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. Author manuscript; available in PMC: 2011 Jul 23.
Published in final edited form as: Cell. 2010 Jul 23;142(2):320–332. doi: 10.1016/j.cell.2010.06.020

Figure 7. SIRT1 suppresses Aβ production by activating ADAM10.

Figure 7

(A) N2A cells stably overexpressing APPswe/PSEN1dE9 transgenes transfected with SIRT1 (mSIRT1). 24 hr after SIRT1 transfection, cells were transfected with 3 different ADAM10-shRNA vectors (1, 2 or 3). N2A cells not overexpressing SIRT1 were also transfected with 3 different ADAM10-shRNA vectors (1, 2 or 3) to silence ADAM10. Separately, SIRT1 was silenced by 3 different shRNAs (1, 2 or 3). 48 hr later, Aβ 1-42 concentration in the conditioned medium was assessed by ELISA (Experimental Procedures). N2A cells without APPswe,PSEN1dE9 transgenes showed no detectable Aβ 1-42 in this assay (not shown). (B) Western blotting of the extracts of these cells using anti-ADAM10 antibody. Quantification of the ADAM10-P (top) and ADAM10 (bottom) were shown on the right as percent of control. (C) Western blotting of the extracts of the cells using anti-NICD antibody. Quantification of the NICD are shown on the right as percent of control. (D) HES1 RNA levels were determined by qPCR from RNA extracted from cells. (E) N2A cells stably overexpressing APPswe/PSEN1dE9 were transfected with SIRT1 (mSIRT1). 24 hr later, cells were incubated with RA (Retinoic Acid), LE-135 or DMSO for an additional 24 hr and assayed by ELISA. (see also Figure S6 and S7)