-induced diabetic rats on alterations in in vitro platelet aggregation and serotonin release. Endothelial damage was assessed by plasma concentrations of von Willebrand factor activity (VIIIR:WF) and factor VIII-related antigen (VIIIR:Ag). Insulin was administered for 5 or 7 days at 9 or 21 days, respectively, after streptozocin. Enhanced platelet aggregation responses to adenosine diphosphate (ADP) and thrombin occurred after both durations of diabetes. Insulin therapy returned ADP-induced, but not thrombin-induced, responses to normal. Enhanced thrombin-induced platelet release of serotonin occurred at both times. Collagen-induced platelet release was enhanced in 28-day diabetic rats. Insulin therapy returned these responses to normal. Plasma concentrations of VIIIR:WF and VIIIR:Ag were elevated in 28-day, but only VIIIR:WF was elevated in 14-day diabetic rats. Insulin therapy reduced the elevated levels of VIIIR:Ag in 28-day diabetic rats, but had little effect on either parameter after the shorter duration of diabetes. In summary, Enhanced platelet aggregation and increased release of serotonin occur shortly after the induction of diabetes by streptozocin in adult rats. These platelet changes precede alterations of endothelial function, as determined by plasma VIIIR:WF and VIIIR:Ag levels. Platelet changes respond more rapidly to insulin therapy than do endothelial changes in diabetic rats. The duration of diabetes before insulin therapy does not affect these relationships.(ABSTRACT TRUNCATED AT 250 WORDS)

The relationship between platelet abnormalities and vessel wall changes in diabetes is not known. We have examined the time course of alterations in in vitro platelet function and endothelial damage, as assessed by measurement of plasma levels of von Willebrand factor (VIIIR:WF) and factor VIII-related antigen (VIIIR:Ag), in streptozotocin-induced diabetic rats. Platelet aggregation and the platelet release reaction in response to ADP, thrombin, and collagen were measured in suspensions of washed platelets prepared from rats 3, 7, 14, or 28 days after induction of diabetes and in control animals. Platelets from diabetic animals showed enhanced aggregation response to ADP as early as 3 days after induction of diabetes and became hyperresponsive to thrombin after 7 days, compared to control platelets. Thrombin-induced release of serotonin was greater in platelets from diabetic animals at 14 days. Collagen-induced responses were not different at any time studied. VIIIR:WF was determined by ristocetin-induced platelet agglutination time in gel-filtered platelets, and VIIIR:Ag was determined by immunoelectrophoretic technique. VIIIR:WF and VIIIR:Ag were significantly enhanced in plasma from rats at 28 days after induction of diabetes and VIIIR:Ag was enhanced in plasma from rats at 14 days after induction of diabetes, but at the earlier times studied, neither were different from values in plasma from control-treated rats. Changes in VIIIR:WF and VIIIR:Ag therefore occurred later than the changes in platelet function. Plasma cholesterol concentrations were not significantly different at any of the times studied, but plasma triglyceride concentrations were significantly increased at 3 days and remained increased with further durations of diabetes. This may have contributed to the observed platelet and vessel wall changes. If these in vitro alterations reflect in vivo behavior, then platelet alterations occur before vessel wall changes and therefore do not appear to be a consequence of such changes in experimental diabetes mellitus.

A simplified approach for estimating free and total anti-insulin antibodies and soluble insulin-anti-insulin immune complexes in serum has been developed and evaluated. For determination of free anti-insulin antibodies, a binding ratio for 125I-labeled insulin at fivefold serum dilution is calculated; the binding ratios obtained are reproducible (run-to-run coefficient of variation, 7.3%) and correlate well with titration of anti-insulin antibodies by other techniques (correlation coefficient, 0.9327). Total anti-insulin antibody concentrations are determined by calculating the [125I]insulin binding ratio for a sample previously acidified, adsorbed with acid dextran-coated charcoal, and neutralized. The difference in binding ratios between two aliquots of the same serum, one diluted 10-fold without manipulation and the second studied at an identical dilution after acidification and removal of free insulin, is taken as an index of the presence of soluble insulin-anti-insulin immune complexes. Because the tests can be performed at a single dilution, both time and materials are conserved without apparent loss of discrimination between sera with high antibody titers and high concentrations of insulin-anti-insulin immune complexes and sera with negative titers or low concentrations.

In view of the tendency toward vascular disease in diabetes mellitus, we studied platelet aggregation in 15 normal, 7 prediabetic, 12 latent, and 20 frankly diabetic subjects. Platelets from latent and frank diabetics showed increased platelet aggregation 4 minutes after adding adenosine 5'-diphosphate (60% verus 29% at 1.0 mu-M), epinephrine (46% versus 14% at 0.25 mu-M), and collagen (72% versus 17% at 0.25 mu-g/ml). Three prediabetics had increased platelet aggregation. Platelet sensitivity to aggregating agents was most marked in frank diabetics, intermediate in latent diabetics, and least in prediabetics. Second-phase platelet aggregation was reversed with acetylsalicylic acid, intravenous tolbutamide, and oral glucose administration. We conclude that platelet aggregation may be increased early in diabetes mellitus and may be involved in the genesis of diabetic microangiopathy. Prospective studies on the effect of therapeutic agents such as acetylsalicylic acid on the natural course of diabetic vascular disease are indicated.

Irreversible second-phase aggregation of platelets in diabetic patients is prevented in vitro by 5, 8, 11, 14-eicosatetraynoic acid (TYA), a competitive inhibitor of the labile aggregation-stimulating substance (LASS) which is formed from arachidonic acid. Thus, inhibition of prostaglandin synthesis inhibits platelet aggregation in diabetic subjects. These findings indicate that platelets from diabetics are subject to control by intracellular mechanisms operative in the regulation of platelet function in normal individuals.

Platelet survival in diabetes mellitus may be decreased or normal, and it is not clear whether altered platelet survival is due to a platelet or to a non-platelet defect. Therefore, platelet survival studies were performed at intervals up to 28 days in streptozotocin-induced diabetic and normal rats, using washed platelets from diabetic or normal animals. When compared to platelets from control rats, there was a significant decrease in platelet survival when platelets from 7 and 14 day diabetic rats were injected into normal controls or into diabetic rats. After 28 days of diabetes, platelet survival in diabetic rats was significantly lengthened, whether the platelets came from control or diabetic rats. Conclusions. Shortened platelet survival in the diabetic rat is caused initially by a platelet defect. Later, non-platelet factors become dominant. These findings may help explain reported discrepancies in results of platelet survival in diabetes mellitus.

Second-phase platelet aggregation induced by adenosine diphosphate (ADP) and epinephrine was measured in fasting platelet-rich plasma in normals, "prediabetics," and diabetics with or without vascular disease. "Plasma factor" potentiation of ADP-induced second-phase platelet aggregation was also estimated, as were megathrombocyte numbers in the same patient groups. There was an increased sensitivity of second-phase platelet aggregation noted with both aggregating agents in all diabetic groups except for the prediabetics. This activity was paralleled by an increase in plasma factor activity. In vivo evidence of an increased turnover of platelets in frank diabetics was suggested by increased numbers of megathrombocytes. These studies demonstrate that platelets from diabetics are sensitive to aggregating agents and that this sensitivity may be related to plasma factor(s) present in diabetics. In vivo platelet aggregation may be present in diabetics. Longitudinal studies will be necessary to establish the relationship of these findings to the genesis of diabetic vascular disease.