Lymphotixin ? Receptor-mediated NF?B Signaling Promotes Glial Lineage Differentiation And Inhibites Neuronal Lineage Differentiation In Mouse Brain Neural Stem/progenitor Cells

BackgroundLymphotoxin(LT)is a lymphokine mainly expressed in lymphocytes.LT? binds one or two membraneassociated LT? to form LT?2?1 or LT?1?2heterotrimers.The predominant LT?1?2 binds to LT? receptor(LT?R)primarily expressed in epithelial and stromal cells.Most studies on LT?R signaling have focused on the organization,development,and maintenance of lymphoid tissues.However,the roles of LT?R signaling in the nervous system,particularly in neurogenesis,remain unknown.Here,we investigated the role of LT?R-mediated NF?B signaling in regulating neural lineage differentiation.Methods: The C57BL/6J wild-type and GFAP-dnI?B? transgenic mice were used.Serum-free embryoid bodies were cultured from mouse embryonic stem cells and further induced into neural stem/progenitor cells(NSCs/NPCs).Primary neurospheres were cultured from embryonic andadult mouse brains followed by monolayer culture for amplification/passage.NF?B activation was determined by adenovirus-mediated NF?B-firefly-luciferase reporter assay and p65/RelB/p52 nuclear translocation assay.LT?R mRNA expression was evaluated by quantitative RT-PCR and LT?R protein expression was determined by immunohistochemistry and Western blot analysis.Multilabeled immunocytochemistry or immunohistochemistry followed by fluorescent confocal microscopy and quantitative analysis of neural lineage differentiation were performed.Graphing and statistical analysis were performed with GraphPad Prism software.ResultsIn cultured NSCs/NPCs,LT?1?2 stimulation induced an activation of classical and non-classical NF?B signaling.The expression of LT?R-like immunoreactivity in GFAP+/Sox2+ NSCs was identified in well-established neurogenic zones of adult mouse brain.Quantitative RT-PCR and Western blot analysis validated the expression of LT?R in cultured NSCs/NPCs and brain neurogenic regions.LT?R expression was significantly increased during neural induction.LT?1?2 stimulation in cultured NSCs/ NPCs promoted astroglial and oligodendrocytic lineage differentiation,but inhibited neuronal lineage differentiation.Astroglial NF?B inactivation in GFAP-dnI?B? transgenic mice rescued LT?R-mediated abnormal phenotypes of cultured NSCs/NPCs.ConclusionThis study provides the first evidence for the expression and function of LT?R signaling in NSCs/NPCs.Activation of LT?R signaling promotes glial lineage differentiation.Our results suggest that neurogenesis is regulated by the adaptive immunity and inflammatory responses.