Background

Oligodendrocytes, which form myelin, enable rapid and efficient nerve conduction. Destruction of myelin causes demyelinating diseases such as multiple sclerosis. Primary oligodendrocyte progenitor cells (OPCs) from postnatal rodents have been utilized to elucidate the developmental mechanism of oligodendrocytes in vitro. However, this process is complicated and takes up to several weeks.

New method

We established a method to culture OPCs from neonatal rat brain in DMEM/F-12 with Stem-Pro, bFGF (10 ng/mL), and rhPDGF (30 ng/mL). The culture, without shaking or immunopanning, became OPC-enriched rather than a mixed glial culture.

Results

Immunofluorescent analysis using cell lineage markers suggested that these cells were initially glial progenitors, which gradually changed to OPCs with a few cells further differentiating into oligodendrocytes. Using compounds that promote OPC differentiation, we confirmed that these cells were compatible for high-throughput screening in a 96-well plate format. In co-culture with dorsal root ganglion neuron, OPCs showed myelin sheath-like morphologies. This method was also applicable to mouse OPCs.

Comparison with existing methods

Although the purity of the OPCs was not comparable to that after immunopanning, most cells were of the oligodendrocyte lineage at 8 DIV, while less than 10% were astrocytes. This method requires mediums with only two growth factors without any specific equipment like antibodies or magnet and takes simple procedures.

Conclusions

The simplicity and high yield of our method make it a good choice when working with oligodendrocytes/OPCs. We believe that this method is an affordable protocol for various biological applications without any special techniques or equipment.

中文翻译：

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从啮齿动物大脑制备少突胶质祖细胞的简单一步法的建立。

背景

形成髓磷脂的少突胶质细胞能够快速有效地传导神经。破坏髓磷脂会引起脱髓鞘疾病，例如多发性硬化症。来自产后啮齿动物的原代少突胶质祖细胞（OPC）已被用于阐明少突胶质细胞的体外发育机制。但是，此过程很复杂，可能需要数周的时间。

新方法

我们建立了一种用Stem-Pro，bFGF（10 ng / mL）和rhPDGF（30 ng / mL）在DMEM / F-12中培养新生大鼠脑中的OPC的方法。该培养物无需摇动或进行免疫淘选就变得富含OPC，而不是混合的神经胶质培养物。

结果

使用细胞谱系标记的免疫荧光分析表明，这些细胞最初是神经胶质祖细胞，逐渐转变为OPC，少数细胞进一步分化为少突胶质细胞。使用促进OPC分化的化合物，我们证实了这些细胞可用于96孔板格式的高通量筛选。在与背根神经节神经元的共培养中，OPCs显示出髓鞘样形态。此方法也适用于鼠标OPC。

与现有方法的比较

尽管OPC的纯度不能与免疫淘洗后的纯度相比，但大多数细胞是8 DIV的少突胶质细胞谱系，而星形胶质细胞不到10％。该方法只需要具有两个生长因子的培养基，而无需任何特异性设备（如抗体或磁铁），并且操作简单。

结论

我们的方法的简便性和高产率使其成为处理少突胶质细胞/ OPC的理想选择。我们认为，这种方法对于各种生物学应用而言都是一种经济实惠的方案，而无需任何特殊技术或设备。