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. 2010 Jun 15;12(12):1339–1353. doi: 10.1089/ars.2009.2791

FIG. 2.

FIG. 2.

Trx1 content, and Trx and TR activities show little if any change with age in heart homogenates and in isolated myocytes from F344 rats. (A) (Left panel) Western blot analysis was performed using anti-Trx1 antibody in heart homogenates from adult (6-mo) and elderly (24-mo) F344 rats. Representative Western blots from two sets of rat homogenates for each age group (6-mo (1), (2) and 24-mo(1), (2)) are shown. Trx1 content is normalized to GAPDH (loading control) and the bar graph is expressed as a ratio of the relative quantities of Trx1 in the heart homogenates from elderly rats as compared to adult rats. Data represent mean ± SEM (n = 11 for 6-mo and n = 13 for 24-mo). (Right panel) Representative Western blot showing unchanged Trx1 content in a pool of six lysates each of cardiomyocytes isolated from young adult (6-mo) and elderly (24-mo) Fischer 344 rats. The bar graph represents the mean Trx1/Src ratio (normalized to 100%) of two separate Western blot analyses of three different sets of pooled lysates corresponding to cardiomyocytes isolated from young adult and elderly rats. One type of pool was made from three individual preparations of cardiomyocytes of each age group, lysed separately in RIPA buffer, frozen at − 80°C, then thawed and combined for analysis. The second type of pool of three different individual cardiomyocyte preparations were lysed in NP-40 buffer and processed analogously. The third type of pool contained equal portions of pool type 1 and pool type 2 for each age group. Although the data represent a composite contributed by six individual biological samples of each age group, they were analyzed as an “n = 3.” (B) Total Trx and TR activities in homogenates isolated from 6-mo and 24-mo F344 rat hearts were determined as described in Materials and Methods. Activities in elderly hearts (mean ± SEM = 1.42 ± 0.39 nmol thiol/min/mg for Trx; 1.63 ± 0.08 μmol NADPH oxidized/min/ml for TR) were normalized to the corresponding activities observed in adult hearts (mean ± SEM = 1.85 ± 0.44 nmol thiol/min/mg total protein for Trx; 1.68 ± 0.13 μmol NADPH oxidized/min/ml for TR). Open bars (□), adult; closed bars (▪), elderly. No significant differences (elderly vs. adult) were found for the Trx or TR activities; n = 4 for Trx, and n = 3 for TR.