背景:微血管并发症与糖尿病患者的不良后果风险明显增加相关,包括冠状动脉微血管损伤,表现为心脏微血管内皮细胞(CMECs)之间的粘附连接破坏。然而,导致糖尿病冠状动脉微血管通透性过高的特定机制仍然难以捉摸。
方法:在具有脂肪组织特异性Adipsin过表达(AdipsinLSL/LSL-Cre)和它们各自的对照(AdipsinLSL/LSL)的小鼠中诱导实验性糖尿病。此外,对培养的CMECs进行高葡萄糖/棕榈酸(HG+PA)处理以模拟糖尿病的机制方法。
结果:结果表明,Adipsin过表达显著降低心脏微血管通透性,保留的冠状动脉微血管完整性,冠状动脉微血管密度增加。Adipsin过表达也减轻了糖尿病小鼠的心脏功能障碍。E/A比,心脏舒张功能的指标,由Adipsin改进。Adipsin过表达延迟左心室不良重塑,增强型LVEF,改善心脏收缩功能.富含脂肪素的外泌体被CMEC吸收,抑制CMECs凋亡,HG+PA处理下CMECs增殖增加。富含脂肪素的外泌体也加速伤口愈合,拯救细胞迁移缺陷,并促进响应HG+PA攻击的管形成。此外,富含Adipsin的外泌体维持内皮细胞边界的粘附连接,并逆转HGPA损伤破坏的内皮高通透性。机械上,Adipsin阻断HG+PA诱导的Src磷酸化(Tyr416),VE-钙粘蛋白磷酸化(Tyr685和Tyr731),和VE-钙黏着蛋白内在化,从而保持CMECs粘附连接的完整性。LC-MS/MS分析和共免疫沉淀分析(Co-IP)揭示了Csk作为Adipsin的直接下游调节剂。Csk敲除增加Src磷酸化(Tyr416)和VE-钙黏着蛋白磷酸化(Tyr685和Tyr731),同时消除Adipsin诱导的VE-cadherin内化抑制。此外,Csk敲低可抵消Adipsin诱导的对体外内皮高通透性和体内冠状动脉微血管内皮屏障完整性的保护作用。
结论:一起,这些发现有利于Adipsin在CMECs粘附连接完整性调节中的重要作用,揭示了其作为糖尿病冠状动脉微血管功能障碍治疗目标的承诺。图形摘要描述了Adipsin诱导的糖尿病冠状动脉微血管功能障碍调节背后的作用机制。
Microvascular complications are associated with an overtly increased risk of adverse outcomes in patients with diabetes including coronary microvascular injury which manifested as disruption of adherens junctions between cardiac microvascular endothelial cells (CMECs). However, particular mechanism leading to diabetic coronary microvascular hyperpermeability remains elusive.
Experimental diabetes was induced in mice with adipose tissue-specific Adipsin overexpression (AdipsinLSL/LSL-Cre) and their respective control (AdipsinLSL/LSL). In addition, cultured CMECs were subjected to high glucose/palmitic acid (HG + PA) treatment to simulate diabetes for a mechanistic approach.
The results showed that Adipsin overexpression significantly reduced cardiac microvascular permeability, preserved coronary microvascular integrity, and increased coronary microvascular density. Adipsin overexpression also attenuated cardiac dysfunction in diabetic mice. E/A ratio, an indicator of cardiac diastolic function, was improved by Adipsin. Adipsin overexpression retarded left ventricular adverse remodeling, enhanced LVEF, and improved cardiac systolic function. Adipsin-enriched exosomes were taken up by CMECs, inhibited CMECs apoptosis, and increased CMECs proliferation under HG + PA treatment. Adipsin-enriched exosomes also accelerated wound healing, rescued cell migration defects, and promoted tube formation in response to HG + PA challenge. Furthermore, Adipsin-enriched exosomes maintained adherens junctions at endothelial cell borders and reversed endothelial hyperpermeability disrupted by HG + PA insult. Mechanistically, Adipsin blocked HG + PA-induced Src phosphorylation (Tyr416), VE-cadherin phosphorylation (Tyr685 and Tyr731), and VE-cadherin internalization, thus maintaining CMECs adherens junctions integrity. LC-MS/MS analysis and co-immunoprecipitation analysis (Co-IP) unveiled Csk as a direct downstream regulator of Adipsin. Csk knockdown increased Src phosphorylation (Tyr416) and VE-cadherin phosphorylation (Tyr685 and Tyr731), while abolishing Adipsin-induced inhibition of VE-cadherin internalization. Furthermore, Csk knockdown counteracted Adipsin-induced protective effects on endothelial hyperpermeability in vitro and endothelial barrier integrity of coronary microvessels in vivo.
Together, these findings favor the vital role of Adipsin in the regulation of CMECs adherens junctions integrity, revealing its promises as a treatment target against diabetic coronary microvascular dysfunction. Graphical abstract depicting the mechanisms of action behind Adipsin-induced regulation of diabetic coronary microvascular dysfunction.