间充质干细胞来源Exosomes通过激活CHSY促进椎间盘退变修复的机制研究

The incidence of low back and neck pain, which is often linked to intervertebral disc degeneration (IDD), is extraordinary high. The common strategies are to remove the degenerated disc by surgical approach. Although such approach provides symptomatic relief, but loss of motion in the operated spine segment and its high financial cost burdens the each patient. With the development of regenerative medicine, Mesenchymal stem cell (MSC) based regenerative therapy brings hope to eliminate such problems. However, there are draw backs such as invasive collection, low cell number harvested, age dependent pluripotency and limited in vitro expansion ability all restricted the use and outcome of MSC based regenerative therapy. Recently, exosome related studies are popular among the regenerative field for its possibility to replace MSC or other stem cells in regenerative medicine. Exosomes are small secreted particles that contain many vital proteins and RNAs, they are small in size but large in number, and they also can be easily transmitted from one cell to another taking advantage of its double layer membrane that resembles the cell membrane. Therefore, our project aims to study the effectiveness of MSC secreted exosomes in promoting the regeneration of degenerated nucleus pulposus. By study the extracellular matrix production of nucleus pulposus cells, and by screening the responsible gene, we try to unveil the function and mechanism of MSC secreted exosomes in nucleus pulposus cells’ extracellular matrix production. This project will not only discover the underlying mechanism of MSC secreted exosomes to degenerated nucleus pulposus cells, but will also show important data on the possibility of exosomes in treating IDD.

由椎间盘退变（IDD）引起的下腰痛及颈肩痛作为现代社会最为常见的疾患之一。由于针对IDD的治疗离不开外科手术对退变椎间盘的切除，这为患者经济以及生活质量上带来严重的负面影响。随着再生医学领域的发展，以间充质干细胞（MSC）为主的再生治疗手段为IDD患者带来希望。然而MSC等干细胞都存在有创采集、分离量少、年龄依赖强及体外扩增能力有限等缺点极大限制其临床应用与疗效。外泌体作为细胞重要分泌物的一种，具有易渗透、易富集、分泌量大、包含递质多等特点是重要的局部微环境调控介质，同时也是干细胞对诸多脏器修复起作用的重要成分。因此，课题组拟针对MSC来源外泌体对椎间盘髓核细胞是否存在修复作用进行研究。通过研究MSC来源外泌体对髓核细胞外基质生成的作用，检测细胞外基质生成关键酶的表达以确定MSC来源外泌体促髓核细胞修复的作用及机制。最终为明确MSC来源外泌体在IDD中的作用机制与应用价值提供重要理论依据。

由椎间盘退变（IDD）引起的下腰痛及颈肩痛作为现代社会最为常见的疾患之一。由于针对IDD的治疗离不开外科手术对退变椎间盘的切除，这为患者经济以及生活质量上带来严重的负面影响。随着再生医学领域的发展，以间充质干细胞（MSC）来源外泌体为主的再生治疗手段为IDD患者带来希望。课题组针对MSC来源外泌体对椎间盘髓核细胞是否存在修复作用进行了本项目的研究。获得利用人髓核细胞系进行增殖实验，明确了MSC外泌体对NP细胞不具备细胞毒性，同时具有提高NP细胞增殖的能力。利用分组定量处理细胞，并使用Western Blot、PCR方法检测，发现间充质干细胞来源外泌体具有促进细胞外基质Aggrecan合成、上调髓核细胞外基质合成酶CHSY以及下调细胞外基质降解酶ADAMTs等表达的作用。而在机制方面，我们发现了外泌体中miR-21、-23、-125、-let7的表达在处理后的髓核细胞中出现显著富集，而通过功能学验证明确了上述miRNA确实能够引起BACH1表达的变化，从而引起HO-1活性的增高，上调CHSY的表达，进而促进椎间盘退变的修复。通过上述研究启发了关于HO-1的调控机制。在前期针对HO-1修复椎间盘退变的基础进一步通过MSC来源外泌体丰富了HO-1的调控机制，此外也通过该研究进一步证明了MSC来源外泌体促进椎间盘修复的机制。最终为明确MSC来源外泌体在IDD中的作用机制与今后将其引用于临床非手术靶向治疗提供重要理论依据。