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Cladribine modifies functional properties of microglia.
Clinical & Experimental Immunology ( IF 3.4 ) Pub Date : 2020-06-03 , DOI: 10.1111/cei.13473 L Ø Jørgensen 1, 2, 3 , K H Hyrlov 1, 2, 3 , M L Elkjaer 1, 2, 3 , A B Weber 1, 2, 3 , A E Pedersen 4, 5 , Å Fex Svenningsen 2, 3 , Z Illes 1, 2, 3
Clinical & Experimental Immunology ( IF 3.4 ) Pub Date : 2020-06-03 , DOI: 10.1111/cei.13473 L Ø Jørgensen 1, 2, 3 , K H Hyrlov 1, 2, 3 , M L Elkjaer 1, 2, 3 , A B Weber 1, 2, 3 , A E Pedersen 4, 5 , Å Fex Svenningsen 2, 3 , Z Illes 1, 2, 3
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Cladribine (CdA), an oral prodrug approved for the treatment of relapsing multiple sclerosis, selectively depletes lymphocytes. CdA passes the blood–brain barrier, suggesting a potential effect on central nervous system (CNS) resident cells. We examined if CdA modifies the phenotype and function of naive and activated primary mouse microglia, when applied in the concentrations 0·1–1 μM that putatively overlap human cerebrospinal fluid (CSF) concentrations. Primary microglia cultures without stimulation or in the presence of proinflammatory lipopolysaccharide (LPS) or anti‐inflammatory interleukin (IL)‐4 were treated with different concentrations of CdA for 24 h. Viability was assessed by MTT [3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide] assay. Phagocytotic ability and morphology were examined by flow cytometry and random migration using IncuCyte Zoom and TrackMate. Change in gene expression was examined by quantitative polymerase chain reaction (qPCR) and protein secretion by Meso Scale Discovery. We found that LPS and IL‐4 up‐regulated deoxycytidine kinase (DCK) expression. Only activated microglia were affected by CdA, and this was unrelated to viability. CdA 0·1–1 μM significantly reduced granularity, phagocytotic ability and random migration of activated microglia. CdA 10 μM increased the IL‐4‐induced gene expression of arginase 1 (Arg1) and LPS‐induced expression of IL‐1β, tumor necrosis factor (TNF), inducible nitric oxide synthase (iNOS) and Arg1, but protein secretion remained unaffected. CdA 10 μM potentiated the increased expression of anti‐inflammatory TNF receptor 2 (TNF‐R2) but not TNF‐R1 induced by LPS. This suggests that microglia acquire a less activated phenotype when treated with 0·1–1 μM CdA that putatively overlaps human CSF concentrations. This may be related to the up‐regulated gene expression of DCK upon activation, and suggests a potential alternative mechanism of CdA with direct effect on CNS resident cells.
中文翻译：
克拉屈滨修饰小胶质细胞的功能特性。
克拉屈滨（CdA）是一种经批准用于治疗复发性多发性硬化症的口服前药，可选择性消耗淋巴细胞。CdA通过血脑屏障，表明对中枢神经系统（CNS）驻留细胞有潜在影响。我们检查了当以0·1-1μM的浓度（假定与人脑脊髓液（CSF）的浓度重叠）施用时，CdA是否能修饰幼稚和激活的原代小鼠小胶质细胞的表型和功能。用不同浓度的CdA处理无刺激或存在促炎性脂多糖（LPS）或抗炎性白介素（IL）-4的原发性小胶质细胞培养物24小时。通过MTT [3-（4,5-二甲基噻唑-2-基）-2,5-二苯基溴化四唑鎓]测定活力。使用IncuCyte Zoom和TrackMate通过流式细胞术和随机迁移检查吞噬能力和形态。通过定量聚合酶链反应（qPCR）和Meso Scale Discovery的蛋白质分泌检查基因表达的变化。我们发现LPS和IL-4上调了脱氧胞苷激酶（DCK）的表达。CdA仅影响激活的小胶质细胞，这与生存能力无关。CdA 0·1-1μM会显着降低活化小胶质细胞的颗粒度，吞噬能力和随机迁移。CdA 10μM增加了IL-4诱导的精氨酸酶1（Arg1）的基因表达和LPS诱导的IL-1β，肿瘤坏死因子（TNF），诱导型一氧化氮合酶（iNOS）和Arg1的表达，但蛋白质分泌仍然不受影响。CdA 10μM增强了LPS诱导的抗炎TNF受体2（TNF-R2）的表达增加，但不增强。这表明当用0·1-1μMCdA处理时，小胶质细胞的活化表型较少，该CdA可能与人类CSF浓度重叠。这可能与激活后DCK的基因表达上调有关，并暗示了直接影响CNS驻留细胞的CdA潜在替代机制。
更新日期：2020-08-12
中文翻译：
克拉屈滨修饰小胶质细胞的功能特性。
克拉屈滨（CdA）是一种经批准用于治疗复发性多发性硬化症的口服前药，可选择性消耗淋巴细胞。CdA通过血脑屏障，表明对中枢神经系统（CNS）驻留细胞有潜在影响。我们检查了当以0·1-1μM的浓度（假定与人脑脊髓液（CSF）的浓度重叠）施用时，CdA是否能修饰幼稚和激活的原代小鼠小胶质细胞的表型和功能。用不同浓度的CdA处理无刺激或存在促炎性脂多糖（LPS）或抗炎性白介素（IL）-4的原发性小胶质细胞培养物24小时。通过MTT [3-（4,5-二甲基噻唑-2-基）-2,5-二苯基溴化四唑鎓]测定活力。使用IncuCyte Zoom和TrackMate通过流式细胞术和随机迁移检查吞噬能力和形态。通过定量聚合酶链反应（qPCR）和Meso Scale Discovery的蛋白质分泌检查基因表达的变化。我们发现LPS和IL-4上调了脱氧胞苷激酶（DCK）的表达。CdA仅影响激活的小胶质细胞，这与生存能力无关。CdA 0·1-1μM会显着降低活化小胶质细胞的颗粒度，吞噬能力和随机迁移。CdA 10μM增加了IL-4诱导的精氨酸酶1（Arg1）的基因表达和LPS诱导的IL-1β，肿瘤坏死因子（TNF），诱导型一氧化氮合酶（iNOS）和Arg1的表达，但蛋白质分泌仍然不受影响。CdA 10μM增强了LPS诱导的抗炎TNF受体2（TNF-R2）的表达增加，但不增强。这表明当用0·1-1μMCdA处理时，小胶质细胞的活化表型较少，该CdA可能与人类CSF浓度重叠。这可能与激活后DCK的基因表达上调有关，并暗示了直接影响CNS驻留细胞的CdA潜在替代机制。
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