Thioredoxin, thioredoxin reductase, and α-crystallin reviveinactivated glyceraldehyde 3-phosphate dehydrogenase in human aged andcataract lens extracts

摘要

Purpose: To investigate whether mammalian thioredoxin (Trx) andthioredoxin reductase (TrxR), with or without α-crystallin canrevive inactivated glyceraldehyde 3-phosphate dehydrogenase (GAPDH) inboth the cortex and nucleus of human aged clear and cataract lenses.Methods: The lens cortex (including capsule-epithelium) and thenucleus were separated from human aged clear and cataract lenses (gradeII and grade IV) with similar average age. The activity of GAPDH in thewater-soluble fraction after incubation with or without Trx or/and TrxRfor 60 min at 30 °C was measured spectrophotometrically. Inaddition, the effect of a combination of Trx/TrxR and bovine lensα-crystallin was investigated.Results: GAPDH activity was lower in the nucleus of clear lensesthan in the cortex, and considerably diminished in the cataractouslenses, particularly in the nucleus of cataract lenses grade IV. Trx andTrxR were able to revive the activity of GAPDH markedly in both thecortex and nucleus of the clear and cataract lenses. The percentageincrease of activity in the cortex of the clear lenses was less thanthat of the nucleus in the presence of Trx and TrxR, whereas it wasopposite in the cataract lenses. The revival of activity in both thecortex and nucleus from the cataract lenses grade II was higher thanthat of the grade IV. Moreover, Trx alone, but not TrxR, efficientlyenhanced GAPDH activity. The combination of Trx and TrxR had greatereffect than that of either alone. In addition, αL-crystallinenhanced the activity in the cortex of cataract grade II with Trx andTrxR present. However, it failed to provide a statistically significantincrease of activity in the nucleus.Conclusions: This is the first evidence to show that mammalian Trxand TrxR are able to revive inactivated GAPDH in human aged clear andcataract lenses, and α-crystallin helped this effect. Theinactivation of GAPDH during aging and cataract development must becaused in part by disulphide formation and in part by unfolding, and canbe recovered by reducing agents and a molecular chaperone.