Nutritional supplementation with copper in the rat. I. Effects on adjuvant arthritis development and on some in vivo- and ex vivo-markers of blood neutrophils.

OBJECTIVE AND DESIGN: The aims of the work were: 1) to confirm the preliminarily observed anti-arthritic potential of a 200 ppm copper-supplemented diet in the rat: 2) to study the impact of the nutritional treatment and of the experimental pathology on neutrophil activity. ANIMALS AND CELLS: Two hundred female Sprague-Dawley rats were used. Polymorphonuclear leukocytes were isolated from these animals for the ex vivo studies. TREATMENT: Control-rats were maintained on a standard diet containing 5 ppm of copper. Supplemented-rats were kept on a diet containing 200 ppm of the metal. METHODS: Mycobacterium butyricum-induced arthritis was studied. Flame atomic absorption spectroscopy was used to assess copper and zinc levels. The "microplate-assay" technique was used to determine serum lysozyme concentration (lysis of Micrococcus lysodeikticus cell walls), as well as neutrophil O2- generation (superoxide dismutase-inhibitable reduction of ferricytochrome-c), and adhesion (activity of the membrane enzyme acid phosphatase). The results were statistically evaluated by the Student's t test. RESULTS: The nutritional copper-supplementation: 1) significantly inhibited the adjuvant-arthritis development (33% +/- 5, P<0.01); 2) did not modify lysozyme secretion or superoxide production; 3) significantly decreased the percentage of cell adhesion by an average of 41% +/- 19 (P<0.01). CONCLUSIONS: The copper-supplemented diet has an anti-arthritic effect which may be also primed by the effect of copper on the expression of the neutrophil cell-adhesion molecules.

Comparison of the effects of the antimetastatic compound ImH[trans-RuCl4(DMSO)Im] (NAMI-A) on the arthritic rat and on MCa mammary carcinoma in mice.

The effects of the new molecule ImH[trans-RuCl4(DMSO)Im] (NAMI-A), administered orally or intraperitoneally to adjuvant-arthritic rats or orally to mice bearing s.c. or i.m. implants of MCa mammary carcinoma, were studied. NAMI-A was not able to modify the progression of chronic inflammation in the complete Freund-adjuvant injected animals. Histology indicated a significant worsening of the inflammatory process, characterised by an increased infiltration of inflammatory cells, as well as by a remarkable deposition of connective tissue fibres around the blood vessels and alveolar walls. NAMI-A had no effect on primary i.m. implanted MCa mammary carcinoma growth and its lung metastasis formation, but significantly interfered with the cell cycle of primary tumor cells following bolus oral administration. On the contrary, NAMI-A caused a significant inhibition of lung metastasis accompanied by a dramatic deposition of connective tissue fibres around the primary tumor mass, when given as medicated food to mice implanted s.c. with MCa tumor. These data indicated that NAMI-A is well absorbed after oral administration although there is no connection between lung concentration and the antimetastatic activity. Conversely, the marked deposition of connective tissues in NAMI-A treated animals is in agreement with the reported effects of the compound on extracellular matrix and tumor blood vessels.

Neutrophil behavior following exposure to in vivo or in vitro zinc in normal and acutely-inflamed rats: studies on lysozyme secretion, superoxide anion release and platelet adhesion.

The mechanism was studied of the anti-inflammatory effect of oral zinc (114 mg/kg/day of elemental metal, given for 14 days) on the development of the carrageenan-induced paw oedema of the rat, and the impact of in vivo treatment on the activity of neutrophils isolated from the blood of inflamed and non-inflamed animals. The effects of the in vitro incubation with the metal on either non-inflamed or inflamed neutrophils coming from zinc-untreated rats were also examined. It was found that the administration of oral zinc inhibited markedly the process of ex vivo adhesion of the cells obtained from the inflamed rats (an observation confirmed by the in vitro experiments). In vitro release of lysozyme and superoxide anion productions were measured: in the absence of zinc, the 30' of pre-incubation carried out before stimulating with PMA did not influence the cell's reactivity of the non-inflamed neutrophils. It was, on the contrary, capable of significantly reducing that of the inflamed ones. As a consequence, it is quite difficult to properly interpret the data obtained studying the activity of the cells exposed to the metal in vitro.

Blood copper and zinc changes in runners after a marathon.

This study reports plasma and total blood cell (TBC) Cu and Zn levels of 16 runners before and after a marathon race. All the pre-race plasma values were in the normal range, while the TBC Cu level was 29.3% (P < 0.01) lower and the TBC Zn level was 29.5% (P < 0.01) higher than mean values of the control group. The run induced a significant decrease in TBC Cu concentration during the days following the race and a light increase in plasma Cu values. It caused a significant increase in TBC Zn concentration at the end of the race (followed by a decrease 24 hours later) without plasma Zn variations. These data show that intense physical exercise could modify trace-element metabolism.

Copper and zinc body levels in inflammation: an overview of the data obtained from animal and human studies.

The development of acute and chronic inflammatory processes induces, in the laboratory animal, a net accumulation of both copper and zinc in many body compartments, the inflamed area included. In rheumatoid arthritis, as well as in animal models, only plasma zinc concentration seems to be significantly correlated with disease severity, while the increase in total plasma copper could be described as an "all or nothing" phenomenon. Moreover, in rheumatoid arthritis, it appears that the disease develops and progresses without being linked to either copper or zinc deficiency conditions. Thus, it seems reasonable to suggest that a rationale for the use of copper and/or zinc in the treatment of inflammatory disorders can only be drawn from the intrinsic pharmacological properties of such trace elements, rather than from the need for their repletion.

Copper and zinc status in rheumatoid arthritis: studies of plasma, erythrocytes, and urine, and their relationship to disease activity markers and pharmacological treatment.

We studied the status of copper and zinc in rheumatoid arthritis (RA). The aims of the work were to ascertain whether or not RA is associated with copper and/or zinc deficiency, to establish the relationship between these trace metals and the main biohumoral and clinical indices of the disease, and to examine the effect on copper and zinc of the drugs normally used by RA patients. Metal levels were measured by atomic absorption spectroscopy in the plasma, whole blood cells and 24 hr urine of 120 RA patients; 70 patients suffering from primary osteoarthritis were used as the control group. In the plasma of RA patients copper and ceruloplasmin levels were found to be significantly increased whereas zinc levels were significantly decreased. No major variations were observed in the blood cell and 24 hr urine copper and zinc levels. Plasma copper was significantly correlated with some of the biohumoral markers of RA, but did not correlate with any of the clinical indices of the disease. Plasma zinc was significantly correlated with numerous of the biohumoral as well as clinical markers of RA. With the exception of an increased urinary excretion of copper in D-penicillamine treated RA patients, drug therapy did not influence the copper status in RA. Conversely, plasma zinc was found to be lower in RA patients taking NSAIDs and/or steroids. These results suggest the following conclusions: i) RA patients do not seem to be deficient in either copper or zinc; ii) plasma copper appears to be a poor index of RA severity; iii) plasma zinc could have some practical value in defining the overall severity of the disease.

Oral zinc as initial therapy in Wilson's disease: two years of continuous treatment in a 10-year-old child.

Two years of continuous therapy promoted a significant overall amelioration in a 10-year-old boy affected by an hepatic form of Wilson's disease in which zinc sulphate was the sole therapy. In particular, liver function returned to normal and hepatic histology also improved. The parameters characterizing copper metabolism were kept under good control, and a decrease in copper concentration was found in both erythrocytes and liver. The copper balance study performed during the 25th month of treatment showed that oral zinc was still efficiently inhibiting the intestinal absorption of copper. No side effects have been reported so far.

[Zinc sulfate in the treatment of psoriatic arthritis]. / Il solfato di zinco nella terapia dell'artrite psoriasica.

We evaluated oral zinc sulphate as a disease-modifying antirheumatic drug (DMARD) in psoriatic arthritis in a preliminary open uncontrolled trial. Twenty patients with psoriatic arthritis were given oral zinc sulphate three times a day at total dose of 600 mg/die, i.e. 120 mg/die of elemental zinc, for 6 months. The 18 patients who completed the trial showed a significant decrease in the number of swollen (p less than 0.01) and tender (p less than 0.05) joints, Ritchie articular index (p less than 0.01), need for nosteroidal anti-inflammatory drugs (p less than 0.001), erythrocyte sedimentation rate (p less than 0.01), and plasma copper level (p less than 0.001). We suggest that zinc sulphate may be an effective and well-tolerated DMARD in psoriatic arthritis. To more strictly verify these preliminary data, a double-blind placebo controlled study is in progress.

Oral zinc sulphate as primary therapeutic intervention in a child with Wilson disease.

An 8-year-old boy with an hepatic form of Wilson disease was treated with oral zinc sulphate as the primary and sole therapy. After 4 months, liver function had dramatically improved, and the parameters characterizing copper metabolism had also normalized.

Copper and zinc status in rats with acute inflammation: focus on the inflamed area.

Status of copper and zinc in plasma, blood cells, liver and hind paws (sectioned at the tibio-tarsal joint) were evaluated in rats with carrageenan-induced paw-oedema; moreover, concentrations of copper and zinc in the supernatant and cell fractions obtained from exudates pooled from rats with carrageenan-induced pleurisy were also determined. The evaluation of copper and zinc status in the blood and in the liver of rats with carrageenan-induced paw oedema, showed that only minor variations differentiated this experimental pathology from the previously studied carrageenan-induced pleurisy in rat. In inflammatory exudates withdrawn from pleural cavity, copper concentrations were found to be higher than the basal values measured in the whole paw, whereas zinc concentrations were found to be dramatically lower. Thus, the induction of the carrageenan paw-oedema determined an increase in copper and a decrease in zinc concentrations in the inflamed paw; however, in the inflamed paw, the total amounts of both copper and zinc were found to be significantly increased.

Copper and zinc status in adjuvant-arthritic rat: studies on blood, liver, kidneys, spleen and inflamed paws.

The status of copper and zinc in plasma, blood cells, kidneys, spleen and hind paws was evaluated in tail-injected adjuvant-arthritic rats, during both the asymptomatic (3 and 7 days after the inoculum) and symptomatic (14, 21 and 30 days after the inoculum) phases of the experimental disease. During the symptomatic phase, inflamed rats were studied divided into two groups on the basis of their arthritic scores (low-score L.S. and high-score H.S. arthritic rats). Copper (both in concentration and total amount) was found significantly increased in plasma, blood cells, liver, spleen and arthritic paws, whereas, in the kidneys, it was found to be lower than normal. Zinc was found to be remarkably increased in the liver. In blood, zinc was found to be decreased in plasma, but almost unchanged in the cellular fraction. Zinc total amount (but not concentration) was increased in the spleen, most likely because of a significant increase in spleen weight. As previously described in the case of acute inflammation, zinc concentration was found to be significantly decreased in arthritic paws, whereas the total amount of the metal present in these inflamed tissues was higher than normal. The status of copper and zinc may well differentiate L.S. from H.S. arthritic rats, especially during the latest phase of the experimental disease, and particularly because of a normalization of the considered parameters in the low-score group. Many of the changes observed in the status of both metals were seen prior the appearance of arthritis. The overall accumulation of copper and zinc which is induced in rat by the development of adjuvant arthritis, is suggested to further sustain the hypothesis of increased body requirements for both metals during inflammation.

Copper and zinc status during acute inflammation: studies on blood, liver and kidneys metal levels in normal and inflamed rats.

The concentrations of copper and zinc in plasma, blood cells, liver and kidneys were determined in a study performed on normal female rats, and in female rats with carrageenan induced pleurisy. In the normal rat, the total amount of both metals increases, from 51 to 79 days of age, in all the compartments examined. This increase was mostly, and in some case exclusively, dependent upon the growth of the animal, although high individual and day to day variations in both copper and zinc values were observed in all the compartments studied. In the blood of inflamed rats a statistically significant increase in copper was measured during the crucial hours of the experiment (i.e. from 6 to 72 h); over 90% of the increase found was attributable to variations in plasma copper concentration values. In the liver of inflamed rats a statistically significant increase in zinc was measured at 6, 22 and 48 h after the carrageenan injection. The induction of the acute non-infective inflammatory process did not cause quantitative changes of both copper and zinc in all the other compartments considered in the present study. These results seem to suggest that, during acute inflammation, the organism increases its requirement for copper and zinc, and that this demand is fulfilled by enhanced intestinal absorption and/or decreased intestinal excretion of both metals.

Copper and inflammation--a possible rationale for the pharmacological manipulation of inflammatory disorders.

Acute and chronic inflammations are characterized, among other features, by changes in the metabolism of copper and by a widespread responsiveness to the therapy with copper-containing molecules. The exact map of inflammation-induced copper movements as well as the role played by the metal in the pathogenesis of inflammatory disorders are, however, far from being clear, and this is especially true in the case of chronic processes. Nevertheless the present knowledge suggests that the "copper approach' may provide a new way for coping with the problem of anti-inflammatory/anti-arthritic therapies. The administration of exogenous copper, and the in vivo manipulation of the endogenous metal levels are proposed as two possible therapeutic strategies, not necessarily mutually exclusive. For a better understanding of the value of such an approach, further research work is needed, especially to attain a more detailed know-how on the involved chemical forms, distribution and functions of copper in both normal as well as inflamed organisms.

Copper metabolism in the acute inflammatory process and its possible significance for a novel approach to the therapy of inflammation.

The concentration and the total content of copper were evaluated in plasma, blood cells, liver and kidney of normal rats and in rats inflamed by intrapleural injection of carrageenan. The experimental disease was followed from zero time up to 10 days after the irritant injection. A statistically significant increase of copper concentration in both plasma and blood cells compartments was measured between zero and 72 h, followed by a prompt return to normal copper levels. On the contrary, copper concentration and total copper content of the liver did not vary throughout the experiment, whereas in the kidney both parameters were found slightly higher than normal 6, 8 and 10 days after the injection of the irritant. These observations are briefly discussed, and proposed as a valuable background to outline new strategies for anti-inflammatory therapy.