Dietary iron concentration may influence aging process by altering oxidative stress in tissues of adult rats.

Iron is an essential element. However, in its free form, iron participates in redox-reactions, leading to the production of free radicals that increase oxidative stress and the risk of damaging processes. Living organisms have an efficient mechanism that regulates iron absorption according to their iron content to protect against oxidative damage. The effects of restricted and enriched-iron diets on oxidative stress and aging biomarkers were investigated. Adult Wistar rats were fed diets containing 10, 35 or 350 mg/kg iron (adult restricted-iron, adult control-iron and adult enriched-iron groups, respectively) for 78 days. Rats aged two months were included as a young control group. Young control group showed higher hemoglobin and hematocrit values, lower levels of iron and lower levels of MDA or carbonyl in the major studied tissues than the adult control group. Restricted-iron diet reduced iron concentrations in skeletal muscle and oxidative damage in the majority of tissues and also increased weight loss. Enriched-iron diet increased hematocrit values, serum iron, gamma-glutamyl transferase, iron concentrations and oxidative stress in the majority of tissues. As expected, young rats showed higher mRNA levels of heart and hepatic L-Ferritin (Ftl) and kidneys SMP30 as well as lower mRNA levels of hepatic Hamp and interleukin-1 beta (Il1b) and also lower levels of liver protein ferritin. Restricted-iron adult rats showed an increase in heart Ftl mRNA and the enriched-iron adult rats showed an increase in liver nuclear factor erythroid derived 2 like 2 (Nfe2l2) and Il1b mRNAs and in gut divalent metal transporter-1 mRNA (Slc11a2) relative to the control adult group. These results suggest that iron supplementation in adult rats may accelerate aging process by increasing oxidative stress while iron restriction may retards it. However, iron restriction may also impair other physiological processes that are not associated with aging.

Vitamin A deficiency increases hepcidin expression and oxidative stress in rat.

OBJECTIVE: The interaction between vitamin A and iron status has been widely reported; however, the exact mechanism involved in this interaction has not been well characterized. The present study investigated the mechanism involved in tissue iron accumulation and changes in the oxidative status in vitamin A-deficient rats. METHODS: Rats were treated with a control diet, a vitamin A-deficient diet, or a vitamin A/iron-deficient diet for 57 d. The animals were sacrificed; the blood, liver, and spleen were collected for biochemical analysis. Analysis of variance or Mann-Whitney tests were used to compare groups and Pearson's or Spearman's tests to investigate the bivariate correlation. RESULTS: Vitamin A deficiency increased liver hepcidin mRNA and iron spleen concentrations; however, iron deficiency in vitamin A-deficient rats deeply inhibits liver hepcidin mRNA expression and significantly increases divalent metal transporter-1 mRNA levels. Liver ferroportin and hereditary hemochromatosis gene mRNA levels did not change in either treatment. In the vitamin A-deficient groups, liver carbonyl protein increased, whereas catalase and glutathione S-transferase activities decreased, suggesting that vitamin A protects the liver against protein oxidation. A significant positive correlation was found between lipid oxidative damage and iron concentration in the liver and spleen (r = 0.611, P = 0.007; r = 0.558, P = 0.025, respectively). CONCLUSION: These results suggest that vitamin A maintains iron homeostasis by the modulation of liver hepcidin expression. The increase of lipid peroxidation in vitamin A deficiency seems to be iron dependent, whereas protein oxidation is not.

Parallel measurement of Ca2+ binding and fluorescence emission upon Ca2+ titration of recombinant skeletal muscle troponin C. Measurement of sequential calcium binding to the regulatory sites.

Calcium binding to chicken recombinant skeletal muscle TnC (TnC) and its mutants containing tryptophan (F29W), 5-hydroxytryptophan (F29HW), or 7-azatryptophan (F29ZW) at position 29 was measured by flow dialysis and by fluorescence. Comparative analysis of the results allowed us to determine the influence of each amino acid on the calcium binding properties of the N-terminal regulatory domain of the protein. Compared with TnC, the Ca(2+) affinity of N-terminal sites was: 1) increased 6-fold in F29W, 2) increased 3-fold in F29ZW, and 3) decreased slightly in F29HW. The Ca(2+) titration of F29ZW monitored by fluorescence displayed a bimodal curve related to sequential Ca(2+) binding to the two N-terminal Ca(2+) binding sites. Single and double mutants of TnC, F29W, F29HW, and F29ZW were constructed by replacing aspartate by alanine at position 30 (site I) or 66 (site II) or both. Ca(2+) binding data showed that the Asp --> Ala mutation at position 30 impairs calcium binding to site I only, whereas the Asp --> Ala mutation at position 66 impairs calcium binding to both sites I and II. Furthermore, the Asp --> Ala mutation at position 30 eliminates the differences in Ca(2+) affinity observed for replacement of Phe at position 29 by Trp, 5-hydroxytryptophan, or 7-azatryptophan. We conclude that position 29 influences the affinity of site I and that Ca(2+) binding to site I is dependent on the previous binding of metal to site II.

A ligação de cálcio à troponina C analisada por diálise de fluxo e por espectroscopia de fluorescência / Calcium binding to troponin C analized by flow dialysis and by fluorescence spectroscopy

A troponina C é o componente do complexo heterotrimérico troponina ao qual o Ca²+ se associa. Pearlstone et al. (Biochemistry 31, 6545, (1992)) utilizaram o cDNA da troponina C de músculo esquelético de galinha (sTnC) para construir um mutante onde a fenilalanina da posição 29 foi substituída por triptofano (mutante F29W). Esse mutante permitiu que a ligação de Ca²+ aos sítios regulatórios amino terminais fosse acompanhada através de técnicas fluorescentes. Entretanto, algumas propriedades da sTnC foram alteradas pela mutação (Li et al. (1995) Biochemistry 34, 8330). No presente estudo, ensaios de ligação direta de Ca²+ bem como titulações de Ca²+ seguidas por fluorescência foram usadas para melhor se entenderem...