Age-related changes in hepatic activity and expression of detoxification enzymes in male rats

Abstract

Process of aging is accompanied by changes in the biotransformation of xenobiotics and impairment of normal cellular functions by free radicals. Therefore, this study was designed to determine age-related differences in the activities and/or expressions of selected drug-metabolizing and antioxidant enzymes in young and old rats. Specific activities of 8 drug-metabolizing enzymes and 4 antioxidant enzymes were assessed in hepatic subcellular fractions of 6-week-old and 21-month-old male Wistar rats. Protein expressions of carbonyl reductase 1 (CBR1) and glutathione S-transferase (GST) were determined using immunoblotting. Remarkable age-related decrease in specific activities of CYP2B, CYP3A, and UDP-glucuronosyl transferase was observed, whereas no changes in activities of CYP1A2, flavine monooxygenase, aldo-keto reductase 1C, and antioxidant enzymes with advancing age were found. On the other hand, specific activity of CBR1 and GST was 2.4 folds and 5.6 folds higher in the senescent rats compared with the young ones, respectively. Interindividual variability in CBR1 activity increased significantly with rising age. We suppose that elevated activities of GST and CBR1 may protect senescent rats against xenobiotic as well as eobiotic electrophiles and reactive carbonyls, but they may alter metabolism of drugs, which are CBR1 and especially GSTs substrates.

Figure 1

Age-related differences in specific activities of biotransformation enzymes with oxidizing activity in rat liver. The white column represents young (6-week-old) rats, and the hatched column indicates old (21-month-old) rats. The activities of individual CYP isoforms were normalized to the total CYP content. Data of each age group represent the mean ± SD (six animals per age group) (*data with P < 0.01, Student's t-test). CYP = cytochrome P450; FMO = flavine monooxygenase.

Figure 2

Specific activities of biotransformation enzymes with reducing activity in rat liver. The white column represents young (6-week-old) rats, and the hatched column indicates old (21-month-old) rats. Values are expressed as mean ± SD (six animals per age group) (*data with P < 0.01, Student's t-test). CBR1 = carbonyl reductase 1; AKR1C = aldo-keto reductase 1C.

Figure 3

Specific activities of biotransformation enzymes with conjugative activity in rat liver. The white column represents young (6-week-old) rats, and the hatched column indicates old (21-month-old) rats. Values are expressed as mean ± SD (six animals per age group) (*data with P < 0.01, Student's t-test). GST = glutathione S-transferase; UGT = UDP-glucuronosyl transferase.

Figure 4

Interindividual differences in specific activities of GST and CBR1 in rat liver. The white columns represent 6-week-old rats (J1–J6), and hatched columns indicate 21-month-old rats (S1–S6). Values are expressed as mean ± SD (six animals per age group). GST = glutathione S-transferase; CBR1 = carbonyl reductase 1.

Figure 5

Age-related changes in the hepatic CBR1 and GSTA protein expression normalized to the amount of β-actin (a). Interindividual differences in protein expression of GST and CBR1 in rat liver (b). Cytosolic fractions (4 μg of protein/lane) of 6-week-old (J1–J6) and 21-month-old (S1–S6) rats were subjected to SDS-PAGE with subsequent immunoblotting. Blots were then reacted with rabbit polyclonal anti-CBR1 antibody or goat polyclonal anti-GSTA antibody. Protein bands were marked by corresponding secondary antibodies using a chemiluminescent detection with an alkaline phosphatase substrate. Protein expression was normalized to the amount of the loading control β-actin using mouse monoclonal anti-beta-actin antibody. GSTA = alpha-glutathione S-transferase; CBR1 = carbonyl reductase 1.

Figure 6

Age-related differences in specific activities of antioxidant enzymes in rat liver. The white column represents young (6-week-old) rats, and the hatched column indicates old (21-month-old) rats. Values are expressed as mean ± SD (six animals per age group) (*data with P < 0.01, Student's t-test). SOD = superoxide dismutase; CAT = catalase; GPx = glutathione peroxidase; nonspecific Px = nonspecific peroxidase.