MDS克隆细胞通过S100A9/TLR4信号重编程MSC的机制研究

Mesenchymal stem cell (MSC) is an important part of the hematopoietic stem cell niche. Studies have shown that MDS cells can establish an MDS bone marrow microenvironment by reprogramming MSC. The mechanism is unclear. In previous studies, we found that MSC senescence was increased in MDS patients accompanying by decreased osteoblastic differentiation and stem cell-supporting capacities. Senescence was increased in normal MSC after co-culture with MDS cells. Inflammatory factor S100A9 was significantly higher expression in MDS cells and bone marrow supernatant. Its receptor TLR4 was expressed on MSC. The senescence of MSC can be induced by rhS100A9. Applicants inferred that MDS clone cells may reprogram MSC through S100A9 / TLR4 signaling to induce senescence for the clone cell growth. To test this hypothesis, we will employ MDS cell lines and primary cell to reveal the role of MDS cells on MSC reprogramming and clarify the molecular mechanism of MSC senescence induced by S100A9/TLR4 signal, using the WB, FCM, RNA interference, gene arrays, CHIP, et al. We will provide empirical evidence from the molecules and cells levels. This study will lay the foundation for the pathogenesis of MDS from tumor cell reprogramming microenvironment, a new viewpoint, and provide new ideas for MDS treatment in the aspect of microenvironment.

MSC是造血干细胞龛的重要成员。研究表明MDS细胞可通过重编程MSC构建MDS骨髓微环境，但机制尚不清楚。申请人在前期研究中发现：MDS骨髓微环境中MSC衰老增加伴成骨分化、造血支持能力下降；MDS细胞与MSC共培养后，后者衰老增加；炎性因子S100A9在MDS细胞及骨髓上清显著高表达，其受体TLR4在MSC富集表达，且rhS100A9可诱导正常MSC发生衰老。申请人推测MDS克隆细胞可能通过S100A9/TLR4信号重编程MSC，诱导细胞衰老以利于克隆生长。为验证这一假说，我们将通过MDS细胞系及原代细胞，采用WB、FCM、RNA干扰、基因芯片、CHIP等手段，从细胞及分子水平探讨MDS克隆细胞对MSC的重编程作用，明确其介导的S100A9/TLR4信号调控MSC衰老的分子机制。本研究将从MDS细胞重编程微环境这个新视点为揭示MDS发病机制奠定基础，为MDS治疗在微环境方面提供新思路。

骨髓增生异常综合征（MDS）患者间充质干细胞呈现衰老表型， 但其机制尚未阐明。恶性克隆细胞和骨髓微环境中的促炎信号被认为 是 MDS 的关键致病因子。我们的研究表明，S100A9 在低危 MDS 中 高表达。此外，正常的原代间充质干细胞（MSCs）和人基质细胞株 HS-27a 与低危 MDS 骨髓单个核细胞共培养后，均获得衰老表型。外 源性 S100A9 也可诱导 MSCs 和 HS-27a 细胞衰老。重要的是，Toll 样 受体 4（TLR4）的抑制或敲除减弱了 S100A9 诱导的细胞衰老。此外， 我们还发现 S100A9 可诱导 NLRP3 炎性小体的形成和 IL-1β的分泌， MDS 患者标本的研究结果进一步证实了这一观点。ROS 和 IL-1β抑制 后可减弱 S100A9 诱导的细胞衰老，而 NLRP3 过表达和外源性 IL-1β 的添加则诱导细胞衰老。我们的研究表明，S100A9 通过 TLR4、NLRP3 炎性小体形成和 IL-1β分泌促进骨髓基质细胞的衰老，我们的发现加深 了对骨髓间充质干细胞 MDS 重编程的分子机制的理解，并表明了 S100A9 在骨髓肿瘤-环境相互作用中的重要作用。