A high throughput drug screening assay to identify compounds that promote oligodendrocyte differentiation using acutely dissociated and purified oligodendrocyte precursor cells.
A high throughput drug screening assay to identify compounds that promote oligodendrocyte differentiation using acutely dissociated and purified oligodendrocyte precursor cells.
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DOI：
10.1186/s13104-016-2220-2
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发表时间：
2016-09-05
影响因子：
1.8
通讯作者：
Leonoudakis D
中科院分区：
文献类型：
--
作者：
Lariosa-Willingham KD;Rosler ES;Tung JS;Dugas JC;Collins TL;Leonoudakis D
多发性硬化症是由导致脱髓鞘和神经变性的自身免疫反应引起的。成人中枢神经系统具有部分通过产生新的少突胶质细胞（OL）来使轴突髓鞘再生的能力。为了鉴定可能促进髓鞘再生的临床候选化合物，我们开发了一种高通量筛选（HTS）测定法来鉴定促进少突胶质细胞前体细胞（OPC）分化为 OL 的化合物。使用急性解离和纯化的大鼠 OPC 与免疫荧光图像定量相结合，我们开发了一种 OL 分化测定法。我们已经用已知的分化促进剂甲状腺激素验证了该测定，并随后使用该测定来筛选 NIH 临床收集库。我们已经确定了 27 种热门化合物，并通过剂量反应分析和半数最大有效浓度 (EC50) 值的生成进行了验证，以便对功效进行排名。该测定鉴定了 OL 分化的新启动子，我们将其归因于 (1) 加入 OL 毒性预筛选以降低有毒化合物的浓度，以及 (2) 使用新鲜纯化的非传代 OPC。这些特征使我们的测定与用于药物发现工作的其他 OL 分化测定不同。这种基于 OL 的急性初级分化测定应该对那些有兴趣筛选大型化合物库以鉴定治疗多发性硬化症和其他脱髓鞘疾病的药物的人有用。本文的在线版本 (doi:10.1186/s13104-016-2220-2) 包含补充材料，可供授权用户使用。
Multiple sclerosis is caused by an autoimmune response resulting in demyelination and neural degeneration. The adult central nervous system has the capacity to remyelinate axons in part through the generation of new oligodendrocytes (OLs). To identify clinical candidate compounds that may promote remyelination, we have developed a high throughput screening (HTS) assay to identify compounds that promote the differentiation of oligodendrocyte precursor cells (OPCs) into OLs. Using acutely dissociated and purified rat OPCs coupled with immunofluorescent image quantification, we have developed an OL differentiation assay. We have validated this assay with a known promoter of differentiation, thyroid hormone, and subsequently used the assay to screen the NIH clinical collection library. We have identified twenty-seven hit compounds which were validated by dose response analysis and the generation of half maximal effective concentration (EC50) values allowed for the ranking of efficacy. The assay identified novel promoters of OL differentiation which we attribute to (1) the incorporation of an OL toxicity pre-screen to allow lowering the concentrations of toxic compounds and (2) the utilization of freshly purified, non-passaged OPCs. These features set our assay apart from other OL differentiation assays used for drug discovery efforts. This acute primary OL-based differentiation assay should be of use to those interested in screening large compound libraries for the identification of drugs for the treatment of MS and other demyelinating diseases. The online version of this article (doi:10.1186/s13104-016-2220-2) contains supplementary material, which is available to authorized users.