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Qi Jiang, John G Flanagan, Darren Chan, Jeremy M Sivak; The Protective Role of AMPK/SIRT1/PGC-1α Signalling in Retinal Astrocytes. Invest. Ophthalmol. Vis. Sci. 2014;55(13):2270.
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The transcriptional co-activator PGC-1α (peroxisome proliferator-activated receptor gamma coactivator-1 alpha) is a key regulator of energy metabolism and oxidative stress response. When activated by upstream regulators such as AMP kinase (AMPK) and sirtuin 1 (SIRT1), PGC-1α modulates the expression of genes including those involved in antioxidant defense. We have previously found that the loss of PGC-1α compromised retinal ganglion cell (RGC) homeostasis and induced pathological astrocyte reactivity. We aim to determine whether promoting activity through the AMPK/SIRT1/PGC-1α pathway in astrocytes can increase their resistance to oxidative stress and ultimately enhance their ability to support the RGCs.
In vitro experiments were performed using primary rat retinal astrocytes and an astrocytic cell line (A7). After exposure to oxidative stress, the level or activation of AMPK, SIRT1, and PGC-1α was determined by Western blot. To evaluate whether stimulating the pathway offers protection, pharmacological agonists of AMPK (AICA-Riboside, AICAR) and SIRT1 (SRT1720) were applied to A7 cells under oxidative stress. Cell viability was then assessed by a colorimetric XTT assay. Subsequently, the expression of several PGC-1α-regulated genes was analyzed after treatment to determine downstream targets that are significantly regulated.
Following oxidative stress, increased phosphorylation of AMPK and elevated nuclear levels of SIRT1 and PGC-1α were evident. Treatment with AICAR and SRT1720 was able to significantly improve cell viability in a dose dependent manner (p<0.05). Combined treatment with both compounds further increased viability compared to either alone. Lastly, analysis of gene expression revealed the upregulation of antioxidant genes, including a two- to three-fold increase in superoxide dismutase 1 (SOD1), in response to AMPK and SIRT1 activation.
The AMPK/SIRT1/PGC-1α axis is stimulated when astrocytes are challenged by oxidative stress. Through pharmacological activation of AMPK and SIRT1, astrocytes exhibit increased resistance against oxidative insult through the expression of antioxidant defense genes.
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