Increased levels of platelet-derived microparticles in patients with diabetic retinopathy

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Abstract

Diabetic retinopathy is caused by capillary occlusions. Platelet-derived microparticles (PMPs) stimulate the coagulation cascade and increase leukocyte and endothelial cell adhesions, both of which are key events in the development of diabetic retinopathy. However, the correlation between the levels of PMPs and diabetic retinopathy has not been precisely determined. The PMPs levels and the expression of platelet CD62P and CD63 were measured in 92 diabetic patients. The level of PMPs was significantly correlated with the expression of CD62P (r = 0.76, P < 0.0001) and CD63 (r = 0.71, P < 0.0001). The mean level of PMPs in diabetics (507 ± 15/104 platelets (plt), mean ± S.E.) was significantly higher than that in normal. The PMPs levels increased with the progression of the diabetic retinopathy; 480 ± 28/104 plt in diabetic patients without retinopathy (n = 25), 504 ± 40/104 plt with mild or moderate non-proliferative diabetic retinopathy (n = 13), 512 ± 29/104 plt with severe non-proliferative diabetic retinopathy (n = 25), and 528 ± 25/104 plt with proliferative diabetic retinopathy (n = 29). The PMPs level in patients with non-perfused retinal areas (582 ± 27/104 plt, n = 24) was significantly higher than patients without non-perfused areas (469 ± 23/104 plt, n = 30; P = 0.0096) and without diabetic retinopathy (P = 0.024). These high correlations indicate that increased levels of PMPs may accelerate diabetic retinopathy.

Introduction

The number of patients with diabetes is increasing dramatically, and diabetic retinopathy resulting in retinal ischemia [1], [2], [3] is a major cause of adult blindness. To prevent the progression of diabetic retinopathy to proliferative diabetic retinopathy (PDR), retinal photocoagulation is performed. Although photocoagulation is the only effective treatment to reduce the retinal ischemia, this treatment destroys retinal cells including photoreceptor cells and can thereby induce severe vision [4]. Therefore, a search for new and better non-invasive therapy for diabetic retinopathy is going on.

Diabetic retinopathy is characterized at its early stage by areas of microvascular damage and capillary non-perfusion. Platelets play an important role in this vascular occlusive process [1], [5], [6], [7], and patients with diabetes develop hypercoagulability and hyperaggreability of platelets [5], [7]. Various platelet abnormalities appear to be involved in the genesis and/or the evolution of diabetic microangiopathy [5], [7], as suggested by the presence of platelet aggregates and thrombi in the small vessels of the retina and the kidneys of diabetic patients [6]. Pro-coagulant activity is expressed as markers of activated platelet, e.g., platelet CD62P (P-selectin), CD63, and platelet-derived microparticles (PMPs) [8], [9].

PMPs were first observed as vesicles released from activated platelets following their adhesion to vessel walls [10]. PMPs contain a platelet-activating factor (PAF), and express high-affinity receptors for factor VIII and platelet/endothelium attachment receptors such as P-selectin [11], [12]. In addition, PMPs increase the level of intracellular adhesion molecule (ICAM)-1, stimulate cytokine secretion, and increase tissue factor expression in endothelial cells [13]. P-selectin is believed to be involved in the initial leukocyte–endothelium interactions [14], [15], [16], and ICAM-1 plays an important role in the adhesion and transmigration of leukocytes [17], [18]. Thus, PMPs activate the coagulation cascade [19], and increase leukocyte and endothelial cell adhesions [12], [20], both of which are key events in the development of capillary occlusion in diabetic patients [21], [22], [23], [24]. Therefore, the release of PMPs has been investigated and determined to be evidence of cellular activation occurring in diseases associated with diabetes and thrombotic risk [12], [21], [22], [23], [24], [25], [26], [27], [28], [29]. However, information on the level of PMPs in patients with diabetic retinopathy is limited [21] and the data have not been well determined.

The purpose of this study was to determine whether the microangiopathic changes in the retina arise from the increased level of PMPs. To accomplish this, we have measured the level of PMPs and activated platelets by determining the expression of CD62P (platelet P-selectin) and CD63. Understanding the role of PMPs in retinal ischemia may provide clues for non-invasive therapies for diabetic retinopathy.

Section snippets

Patients

Ninety-two type 2 diabetic patients (37 men and 55 women, ages 31–83 years, average 60.5 years) were studied. All patients who were diagnosed as having type 2 diabetes and agreed to have additional examinations were investigated. An informed consent was obtained from all patients after an explanation of the purpose and procedures of the study, and the procedures were performed to conform to the tenets of the declaration of Helsinki. Patients with severe nephropathy, under treatment by

Correlations between PMPs, and CD62P and CD63

The level of PMPs was strongly correlated with the level of CD62P (r = 0.76, P < 0.0001, Pearson's product moment coefficient of correlation, Fig. 1A). The Spearman's rank correlation coefficient was also significant (P < 0.0001). The level of PMPs was significantly higher in patients with higher concentration of CD63 (r = 0.71 and P < 0.0001, Fig. 1B). These results demonstrated that the platelets were indeed activated, and in all likelihood, accounted for the higher levels of PMPs.

Correlation between PMPs and HbA1c

There was a

Discussion

Diabetic retinopathy is caused by capillary occlusions, and capillary occlusions result from microvascular thrombi in which erythrocytes, platelets, and leukocytes play a role [1], [31]. The role of platelets in the pathogenesis of diabetic retinopathy has been pointed out [5] and diabetic patients have activated platelets [5], [6], [31], but the role of activated platelets in diabetic retinopathy has not been determined. Activated platelets express adhesive molecules, e.g., CD62P (platelet

Acknowledgement

This study was supported in part by a Grant-in Aid for Scientific Research from the Ministry of Education in Japan.

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