组织干细胞的神经保护机理研究

Mesenchymal stem cells (MSCs) are considered one of the most promising candidates for cell-based therapies in various neurodegenerative diseases, including Parkinson's disease (PD). Despite advances in MSC transplantation that aim to prevent neurodegeneration and motor dysfunction of PD patients in preclinical trials, the exact mechanism of MSC therapy in PD have not yet been ascertained. The aim of this study was therefore to address the molecular mechanism underlying the neuroprotective properties of human MSCs transplanted in 6-OHDA-induced rat PD model. Our previous report showed that MSC-mediated immunosuppression occurrs through inflammatory cytokine-upregulation of iNOS/IDO and chemokines, suggesting that MSCs are able to response to inflammatory signals and exhibit the immunoregulatory properties. Prompted by this finding, we hypothesize that upon the influence by in vivo neuro-inflammatory microenvironment, MSCs exert their neuroprotective effect in PD model through certain secreted factor(s) or metabolite(s). We propose three specific procedures to prove this hypothesis using primary human neural cell models and the rat PD model. We will first investigate the interaction between transplanted MSCs and endogenous neural cells to identify the target cells of MSC function in vivo. Next, we will examine whether MSCs are able to affect neuroinflammation/oxidative stress/neural apoptosis of PD to elucidate the molecular mechanisms of MSCs in PD. Lastly, we will perform gene array and metabolism analysis and screen a panel of cytokines or metabolites secreted by MSCs to identify the key factor/metabolite directly accountable for the treatment efficacy of MSCs in PD model. This study will provide insights into the mechanisms of action of MSC therapy in PD and lead to improvements in MSC-based cell therapies for various neurodegenerative diseases.

组织干细胞已成为目前干细胞治疗神经退行性疾病的最佳选择之一，然而其具体的作用机制尚不明确。为切实接近临床应用，本项目以人源的组织干细胞为研究对象，将深入阐明人源组织干细胞在帕金森疾病中的神经保护机制。我们将通过人神经瘤母细胞系SH-Sy5y,人原代神经细胞和大鼠帕金森疾病模型，采用real time PCR、Western blot、腺病毒载体转染、RNA干扰等手段，从分子、细胞、组织以及动物整体水平等多方面探讨组织干细胞治疗帕金森病的作用机制，揭示组织干细胞与机体微环境的交互作用；明确其体内作用的靶细胞；深入探讨组织干细胞在抗神经炎症/氧化应激/神经细胞凋亡等方面的作用；通过细胞因子表达谱和代谢组学等检测和分析，探寻组织干细胞在治疗中发挥关键作用的细胞因子或代谢产物。本研究将为揭示组织干细胞的治疗机制奠定基础，为组织干细胞在神经退行性疾病的临床防治提供新的思路。

组织干细胞又名间充质干细胞（MSCs)，已成为目前干细胞治疗神经退行性疾病的最佳细胞来源之一，然而其具体的作用机制尚不明确。此研究中，我们发现，炎症因子IFN-g和TNF-a预处理的人源MSC（cytokine-primed MSCs, pMSCs)移植入小鼠脑纹状体区后能够显著缓解6-OHDA诱导的小鼠帕金森疾病（Parkinson's disease, PD)。体内细胞示踪实验发现pMSCs在小鼠脑内的迁移和存活能力非常有限，在移植两周后几乎检测不到外源MSCs，由此排除了其通过神经分化进行组织修复的可能性。研究表明，pMSC移植能够显著降低PD小鼠脑损伤区的氧化应激水平，同时通过上调抗氧化应激相关酶的酶活和含量提高PD小鼠的抗氧化应激能力。我们前期研究发现，炎症因子刺激下的MSCs能够高表达吲哚胺2,3 - 双加氧酶（IDO）。IDO是色氨酸分解代谢（犬尿氨酸途径）中催化起始反应的酶，通过对MSCs的犬尿氨酸途径的鉴定，我们发现人源MSC能够将色氨酸几乎完全转化为代谢产物kynurenine(KYN)、kynurenic acid(KYNA)和anthranilic acid(AA)。实验表明，IDO在MSC的神经保护作用中发挥关键作用，由IDO催化色氨酸分解所产生的KYNA及其前体KYN，介导了pMSC的抗氧化应激及神经保护作用。此研究中的发现将为MSCs治疗PD的临床应用和机制研究提供新的视角和依据。