SDX抑制NLRP3炎症小体激活以减轻糖尿病视网膜微血管渗漏(1)
【摘要】 目的 探討舒洛地特(SDX)治疗糖尿病视网膜病变的可能性及分子机制。方法 将40只C57BL/6J小鼠随机分成4组各10只，分别为对照+生理盐水(Con + NS)组、对照+舒洛地特(Con + SDX)组、糖尿病+生理盐水(DM+NS)组和DM + SDX组，SDX的用量为腹腔注射10 mg/(kg·d)、持续12周，对照给予等量生理盐水。采用高糖(25 mmol/L葡萄糖)处理人视网膜微血管内皮细胞(HRMEC)以模拟糖尿病刺激，并与一系列浓度的SDX(0.125、0.25、0.5、1.0 lsu/ml)共处理。用伊文思蓝法检测小鼠视网膜的渗漏情况，用蛋白免疫印迹法、激光共聚焦等技术检测小鼠视网膜组织和HRMEC中的核苷酸结合寡聚域样受体蛋白3(NLRP3)炎症小体的表达和激活情况，以及凋亡相关斑点样蛋白(ASC)、Caspase-1的变化。结果 DM + NS组伊文思蓝吸光度、NLRP3炎症小体及ASC的表达均较Con + NS组高，而DM + SDX组则低于DM + NS组(P均< 0.05)。SDX抑制糖尿病小鼠视网膜渗漏、NLRP3炎症小体激活和视网膜微血管渗漏。HRMEC的NLRP3炎症小体组分和cleaved Caspase-1在高糖刺激下表达上调(P均< 0.05);经不同浓度的SDX处理后，NLRP3炎症小体的表达被抑制，抑制程度随SDX浓度的增加而增加(P均< 0.05)。结论 舒洛地特能抑制糖尿病诱导的NLRP3炎症小体激活和视网膜微血管渗漏，可用于治疗糖尿病视网膜病。
【关键词】 舒洛地特;糖尿病视网膜病变;核苷酸结合寡聚域样受体蛋白3炎症小体;
微血管渗漏;人视网膜微血管内皮细胞
Sulodexide alleviates retinal microvascular leakage by inhibiting the activation of NLRP3 inflamm-asome in diabetic retinopathy Li Jingren， Li Ting， He Xuemin， Zhou Li， Tang Xixiang， Chen Yanming. Department of Endocrinology & Metabolism， the Third Affiliated Hospital of Sun Yat-sen University， Guangzhou 510630， China
Corresponding author， Chen Yanming
【Abstract】 Objective To investigate the feasibility and molecular mechanism of sulodexide in the treatment of diabetic retinopathy. Methods Forty C57BL/6J mice were randomly into the control + normal saline (Con + NS group)， control + sulodexide (Con + SDX group)， diabetes mellitus + normal saline (DM + NS group) and diabetes mellitus + sulodexide groups (DM + SDX group). Sulodexide was intraperitoneal injected at a dose of 10 mg/(kg·d) for 12 weeks， and an equivalent quantity of phosphate buffer saline (PBS) was given in the control group. Human retinal microvascular endothelium cells (HRMECs) were stressed by high glucose (25 mmol/L) to mimic the diabetic conditions， and co-treated with a serial doses of sulodexide (0.125， 0.25， 0.5 and 1.0 lsu/ml). The mouse retinal vascular leakage was evaluated by the Evans blue assay. The expression and activation of nucleotide-binding and oligomerization domain-like receptor protein 3 (NLRP3) inflammasome in the retinal tissues and HRMEC of mouse models， and the changes of the apoptosis-associated speckle-like protein containing CARD (ASC) and caspase-1 were measured by Western blot and laser confocal imaging， etc. Results The absorbance value of Evans blue assay and the expression levels of NLRP3 inflammasome and ASC in the DM + NS group were significantly higher compared with those in the Con + NS group， whereas the values in the DM + SDX group were considerably lower than those in the DM + NS group (all P < 0.05). Sulodexide significantly inhibited the increased retinal leakage， the activation of NLRP3 inflammasome and attenuated the retinal vascular leakage in mouse models. The expression levels of NLRP3， Caspase-1 and cleaved Caspase-1 of HRMEC were significantly up-regulated under the condition of high glucose (all P < 0.05). After treatment with different concentrations of sulodexide， the expression levels of NLRP3 and ASC were significantly down-regulated in a dose-dependent manner (both P < 0.05). Conclusion Sulodexide can inhibit the DM-induced activation of NLRP3 inflammasome and retinal microvascular leakage， which can be applied to treat diabetic retinopathy., http://www.100md.com(李静仁?李婷?何学敏?周丽?唐喜香?陈燕铭)
【关键词】 舒洛地特;糖尿病视网膜病变;核苷酸结合寡聚域样受体蛋白3炎症小体;
微血管渗漏;人视网膜微血管内皮细胞
Sulodexide alleviates retinal microvascular leakage by inhibiting the activation of NLRP3 inflamm-asome in diabetic retinopathy Li Jingren， Li Ting， He Xuemin， Zhou Li， Tang Xixiang， Chen Yanming. Department of Endocrinology & Metabolism， the Third Affiliated Hospital of Sun Yat-sen University， Guangzhou 510630， China
Corresponding author， Chen Yanming
【Abstract】 Objective To investigate the feasibility and molecular mechanism of sulodexide in the treatment of diabetic retinopathy. Methods Forty C57BL/6J mice were randomly into the control + normal saline (Con + NS group)， control + sulodexide (Con + SDX group)， diabetes mellitus + normal saline (DM + NS group) and diabetes mellitus + sulodexide groups (DM + SDX group). Sulodexide was intraperitoneal injected at a dose of 10 mg/(kg·d) for 12 weeks， and an equivalent quantity of phosphate buffer saline (PBS) was given in the control group. Human retinal microvascular endothelium cells (HRMECs) were stressed by high glucose (25 mmol/L) to mimic the diabetic conditions， and co-treated with a serial doses of sulodexide (0.125， 0.25， 0.5 and 1.0 lsu/ml). The mouse retinal vascular leakage was evaluated by the Evans blue assay. The expression and activation of nucleotide-binding and oligomerization domain-like receptor protein 3 (NLRP3) inflammasome in the retinal tissues and HRMEC of mouse models， and the changes of the apoptosis-associated speckle-like protein containing CARD (ASC) and caspase-1 were measured by Western blot and laser confocal imaging， etc. Results The absorbance value of Evans blue assay and the expression levels of NLRP3 inflammasome and ASC in the DM + NS group were significantly higher compared with those in the Con + NS group， whereas the values in the DM + SDX group were considerably lower than those in the DM + NS group (all P < 0.05). Sulodexide significantly inhibited the increased retinal leakage， the activation of NLRP3 inflammasome and attenuated the retinal vascular leakage in mouse models. The expression levels of NLRP3， Caspase-1 and cleaved Caspase-1 of HRMEC were significantly up-regulated under the condition of high glucose (all P < 0.05). After treatment with different concentrations of sulodexide， the expression levels of NLRP3 and ASC were significantly down-regulated in a dose-dependent manner (both P < 0.05). Conclusion Sulodexide can inhibit the DM-induced activation of NLRP3 inflammasome and retinal microvascular leakage， which can be applied to treat diabetic retinopathy., http://www.100md.com(李静仁?李婷?何学敏?周丽?唐喜香?陈燕铭)