Department of Molecular and Cellular Physiology, Louisiana State University Health Sciences Center-Shreveport, Shreveport, LA 71103, USA.
Department of Chemistry and Chemical Biology, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
Glycobiology. 2022 Jul 13;32(8):720-734. doi: 10.1093/glycob/cwac029.
The endothelial glycocalyx regulates vascular permeability, inflammation, and coagulation, and acts as a mechanosensor. The loss of glycocalyx can cause endothelial injury and contribute to several microvascular complications and, therefore, may promote diabetic retinopathy. Studies have shown a partial loss of retinal glycocalyx in diabetes, but with few molecular details of the changes in glycosaminoglycan (GAG) composition. Therefore, the purpose of our study was to investigate the effect of hyperglycemia on GAGs of the retinal endothelial glycocalyx.
GAGs were isolated from rat retinal microvascular endothelial cells (RRMECs), media, and retinas, followed by liquid chromatography-mass spectrometry assays. Quantitative real-time polymerase chain reaction was used to study mRNA transcripts of the enzymes involved in GAG biosynthesis.
Hyperglycemia significantly increased the shedding of heparan sulfate (HS), chondroitin sulfate (CS), and hyaluronic acid (HA). There were no changes to the levels of HS in RRMEC monolayers grown in high-glucose media, but the levels of CS and HA decreased dramatically. Similarly, while HA decreased in the retinas of diabetic rats, the total GAG and CS levels increased. Hyperglycemia in RRMECs caused a significant increase in the mRNA levels of the enzymes involved in GAG biosynthesis (including EXTL-1,2,3, EXT-1,2, ChSY-1,3, and HAS-2,3), with these increases potentially being compensatory responses to overall glycocalyx loss. Both RRMECs and retinas of diabetic rats exhibited glucose-induced alterations in the disaccharide compositions and sulfation of HS and CS, with the changes in sulfation including N,6-O-sulfation on HS and 4-O-sulfation on CS.
内皮糖萼调节血管通透性、炎症和凝血，并充当机械感受器。糖萼的丢失可导致内皮损伤，并有助于几种微血管并发症的发生，因此可能促进糖尿病性视网膜病变。研究表明，糖尿病患者的视网膜糖萼存在部分丢失，但糖胺聚糖 (GAG) 组成的变化缺乏分子细节。因此，我们研究的目的是研究高血糖对视网膜内皮糖萼 GAG 的影响。
从大鼠视网膜微血管内皮细胞 (RRMEC)、培养基和视网膜中分离 GAG，然后进行液相色谱-质谱分析。实时定量聚合酶链反应用于研究参与 GAG 生物合成的酶的 mRNA 转录本。
高血糖显着增加了肝素硫酸盐 (HS)、软骨素硫酸盐 (CS) 和透明质酸 (HA) 的脱落。在高葡萄糖培养基中生长的 RRMEC 单层中 HS 水平没有变化，但 CS 和 HA 水平显着下降。同样，尽管糖尿病大鼠视网膜中的 HA 减少，但总 GAG 和 CS 水平增加。RRMEC 中的高血糖导致参与 GAG 生物合成的酶的 mRNA 水平显着增加（包括 EXTL-1、2、3、EXT-1、2、ChSY-1、3 和 HAS-2、3），这些增加可能是对整体糖萼丢失的代偿反应。糖尿病大鼠的 RRMEC 和视网膜均表现出葡萄糖诱导的 HS 和 CS 二糖组成和硫酸化改变，硫酸化改变包括 HS 上的 N,6-O-硫酸化和 CS 上的 4-O-硫酸化。
