Redondo-Castro Elena, Cunningham Catriona, Miller Jonjo, Martuscelli Licia, Aoulad-Ali Sarah, Rothwell Nancy J, Kielty Cay M, Allan Stuart M, Pinteaux Emmanuel
Faculty of Biology, Medicine and Health, University of Manchester, Manchester, UK.
Wellcome Trust Centre for Cell-Matrix Research, Faculty of Biology, Medicine and Health, University of Manchester, Manchester, UK.
Stem Cell Res Ther. 2017 Apr 17;8(1):79. doi: 10.1186/s13287-017-0531-4.
Inflammation is a key contributor to central nervous system (CNS) injury such as stroke, and is a major target for therapeutic intervention. Effective treatments for CNS injuries are limited and applicable to only a minority of patients. Stem cell-based therapies are increasingly considered for the treatment of CNS disease, because they can be used as in-situ regulators of inflammation, and improve tissue repair and recovery. One promising option is the use of bone marrow-derived mesenchymal stem cells (MSCs), which can secrete anti-inflammatory and trophic factors, can migrate towards inflamed and injured sites or can be implanted locally. Here we tested the hypothesis that pre-treatment with inflammatory cytokines can prime MSCs towards an anti-inflammatory and pro-trophic phenotype in vitro.
Human MSCs from three different donors were cultured in vitro and treated with inflammatory mediators as follows: interleukin (IL)-1α, IL-1β, tumour necrosis factor alpha (TNF-α) or interferon-γ. After 24 h of treatment, cell supernatants were analysed by ELISA for expression of granulocyte-colony stimulating factor (G-CSF), IL-10, brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), IL-1 receptor antagonist (IL-1Ra) and vascular endothelial growth factor (VEGF). To confirm the anti-inflammatory potential of MSCs, immortalised mouse microglial BV2 cells were treated with bacterial lipopolysaccharide (LPS) and exposed to conditioned media (CM) of naïve or IL-1-primed MSCs, and levels of secreted microglial-derived inflammatory mediators including TNF-α, IL-10, G-CSF and IL-6 were measured by ELISA.
Unstimulated MSCs constitutively expressed anti-inflammatory cytokines and trophic factors (IL-10, VEGF, BDNF, G-CSF, NGF and IL-1Ra). MSCs primed with IL-1α or IL-1β showed increased secretion of G-CSF, which was blocked by IL-1Ra. Furthermore, LPS-treated BV2 cells secreted less inflammatory and apoptotic markers, and showed increased secretion of the anti-inflammatory IL-10 in response to treatment with CM of IL-1-primed MSCs compared with CM of unprimed MSCs.
Our results demonstrate that priming MSCs with IL-1 increases expression of trophic factor G-CSF through an IL-1 receptor type 1 (IL-1R1) mechanism, and induces a reduction in the secretion of inflammatory mediators in LPS-activated microglial cells. The results therefore support the potential use of preconditioning treatments of stem cells in future therapies.
炎症是导致中风等中枢神经系统（CNS）损伤的关键因素，也是治疗干预的主要靶点。中枢神经系统损伤的有效治疗方法有限，且仅适用于少数患者。基于干细胞的疗法越来越多地被考虑用于治疗中枢神经系统疾病，因为它们可以作为炎症的原位调节剂，促进组织修复和恢复。一种有前景的选择是使用骨髓间充质干细胞（MSC），其可分泌抗炎和营养因子，能向炎症和损伤部位迁移或可局部植入。在此，我们测试了如下假设：用炎性细胞因子预处理可使间充质干细胞在体外转变为抗炎和促营养表型。
来自三个不同供体的人骨髓间充质干细胞在体外培养，并用以下炎性介质处理：白细胞介素（IL）-1α、IL-1β、肿瘤坏死因子α（TNF-α）或干扰素-γ。处理24小时后，通过酶联免疫吸附测定（ELISA）分析细胞上清液中粒细胞集落刺激因子（G-CSF）、IL-10、脑源性神经营养因子（BDNF）、神经生长因子（NGF）、IL-1受体拮抗剂（IL-1Ra）和血管内皮生长因子（VEGF）的表达。为确认间充质干细胞的抗炎潜力，用细菌脂多糖（LPS）处理永生化小鼠小胶质细胞BV2，并使其暴露于未处理或经IL-1预处理的间充质干细胞的条件培养基（CM）中，通过ELISA测定分泌的小胶质细胞源性炎性介质（包括TNF-α、IL-10、G-CSF和IL-6）水平。
未受刺激的间充质干细胞组成性地表达抗炎细胞因子和营养因子（IL-10、VEGF、BDNF、G-CSF、NGF和IL-1Ra）。用IL-1α或IL-1β预处理的间充质干细胞显示G-CSF分泌增加，这被IL-1Ra阻断。此外，与未预处理的间充质干细胞的条件培养基相比，LPS处理的BV2细胞分泌的炎性和凋亡标志物减少，且在用经IL-1预处理的间充质干细胞的条件培养基处理后，其抗炎性IL-10的分泌增加。
我们的结果表明，用IL-1预处理间充质干细胞可通过1型IL-1受体（IL-1R1）机制增加营养因子G-CSF的表达，并诱导LPS激活的小胶质细胞中炎性介质分泌减少。因此，这些结果支持了干细胞预处理疗法在未来治疗中的潜在应用。
