Novel insights into mitochondrial molecular targets of iron-induced neurodegeneration: Reversal by cannabidiol.

Evidence has demonstrated iron accumulation in specific brain regions of patients suffering from neurodegenerative disorders, and this metal has been recognized as a contributing factor for neurodegeneration. Using an experimental model of brain iron accumulation, we have shown that iron induces severe memory deficits that are accompanied by oxidative stress, increased apoptotic markers, and decreased synaptophysin in the hippocampus of rats. The present study aims to characterize iron loading effects as well as to determine the molecular targets of cannabidiol (CBD), the main non-psychomimetic compound of Cannabis sativa, on mitochondria. Rats received iron in the neonatal period and CBD for 14 days in adulthood. Iron induced mitochondrial DNA (mtDNA) deletions, decreased epigenetic modulation of mtDNA, mitochondrial ferritin levels, and succinate dehydrogenase activity. CBD rescued mitochondrial ferritin and epigenetic modulation of mtDNA, and restored succinate dehydrogenase activity in iron-treated rats. These findings provide new insights into molecular targets of iron neurotoxicity and give support for the use of CBD as a disease modifying agent in the treatment of neurodegenerative diseases.

Effects of carbonyl iron powder on iron deficiency anemia and its subchronic toxicity.

Carbonyl iron powder (CIP) has been used as a food additive or mineral supplement. However, the effects of CIP on iron deficiency anemia (IDA) and its subchronic toxicity have not been investigated. We found that oral administration of CIP at a dose of 2.96 mg/kg recovered the hemoglobin concentration of erythrocytes of IDA rats to the normal level after 8 days. The no observed adverse effect level of CIP in rats was considered to be > 200 mg/kg. The hematological and serum biochemical parameters of the rats did not differ significantly between the control and treated groups. There were no morphological changes observed in the organs including liver, kidneys, spleen, testes, stomach and intestine. Therefore, CIP might be a safe iron supplement.

Quercetin prevents ethanol-induced iron overload by regulating hepcidin through the BMP6/SMAD4 signaling pathway.

Emerging evidence has demonstrated that chronic ethanol exposure induces iron overload, enhancing ethanol-mediated liver damage. The purpose of this study was to explore the effects of the naturally occurring compound quercetin on ethanol-induced iron overload and liver damage, focusing on the signaling pathway of the iron regulatory hormone hepcidin. Adult male C57BL/6J mice were pair-fed with isocaloric-Lieber De Carli diets containing ethanol (accounting for 30% of total calories) and/or carbonyl iron (0.2%) and treated with quecertin (100 mg/kg body weight) for 15 weeks. Mouse primary hepatocytes were incubated with ethanol (100 mM) and quercetin (100 µM) for 24 h. Mice exposed to either ethanol or iron presented significant fatty infiltration and iron deposition in the liver; these symptoms were exacerbated in mice cotreated with ethanol and iron. Quercetin attenuated the abnormity induced by ethanol and/or iron. Ethanol suppressed BMP6 and intranuclear SMAD4 as well as decreased hepcidin expression. These effects were partially alleviated by quercetin supplementation in mice and hepatocytes. Importantly, ethanol caused suppression of SMAD4 binding to the HAMP promoter and of hepcidin messenger RNA expression. These effects were exacerbated by anti-BMP6 antibody and partially alleviated by quercetin or human recombinant BMP6 in cultured hepatocytes. In contrast, co-treatment with iron and ethanol, especially exposure of iron alone, activated BMP6/SMAD4 pathway and up-regulated hepcidin expression, which was also normalized by quercetin in vivo. Quercetin prevented ethanol-induced hepatic iron overload different from what carbonyl iron diet elicited in the mechanism, by regulating hepcidin expression via the BMP6/SMAD4 signaling pathway.

It is still not for the old iron: adjuvant effects of carbonyl iron in experimental autoimmune encephalomyelitis induction.

Experimental autoimmune encephalomyelitis (EAE) is a model of multiple sclerosis. Dark Agouti rats immunized with spinal cord homogenate (SCH) and carbonyl iron (CI), as an adjuvant, develop severe hyperacute form of EAE. They succumb to EAE earlier and have higher clinical scores and lethality rate in comparison to counterparts immunized with SCH + complete Freund's adjuvant. There is no difference in the number of cells or in histological presentation of the CNS infiltrates of rats immunized with the two adjuvants. However, there are more granulocytes, NK and NKT cells, and less CD4(+) T cells in the spinal cord infiltrates of SCH + CI-immunized animals. Nitric oxide (NO)-generating enzyme inducible NO synthase have higher expression in spinal cord of SCH + CI-immunized rats, and this corresponds to more intensive nitrotyrosine formation in the CNS tissue of these rats. Abundant infiltration of granulocytes and NK cells into the CNS and excessive generation of peroxynitrite within the CNS of SCH + CI-immunized rats might account for the severe neurological deficits induced by immunization with CI. These factors should be closely examined in the fulminant forms of multiple sclerosis and acute disseminated encephalomyelitis, as they could represent a promising targets for therapy.

Variegate porphyria induces plasma and neutrophil oxidative stress: effects of dietary supplementation with vitamins E and C.

Our aim was to analyse the influence of variegate porphyria (VP) on the antioxidant defenses and markers of oxidative damage and inflammation in plasma and neutrophils and the effects of dietary supplementation with vitamins E and C on these parameters in plasma, neutrophils and erythrocytes. Twelve women affected by VP and twelve pair-matched healthy control women participated in a double-blind crossover study. Each participant took 50 mg/d of vitamin E and 150 mg/d of vitamin C, or a placebo, for 6 months, by consuming an almond-based beverage as the vehicle. Women affected by VP presented higher C-reactive protein and malondialdehyde (MDA) circulating levels. Plasma antioxidant defenses were not different between porphyric and control women. Neutrophils from VP women presented decreased catalase (CAT) and glutathione reductase (GR) activities together with increased protein carbonyl levels. Reactive oxygen species (ROS) production from stimulated neutrophils was also higher in porphyric women than their controls. Dietary supplementation was effective in increasing alpha-tocopherol levels in neutrophils and in reducing MDA levels in plasma. Erythrocyte CAT and GR activities were enhanced by the enriched beverage only in the control subjects. In conclusion, women affected by VP present a situation of inflammation, plasma oxidative damage and neutrophils more primed to the oxidative burst, with decreased antioxidant activities and increased ROS production capabilities and protein oxidative damage. Dietary supplementation with vitamin E (50 mg/d) and vitamin C (150 mg/d) for 6 months decreased plasma oxidative damage and enhanced the erythrocyte activities of CAT and GR.

Interplay among tetrahedrane, butterfly diradical, and planar rhombus structures in the chemistry of the binuclear iron carbonyl phosphinidene complexes Fe2(CO)6(PX)2.

Density functional theory studies on a series of Fe2(CO)6(PX)2 derivatives show the tetrahedrane to be the most stable for the alkyl (X = Me, tBu), P-H (X = H), and chloro (X = Cl) derivatives. However, butterfly diradical and planar rhombus structures are found to be more stable than tetrahedranes for the amino (X = NH2, NMe2, and NiPr2) and aryloxy (R = 2,6-tBu2-4-Me-C6H2O) derivatives. For the chloro (X = Cl) and methoxy (X = OMe) derivatives energetically accessible bishomotetrahedrane Fe2(CO)6P2(mu-X)2 isomers are observed in which the X substituents on the phosphorus atoms interact with the iron atom to form two direct Fe-X bonds at the expense of two of the four Fe-P bonds. In addition, the global minimum for the hydroxy (X = OH) derivative is an unusual FeP-butterfly structure with a central Fe-P bond as well as two external Fe-P bonds, one external P-P bond, and one external Fe=Fe double bond. Comparison of calculated with experimental nu(CO) frequencies shows that low-temperature Nujol matrix photolysis of (iPr2NP)2COFe2(CO)6 leads to a planar rhombus rather than a tetrahedrane isomer of Fe2(CO)6(PNiPr2)2.

Influence of purified saponin mixture from Astragalus corniculatus Bieb. on phagocytic cells in Graffi-tumor bearing hamsters.

A purified saponin mixture (PSM) from Astragalus corniculatus Bieb. protected significantly hamsters against the experimental Graffi myeloid tumor. The application of PSM increased the survival rates, prolonged mean survival time and the tumor growth was markedly reduced. A purified saponin mixture (PSM) of Astragalus corniculatus Bieb. was evaluated for its immunostimulating potentials on the phagocytic cells in Graffi-tumor bearing hamsters. The number, migration and phagocytic indexes of peritoneal macrophages (pMøs) and of blood polymorphonuclear leukocytes (PMNs) were evaluated in healthy and Graffi-tumor bearing hamsters (G-TBH) treated with PSM. It was established that the Graffi myeloid tumor induced suppression of the phagocytic abilities of pMøs and PMNs. The number and migration of pMøs was significantly decreased during the whole period of observation. All tested parameters-number, migration and phagocytic activities of pMøs, as well as phagocytic ability of PMNs increased significantly in healthy and G-TBH after i.p. application of the 50 mg/kg body weight PSM. The PMS extracts from Astragalus corniculatus Bieb. are isolated and examined and their immunostimulating and immunorestorating impact on phagocitic cells was proven for the first time. This effect could be due to their high content of purified saponins.

Comparative assessment of the bioavailability, efficacy and safety of a modified-release (MR) carbonyl iron tablet and oral conventional iron preparation in adult Indian patients with nutritional iron deficiency anaemia.

The objective of the study is to evaluate the bioavailability, efficacy and safety of a new modified-release (MR) formulation of carbonyl iron (45 mg) relative to a commercially available conventional formulation of ferrous fumarate (300 mg) in adult Indian patients with clinical and laboratory diagnosis of nutritional iron deficiency anaemia. This prospective, comparative, randomised, double-blind study was carried out among 60 patients received a single daily dose of either MR carbonyl iron or ferrous fumarate for 12 weeks. The effect of therapy on haematological parameters and iron status and estimation of bioavailability were the main efficacy outcomes. There was a significant (p<0.05) increase in mean haemoglobin levels, reticulocyte counts, haematocrit and mean corpuscular volume in MR carbonyl iron group compared to ferrous fumarate group. There was also an increase in mean serum iron and ferritin levels and a corresponding decrease in total iron binding capacity in MR carbonyl iron group compared to ferrous fumarate group at the end of 12 weeks therapy. The estimated overall bioavailability of MR carbonyl iron was about 147% that of ferrous fumarate. Both the formulations were equally well-tolerated and adverse events were mainly gastrointestinal in nature. The prevalence of adverse events was slightly more in the ferrous fumarate group. It can be concluded that the MR formulation of carbonyl iron was more efficacious than ferrous fumarate in correcting haematologic abnormalities and improving iron status in patients with nutritional iron deficiency anaemia. In conditions where efficacy is an important consideration, the higher bioavailability of MR carbonyl iron may make it the treatment of choice for nutritional iron deficiency anaemia.

Acute toxicity of carbonyl iron and sodium iron EDTA compared with ferrous sulfate in young rats.

According to the American Association of Poison Control Centers, exposures to excessive doses of iron supplements still occur in children less than 6 years of age. Since 1998, there has been one death among U.S. children in this age group. Exposures, including adverse events, to iron supplements and iron-containing vitamins for the years 1999 and 2000 were 23,215 and 24,249, respectively. To reduce the potential seriousness of such exposures, carbonyl iron (Fe(0)) has been suggested as a possible replacement for ferrous sulfate (FeSO(4)). Carbonyl Fe is a unique form of elemental iron because of its small particle size. It is highly bioavailable when used to correct iron deficiency anemia. There is also current interest in using sodium iron(III) ethylenediaminetetraacetate (NaFeEDTA) for food fortification. In this study both NaFeEDTA and carbonyl Fe were compared with FeSO(4), the most common form of iron for dietary supplements, to obtain information relevant to the acute toxicological profile in young rats. With FeSO(4) and NaFeEDTA, total liver nonheme iron increased with increasing dose, but the response was approximately 50% lower with NaFeEDTA compared with FeSO(4). Serum iron peaked at approximately 0.5 to 1 h for both FeSO(4) and carbonyl Fe, while NaFeEDTA was elevated up to 4 h. FeSO(4) had an LD(50) of 1.1 g Fe/kg and was approximately 45 times more toxic than carbonyl Fe, which had an LD(50) greater then 50 g Fe/kg. NaFeEDTA had an LD(50) of 1.3 g Fe/kg and, when compared with FeSO(4), had approximately the same level of toxicity.

Carbonyl-iron supplementation induces hepatocyte nuclear changes in BALB/CJ male mice.

BACKGROUND/AIMS: In humans, chronic iron excess may induce hepatic fibrosis and/or hepatocellular carcinoma. This work was undertaken to investigate hepatic iron overload outcome in iron-overloaded mice. METHODS: BALB/cJ male mice received supplements of 0, 0.5, 1.5 and 3% carbonyl-iron for 2, 4, 8 and 12 months. Histological staining, immunohistochemistry using ferritin antibodies and electron microscopic studies were performed on liver. Liver iron concentration was measured biochemically. Mitotic index and hepatocyte nuclear size were evaluated on Feulgen-stained slides. RESULTS: Liver iron concentration was increased, reaching 13 times control value after 12 months in 3% iron-overloaded mice, and iron was found predominantly in hepatocytes, with a porto-centrolobular decreasing gradient. Neither hepatic fibrosis nor hepatocellular carcinoma was found. Perls' stain positive inclusions containing ferritin were found within hepatocyte nuclei in 3%-overloaded mice. Electron microscopy disclosed that inclusions consisted of ferritin particle aggregates without a limiting membrane. Mice overloaded with 3% iron for 12 months showed larger hepatocyte nuclei than control mice and a mitotic index increase with presence of abnormal tripolar mitotic figures. In addition, some iron-free hepatocytes were observed. CONCLUSIONS: Carbonyl-iron supplementation produces significant iron overload in mice but does not result in liver fibrosis or hepatocellular carcinoma after 12 months. However, nuclear changes were produced in hepatocytes, and occasional iron-free hepatocytes were observed: these may represent preneoplastic changes caused by iron overload.

Inhalation of high concentrations of low toxicity dusts in rats results in impaired pulmonary clearance mechanisms and persistent inflammation.

This study was carried out to assess the time course of pulmonary clearance impairment and persistence of inflammation following high-dose inhalation exposures to titanium dioxide (TiO2) or carbonyl iron (CI) particles. Male rats were exposed to air, TiO2 or CI particles 6 hr/day, 5 days/week, for 4 weeks at concentrations of 5, 50, and 250 mg/m3 and evaluated at selected intervals through 6 months postexposure. Indices of pulmonary inflammation as well as alveolar macrophage clearance functions (i.e., morphology, in vivo and in vitro phagocytosis, and chemotaxis), cell proliferation, and histopathology endpoints were measured at several postexposure time periods through 6 months. In addition, amounts of TiO2 or CI in lungs and tracheobronchial lymph nodes were measured to allow an evaluation of particle clearance and translocation patterns. Four-week exposures to TiO2 or CI particles at concentrations of 250 mg/m3 resulted in lung burdens of 12 mg titanium and 17 mg iron, respectively, with particle retention half-times ranging from 68 days for 5 mg/m3 TiO2 to approximately 330 days for 250 mg/m3. The impact of this TiO2 dust load and similar lung burdens of CI particles produced a sustained pulmonary inflammatory response measured through a period of 3-6 months postexposure concomitant with increases in BrdU cell labeling of terminal airway and pulmonary parenchymal cells. The impairment of particle clearance mechanisms was accounted for by deficits in in vitro phagocytic and chemotactic potential of alveolar macrophages recovered from the lungs of high-dose, TiO2- or CI-exposed rats. Free granular pigment (TiO2 or CI) was present on the hypertrophic mucosal surfaces of bronchioles and bronchi, and particle-laden macrophages, found individually, were numerous throughout alveoli and within lymphoid tissues immediately after exposure. Aggregates of particle-laden macrophages were present within alveoli and alveolar ducts from 1 week postexposure through the entire 6-month recovery period. Macrophage accumulations increased in size and number from 1 week through 1 month postexposure and then appeared to remain constant through the remaining 5-month postexposure period. Minimal cellular hypertrophy and hyperplasia were evident at alveolar duct bifurcations adjacent to macrophage aggregates, and this effect was most prominent at 3 to 6 months postexposure. The results of this study clearly demonstrate that exposure to high dust concentrations of two different innocuous particle types produced sustained pulmonary inflammation, enhanced proliferation of pulmonary cells, impairment of particle clearance, deficits in macrophage function, and the appearance of macrophage aggregates at sites of particle deposition. In addition, the mass deposition rate determination appears to be a less sensitive indicator of "overload" when compared to biomarkers of pulmonary toxicity, such as macrophage function and cellular inflammation and proliferation indices.

Giant mitochondria induced in rat pancreatic exocrine cells by ethanol and iron.

BACKGROUND: Mitochondria are sensitive indicators of cellular pathology. Under certain circumstances, these organelles respond to cellular insult by a marked increase in size, resulting in the formation of giant mitochondria (megamitochondria). Ethanol has been implicated in the induction of giant mitochondria in rat hepatocytes. Since ethanol is reported to affect pancreatic mitochondria, we examined this organ for evidence of mitochondrial giantism in rats administered ethanol and a relatively small amount of supplementary iron. METHODS: Diets were administered via a chronically implanted gastrostomy catheter. Rats were segregated in four groups: 1) basic high-fat diet, 2) ethanol and a high fat diet, 3) carbonyl iron and a high-fat diet, and 4) ethanol and carbonyl iron combined with a high-fat diet. After the animals were on their respective diets for 16 weeks, specimens of pancreas were extirpated and processed for and examined by transmission electron microscopy. RESULTS: Mitochondria in rats on the basic high fat diet resemble those in untreated controls. With ethanol and a high-fat diet, some mitochondria in virtually every exocrine cell are profoundly altered. Such organelles, which are of normal size, have undergone rearrangement of their internal membranes, with three of four parallel cristae residing at one or both poles of spherical to ovate mitochondria. In rats receiving both ethanol and carbonyl iron, giant mitochondria are present in pancreatic exocrine cells. Except for their size, megamitochondria in many cases are virtual likenesses of the altered mitochondria in the alcohol-high-fat rats, having several stacked cristae at either pole and an enhanced matrix compartment. Many of the giant mitochondria have at least one expanded crista, which contains packets of helically coiled filaments. When control rats are fed carbonyl iron without ethanol, the pancreatic mitochondria display a marked propensity for forming clusters of tightly interlocked organelles, formations that may be a prelude to mitochondrial fusion. CONCLUSIONS: It appears that iron supplementation sets the stage for organelle fusion, hence enlargement, with the ethanol providing the stimulus for fusion to actually take place, and controlling the final morphology of the resultant megamitochondria.

Accelerated response to reinoculation in experimental allergic encephalomyelitis: histopathologic study.

Experimental allergic encephalomyelitis (EAE) was induced in Lewis rats by the injection of spinal cord tissue or myelin basic protein and adjuvants (Freund's or carbonyl iron or pertussis vaccine), or by adoptive immunization. After an interval of five to 12 weeks, the recovered rats were reinoculated by a different route and usually with a different adjuvant. The onset of the second attack was determined by the histologic detection of EAE lesions at intervals during the incubation period. In each of ten experiments, the second attack of EAE occurred one or two days earlier than in naive controls injected at the same time. Residual EAE lesions left over from the first attack could not explain the findings in the reinoculated rats. The accelerated response to the second inoculation may be related to the anamnestic response of classical immunology or to residual damage to the blood-brain barrier. Resistance to a second attack was not encountered in this histopathologic study.

Development of a short-term inhalation bioassay to assess pulmonary toxicity of inhaled particles: comparisons of pulmonary responses to carbonyl iron and silica.

This paper describes a short-term inhalation bioassay for evaluating the lung toxicity of inhaled particulate materials. To validate the method, rats were exposed for 6 hr or 3 days to various concentrations of either aerosolized alpha-quartz silica or carbonyl iron particles. Cells and fluids from groups of sham- and dust-exposed animals were recovered by bronchoalveolar lavage (BAL). Alkaline phosphatase, lactate dehydrogenase (LDH), and protein values were measured in BAL fluids at several time points postexposure. Cells were identified, counted, and evaluated for viability. Pulmonary macrophages (PM) were cultured and studied for morphology, chemotaxis, and phagocytosis by scanning electron microscopy. The lungs of additional exposed animals were processed for histopathology and transmission electron microscopy. Brief exposures to silica elicited a sustained granulocytic inflammatory response (primarily neutrophils) with concomitant increases in alkaline phosphatase, LDH, and protein in the lavage fluids (p less than 0.05). In addition, PM functional capacity was depressed (p less than 0.05) and histopathologic lesions were observed within 1 month after exposure. In contrast, 6-hr or 3-day exposures to CI produced no cellular, cytotoxic, or alveolar/capillary membrane permeability changes at any time postexposure. PM function was either enhanced or unchanged from controls. These data demonstrate that short-term, high-dose inhalation exposures of silica produce effects similar to those previously observed using intratracheal instillation or chronic inhalation models, and lend support to this method as a reliable short-term bioassay for evaluating the pulmonary toxicity and mechanisms associated with exposures to new and untested materials.

Effect of dietary iron overload on lipid peroxidation, prostaglandin synthesis and lymphocyte proliferation in young and old rats.

The effect of dietary iron overload on lipid peroxidation (LP), prostaglandin (PG) synthesis and lymphocyte proliferation was examined in young and old F344 rats. Rats 4 and 19-22 mo old were fed AIN-76 diet for 11-12 wk supplemented with 2.5% carbonyl iron obtained from two sources (Type A and B). Animals supplemented with Type A iron showed reduced food intake and weight gain associated with marked increases in extrahepatic and hepatic iron concentration. Rats receiving Type B iron had food intakes and body weights similar to those of controls but exhibited small increases in tissue iron concentration. Old control rats compared to young had significantly higher conjugated dienes (CD) in hepatic microsomes. Feeding Type A iron diets induced a significantly higher level of CD in hepatic microsomes from old rats compared to young rats. Iron overloaded rats also showed highly correlated (r = 0.94) increases in the urinary excretion of thiobarbituric acid-reactive substances and PG metabolites indicating increased in vivo LP and PG synthesis. Mitogen-stimulated PGE2 synthesis in young rats was increased at 4 wk in association with enhanced T-cell proliferation stimulated by Concanavalin A. Lymphocyte proliferation was significantly lower in old than in young control or iron-treated rats. The lack of efficacy of Type B vs. Type A iron appears due to a larger particle size and lower bioavailability. In conclusion, iron overloading increases in vivo LP and PG metabolism. Furthermore, the mitogenic response to Concanavalin A in young rats is enhanced after 4 wk of iron overloading.

[Antimetastatic activity of spleen macrophages and T-lymphocytes in mice with transplantable tumors]. / Antimetastaticheskaia aktivnost' makrofagov i T-limfotsitov selezenki myshei s perevivnymi opukholiami.

The antimetastatic activity (AA) of spleen macrophages and T-lymphocytes of mice bearing four syngeneic tumours were tested in adoptive transfer system. Elimination of phagocytic cells by treatment with carrageenan or by carbonyl iron resulted in a complete (tumours LS, MMT1, MMT-T6I) or partial (MMT-T6I) AA decrease. Spleen macrophages (adherent cells) of tumour-bearers possessed a significant AA. The treatment of spleen cells both by anti-Thy-1-serum and by complement enhanced AA of spleen cells of LS and MMT1 tumour-bearers, but led to a partial AA suppression of spleen cells of MMT-T6I tumour-bearers. Thus, the efficiency of antimetastatic defence may considerably depend on the presence of synergism between macrophages and T-lymphocytes in realization of their AA.

Ultrastructural changes in the pancreas of carbonyl iron-fed rats.

Diabetes mellitus is found with increased frequency in patients with both primary and secondary hemochromatosis. In these conditions, the pancreas shows fibrosis and iron overload of acini, interstitium, and islet B cells. Previous morphological studies have only described changes found in advanced stages of disease, while abnormalities of the initial stage of iron overload have, as yet, not been reported. Rats fed a carbonyl iron-supplemented diet for 4-15 months showed storage iron deposition (ferritin and hemosiderin) in many organs, in a pattern similar to primary human hemochromatosis. Electron microscopic examination of the pancreas showed ferritin particles segregated in lysosomes of acinar cells, as well as diffuse cytosiderosis of macrophages in the interstitial septa. In the islets, iron deposits were discrete and only in B cells. In the absence of electron-microscopic studies of incipient pancreatic cytosiderosis in human subjects, the present experimental animal study may contribute to a better understanding of the pathway leading to the extensive lesions found in the advanced stages of the human iron overloading diseases.

Hepatic mitochondrial oxidative metabolism and lipid peroxidation in experimental hexachlorobenzene-induced porphyria with dietary carbonyl iron overload.

Both human porphyria cutanea tarda and experimental hexachlorobenzene-induced porphyria are associated with hepatic injury and are potentiated by excess hepatic iron. The mechanisms whereby cellular injury occurs and the synergistic role of iron overload are unknown. In the present experiments, we studied hepatic mitochondrial function and lipid peroxidation in rats with hexachlorobenzene-induced porphyria in which iron loading was achieved by dietary carbonyl iron supplementation. Female rats were treated for 8 weeks, receiving a chow diet supplemented with hexachlorobenzene (0.2%, w/w), carbonyl iron (1.0%, w/w) or hexachlorobenzene + iron. Hepatic total porphyrins were increased 100-fold in rats receiving hexachlorobenzene (hexachlorobenzene alone and hexachlorobenzene + Fe), and total hepatic iron was increased approximately 10-fold in rats receiving iron supplementation (Fe alone and hexachlorobenzene + Fe). There was a significant increase in mitochondrial lipid peroxidation in rats treated with hexachlorobenzene alone and hexachlorobenzene + Fe. A significant reduction in mitochondrial respiratory control ratios and in oxidative phosphorylation (ADP/O ratios) using glutamate and succinate as substrates was demonstrated when rats were treated with hexachlorobenzene + iron. The reductions in respiratory control ratios were due to a combination of an inhibitory defect in electron transport as evidenced by an irreversible decrease in State 3 respiration and an uncoupling effect as evidenced by an increase in State 4 respiration. These findings suggest that lipid peroxidation and mitochondrial dysfunction may contribute to the hepatotoxicity seen in hexachlorobenzene-induced porphyria.

The hyperacute form of allergic encephalomyelitis produced in rats without the aid of pertussis vaccine.

The hyperacute form of experimental allergic encephalomyelitis (EAE), characterized by a short incubation period, severe paralysis, high mortality, and abundant polymorphonuclear leukocytes and fibrin in the lesions, was produced in rats without the use of pertussis vaccine (previously considered an essential requirement) or Freund's adjuvant. Carbonyl iron or mineral oil without mycobacteria were effective adjuvants and whole rat spinal cord was the best antigen. Hyperacute EAE was produced in this manner in some Lewis rats, most dark agouti (DA) rats and most F1 hybrids of these two strains. Clinical signs were earlier in onset and more severe in the DA strain than in the Lewis strain in all adjuvant-antigen combinations that were tested. Dark agouti rats developed clinical signs in six days, histological lesions in five days, and localized EAE lesions could be induced in four days. The data support the hypothesis that hyperacute type lesions (neutrophils and fibrin) can be caused by an exceptionally strong immune response to neural antigen, whether that response is engendered by a particular adjuvant (pertussis vaccine) or by an unusual degree of genetic susceptibility (DA rats).

High-dose carbonyl iron for iron deficiency anemia: a randomized double-blind trial.

To determine if high doses of oral iron could shorten the duration of therapy necessary to treat Fe deficiency anemia, high-dose Fe 600 mg three times per day (given as nontoxic carbonyl Fe) was compared with standard ferrous sulfate 60 mg Fe++ three times per day in a randomized, double-blind, 3-wk trial involving 36 female blood donors with mild Fe deficiency anemia. In animal studies, both forms of Fe have similar bioavailability when administered in equal amounts. High-dose carbonyl Fe was well tolerated with gastrointestinal side effects similar those observed with standard FeSO4 therapy. The 10-fold larger amount of Fe resulted in a mean 1.5-fold increase in estimated Fe absorption. Both regimens corrected anemia but neither replenished storage Fe. These results suggest that the principal advantage to the use of carbonyl Fe would derive from its safety rather than from the large doses that can be given.