基于超声多模态评价技术探讨肝脏靶向递送ABCA1新策略在动脉粥样硬化防治中的应用

Atherosclerosis (AS) is a prevalent and life-threaten disease. Currently, there are still no effective therapeutic strategies. Developing the prevention and therapeutic strategies is essentially important for both the patients and the society. Recent studies have revealed that the HDL (high density lipoprotein) synthesized by the liver plays an important role in the reverse cholesterol transport and thus AS development. Moreover, the ATP-binding cassette transporter ABCA1 has been found to be essential for the HDL synthesis and maturation, while aberrant low expression of ABCA1 in the liver has been linked to AS. In view of these data, we assumed that targeted increasing the expression of ABCA1 in the liver would be a promising to enhance the HDL level and thus be beneficial for AS prevention and therapy. With the funding support of NSFC, we found that exosomes preferentially circulate into the liver. In addition, we have developed an exosome based strategy for specific enhancing gene expression in targeted cells or tissues. Briefly, we have engineered the exosomes to carry the target protein on the surface and encapsulate the inhibitors for the inhibitory endogenous miRNAs inside. The engineered exosomes thus are more potent in enhancing target protein expression in the recipient cells. In the proposed project, we would develop a liver specific ABCA1 delivery strategy based on the previous experience on exosome engineering. Next, we would systemically analyze the efficacy and side effects of the new ABCA1 delivery strategy on the AS prevention and therapy in the mouse model by ultrasound based evaluation. Moreover, we would also analyze the possible molecular mechanisms responsible for the deregulated ABCA1 in AS patients, which would provide clues for the selection of the candidates for the strategy. Taken together, the current proposal would be promising in decoding the mechanism of atherosclerosis and establishing new treatments for the disease.

截至目前，动脉粥样硬化（AS）的防治依然缺乏有效手段。既往发现肝脏合成的高密度脂蛋白（HDL）对胆固醇逆向运输及AS防治至关重要；而转运蛋白ABCA1是HDL加工成熟的关键分子，AS时多伴有ABCA1功能降低。因此，增强肝脏ABCA1功能，有望提升HDL合成能力，从源头上防治AS。可见，肝脏递送ABCA1这一膜分子是解决该问题的关键。课题组发现，外泌体具有良好的肝脏靶向性，是肝脏递送ABCA1的理想载体。本项目拟在此基础上，利用光遗传和基因工程技术等改构外泌体，使其外表面携带ABCA1，内部加载能够增强内源ABCA1表达的小核酸，实现双途径增强肝脏ABCA1的表达；然后结合超声评价AS的优势，利用AS小鼠模型探讨外泌体递送ABCA1防治AS的效果；最后，通过临床样本分析AS患者ABCA1表达异常的分子机制，为新策略的优势提供临床依据。本项目在加深AS机制认识的同时，有望为其防治提供新思路。

探讨动脉粥样硬化（AS）防治策略具有重要的临床意义。本项目创建了基于外泌体的靶向输送核酸药物的方法，为动脉粥样硬化的防治提供全新的靶向干预策略和新思路，主要取得了以下几方面的结果：1）我们利用基因工程技术构建富含ABCA1 mRNA的外泌体，并在体外验证其在体外可高效将mRNA递送到肝细胞，发挥作用；2) 富含ABCA1的外泌体可升高HDL的水平，延缓动脉粥样硬化的进程；3）超声监测及组织学染色，证实该策略无明显毒性，具有良好的安全性。在本项目的支持下，围绕项目的中心目标，课题组还探讨了超声联合外泌体治疗动脉粥样硬化的其它策略，主要包括：1）在药物递送方面：①建立了基于超声靶向爆破技术（UTMD）提高外泌体靶向递送效率的新策略；②建立了利用超声造影剂示卓安封闭巨噬细胞提高外泌体肝实质细胞的递送效率的新策略；2）在外泌体来源和装载基因选择方面：①肝脏靶向递送Ldlr mRNA逆转家族性高胆固醇血症小鼠AS情况，并注册临床试验，有望在临床上用于家族性高胆固醇血症患者的治疗；②发现棕色脂肪组织外泌体（BAT-Exos）能够缓解肥胖小鼠的代谢综合征；③外泌体递送炎症响应型IL10翻译系统，可以按需抗炎，减轻AS严重程度;○4构建了一种工程化外泌体可以靶向降解细胞内炎性分子，发挥抗炎功能。上述工作创建了多个基于外泌体的AS防治策略，相关研究结果已经发表SCI期刊收录论文9篇，授权国家发明专利4项。

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