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. Author manuscript; available in PMC: 2012 Jan 26.
Published in final edited form as: Mol Cell. 2010 Dec 22;40(6):893–904. doi: 10.1016/j.molcel.2010.12.013

Figure 7. MnSOD dismutase activity, lysine 122 acetylation, and the cellular metabolic and damage/cytotoxic response are altered by IR.

Figure 7

(a) Sirt3+/+ and Sirt3−/− mice at 12 weeks were exposed to sham or 2 × 2 Gy exposure every 24 hours. Livers were harvested 20 hours post-exposure, and mitochondria were isolated. MnSOD activity in MEFs was measured via a competitive inhibition assay as described (Spitz and Oberley, 1989). Activity data for MnSOD is presented as units of SOD activity per milligram of protein. (b) Mitochondrial extracts from above were analyzed for acetylation of MnSOD lysine 122 via mass spectrometry. Results are presented as fold change from the untreated, wild-type mouse livers. (c) Sirt3−/− mouse livers exposed to IR exhibit marked cytoplasmic vacuolation of periportal to midzonal hepatocytes. The wild-type and Sirt3−/− livers without and with exposure to IR were H & E stained and scored for degree of hepatocellular cytoplasmic vacuolation; low, medium and high by a pathologist (AKO). Representative micrographs are shown. Scale bar = 80 μm. (d) Quantification of the H & E for liver cells. Liver cells were scored as low = no detectable cytoplasmic vacuolation, med = moderate dilation of the cytoplasm by clear space primarily affecting periportal hepatocytes, high = severe dilation of the cytoplasm by clear space and poorly defined clear vacuoles primarily affecting periportal to midzonal hepatocytes. (e) Apoptosis was determined in the Sirt3+/+ and Sirt3−/− livers exposed to IR. Apoptotic cells were identified in liver sections by TUNEL assay. Sections were scored by a pathologist blinded to the groupings. TUNEL-positive cells were counted in 10 randomly selected 200X fields per liver section. Results in this figure are reported as the average number of positive cells per field. Data are presented as the average +/− SD. * indicates P < 0.05 and ** indicates P < 0.01. (f) Irradiated Sirt3−/− mouse liver cells exhibit increased anti-nitrotyrosine IHC staining. Liver tissue from wild-type and Sirt3−/− mice were stained with an anti-nitrotyrosine antibody (StressMarq Biosciences Inc.). A representative micrograph is shown. Scale bar = 80 μm. See Figure S7E for quantification.