|Budget Amount *help
¥3,100,000 (Direct Cost : ¥3,100,000)
Fiscal Year 1999 : ¥2,000,000 (Direct Cost : ¥2,000,000)
Fiscal Year 1998 : ¥1,100,000 (Direct Cost : ¥1,100,000)
This study investigates the role of reactive oxygen species (ROS) in anuran tadpole tails undergoing atrophy. Changes in catalase, superoxide dismutase (SOD) and caspase activities, genomic DNA, and nitric oxide (NO) generation were investigated biochemically using Rana rugosa and Rana japonica tadpole tails undergoing regression during thyroid hormone (TH) enhancement. From the results obtained so far, the following process is proposed for apoptosis in the tadpole tail : an increase in TH induces NO generation (by NO synthase (NOS) activation?) , causing cyiochrome oxidase inhibition, which results in superoxide radical (OィイD22ィエD2) generation. This leads to an increase in hydrogen peroxide (HィイD22ィエD2OィイD22ィエD2) after catalase activity inhibition by NO, followed by caspase-3 activation by lysosomal enzyme release, which induces DNA fragmentation, resulting in apoptosis. At present, we are conducting investigations aimed at clarifying the following : 1. Evidence of HィイD22ィエD2O, accumulation in tail cells ; 2. Gene expression of iNOS ; 3. The mechanism for generation of OィイD22ィエD2 and TH activation of caspase ; 4. Gene expression of catalase. Using a cell free system, we biochemically demonstrated for the first time that lysosomes are another important player in apoptosis. Here ROS disrupts lysosomal membranes. This is followed by the release of lysosomal enzymes, followed by the activation of caspase-3. The following problems should be clarified at cellular level : l. The formation of apoptotic bodies and DNA fragmentation into nucleosomal fragments in apoptotic cells ; 2. A mechanism for apoptosis by way of lysosomes ; 3. The kinds of lysosomal enzymes directly involved in apoptosis.