AAV-p65shRNA和AAV-BMP4联合应用抑制早期骨性关节炎软骨细胞退变的实验研究

项目名称： AAV-p65shRNA和AAV-BMP4联合应用抑制早期骨性关节炎软骨细胞退变的实验研究

项目编号： No.81472136

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 医药、卫生

项目作者： 李广恒

作者单位： 郑州大学

项目金额： 72万元

中文摘要： 软骨细胞的炎症、修复是骨性关节炎早期病程中软骨细胞退变的两个重要机制，分别起着诱发、减缓该病程的作用。抑制炎症和促进修复均可减缓软骨细胞的退变，但目前尚不清楚炎症和修复两者的内在联系对其退变的影响。我们前期研究发现，BMP4基因治疗可促进软骨细胞修复而抑制其退变。本次研究中，我们提出三个假说：1）体外抑制NF-KB炎症信号并联合应用BMP4对软骨细胞的退变可产生协同抑制作用；2）体外协同抑制作用发挥最大化的条件是两者具有合理比例的作用强度；3）基于体外试验结果，关节腔内注射合理比例的两种病毒颗粒可减缓羊骨性关节炎的软骨细胞退变。我们拟采用体外转染人骨性关节炎软骨细胞AAV-p65shRNA和AAV-BMP4的方法， 经软骨细胞3D培养和羊关节腔内病毒颗粒注射试验，分别从体外和体内实验观察软骨细胞的退变，从而进一步揭示软骨细胞炎症和修复机制对软骨退变影响，探索治疗早期骨性关节炎的新方法。

中文关键词： 骨性关节炎；软骨细胞；信号通路；软骨组织工程；干细胞

英文摘要： Inflammation and repair of chondrocytes are the two main mechanisms underlying the cartilage degeneration of early phase of osteoarthritis. They play important role in triggering and inhibiting the process, respectively. Both inhibiting the inflammation and improving the repair of chondrocytes will slow down the process of cartilage degeneration. However, the relation between the inflammation and repair of OA chondrocytes is not clear as well as its' effect on cartilage degeneration. Our previous studies showed that BMP4 gene therapy can promote the repair of OA chondrocytes and deter the OA process. In the present study, we hypothesize that combined inhibiting the NF-KB inflammation signal by AAV-p65shRNA and promoting repair by AAV-BMP4 will yield the synergistic inhibiting effect of OA chondrocytes degeneration; to maximize the synergistic inhibiting effect needs certain ratio of AAV-p65shRNA and AAV-BMP4 transduction; based on the in vitro result, combined injection of certain ratio of AAV-p65shRNA and AAV-BMP4 particles into the sheep knee joint will inhibit the degeneration process of OA chondrocytes. In the present study, we will use AAV-p65shRNA and AAV-BMP4 to transduce human OA chondrocytes in vitro, and use 3D cell pellet culture to induce the transduced chondrocytes and study their chondrogenic differentiation. In vivo, we will inject both AAV-p65shRNA and AAV-BMP4 particles at certain ratio into the sheep OA knee and examine the cartilage degeneration at different time points. We believe that the combined application of certain ratio of AAV-p65shRNA and AAV-BMP4 will have an inhibiting synergistic effect on OA cartilage chondrocytes degeneration, which may enlighten a new therapy for the early stage of OA.

英文关键词： Osteoarthritis;chondrocyte;signal pathway;cartilage tissue engineering;stem cell