Department of Veterinary Clinical Sciences, University of Copenhagen, Agrovej 8, 2630, Taastrup, Denmark.
Department of Health Science and Technology, Aalborg University, Fredrik Bajers Vej 7E, 9220, Aalborg Ø, Denmark.
Stem Cell Res Ther. 2020 May 20;11(1):187. doi: 10.1186/s13287-020-01706-7.
Similar to humans, the horse is a long-lived, athletic species. The use of mesenchymal stromal cells (MSCs) is a relatively new frontier, but has been used with promising results in treating joint diseases, e.g., osteoarthritis. It is believed that MSCs exert their main therapeutic effects through secreted trophic biomolecules. Therefore, it has been increasingly important to characterize the MSC secretome. It has been shown that the effect of the MSCs is strongly influenced by the environment in the host compartment, and it is a crucial issue when considering MSC therapy. The aim of this study was to investigate differences in the in vitro secreted protein profile between naïve and chondrogenic differentiating bone marrow-derived (BM)-MSCs when exposed to an inflammatory environment.
Equine BM-MSCs were divided into a naïve group and a chondrogenic group. Cells were treated with normal expansion media or chondrogenic media. Cells were treated with IL-1β for a period of 5 days (stimulation), followed by 5 days without IL-1β (recovery). Media were collected after 48 h and 10 days. The secretomes were digested and analyzed by nanoLC-MS/MS to unravel the orchestration of proteins.
The inflammatory proteins IL6, CXCL1, CXCL6, CCL7, SEMA7A, SAA, and haptoglobin were identified in the secretome after 48 h from all cells stimulated with IL-1β. CXCL8, OSM, TGF-β1, the angiogenic proteins VCAM1, ICAM1, VEGFA, and VEGFC, the proteases MMP1 and MMP3, and the protease inhibitor TIMP3 were among the proteins only identified in the secretome after 48 h from cells cultured in normal expansion media. After 10-day incubation, the proteins CXCL1, CXCL6, and CCL7 were still identified in the secretome from BM-MSCs stimulated with IL-1β, but the essential inducer of inflammation, IL6, was only identified in the secretome from cells cultured in normal expansion media.
The findings in this study indicate that naïve BM-MSCs have a more extensive inflammatory response at 48 h to stimulation with IL-1β compared to BM-MSCs undergoing chondrogenic differentiation. This extensive inflammatory response decreased after 5 days without IL-1β (day 10), but a difference in composition of the secretome between naïve and chondrogenic BM-MSCs was still evident.
与人类相似，马是一种长寿且运动能力强的物种。间充质基质细胞（MSCs）的使用是一个相对较新的领域，但已在治疗关节疾病（如骨关节炎）方面取得了有希望的效果。据信，MSCs 通过分泌营养生物分子发挥其主要治疗作用。因此，表征 MSC 分泌组变得越来越重要。已经表明，MSCs 的作用受宿主环境中环境的强烈影响，当考虑 MSC 治疗时，这是一个关键问题。本研究的目的是研究在暴露于炎症环境时，幼稚和软骨分化骨髓源性（BM）-MSCs 之间体外分泌蛋白谱的差异。
将马 BM-MSCs 分为幼稚组和软骨分化组。细胞用正常扩增培养基或软骨分化培养基处理。细胞用 IL-1β 处理 5 天（刺激），然后无 IL-1β 处理 5 天（恢复）。在 48 小时和 10 天后收集培养基。分泌组通过 nanoLC-MS/MS 消化和分析以揭示蛋白质的协调作用。
在所有用 IL-1β 刺激的细胞的 48 小时后，在分泌组中鉴定出炎症蛋白 IL6、CXCL1、CXCL6、CCL7、SEMA7A、SAA 和触珠蛋白。在正常扩增培养基中培养的细胞的 48 小时后，分泌组中仅鉴定出 CXCL8、OSM、TGF-β1、血管生成蛋白 VCAM1、ICAM1、VEGFA 和 VEGFC、蛋白酶 MMP1 和 MMP3 以及蛋白酶抑制剂 TIMP3。孵育 10 天后，在 IL-1β 刺激的 BM-MSCs 的分泌组中仍鉴定出 CXCL1、CXCL6 和 CCL7，但在正常扩增培养基中培养的细胞的分泌组中仅鉴定出炎症的主要诱导剂 IL6。
本研究的结果表明，与正在进行软骨分化的 BM-MSCs 相比，幼稚 BM-MSCs 在 48 小时时对 IL-1β 的刺激有更广泛的炎症反应。在没有 IL-1β 的 5 天后（第 10 天），这种广泛的炎症反应会减弱，但幼稚和软骨分化的 BM-MSCs 之间的分泌组组成仍存在差异。
