Low Ceruloplasmin Levels Research Articles

Roles of copper accumulation in hepatocarcinogenesis of LEC rats

The aim of this study was to discover possible mechanisms of hepatocarcinogenesis of the Long-Evans cinnamon (LEC) rat, a new mutant inbred strain that manifests severe jaundice and anemia a few months after birth. The animals have an abnormal accumulation of copper in the liver and a low level of ceruloplasmin in the plasma. The LEC rat may be considered a useful animal model for Wilson's disease and hepatocellular carcinoma. J. Trace Elem. Exp. Med. 10:73–79, 1997. © 1997 Wiley-Liss, Inc.

Wilson Disease

Wilson disease (hepatolenticular degeneration) is an autosomal recessive, inherited disorder of copper metabolism resulting in excessive accumulation of copper in the liver, brain, and other organs of the body. The manifestations of the disease are related directly to this accumulation of copper. Copper homeostasis normally is a product of the balance between intestinal absorption of dietary copper and hepatic biliary excretion of excess copper. In Wilson disease, incorporation of hepatic copper into ceruloplasmin is defective and excretion of copper in the bile is reduced. A low level of ceruloplasmin, which until a few years ago was erroneously considered to be the basis for the disease, is a consequence of the underlying metabolic defect.

Low ceruloplasmin levels during recovery from major burn injury: Influence of open wound size and copper supplementation

Copper (Cu) status is often judged by the plasma level of its chief transport protein, ceruloplasmin (Cp). Only copper deficiency and heredity are known to decrease circulating Cp. Cp is an acute-phase responsive protein in trauma and it is also induced by Cu supplementation. Despite this, plasma concentrations of Cp remain low during the acute recovery from major burn injury. The high provision of vitamin C typically used in burn patients may influence these observations when an indirect oxidase activity assay is used. We employed a radial immunodiffusion (RID) assay specific for the Cp protein as well as an indirect oxidase assay for Cp in a series of 11 burned children who were supplemented with both Cu and vitamin C, either enterally or parenterally. Our findings confirm that low Cp is a characteristic of the acute recovery from major burns. The oxidase assay is shown to be valid for very low Cp levels even during high vitamin C provision. When these data are combined with our previously reported series, a strong relationship between the size of the open wound area and the amount of circulating Cp is demonstrated. Copper supplementation by either the enteral or parenteral routes is only marginally successful in restoring Cp toward normal levels.

Excess Copper and Ceruloplasmin Biosynthesis in Long-term Cultured Hepatocytes from Long-Evans Cinnamon (LEC) Rats, a Model of Wilson Disease

Immortalized hepatic cell lines obtained from laboratory animals or patients with defects in copper metabolism in the liver provide new approaches to examine related metabolism and toxicity. We established a series of hepatic cell lines from the liver of Long-Evans Cinnamon (LEC) rats, using recombinant adenovirus which expresses SV40 large T. Cells from the LEC rats were cultured and accumulated larger amounts of copper than did control cells, when the concentrations of copper in the culture medium exceeded 5 microM. The secretion of ceruloplasmin (CP) from the cultured cells was not reduced in hepatocytes from LEC cells, as compared with the control cells. As accumulation of copper did not affect CP secretion, CP production was not likely to be affected by the accumulation of copper in LEC rat hepatocytes. The production of holo-CP was further investigated by transfection of human CP cDNA and detection of human holo-CP by immunological procedures and use of a monoclonal antibody (mAb CP60) which recognizes human holo-CP but not human apo-CP and rat CP. Hepatocytes from the LEC rats processed and secreted holo-CP into the medium, even with excess copper present in the medium. These observations suggest that the genetic defect in LEC rats did not alter biosynthetic and secretory pathways of CP and that the intracellular copper concentration did not regulate the synthesis and processing of CP in the cultured hepatocytes. Low ceruloplasmin levels are observed in most, but not all, patients with Wilson disease, as well as in LEC rats. Our results do suggest that the copper transporting ATPase encoded in the Wilson disease gene is not a integral part of the biochemical mechanism of copper incorporation into apoprotein. The cell lines and immunological procedures we used are expected to add to information on biologically important process related to copper metabolism and to CP biosynthesis.

Plasma Copper and Antioxidant Status in Wilson's Disease

It has been demonstrated that the level of serum copper unbound to ceruloplasmin (loosely bound copper) is increased in Wilson's disease, although the total serum copper concentration is usually low, reflecting a low ceruloplasmin level. To assess the contribution of free radical reactions catalyzed by nonceruloplasmin copper to the development of complications in this disease, we investigated copper and antioxidant status in four untreated patients who had hepatic dysfunction with or without hemolytic anemia and made a comparison with five patients controlled on penicillamine therapy and 19 age-matched healthy children. We found that loosely bound copper in plasma measured by the phenanthroline assay was detectable in three of four untreated patients with Wilson's disease, but was not detectable in the patients during therapy or in the healthy controls. Among the various antioxidants, the ascorbate and urate levels were markedly reduced before treatment (mean +/- SD, 23 +/- 16 microM for ascorbate and 90 +/- 59 microM for urate) compared with the values in the patients during treatment with penicillamine (67 +/- 19 and 302 +/- 78 microM, p < 0.05) and in control children (60 +/- 8 and 254 +/- 48 microM, p < 0.05). We also demonstrated that the plasma concentration of allantoin, an oxidation product of uric acid and a possible marker of radical generation in vivo, was markedly elevated in the untreated patients (11.0 +/- 1.8 versus 4.3 +/- 0.5 microM in patients on therapy and 6.5 +/- 0.8 microM in controls, p < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Studies on the relationship between sway disease of bactrian camels and copper status in Gansu province.

The clinical signs of camel sway disease in the Hexi Corridor of Gansu province were defined. The contents of eight minerals in soils, in forage and in the blood and hair of bactrian camels from this region were determined. The related blood indices were also measured. The concentration of molybdenum in soils and forage was 4.8 +/- 0.02 and 4.8 +/- 0.25 micrograms/g (dry matter), respectively, the copper to molybdenum ratio in the forage being only 1.3. The concentration of copper in blood and hair from the camels was 0.28 +/- 0.17 micrograms/ml and 3.50 +/- 1.00 micrograms/g, respectively. There was a hypochromic microcytic anaemia and a low level of ceruloplasmin in the blood. It is therefore suggested that sway disease of bactrian camels in this region is caused by secondary copper deficiency, mainly due to the high molybdenum content in soils and forage. The copper deficiency in the camels was aggravated during reproduction. Oral administration of copper sulphate can prevent and cure the disease.

Copper and zinc metabolism in aminonucleoside-induced nephrotic syndrome.

Copper (Cu) and zinc (Zn) were measured in urine, serum and tissues from rats with nephrotic syndrome (NS) induced with a single subcutaneous dose of puromycin aminonucleoside (PAN; 15 mg/100 g BW). Control animals were pair-fed. Urine was collected daily, and the rats were sacrificed on day 10. PAN-nephrotic rats had proteinuria (days 3-10), high urinary Cu (days 1, 2, 4-10) and Zn (days 3-10) excretion. On day 10, nephrotic rats had: (a) albuminuria, hypoalbuminemia, hypoproteinemia, high urine and low serum levels of ceruloplasmin; (b) low Cu and Zn serum levels; (c) high clearance and fractional excretion of Cu and Zn, and (d) low kidney and liver Cu content and essentially normal tissue Zn levels. The alterations in Cu metabolism were more intense than those in Zn metabolism. Urine Cu and Zn showed a positive correlation with urine total protein on days 3-10 which suggests that high urinary excretion of Cu and Zn may be due to the excretion of its carrier proteins. In conclusion, these rats did not show a typical Zn deficiency but a clear decrease in Cu in the liver and kidney.

Hereditary Low Level of Plasma Ceruloplasmin in LEC Rats Associated with Spontaneous Development of Hepatitis and Liver Cancer

Both young (5 weeks old) and old (61 100 weeks old) hereditary hepatitis LEC rats showed a markedly low level of plasma ceruloplasmin (Cp) ferroxidase activity as compared with that of age‐matched LEA and BN strain rats. This trait was genetically examined hy the use of (BN × LEC) F1 hybrid and (F1× LEC) backcross rats. The F1 hybrids never developed hepatitis and showed a similar level of Cp to that found in the parental BN rats. Among the backcross rats with about 1:1 segregation rate for hepatitis, affected rats had a remarkably decreased level of Cp, as found in LEC rats, whereas unaffected rats exhibited a similar level of Cp to that of BN, F1 and LEA rats. These results indicate that the low level of Cp is heritable in a single autosomal recessive mode in LEC rats. The observed tight link between the low Cp level and the hepatitis in LEC rats suggests that defective copper metabolism may he associated with the occurrence of hepatitis in LEC rats, since Cp is a copper‐binding protein primarily involved in copper transport from the liver.

Copper Deficiency during Total Parenteral Nutrition: Clinical Analysis of Three Cases

Three adult cases in which copper deficiency developed during long-term total parenteral nutrition (TPN) without copper supplementation have been described, together with a brief review of the literature. All three patients were suffering from malabsorption when TPN was instituted, and overt symptoms of copper deficiency developed an average of 5.8 months after the start of TPN. Clinically, leukopenia with neutropenia and low plasma levels of copper and ceruloplasmin were seen in all cases. The dosage of copper administration in these cases was 0.3 to 7.2 mg of copper/day, or 5.3 to 133 micrograms of copper/kg/day, with total doses of 7 to 176 mg of copper.

Development of Anemia in Copper-Deficient Rats Fed High Levels of Dietary Iron and Sucrose

The effects of dietary carbohydrate and iron on the development of copper deficiency were examined. Male Sprague-Dawley rats (n = 48) were limit-fed one of eight diets in a 2 × 2 × 2 factorial design for 19 d. Two levels of copper (0.85 or 8.6 µg Cu/g diet) and iron (54 or 226 µg Fe/g diet) and two types of carbohydrate (sucrose or cornstarch, 65.3%) were fed. Compared with control rats, copper-deficient rats had lower hematocrits, lower ceruloplasmin levels, lower tissue levels of copper and increased hepatic iron levels. Copper-deficient rats fed sucrose had significantly lower hematocrits, lower apparent absorption of copper, lower liver iron levels and higher plasma triglyceride levels than copper-deficient rats fed cornstarch. Copper-deficient rats fed sucrose with 226 µg Fe/g diet had hematocrit levels that were 15% lower than all other copper-deficient levels and 23% lower than control levels. Tissue levels of copper among copper-deficient rats were not affected by the type of carbohydrate or by the level of dietary iron. These data indicate that both high iron and sucrose can affect the development of the copper deficiency.

Serum copper, serum ceruloplasmin, and erythrocyte sedimentation rate measurements in children with Hodgkin's disease, non-Hodgkin's lymphoma, and nonmalignant lymphadenopathy.

Between 1975 and 1981, 29 children with Hodgkin's disease, 22 with non-Hodgkin's lymphoma, and 13 with lymphadenopathy from various nonmalignant causes were studied with respect to the usefulness of serum copper, ceruloplasmin, and erythrocyte sedimentation rate (ESR) as markers of disease activity. Although elevated copper and ceruloplasmin levels were indicators of disease activity in the childhood lymphomas, and in patients with Hodgkin's disease the levels tended to be higher in those with advanced clinical stage, they were no better markers of disease activity than the ESR. Moreover, elevated serum copper and ceruloplasmin were nonspecific, since one child with benign lymphadenopathy and one Hodgkin's disease patient with an infection had raised levels. Low levels of copper and ceruloplasmin were found in four jaundiced patients probably because of impaired hepatic synthesis of ceruloplasmin. The authors conclude that serum copper and ceruloplasmin levels are no better markers of disease activity in children with lymphomas than is the ESR.

Future of cardiothoracic surgery.

<h3>Objective.</h3> To delineate urine biomarkers that forecast response to therapy of lupus nephritis (LN). <h3>Methods.</h3> Starting from the time of kidney biopsy, patients with childhood-onset systemic lupus erythematosus who were diagnosed with LN were studied serially. Levels of 15 biomarkers were measured in random spot urine samples, including adiponectin, α-1-acid glycoprotein (AGP), ceruloplasmin, hemopexin, hepcidin, kidney injury molecule 1, monocyte chemotactic protein-1, lipocalin-like prostaglandin D synthase (LPGDS), transforming growth factor-β (TGF-β), transferrin, and vitamin D binding protein (VDBP). <h3>Results.</h3> Among 87 patients (mean age 15.6 yrs) with LN, there were 37 treatment responders and 50 nonresponders based on the American College of Rheumatology criteria. At the time of kidney biopsy, levels of TGF-β (p &lt; 0.0001) and ceruloplasmin (p = 0.006) were significantly lower among responders than nonresponders; less pronounced differences were present for AGP, hepcidin, LPGDS, transferrin, and VDBP (all p &lt; 0.05). By Month 3, responders experienced marked decreases of adiponectin, AGP, transferrin, and VDBP (all p &lt; 0.01) and mean levels of these biomarkers were all outstanding (area under the receiver-operating characteristic curve ≥ 0.9) for discriminating responders from nonresponders. Patient demographics and extrarenal disease did not influence differences in biomarker levels between response groups. <h3>Conclusion.</h3> Low urine levels of TGF-β and ceruloplasmin at baseline and marked reduction of AGP, LPGDS, transferrin, or VDBP and combinations of other select biomarkers by Month 3 are outstanding predictors for achieving remission of LN. If confirmed, these results can be used to help personalize LN therapy.

Identification and Purification of a Non-Ceruloplasmin Ferroxidase of Human Serum

A non-ceruloplasmin ferroxidase (ferroxidase-II) protein was isolated from human serum and completely resolved from ceruloplasmin by DEAE-Sephadex A-50 chromatography. This protein was isolated and purified approximately 500-fold by utilization of the Cohn IV-1 fraction, DEAE-Sephadex A-50 chromatography, and gel filtration on Agarose A-15m, followed by gel filtration on Agarose A-50m. Ferroxidase-II differed from ceruloplasmin in many respects. Three of the most important differences were that ferroxidase-II (a) was yellow rather than blue as ceruloplasmin, (b) it was not inhibited by azide, and (c) it exhibited no p-phenylenediamine oxidase activity. Ferroxidase-II also differed significantly from the known plasma amine oxidases. The ferroxidase activity of ferroxidase-II was not lost by dialysis or ultrafiltration, but was inactivated by heat treatment and was proportional to the protein concentration at all stages of purification. Oxygen consumption with ferroxidase-II was observed simultaneously with iron oxidation. The ferroxidase-II activity in Wilson's disease serum represented a much larger percentage of the total ferroxidase activity than in normal serum. Although ferroxidase-II was decreased in Wilson's disease serum, it was reduced to a lesser extent than the ceruloplasmin ferroxidase activity. Thus ferroxidase-II may account for the lack of correlation of ferroxidase activity with p-phenylenediamine oxidase activity of Wilson's disease serum and may be responsible for the maintenance of near normal iron metabolism despite the low levels of ceruloplasmin.

The behaviour of ceruloplasmin fractions in liver diseases.

Abstract. Ceruloplasmin fractions I(C‐C) and II(C‐D) have been estimated in patients with infectious hepatitis, liver metastases and cirrhosis of the liver. In infectious hepatitis and metastases serum ceruloplasmin content was increased and the percentage of fraction I(C‐C) was decreased; no correlation between these parameters could be found. In the group of cirrhotics mean serum ceruloplasmin concentration was normal but the mean percentage of fraction I(C‐C) was decreased. A positive correlation between these values could be demonstrated. Very low ceruloplasmin levels were found in some patients with the most severe form of liver cirrhosis; if this is the result of an impaired synthesis, it seems probable that especially fraction I(C‐C) is affected by this disturbance. The possibility that only fraction I(C‐C) is synthesized by the liver is discussed.

Factors influencing serum ceruloplasmin levels in normal individuals.

Abstract Serum ceruloplasmin levels are known to be altered by pregnancy and by disease, particularly Wilson's disease in which this protein is usually deficient. Normal healthy individuals show considerable variation in their ceruloplasmin levels. Some of the factors responsible for this variation have now been investigated. The ceruloplasmin levels of 309 normal unrelated individuals were determined by measuring the rate of oxidase activity of serum with paraphenylenediamine dihydrochloride. The method described has high precision and reproducibility. The ceruloplasmin levels varied with age, with the high levels of early childhood gradually declining to adult levels by 12 years of age. Sex differences in ceruloplasmin levels were found only during adolescence. The levels of adults showed little variation from day to day and were not influenced by body weight or the menstrual cycle. Familial factors were found to influence the ceruloplasmin levels. Two families were found in which low ceruloplasmin levels showed a regular pattern of inheritance.

Wilson's Disease

Whatever the cause of Wilson's disease there is one clear cut biochemical abnormality: an increased total body copper. It seems therapeutically plausible to remove the excess copper as quickly and completely as possible. This can be achieved by oral administration of penicillamine. With judicious but humane advice concerning the avoidance of foods containing a high copper content and the regular administration of penicillamine, the copper content of the body returns toward normal and in some cases is associated with undoubted clinical improvement. Replacement of a low ceruloplasmin level either by estrogen administration or by infusion of ceruloplasmin seems of little value. The regular infusion of apoceruloplasmin, to prevent absorption of copper and penicillamine to eliminate the copper already present, would be logical therapy for this particular inborn error of metabolismn and already the therapeutic nihilism epitomized in the statement that inherited diseases are cured only by the grave is happily mistaken.