Ascorbic acid deficiency affects the metabolism of cytosolic ferritin but not lipid-associated ferritin in livers of guinea pigs.

Two physicochemically and metabolically separate pools of ferritin, namely cytosolic ferritin and lipid-associated ferritin, are present in the livers of guinea pigs. In this paper we establish that the iron content of cytosolic ferritin is dependent on and linearly related to ascorbic acid concentration, whereas changes in concentration of this vitamin do not affect the iron content of lipid-associated ferritin. In livers of ascorbic acid-deficient guinea pigs both synthesis and degradation of cytosolic ferritin are diminished equally. Consequently cytosolic ferritin is metabolized more slowly without changes in its pool size. In contrast with cytosolic ferritin, the metabolism of lipid-associated ferritin is unaffected by ascorbic acid deficiency. The differential effects of ascorbic acid deficiency on the physicochemical characteristics as well as on the metabolism of cytosolic ferritin and lipid-associated ferritin suggest that the two forms of ferritin have different functional roles.

Cytosolic ferritin and lipid-associated ferritin are metabolically different in guinea-pig livers.

A distinct pool of liver ferritin has been described in man, guinea pigs and rats [Cham, Roeser, Nikles & Ridgway (1986) Clin. Chim. Acta 158, 71-79]. This ferritin accounts for approx. 30% of total intracellular ferritin. It differs from previously described cytosolic and 'microsomal-fraction' ferritin by its firm association with lipid and by the absence of heat-stability at 75 degrees C. The present study demonstrates that cytosolic ferritin and lipid-associated ferritin in guinea-pig livers have distinctly different rates of turnover. Cytosolic ferritin has a rate of turnover approx. 3.5 times as high as lipid-associated ferritin. The apparent metabolic heterogeneity suggests that the two forms of ferritin may have different functional roles.

Effects of non-ionic and anionic detergents on lipid-associated tissue ferritin.

Lipid-associated ferritin from homogenates of guinea pig liver is released from its conjugate(s) by incubation with the non-ionic detergents Triton X-100 and Nonidet P-40 but not by incubation with the anionic detergent deoxycholate. The amount of lipid-associated ferritin released from its conjugate(s) depends on the concentration of the non-ionic detergents. At a final non-ionic detergent concentration of about 20 g/L, all lipid-associated ferritin is released from its conjugate(s) in a liver homogenate. The amount released is identical with the amount of the lipid-associated ferritin obtained by extraction of the same liver homogenate with a mixture of butanol and diisopropyl ether.

Lipid associated tissue ferritin.

Delipidation of liver homogenates, using an organic solvent system which does not denature proteins, increases measurable ferritin by 25-33%, compared to ferritin concentrations by standard heat supernatant assays. When applied to polyacrylamide gradient gels, lipid-associated ferritin does not enter the gel but after delipidation, this ferritin co-migrates with cytosolic ferritin and with purified liver ferritin. The biological significance of the association of ferritin with lipid has yet to be examined.

A procedure for the purification of ferritin from human liver by heating a methanol-treated homogenate.

A simple, rapid technique for purification of ferritin from human liver tissue is described. Methanol, at a final concentration of 40% (v/v) in liver homogenate, precipitates the majority of proteins but does not affect ferritin. Subsequent heating of this homogenate at 75 degrees C for 10 min results in a purified ferritin preparation as judged by immunoelectrophoresis and polyacrylamide gel electrophoresis. The resultant purified ferritin contained the same amount of iron as the original endogenous ferritin. There were no significant differences (paired t tests) in the amount of protein in the purified ferritin preparation when measured by rocket immunoelectrophoresis and by the Lowry procedure, suggesting that the antigenecity of ferritin was unaffected by the methanol and heat treatment. Both endogenous liver ferritin and radiolabeled human liver ferritin added to liver homogenates were recovered after methanol and heat treatment with similar yields (77 +/- 7% and 70 +/- 2%, respectively) when compared with the standard treatment of heating a homogenate at 75 degrees C. The overall ferritin yield with this rapid procedure was 40%.

Tissue ferritin in scorbutic guinea-pigs.

Tissue ferritin metabolism was compared in control and ascorbic acid (AA) deficient guinea-pigs. Concentrations of ferritin protein in the liver (0.98 +/- 0.61 mg/g wet weight) and spleen (0.48 +/- 0.23 mg/g) of control animals did not change after tissue depletion of AA. Iron dextran (75 mg/kg weight, i.m.) caused a 4-5-fold increase in tissue ferritin concentrations in controls whereas no increase in tissue ferritin occurred in scorbutic animals. The rise in tissue total iron concentration was similar in the two groups. Liver ferritin synthesis was similar in control and scorbutic animals. After stimulation with iron (8 mg/kg iron dextran i.v.), ferritin synthesis rose in both groups of animals. However, the pattern of response differed. At 24 h after iron dextran, ferritin synthesis in controls was still significantly elevated (P less than 0.001) and liver ferritin protein continued to rise, whereas in scorbutic animals, ferritin synthesis had declined to pre-iron injection levels, and no rise in ferritin protein values occurred. It is concluded that the ferritin synthetic apparatus in AA deficient tissues remained intact and capable of responding to added iron. The absence of a sustained elevation in tissue ferritin protein after an iron load appeared to be due to inadequate stimulation of ferritin synthesis by intracellular iron. It is suggested that AA has a physiological role in the reduction of intracellular iron and that it is the reduced form of iron which stimulates ferritin synthesis. Abnormalities of iron metabolism occur in AA depleted tissues when the quantity of Fe3+ entering cells exceeds the residual reducing capacity of those cells.

A comparison of the antifertility effects of alkylating agents and vinca alkaloids in male rats.

1 The anti-fertility effects of cyclophosphamide, nitrogen mustard, vincristine and vinblastine were studied and compared in male rats. 2 The effects of the drugs on body weight and haematological values were used to monitor the pharmacological actions of the drugs. 3 All four drugs impaired fertility, the severity of the impairment depending on dose and duration of treatment. 4 Testicular size and histological appearances remained mostly normal, even in infertile animals, but seminiferous tubules were fewer in number and maturation arrest at the spermatid level was evident in some sections. 5 Recovery of drug-induced infertility occurred in 64% of treated animals, 9 to 40 weeks after cessation of treatment. 6 Morbidity and mortality were much higher with alkylating agents than with vinca alkaloids for approximately similar degrees of impairment in fertility.