Alzheimer’s Disease and Copper Biochemistry

23 May 2017

Siotto et al. [1] studied 84 patients with Alzheimer’s disease and observed that a one-unit increase in the specific activity of ceruloplasmin (the ratio of ceruloplasmin measured enzymatically to ceruloplasmin measured by immunoassay) was associated with a fall in disease risk of 89% in comparison to 58 healthy people.

Golden [2] suggests that decreased concentrations of copper in organs or serum along with decreased activities of enzymes dependent on copper can be used to assess copper malnutrition. Because some of these tests are rather insensitive indices of nutritional status, several were compared based on experiments with animals and people. It was concluded that this specific activity is a moderately sensitive measure of copper deficiency [3].

Eight women in early middle age were fed a conventional diet low in copper in a metabolic ward for 135 days. Copper depletion was detected and normal copper status was restored later. The specific activity of ceruloplasmin was one of the more sensitive measurements of deficiency [4]. Perhaps the patients of Siotto et al. [1] with higher specific activities were less deficient than those with lower values.

It is hypothesized that Alzheimer’s disease is caused by copper deficiency for many reasons [5]. Most important: the hypothesis explains why people with Down’s syndrome get Alzheimer’s disease earlier in life than others. The trisomy of chromosome 21 induces an excess of copper/zinc superoxide dismutase which depends on copper for activity [6, 7] and thus causes a concomitant increase in the requirement for dietary copper.

The authors [1] also noted smaller associations with iron metabolism and that non-ceruloplasmin copper (also known as “free” copper or labile copper) predicted risk. According to Linder and Goode [6], copper in plasma is found in ceruloplasmin, albumin, transcuprein and smaller molecules in descending order. Ceruloplasmin is a copper source for the other fractions. As noted above [3], copper in these fractions changes unevenly in deficiency. Large effects on iron metabolism from copper deficiency have been known for nearly a century [8].

Measurements of biochemistry and cognitive function should be made on patients with Alzheimer's disease who are supplemented with copper. At least 4 mg of copper should be given daily as a well-absorbed copper compound such as copper gluconate. More biochemical measurements on animals deficient in copper also will be helpful. Erythrocyte and extracellular superoxide dismutase, cytochrome c oxidase in platelets and lysyl oxidase in serum are suggested [9], inter alia. The proposed experiments may help to clarify the utility of ceruloplasmin specific activity and other aspects of copper metabolism in the study of Alzheimer’s disease.

Leslie M. Klevay
Professor Emeritus of Internal Medicine, University of North Dakota, School of Medicine and Health Sciences, 1301 North Columbia Rd., Grand Forks, ND, USA. Email: leslie.klevay(a)med.und.edu

References
[1] Siotto M, Simonelli I, Pasqualetti P, Mariani S, Caprara D, Bucossi S, Ventriglia M, Molinario R, Antenucci M, Rongioletti M, Rossini PM, Squitti R (2016) Association between serum ceruloplasmin specific activity and risk of Alzheimer's disease. J Alzheimers Dis 50, 1181-1189.
[2] Golden MHN (1996) Severe malnutrition. In Oxford Textbook of Medicine, Weatherall DJ, Ledingham JG, Warrell DA, eds. Oxford University Press, Oxford, pp. 1278-1296.
[3] Klevay LM (2014) Diagnosis of copper deficiency. BMJ 348, g3691.
[4] Milne DB, Klevay LM, Hunt JR (1988) Effects of ascorbic acid supplements and a diet marginal in copper on indices of copper nutriture in women. Nutr Res 8, 865-873.
[5] Klevay LM (2008) Alzheimer's disease as copper deficiency. Med Hypotheses 70, 802-807.
[6] Linder MC, Goode CA (1991) Biochemistry of Copper, Plenum Press, New York, pp. 119, 122, 194.
[7] Owen CA, Jr. (1982) Biochemical aspects of copper, Noyes Publications, Park Ridge, NJ, pp. 78-9.
[8] Hart EB, Steenbock H, Waddell J, Elvehjem CA (1928) Iron in nutrition. VII. Copper as a supplement to iron for hemoglobin building in the rat. J Biol Chem 77, 797-812.
[9] Klevay LM (2011) Is the Western diet adequate in copper? J Trace Elem Med Biol 25, 204-212.

Comments

Submitted by Mariacristina Siotto, PhD on

I would like to thank Professor Emeritus Leslie M. Klevay for having read the article carefully and for allowing an open debate on copper involvement in Alzheimer’s disease (AD).

Our approach to ceruloplasmin specific activity (eCp/iCp) was inspired from a study by Professor Brewer and colleagues [1] that described the eCp/iCp to be lower in a group of patients with respect to controls, while the percentage of free copper was higher in AD group than in controls. The ratio eCp/iCp was proposed by Milne and Johnson [2] as an indicator of copper status not affected by age, gender, or hormone use. In our previous study [3], we measured eCp/iCp in Wilson disease (WD), demonstrating that it was lower than in the control group. A previous study [4] on WD showed that ceruloplasmin values measured immunologically could give higher values than those measured enzymatically, as also suggested by Walshe [5]. Merle et al. [6] proposed serum ceruloplasmin oxidase activity as a sensitive and highly specific diagnostic marker for WD. In fact, the commercial kits usually employed do not discriminate between apo-ceruloplasmin and the completely active form of the protein: the holo-ceruloplasmin. Therefore, in WD patients, an apparent normal value of ceruloplasmin concentration could camouflage an inactive ceruloplasmin. Our study [3] confirmed eCp/iCp as a good index of the real status of circulating ceruloplasmin in WD.

A lower activity of ceruloplasmin in WD derives from a defective ATP7B copper pump, which does not excrete the copper from the liver and does not load copper into the nascent apo-ceruloplasmin. This is the reason why, in WD, mutations of the ATP7B gene modify copper incorporation into the protein, producing apo-ceruloplasmin that is less active. Moreover, this dysfunction produces a higher fraction of copper not bound to ceruloplasmin (Non Cp-Cu), which is one typical altered parameter in WD [7].

The same concept can be applied to copper dismetabolism in AD. In the course of the last ten years, a number of studies demonstrated that increased concentrations of Non Cp-Cu and other copper abnormalities are also evident in AD (meta-analysis of copper in AD: [8-10]; meta-analysis of Non Cp-Cu in AD: [11]). Moreover, specific ATP7B gene variants have been reported to independently increase the risk of AD [12-17]. It is also important to note that a meta-analysis on brain metals in AD [10] demonstrated that copper was significantly depleted in AD. Thus, an in increase in copper in general circulation corresponds to a copper depletion in the brain.

In our opinion, the problem of copper dismetabolism is strictly related to ATP7B imbalance. It is clear that a low copper diet could be recommended only to patients with evidence of copper dismetabolism, both obtained from genetic evaluation of ATP7B gene or from biochemical copper panel measurements (copper, eCp/iCp, Non Cp-Cu). Copper, in fact, is an essential metal and, in normal metabolism, it is fundamental to intake the appropriate quantity from diet.

We agree that the eCp/iCp index measured in subjects without ATP7B impairment is a sensitive measurement of copper deficiency, because the lower the copper intake in the diet, the lower the biosynthesis of ceruloplasmin and the lower the eCp/iCp; even so, this is not the case in AD, as clearly demonstrated in our article, showing increased levels of Non Cp-Cu associated with eCp/iCp decrement.

So, finally, we do not agree with Prof. L.M. Klevay’s interpretation of our data, and strongly advise against taking copper supplements as stated in [18]. In fact, supplements have been associated with an 18% increase in mortality [19] and cognitive decline [20] in large population studies: IOWA women (40,000 subjects) and CHAP study (~4,000 subjects follow up for 9 years), respectively.

Mariacristina Siotto and Rosanna Squitti

References
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[2] Milne DB, Johnson PE (1993) Assessment of copper status: Effect of age and gender on reference ranges in healthy adults. Clin Chem 39, 883–887.
[3] Siotto M, Pasqualetti P, Marano M, Squitti R (2014) Automation of o-dianisidine assay for ceruloplasmin activity analyses: usefulness of investigation in Wilson’s disease and in hepatic encephalopathy. J Neural Transm 121, 1281–1286.
[4] Macintyre G, Gutfreund KS, Martin WRW, Camicioli R, Cox DW (2004) Value of an enzymatic assay for the determination of serum ceruloplasmin. J Lab Clin Med 144, 294–301.
[5] Walshe J (2003) Wilson’s disease: the importance of measuring serum caeruloplasmin non-immunologically. Ann Clin Biochem 40, 115–121.
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[7] Easl (2012) EASL Clinical Practice Guidelines: Wilson’s disease. J Hepatol 56, 671–685.
[8] Ventriglia M, Bucossi S, Panetta V, Squitti R (2012) Copper in Alzheimer’s disease: a meta-analysis of serum, plasma, and cerebrospinal fluid studies. J Alzheimers Dis 30, 981–984.
[9] Wang ZX, Tan L, Wang HF, Ma J, Liu J, Tan MS, Sun JH, Zhu XC, Jiang T, Yu JT (2015) Serum iron, zinc, and copper levels in patients with Alzheimer’s disease: a replication study and meta-analyses. J Alzheimers Dis 47, 565–581.
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[11] Squitti R, Simonelli I, Ventriglia M, Siotto M, Pasqualetti P, Rembach A, Doecke J, Bush AI (2014) Meta-analysis of serum non-ceruloplasmin copper in Alzheimer’s disease. J Alzheimers Dis 38, 809–822.
[12] Bucossi S, Mariani S, Ventriglia M, Polimanti R, Gennarelli M, Bonvicini C, Pasqualetti P, Scrascia F, Migliore S, Vernieri F, Rossini PM, Squitti R (2011) Association between the c. 2495 A>G ATP7B polymorphism and sporadic Alzheimer’s disease. Int J Alzheimers Dis 2011, 973692.
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[14] Squitti R, Polimanti R, Siotto M, Bucossi S, Ventriglia M, Mariani S, Vernieri F, Scrascia F, Trotta L, Rossini PM (2013) ATP7B variants as modulators of copper dyshomeostasis in Alzheimer’s disease. Neuromolecular Med 15, 515–522.
[15] Liu H, Lin W, Wang W, Tsai C, Wu W, Chiou M, Shen C, Wu B, Tsai F (2013) Genetic variability in copper-transporting P-type Triphosphatase (ATP7B) is associated with Alzheimer’s disease in a Chinese population. J Biol Regul Homeost Agents 27, 319–327.
[16] Squitti R, Ventriglia M, Gennarelli M, Colabufo NA, El Idrissi IG, Bucossi S, Mariani S, Rongioletti M, Zanetti O, Congiu C, Rossini PM, Bonvicini C (2017) Non-ceruloplasmin copper distincts subtypes in Alzheimer’s disease: a genetic study of ATP7B frequency. Mol Neurobiol 54, 671-681.
[17] Squitti R, Ventriglia M, Gennarelli M, Colabufo NA, El Idrissi IG, Bucossi S, Mariani S, Rongioletti M, Zanetti O, Congiu C, Rossini PM, Bonvicini C (2017) Erratum to: Non-ceruloplasmin copper distincts subtypes in Alzheimer’s disease: a genetic study of ATP7B Frequency. Mol Neurobiol 54, 682-683.
[18] Barnard ND, Bush AI, Ceccarelli A, Cooper J, de Jager CA, Erickson KI, Fraser G, Kesler S, Levin SM, Lucey B, Morris MC, Squitti R (2014) Dietary and lifestyle guidelines for the prevention of Alzheimer’s disease. Neurobiol Aging 35, 1–5.
[19] Mursu J, Robien K, Harnack LJ, Park K, Jacobs Jr. DR (2011) Dietary supplements and mortality rate in older women: the Iowa Women’s Health Study. Arch Intern Med 171, 1625–1633.
[20] Morris MC, Evans DA, Tangney CC, Bienias JL, Schneider JA, Wilson RS, Scherr PA (2006) Dietary copper and high saturated and trans fat intakes associated with cognitive decline. Arch Neurol 63, 1085–1088.
[21] Tecchio F, Vecchio F, Ventriglia M, Porcaro C, Miraglia F, Siotto M, Rossini PM, Rongioletti M, Squitti R (2016) Non-ceruloplasmin copper appears a distinct subtype of Alzheimer`s disease: a study of EEG-derived brain activity. Curr Alzheimer Res 13, 1374-1384.