Senescenceassociated Sa0galactosidase As An Enzymatic Aging Marker

Acidic senescence-associated-0-galactosidase (SA-0-gal) is histochemically detectable at pH 6 upon senescence in cultured cells in vitro and aged skin in vivo (Dimri et al., 1995). This SA-0-gal marker can differentiate between the replicatively senescent state and the simply quiescent state during growth-arrest. This marker has been used to identify senescent cells in selected tissues from several aging animal models in vivo (Cao et al., 2003;

9-Month Old 17-Month Old 22-Month Old 31-Month Old

9-Month Old 17-Month Old 22-Month Old 31-Month Old

Figure 28.1. Whole-body detection of SA-J-gal activity in adult zebrafish of different ages (Kishi et al., 2003). After whole-mount fixation of adult zebrafish with whole body, SA-J-gal staining was performed at pH 6 in 9, 17, 22, and 31-month-old zebrafish samples. As shown in this figure, younger 9-month-old fish exhibited very faint background staining in some abdominal regions, whereas several different levels of staining in individual fish were observed in 17- and 22-month-old fish. The staining patterns of each fish varied in these age groups. Some individuals showed fairly strong staining, while others were relatively variable. On the other hand, all examined 31-month-old fish exhibited notably intense staining over their entire bodies.

Figure 28.1. Whole-body detection of SA-J-gal activity in adult zebrafish of different ages (Kishi et al., 2003). After whole-mount fixation of adult zebrafish with whole body, SA-J-gal staining was performed at pH 6 in 9, 17, 22, and 31-month-old zebrafish samples. As shown in this figure, younger 9-month-old fish exhibited very faint background staining in some abdominal regions, whereas several different levels of staining in individual fish were observed in 17- and 22-month-old fish. The staining patterns of each fish varied in these age groups. Some individuals showed fairly strong staining, while others were relatively variable. On the other hand, all examined 31-month-old fish exhibited notably intense staining over their entire bodies.

Keyes et al., 2005; Varela et al., 2005). We previously reported on SA-J-gal induction during the aging process in zebrafish in vivo (Kishi, 2004; Kishi et al., 2003). We demonstrated significant increases of SA-J-gal activity in skin from aged (more than 31 months) versus young (less than 17 months) zebrafish (Kishi et al., 2003) (Figure 28.1).

The presence of SA-J-gal has been determined further in 6, 18, 30, and 42-month-old fish for more than 10 fish in each age group. We have newly developed a quantitative approach through conversions of captured actual images into pixel images scored in zebrafish embryos and adults. Based on scanning of the fixed parts of the trunk region in adult fish, the images were converted into pixels, and a prominent increase of SA-J-gal intensity was found with age. In our studies, SA-J-gal activity of zebrafish appeared to correlate well with their chronological ages.

On the other hand, in zebrafish embryos under genotoxic or oxidative stress, induction of SA-J-gal was successfully measured qualitatively and quantitatively (Figure 28.2).

This will open up a new avenue for screening of potential aging mutants by the embryonic senescence phenotype in zebrafish without spending lengthy time-of-lifespan analyses.

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