Excessive Oxidative Stress Contributes to Increased Acute ER Stress Kidney Injury in Aged Mice

Figure 3

Abnormalities in UPR in the kidneys of old mice: renal cortex RNA was obtained from young and old mice at baseline, 24 hours, 48 hours, and 72 hours after high dose of tunicamycin (0.8 μg/g) injection. The levels of GRP78, GRP94, phosphorylated PERK (PERK-p), phosphorylated eIF2α (eIF2α-p), total eIF2α, CHOP, caspase 12, and PARP were measured in eight animals from each group by Western blots. XBP-1 and GAPDH mRNA expression in kidneys at baseline and 48 hours after tunicamycin injection was determined by RT-PCR (). Results from two representative animals of baseline and tunicamycin-treated young and old mice were shown. The intensity of each blot band was quantitated using a densitometer. Data from the untreated kidneys of young mice was arbitrarily defined as 1 after correcting with the intensity of the individual β-actin band of the same sample. Lanes Y (baseline) and YT (tunicamycin treated) were samples from young mice. Lanes O (baseline) and OT (tunicamycin treated) were samples from old mice. Both GRP78 and GRP94 were significantly increased in the kidneys of both young and old mice at 24 (a, b) and 72 hours (c, d) after tunicamycin treatment. No differences were found between young and old mice. At baseline, phosphorylated PERK levels were low (a, e). The levels were significantly elevated in both old and young mice 24 hours after high dose of tunicamycin injection (a). Phosphorylated PERK remained high in young mice but was largely lost in old mice at 72 hours (e). Subsequently, at 72 hours, phosphorylated eIF2α was increased in young mice while it was decreased in old mice (e). High-dose tunicamycin treatment was associated with more increased kidney CHOP protein levels in old mice (f). XBP-1 mRNA splicing occurred only in the kidneys of young mice after treatment (g). Cleaved caspase 12 levels were increased in old mice at baseline and were further increased after tunicamycin treatment (h). The increase in cleaved PARP was also more robust in old mice (h). vs. young mice at baseline. vs. old mice at baseline.

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