RESUMO
Apoptosis and necrosis are two distinct forms of cell death that can occur in response to different agents and stress conditions. In order to verify if the oxidative stress induced by dietary selenium and vitamin E deficiencies can lead muscle cells to apoptosis, one-day-old chicks were reared using diets differing in their vitamin E (0 or 10 IU/kg) and selenium (0 or 0.15 ppm) supplementation. Chick skeletal muscle tissue was obtained from 28-day-old animals and used to verify apoptosis occurrence based on caspase activity detection and DNA fragmentation. Antioxidant deficiency significantly increased caspase-like activity assessed by the hydrolysis of fluorogenic peptide substrates (Abz-peptidyl-EDDnp) at lambdaexc = 320 nm and lambdaem = 420 nm. Proteolytic activation was not accompanied by typical internucleosomal DNA fragmentation detected by field inversion gel electrophoresis. Although the general caspase inhibitor N-benzyloxycarbonyl-Val-Ala-Asp(O-Me) fluoromethyl ketone (Z-VAD-fmk) (0 to 80 muM) did not block caspase-like activity when preincubated for 30 min with muscle homogenates, the hydrolyzed substrates presented the same cleavage profile in HPLC (at the aspartic acid residue) when incubated with the purified recombinant enzyme caspase-3. These data indicate that oxidative stress causes caspase-like activation in muscle cells and suggest that cell death associated with exudative diathesis (dietary deficiency of selenium and vitamin E) can follow the apoptotic pathway
Assuntos
Animais , Apoptose , Caspases , Músculo Esquelético , Deficiência de Vitamina E , Galinhas , Fragmentação do DNA , Ativação Enzimática , Músculo EsqueléticoRESUMO
One-day-old chicks were reared using diets that differed in their vitamin E and/or selenium content. In chicks depleted of both selenium and vitamin E, signs of exudative diathesis on the superficial pectoralis muscle were observed. The purpose of this research was to determine the defective points of the antioxidant defense system, which made this tissue highly susceptible to nutritionally-induced oxidative stress. Vitamin E, and selenium in lower magnitude, were the factors that strikingly affected the course of mitochondrial lipid peroxidation. Animals fed diets deficient in vitamin E and selenium displayed the lowest reduced glutathione level and glutathione peroxidase activity. The decreased levels of reduced glutathione were not due to a defective activity of glutathione reductase, which was increased in both mitochondria and cytosol. The absence of vitamin E was linked to lowering of mitochondrial thiol levels. The Glutathione peroxidase/Cu,Zn-superoxide dismutase ratio was 2.8 in animals fed selenium and vitamin E, and decreased to 0.13 in animals deficient in both nutrients. This change was indicative of oxidant-induced damage mediated by hydrogen peroxide. Catalase activity increased in an attempt to counteract the decrease in glutathione peroxidase activity. The results obtained showed that alpha-tocopherol and Se deficiencies caused multiple alterations in the antioxidant system and adversely affected the redox state of chicken superficial pectoralis muscle.