Loss of the retinal glycocalyx in diabetes

糖尿病视网膜病变是成人失明的主要原因，被认为是一种微血管并发症 这种疾病的视网膜微血管以微动脉瘤、白细胞-内皮细胞粘连、 出血、通透性增加、毛细血管闭塞、新生血管等。 视网膜病理的潜在因素是内皮细胞表面层的丢失(糖萼)。这个 糖萼是一层致密的分子，包括蛋白多糖(如Syndecan-1)和糖胺多糖 (GAGS，例如硫酸乙酰肝素)。糖基化促进了血浆分子和它们之间的相互作用 内皮受体，抑制血栓形成，保护白细胞-内皮细胞黏附，转导 剪切力，起到渗透屏障的作用。由于硫酸乙酰肝素可以被肝素酶裂解， Syndecan-1在其胞外区上有多个易被基质切割的位点 金属蛋白酶(MMPs)，我们假设这些酶在糖尿病的糖基化缺失中起作用。 在联合攻击Syndecan-1和硫酸乙酰肝素的情况下，糖萼的功能可能是 大量丢失，已在许多模型中显示可增强白细胞与内皮细胞的黏附。 这种粘连被认为促进了糖尿病视网膜中毛细血管的闭塞，导致 随后形成不再灌流的“脱细胞”毛细血管，形成缺血区。 最终，由此产生的缺氧促进了不受控制的血管生成，从而干扰了清晰的视力。当一个 白细胞附着于血管内皮细胞表面，最终会被剪切力强迫回到血流中 力，或替代地，以一种机制介导的穿过内皮(移民)进入周围组织 通过血小板内皮细胞黏附分子-1(PECAM-1)表达。然而，PECAM-1的丢失阻止了白细胞 迁移，黏附的白细胞可以被困在毛细血管内，导致持续的闭塞和 最终毛细血管死亡。我们和其他人已经注意到糖尿病视网膜中PECAM-1的丢失 微循环，这种损失可能是由基质金属蛋白酶介导的切割，细胞因子- 介导的产量下降，和/或蛋白酶体降解。以下具体目标涉及 总的假设是糖尿病导致关键的内皮表面分子丢失，从而导致白细胞 无细胞毛细血管的黏附、毛细血管闭塞和最终发展：(1)研究假说 糖尿病视网膜微循环中硫酸乙酰肝素的丢失是由肝素酶直接介导的 并间接通过基质金属蛋白酶促进Syndecan-1的裂解，(2)确定其作用机制(S) 糖尿病视网膜微循环中PECAM-1的丢失，通过检测表达降低，增加 蛋白酶体的降解和从内皮细胞表面的切割，以及(3)验证假说 白细胞对毛细血管的阻塞和无细胞毛细血管的发育可以通过以下方法减弱 保护内皮细胞表层。

Diabetic retinopathy, the leading cause of blindness in adults, is recognized as a microvascular complication of the disease, with retinal microvessels characterized by microaneurysms, leukocyte-endothelial cell adhesion, hemorrhages, increased permeability, capillary occlusion, neovascularization, etc. We propose that an underlying contributor to the retinal pathology is the loss of the endothelial surface layer (glycocalyx). The glycocalyx is a dense layer of molecules including proteoglycans (e.g., syndecan-1) and glycosaminoglycans (GAGs, e.g., heparan sulfate). The glycocalyx facilitates the interactions between plasma molecules and their endothelial receptors, inhibits thrombosis, shields against leukocyte-endothelial cell adhesion, transduces shear force, and acts as a permeability barrier. Inasmuch as heparan sulfate can be cleaved by heparanase, and syndecan-1 has multiple sites on its extracellular domain susceptible to cleavage by matrix metalloproteinases (MMPs), we hypothesize a role for these enzymes in the loss of the glycocalyx in diabetes. With a combined attack on both syndecan-1 and heparan sulfate, the functions of the glycocalyx could be substantially lost, which has been shown in many models to enhance leukocyte-endothelial cell adhesion. Such adhesion is thought to promote the occlusion of capillaries in the diabetic retina, leading to the subsequent development of “acellular” capillaries that are no longer perfused, creating ischemic zones. Ultimately, the resulting hypoxia promotes uncontrolled angiogenesis that interferes with clear vision. When a leukocyte adheres to the endothelial surface, it can eventually be coerced back into the bloodstream by shear forces, or alternatively, cross the endothelium (emigrate) into the surrounding tissue in a mechanism mediated by platelet endothelial cell adhesion molecule-1 (PECAM-1). However, loss of PECAM-1 prevents leukocyte emigration, and adherent leukocytes can be trapped within capillaries causing continued occlusion and eventual capillary death. We and others have noted a loss of PECAM-1 from the diabetic retinal microcirculation, with this loss possibly caused by mechanisms such as MMP-mediated cleavage, cytokine- mediated decrease in production, and/or proteasomal degradation. The following specific aims address the overall hypothesis that diabetes induces the loss of key endothelial surface molecules that results in leukocyte adhesion, capillary occlusion, and eventual development of acellular capillaries: (1) Investigate the hypothesis that the loss of heparan sulfate from the diabetic retinal microcirculation is mediated directly by heparanase and indirectly by MMP-facilitated cleavage of syndecan-1, (2) Determine the mechanism(s) responsible for the loss of PECAM-1 from the diabetic retinal microcirculation, by testing for decreased expression, increased proteasomal degradation, and cleavage from the endothelial cell surface, and (3) Investigate the hypothesis that capillary occlusion by leukocytes and the development of acellular capillaries can be attenuated by protection of the endothelial surface layer.