脂质纳米(LNP)-RNAi技术治疗骨性关节炎软骨退变的实验研究

OA has remained as the major cause of human body joint dysfunction, unfortunately for which there has not existed an effective treatment. Previous research has demonstrated that Ihh has a major role in the early pathogenesis of articular cartilage degeneration disease through mediating MMP-13. Therefore, it may reverse, or even block the pathological process by targeting silence of the genes of Ihh and MMP-13. To achieve that, we should first ascertain the concentration of Ihh and MMP-13 during the cartilage damage in the OA animal model and then choose an optimal time course for administration of RNAi on OA. However, due to the lack of blood vessel at the joint cartilage and its compact structure, it is hard for siRNA to enter the joint cartilage and achieve the targeting gene silence. To solve the problem, a new lipid nanoparticles LNP-RNAi delivery system loading specific siRNA which may enter the cartilage surface was created. Through endocytosis, the novel delivery system further forwarded the siRNA to the cartilage cell, as result in restraining the cartilage cell hypertrophy and inhibiting the cartilage cell apoptosis.This study aims to pinpoint the correlation between Ihh,MMP-13 gene and OA cartilage damage, assess the mechanism and effect of lipid nanoparticles LNP-RNAi in treating OA and finally discover new treatment for OA cartilage degeneration disease.

骨关节炎（OA）是引起关节功能障碍的主要原因，目前尚无有效治疗方法。前期研究表明，印度刺猬因子（Ihh）通过介导金属蛋白酶-13(MMP-13)的表达在关节软骨退变疾病早期发病机制中发挥重要作用。故：靶向沉默Ihh及MMP-13基因可能阻断甚至逆转OA病理生理进程。为此，我们首先明确OA动物模型中Ihh及MMP-13浓度在OA软骨损害进程中的时间变化规律，进而选择最佳时间点运用RNAi技术实现靶基因的沉默，达到治疗OA的效果。但由于软骨缺乏血管且结构致密，siRNA很难进入软骨表层。因此，我们构建新型脂质纳米LNP-RNAi递送系统并荷载特定siRNA，通过胞吞作用将siRNA递送至软骨细胞，抑制其肥大样改变并降低细胞凋亡。本研究旨在明确Ihh及MMP-13与OA软骨损害的时间变化规律，进而探讨脂质纳米LNP-RNAi技术治疗OA的作用机制及效果，有望为OA软骨退变疾病寻找新的治疗方法。

研究背景：之前研究表明，早期干细胞中印度刺猬因子(Ihh)缺失后生长板骨化加速进而导致生长板迅速闭合。生长板(或骨骺)损伤占儿童骨折的30%左右，导致骨骼生长畸形，如损伤部位异位骨桥进而形成成角畸形和肢体短缩。异位骨化和骨桥的形成是生长板修复术后面临的主要挑战。主要内容：本研究拟解决两个问题：1）明确特异性敲除Ihh的Col2a1-CreERT2;Ihhfl/fl系和Prx1-Cre;Ihhfl/fl;Rosa26-ZsGreen1系小鼠生长板是否发生异位骨化；2）外源性过表达Ihh是否能抑制生长板异位骨化进而修复生长板。重要结果：Ihh控制生长板发育的骨化过程，Ihh在体内的缺失加速生长板异常骨化程序进而导致生长板缺失，而过表达Ihh则通过抑制骨桥的形成进而修复受损的生长板结构。本研究表明过表达Ihh可修复生长板损伤，促进生长板软骨再生，可为生长板损伤提供新的治疗手段。科学意义：这是首次明确表明过表达Ihh可以通过抑制骨桥的形成来修复生长吧损伤结构。这一发现对治疗生长板损伤有重要的临床应用价值！关键数据见正文报告！

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