Little is known about the biological effect of folate in the protection against mitochondrial (mt) oxidative decay. The objective of the present study was to examine the consequence of folate deprivation on mt oxidative degeneration, and the mechanistic link underlying the relationship. Male Wistar rats were fed with an amino acid-defined diet containing either 8 (control) or 0 (folate-deficient, FD) mg folic acid/kg diet. After a 4-week FD feeding period, significant elevation in oxidative stress was observed inside the liver mitochondria with a 77 % decrease in mt folate level (P,0·001), a 28 % reduction in glutathione peroxidase activity (P¼0·0333), a 1·2-fold increase of mt protein carbonyls (P¼0·0278) and an accumulated 4834 bp large-scale deletion in mtDNA. The elicited oxidative injuries in FD liver mitochondria were associated with 30 % reduction of cytochrome c oxidase (CcOX) activity (P¼ 0·0264). The defective CcOX activity in FD hepatocytes coincided with mt membrane potential dissipation and intracellular superoxide elevation. Exposure of FD hepatocytes to pro-oxidant challenge (32 mM-copper sulphate for 48 h) led to a further loss in CcOX activity and mt membrane potential with a simultaneous increase in superoxide production. Preincubation of pro-oxidanttreated FD hepatocytes with supplemental folic acid (10-1000 mM) reversed the mt oxidative defects described earlier and diminished superoxide overproduction. Increased supplemented levels of folic acid strongly correlated with decreased lipid peroxidation (g 20·824, P¼0·0001) and protein oxidative injuries (g 2 0·865, P¼ 0·0001) in pro-oxidant-challenged FD liver mitochondria. Taken together, the results demonstrated that folate deprivation induces oxidative stress in liver mitochondria, which is associated with CcOX dysfunction, membrane depolarization and superoxide overproduction. The antioxidant activity of supplemental folic acid may partially, if not fully, contribute to the amelioration of pro-oxidantelicited mt oxidative decay. The biochemical function of folate involved in one-carbon metabolism critical for cellular proliferation and nuclear DNA repair has been well documented (Shane, 1995). In the last decade, growing literature evidence suggests another potential role of folate on antioxidant action (Nakano et al. However, the basis for folate status and oxidative insults is unclear. Folic acid has been proposed to scavenge peroxyl radicals, azide radicals and hydroxyl radicals in an in vitro radical reaction model system (Joshi et al. 2001). Particularly, the intracellular superoxide-scavenging capability of folic acid was observed in various pro-oxidant-challenged cells such as homocysteine thiolactone-treated HL60 cells (Huang et al. 2002) and 7-ketocholesterol-treated U937 cells (Huang et al. 2004). Increased folate intake was found to improve endothelial function in patients with coronary artery disease, which is largely independent of homocysteine lowering action. Instead, reduction of intracellular endothelial superoxide may have ...
Cordyceps sinensis contains a factor that stimulates corticosteroid production in the animal model. However, it is not known whether this drug acts directly on the adrenal glands or indirectly via the hypothalamus-pituitary axis. In the present study, we used primary rat adrenal cell cultures to investigate the pharmacological function of a water-soluble extract of Cordyceps sinensis (CS) and the signaling pathway involved. Radioimmunoassay of corticosterone indicated that the amount of corticosterone produced by adrenal cells is increased in a positively dose-dependent manner by CS, reaching a maximum at 25 microg/ml. This stimulating effect was seen 1 h after CS treatment and was maintained for up to 24 h. Concomitantly, the lipid droplets in these cells became small and fewer in number. Immunostaining with a monoclonal antibody, A2, a specific marker for the lipid droplet capsule, demonstrated that detachment of the capsule from the lipid droplet occurs in response to CS application and that the period required for decapsulation is inversely related to the concentration of CS applied. The mechanism of CS-induced steroidogenesis is apparently different from that for ACTH, since intracellular cAMP levels were not increased in CS-treated cells. However, combined application with calphostin C, a PKC inhibitor, completely blocked the effect of CS on steroidogenesis, suggesting that activation of PKC may be responsible for the CS-induced steroidogenesis.
Enzyme kinetic data has indicated the probability of “soluble” enzymes existing in situ as macromolecular complexes. These enzymes often function in sequential reactions, e. g., mitochondrial Krebs cycle. Therefore, it is important to define morphologically their functional in situ localization. One such enzyme of interest is ATP-citrate lyase (citrate cleavage enzyme). Biochemically, it has been characterized as a cytoplasmic enzyme. However, there are teleological as well as other indications that it may be concentrated in the vicinity of mitochondria.
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