The Effects Of M1/M2Inflammatory Environment On The Differentiation Of Neural Stem Cells From Spinal Cord

The endogenous environment caused by spinal cord injury (SCI) is notsufficient to support the neural stem cells (NSCs) differentiation to neurons, sothat the replacement of dead neurons with NSC transplantation at the lesion sitedoes not normally occur. Different from the brain which NSCs mainly reside insubventricular zone (SVZ) and subgranular zone (SGZ), NSCs in spinal cordlocate in ependymal of central canal. After SCI, most of the spinal cord NSCsdifferentiate into astrocytes which lead to the formation of the glial scarinhibiting the axon growth and functional recovery.Inflammation is one of the most important pathological processes in SCI.Macrophage/microglia is the most important inflammatory cells whichdetermines mainly the injured environment after SCI. Macrophages can beactivated and differentiated into M1/M2macrophages which possess differentfunctions. M1macrophages release pro-inflammatory factors, free radical and soon which are detrimental to neurons and glial cells. However, M2macrophages can promote tissue recovery by secreting inhibiting inflammatory factors whichsuppress pro-inflammation cascades. So, different inflammatory environmentmay influence the recovery of SCI.Whether has inflammation effects on the differentiation of endogenousNSCs in spinal cord? So our experiments pay more attention to the role ofdifferent types of inflammation in NSCs differentiation in vitro. By doing so,we’re looking forward to find enrichment environment for spinal cord NSC toneuronal differentiation in vivo.Firstly, we polarized macrophage, N9microglia cell line and spinal cordprimary microglia with LPS+INF-γ and IL-4through “classical activation” and“alternative activation”, respectively. Then we detect the specific genes orproteins expression of M1/M2type cells by qPCR and Western blot. So we canjudge their polarized states by two different polarization pathways. Third, inensuring the successful polarization, we prepared M1/M2conditioned mediumrespectively. After that, we cultured spinal cord NSCs in vitro with M1or M2conditioned medium for5days. Finally, using immunohistochemistry staining,qPCR and Western blot to evaluate the differentiation of spinal cord NSCs indifferent inflammation environment.Our present date shows that primary bone marrow macrophages, N9microglia and spinal cord primary microglia all can be polarized towards M1macrophages stimulated24hours by “classical activation” pathway. We alsofind that spinal cord NSCs mainly differentiate into astrocytes in the M1inflammatory environment instead of neurons. By contrast, spinal cord stemcells differentiate significantly towards neurons and oligodendrocytes. Toconclusion, our study hints that M1inflammation environment promotes thedifferentiation of spinal cord NSCs mainly towards astrocytes, wheares, M2 inflammation environment principally contributes to the differentiation ofneurons and oligodendrocytes.