去白细胞优化富血小板血浆促进软骨修复的机制研究

Clinical trials and animal studies indicated considerable potential of autologous platelet-rich plasma (PRP) in the treatment of osteoarthritis. However, leukocyte-rich PRP (L-PRP) may decrease the therapeutic effect of pure PRP (P-PRP) and induce more adverse events, including pain and swelling. It was reported that pro-inflammatory cytokines, the same as those released from leukocytes, injured chondrocytes and probably brought about pain. NF-κB pathway activation was believed to play an important role in those pathophysiological processes. Our preliminary results showed that leukocytes inhibited P-PRP-induced-upregulation of Col II and aggrecan in chondrocytes. Therefore, we aim to optimize PRP through the depletion of leukocytes to avoid NF-κB activation, thereby promoting cartilage repair and reducing pain. Firstly, we are planning to compare the effects of L-PRP and P-PRP on osteoarthritic chondrocytes in vitro and on cartilage repair in vivo to analyze the role of leukocytes. Secondly, the effect of L-PRP on NF-κB signaling pathway will be explored in vitro and in vivo. Specific inhibitors and siRNA will also be adopted to further verify the role of NF-κB when leukocytes are present. Based on the final results, the mechanism of optimized PRP enhancing cartilage repair and reducing adverse events will be elucidated. We will thus have a better understanding of PRP in the treatment of OA, and provide more basic and pre-clinical evidence for clinicians.

动物研究和临床试验均发现关节内注射富血小板血浆（PRP）治疗OA有良好的应用前景，但PRP中含有白细胞会降低治疗效果，引起关节疼痛和肿胀。白细胞分泌的炎性因子会损伤软骨细胞，促进疼痛介质分泌，这些作用与NF-κB的活化有关。前期结果显示PRP中的白细胞会抑制软骨细胞Col II和aggrecan的表达。所以，本课题拟通过去除白细胞来优化PRP，避免活化NF-κB，从而促进软骨修复和抑制疼痛介质分泌。先在细胞水平比较含白细胞PRP和去白细胞PRP对OA软骨细胞生物学行为的影响，并在动物水平验证，分析白细胞对软骨修复的影响；再从体外和体内深入探讨PRP中白细胞对NF-κB活化的影响，并用阻断剂和siRNA干扰进一步验证，明确白细胞影响软骨细胞代谢的分子机制。最后，总结归纳分析，阐明优化PRP治疗OA的具体作用机制，丰富对PRP有效成分的认识，为临床上优化PRP治疗OA提供理论依据。

动物研究和临床试验均发现关节内注射富血小板血浆（PRP）治疗OA有良好的应用前景，但PRP中含有白细胞会降低治疗效果，引起关节疼痛和肿胀。白细胞分泌的炎性因子会损伤软骨细胞，促进疼痛介质分泌，这些作用与NF-κB的活化有关。所以，本课题通过去除白细胞来优化PRP，避免活化NF-κB，从而促进软骨修复和抑制疼痛介质分泌。结果发现PRP中含有白细胞会降低软骨细胞的增殖、分泌胞外基质和迁移的能力，影响骨髓间充质干细胞修复软骨损伤的能力，这主要与白细胞会释放IL-1b和TNFa等炎性因子激活NF-kB通路有关，因为在向含白细胞的PRP中添加了该通路阻断剂，这种抑制效果就会得到显著缓解。因而，在促进关节软骨修复方面，采用去除白细胞的PRP效果可能更好，这对临床上采用PRP治疗OA或关节软骨损伤具有一定的指导意义。此外，本项目还改良了制作不含白细胞PRP的离心方法，并研发了相关装置，获得国家授权专利。