Sarkar Souvarish, Malovic Emir, Plante Brandon, Zenitsky Gary, Jin Huajun, Anantharam Vellareddy, Kanthasamy Arthi, Kanthasamy Anumantha G
Biomedical Sciences & Iowa Center for Advanced Neurotoxicology, Iowa State University.
Biomedical Sciences & Iowa Center for Advanced Neurotoxicology, Iowa State University;
J Vis Exp. 2017 Apr 13(122):55364. doi: 10.3791/55364.
Microglia are the primary responders to central nervous system insults; however, much remains unknown about their role in regulating neuroinflammation. Microglia are mesodermal cells that function similarly to macrophages in surveying inflammatory stress. The classical (M1-type) and alternative (M2-type) activations of macrophages have also been extended to microglia in an effort to better understand the underlying interplay these phenotypes have in neuroinflammatory conditions such as Parkinson's, Alzheimer's, and Huntington's Diseases. In vitro experimentation utilizing primary microglia offers rapid and reliable results that may be extended to the in vivo environment. Although this is a clear advantage over in vivo experimentation, isolating microglia while achieving adequate yields of optimal purity has been a challenge. Common methods currently in use either suffer from low recovery, low purity, or both. Herein, we demonstrate a refinement of the column-free CD11b magnetic separation method that achieves a high cell recovery and enhanced purity in half the amount of time. We propose this optimized method as a highly useful model of primary microglial isolation for the purposes of studying neuroinflammation and neurodegeneration.
小胶质细胞是中枢神经系统损伤的主要应答者;然而,关于它们在调节神经炎症中的作用仍有许多未知之处。小胶质细胞是中胚层细胞,在监测炎症应激方面的功能与巨噬细胞相似。巨噬细胞的经典(M1型)和替代性(M2型)激活也已扩展到小胶质细胞,以便更好地理解这些表型在帕金森病、阿尔茨海默病和亨廷顿病等神经炎症性疾病中的潜在相互作用。利用原代小胶质细胞进行的体外实验提供了快速可靠的结果,这些结果可能适用于体内环境。虽然这相对于体内实验是一个明显的优势,但在获得足够产量的最佳纯度的同时分离小胶质细胞一直是一个挑战。目前常用的方法要么回收率低,要么纯度低,要么两者都存在问题。在此,我们展示了一种无柱CD11b磁性分离方法的改进,该方法能在一半的时间内实现高细胞回收率和更高的纯度。我们提出这种优化方法作为研究神经炎症和神经退行性变的原代小胶质细胞分离的高度有用模型。