ND02
Role of Histone Deacetylase SIRT1 in C5b-9 Mediated Oligodendrocyte Cell Cycle Activation
SIRT1 is an NAD-dependent histone deacetylase that is involved in the regulation of transcription, apoptosis, and metabolism. Previous studies have shown that SIRT1 regulates the differentiation of neural progenitor cells; however, the exact mechanism is still unknown. We have previously demonstrated that sublytic levels of complement C5b- 9 terminal complex increased the survival of oligodendrocytes (OLGs) and induced their dedifferentiation. We have now investigated the role of SIRT1 in sublytic C5b-9 induced cell cycle activation.
Objectives:
Our aim was to investigate the role of SIRT1 in OLGs differentiation and to investigate if sublytic levels of C5b-9 modulate SIRT1 expression and cell cycle activation. We also investigated the downstream effects of SIRT1 by measuring histone H3 Lysine 9 trimethylation (H3K9me3) and expression of cyclin D1.
Methods:
OLG progenitor cells were purified from the brain of rat pups and differentiated in vitro using differentiation media. We stimulated OLGs with C5b-9 or C5b6 for 3, 6 and 18 h to determine the effect of the sublytic C5b-9 terminal complement on SIRT1 expression. We measured SIRT1 mRNA using real-time PCR. SIRT1, cyclin D1, and H3K9me3 protein expression in OLGs was measured using western blotting.
Results:
SIRT1 mRNA and protein expression decreased during in vitro differentiation of OLGs. H3K9me3 levels were higher in progenitor cells and declined during differentiation. Cyclin D1 expression transiently increased during differentiation. Stimulation of mature OLGs with sublytic C5b-9 for 3h resulted in a significant decrease in SIRT1 mRNA expression and protein levels while stimulation with C5b6 had no effect. SIRT1 expression levels in OLGs after 8h of exposure to C5b-9 were significantly lower than in C5b6 treated cells. H3K9me3 expression levels were high in unstimulated OLGs and decreased significantly after stimulation with C5b-9. C5b6 treatment had no effect on H3K9me3, with levels being similar to unstimulated OLGs. Cyclin D1 expression levels in OLGs increased after stimulation with C5b-9, indicating cell cycle activation.
Conclusions:
Our data indicate that C5b-9 stimulation of OLGs reduces SIRT1 expression. Decreased levels of SIRT1 contribute to cell cycle activation by decreasing trimethylation of histone H3 lysine 9.