Blockade by phosphorothioate aptamers of advanced glycation end products-induced damage in cultured pericytes and endothelial cells

详细信息    查看全文

• 作者：Yuichiro Higashimoto ; Takanori Matsui ; Yuri Nishino ; Junichi Taira ; Hiroyoshi Inoue ; Masayoshi Takeuchi ; Sho-ichi Yamagishi
• 关键词：AGEs ; advanced glycation end products ; EC ; endothelial cells ; VEGF ; vascular endothelial growth factor ; PAI-1 ; plasminogen activator inhibitor-1 ; RAGE ; receptor for AGEs ; SELEX ; systematic evolution of ligands by exponential enrichment ; AGEs&ndash ; thioaptamers ; phosphorothioate aptamers directed against AGEs ; HUVECs ; human umbilical vein endothelial cells ; HSA ; human serum albumin ; BSA ; bovine serum albumin ; ssDNA ; single-stranded DNA ; PCR ; polymerase chain reactions ; dATP(伪S) ; 2&prime ; -deoxyad
• 刊名：Microvascular Research
• 出版年：November, 2013
• 年：2013
• 卷：90
• 期：Complete
• 页码：64-70
• 全文大小：644 K

文摘

Advanced glycation end products (AGEs) not only inhibit DNA synthesis of retinal pericytes, but also elicit vascular hyperpermeability, pathological angiogenesis, and thrombogenic reactions by inducing vascular endothelial growth factor (VEGF) and plasminogen activator inhibitor-1 (PAI-1) through the interaction with the receptor for AGEs (RAGE), thereby being involved in the pathogenesis of diabetic retinopathy. In this study, we screened novel phosphorothioate-modified aptamers directed against AGEs (AGEs-thioaptamers) using a combinatorial chemistry in vitro, and examined whether these aptamers could inhibit the AGE-induced damage in both retinal pericytes and human umbilical vein endothelial cells (HUVECs). We identified 11 AGEs-thioaptamers; among them, clones #4, #7s and #9s aptamers had higher binding affinity to AGEs-human serum albumin (HSA) than the others. Surface plasmon resonance analysis revealed that K_D values of #4s, #7s and #9s were 0.63, 0.36, and 0.57 nM, respectively. Furthermore, these 3 clones dose-dependently restored the decrease in DNA synthesis in AGE-exposed pericytes. AGEs significantly increased RAGE, VEGF and PAI-1 mRNA levels in HUVEC, all of which were completely blocked by the treatment with 20 nM clone #4s aptamer. Quartz crystal microbalance analysis confirmed that #4s aptamer dose-dependently inhibited the binding of AGEs-HSA to RAGE. Our present study demonstrated that AGEs-thioaptamers could inhibit the harmful effects of AGEs in pericytes and HUVEC by suppressing the binding of AGEs to RAGE. Blockade by AGEs-thioaptamers of the AGEs-RAGE axis might be a novel therapeutic strategy for diabetic retinopathy.