Microglia-derived TNF-α mediates Muller cell activation by activating the TNFR1-NF-κB pathway

Microglia and its interaction with Muller cells are responsible to retinal surveillance during retinal neuro-degeneration, however, the role and mechanism of microglia-derived tumor necrosis factor (TNF)-alpha in the activation of retinal Muller cells have not been fully elucidated. In the present study, primary microglia and Muller cells were isolated from newborn Sprague-Dawley (SD) rats with purities of 88.2 +/- 6.2% and 92.2 +/- 2.2%, respectively. By performing immunofluorescence and Western blot analysis, we found that TNF receptor (TNFR)-1 and TNFR2 were expressed in Muller cells. After co-cultured with microglia-conditioned medium (MCM), the elevated mRNA levels of glial fibrillary acidic protein (GFAP), proinflammatory factors (TNF-alpha, IL-1 beta, CXCL-1, CSF-1, NOS2, COX2) and decreased CNTF mRNA levels were found in Muller cells. However, pretreatment with R-7050 (a TNF-alpha receptor inhibitor) or anti-TNFR1 significantly abrogated the changes. Simultaneously, pretreatment with anti-TNFR2 slightly inhibited the expression of GFAP in MCM-incubated Muller cells. Meanwhile, anti-TNFR1 treatment reversed the increased expression of CSF-1 and IL-1 beta induced by TNF-alpha. Compared to the control groups, the phosphorylation of NF-kappa B P65, MAPK P38 and ERK1/2 in TNF-alpha-treated Muller cells was significantly increased. Nevertheless, pretreatment with anti-TNFR1 inhibited the phosphorylation of NF-kappa B P65 and MAPK p38, especially NF-kappa B P65. Additionally, pretreatment with Bay117082 (an NF-kappa B inhibitor) also significantly inhibited NF-kappa B P65 phosphorylation and GFAP expression. Moreover, anti-TNFR1 and Bay117082 treatment reduced NF-kappa B P65 phosphorylation of Muller cells induced by MCM. These results suggested that microglia-derived TNF-alpha served as a vital role in regulating Muller cells activation during retinal neurodegeneration.

Automated summary

The study revealed that microglia-derived TNF-α plays a crucial role in regulating the activation of Muller cells during retinal neurodegeneration. TNF-α led to increased expression of GFAP and proinflammatory factors in Muller cells, while anti-TNFR1 and NF-κB inhibitor could reverse these changes.