Poly(I:C) promotes TNFa/TNFR1-dependent oligodendrocyte death in mixed glial cultures

Steelman, Andrew J; Li, Jianrong

To further determine whether microglia were indeed the TNFα producers and to test whether microgliaderived factors were capable of inducing astroglial TNFα, we performed intracellular cytokine staining for TNFα in mixed cultures treated with poly(I:C). We found that TNFα + cells with microglia morphology were abundant upon stimulation with poly(I:C) or LPS, but were absent in vehicle control treated mixed glial cultures ( Figure 5D ). Immunostaining of poly(I:C)-stimulated cultures with an isotype control antibody did not reveal any non-specific signal (not shown). Importantly, dual staining for TNFα and either GFAP or Iba-1 demonstrated that astrocytes did not produce TNFα following stimulation with LPS or poly(I:C) as TNFα + cells never localized to GFAP + cells ( Figure 5D ). Instead, TNFα immunoreactivity was confined to Iba-1 + cells, indicating microglia as being the sole source of TNFα under these conditions ( Figure 5E ).