SELENIUMDr.Arun Babu.N.B.MD (Biochemistry)

Overview… History & Chemistry

… physiological functions, dietary reference intakes and sources

… absorption, metabolism & excretion

… interactions, Se deficiency & toxicity, treatment

… analysis of tissue levels and Se status

… its health benefits

… importance to human health

… Se: a double edged sword2

HISTORY & CHEMISTRY

Metal element present naturally in large quantities in earth’s crust & in soil.

Discovered by Jons Berzelius in 1817 (Selene in Greek means Moon)

Located in same group as Oxygen, Sulphur & Te in periodic table.

Exist in elemental, organic & inorganic forms Elemental 7 Inorganic forms (Selenide/Selenite/Selenate) – more

Toxic, but have less oral absorption Most of Se consumed as normal diet – Organic

(Dimethylselenide/Selenobetaine) Multivitamins & Dietary supplements – Inorganic (Sodium

selenate) Exist in 4 oxidation states (-2, +1, +2 & +6)

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CHEMISTRY Enzymes containing Selenocysteine in their active site are

called selenoproteins. Glutathione peroxidase

Most familiar selenoprotein in our body Using reduced glutathione (GSH) as a substrate, it reduces H2O2 to

water. Limits oxidative stress by detoxifying Reactive Oxygen Species (ROS).

Iodothyronine deiodinase Important role in converting T4 to T3. Allows metabolic adaptation to severe stress or injury.

Thioredoxin reductase Important role in regenerating antioxidants like Vit-C Helps to regulate cell growth & viability Immunomodulator & Chemotherapeutic agent

Selenomethionine A storage protein of selenium

Other storage sites – Plasma proteins (like albumin) – known as selenium binders.

Biologically active form of Selenium – Selenocysteine.4

Physiological Function of Selenoproteins (Examples)

glutathione peroxidases (GPx)

iodothyronine deiodinases

thioredoxin reductases

selenoprotein P

selenoprotein W

DNA-bound spermatic selenoprotein

R-O-O-H

R-O-H

T4

T3 rT3

3,3’T2

NADPH NADP+

oxidizedthioredoxin

reducedthioredoxin cell growth and survival

oxidized ascorbatereducedascorbate

Antioxidant and transport function.probably involved in Se-homeostasis

needed for muscle function

mitochondrial capsule selenoprotein

antioxidants

may protect developing sperm

protects developing sperm from ox. damage,polymerizes into structural protein required for stability/motility of mature sperm

selenophosphate synthetaserequired for selenoprotein synthesis

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Dietary Reference Intakes and Sources Recommended daily intake of 30 μg found in:

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lobstertunamusselskidney (calf)codfishoystersliver (cow)egg pastapork meateggswhole grain bread

In foods, Se is usually present as amino acid derivatives – selenomethionine

COMA set RNIs of 0.075 and 0.060 mg Se per day for males & females respectively0.075 mg Se per day for lactating women (COMA,1991)Lower limit of WHO safe range of the population mean intake to meet requirements - 0.040 mg Se per day

COMA – Committee On Medical Aspects of food & nutrition policy.

Recommended daily intakes (SGE)children 25-60 μg/dadolescents 30-70 μg/dadults 30-70 μg/d (U.S. RDA: 55 μg/d)

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Dietary Reference Intakes

Safe upper limit (SGE) adults 400 μg/d

(symptoms of chronic overload above 800 μg/d)

narrow window between ingested amounts resulting in deficiency and those resulting in toxicity!

ABSORPTION & BIOAVAILABILITY Readily absorbed from small intestine. Extent of absorption depends on nature of compound. Soluble selenate & selenomethionine – Most readily absorbed.

DISTRIBUTION & METABOLISM Widely distributed throughout the body Can be detected in breast milk Cross placenta in animals Levels – Slightly higher in Liver & Kidneys than in other

tissues. Can be incorporated into selenoproteins Bind to selenium binding proteins Incorporated into volatile methylated metabolites, which are

subsequently excreted.

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EXCRETION Largely excreted in urine Excreted in breath along with some volatile metabolites Some faecal excretion occurs, usually following chronic

administration.

INTERACTIONS Complex interaction between selenium & iron Severe Se deficiency – May Increase hypothyroid stress caused

by iodine deficiency Selenium supplementation alone – May aggravate iodine

deficiency by accelerating iodine loss. Interaction significant only in cases of severe deficiency Selenium interacts with other metals & Ascorbic acid Selenium metabolism affects the metabolism & thus toxicities of

some xenobiotics9

SELENIUM DEFICIENCY 3 diseases associated with it namely:

Keshan’s Disease Kashin – Beck disease. Myxoedematous Endemic Cretinism.

KESHAN’S DISEASE Found out in China Affect children (2 to 10yrs) and women of child bearing age. Fatigue (even after mild exercise) Cardiac arrythmia Palpitation Loss of appetite Cardiac insufficiency Cardiomegaly Congestive cardiac failure Coronaries - Unaffected

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Cardiomyopathy

Mitochondrial dysfunction Sarcolema Earliest affected. Multifocal myocardial necrosis Fibrosis Periacinar Pancreatic fibrosis

KASHIN-BECK DISEASE In China, Russia & Korea Selenium responsive bone & joint disease Severe joint pain Short stature Joint Necrosis (Epiphyseal degeneration of the arm and leg

joints – Result in structural shortening of the fingers & long bones with consequent growth retardation and stunting).

Chondrocyte Necrosis – Multifactorial in origin TREATMENT

Supportive Therapy Selenium Supplementation.

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MYXOEDEMATOUS ENDEMIC CRETINISM Seen in Africa Partially related to selenium deficiency Iodine deficiency also occur in this area of Africa. Decreased function of Iodothyronine deiodinases

decreased T3 production Selenium dependent Glutathione peroxidase detoxifies

injurious H2O2, which abounds in Thyroid. Manifest symptoms of Mental Retardation &

Hypothyroidism. Prolonged use of Total Parenteral Nutrition (TPN) – Assoc.

with Se & other nutritional deficiencies. Severe Crohn’s disease & Ulcerative colitis – Assoc. with Se

deficiency & other micronutrients. Usually corrected once the patient is transitioned to

traditional diets.

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Myxoedematous endemic cretinism in

the Democratic Republic of Congo

SELENIUM TOXICITY / SELENOSIS ACUTE SELENIUM TOXICITY

Appear to occur at doses of ≥ 0.5 mg/kg body wt. Hypersalivation Nausea Emesis Stomach Ache Garlic aroma on the breath (due to excretion of volatile

selenium metabolites) GI effects (severe vomiting & diarrhoea) Hair Loss (Due to disruption of structural proteins in Keratin) Fatigue Irritability Neurological Disturbances (Restlessness, Spasms,

Tachycardia) Paraesthesias

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CHRONIC SELENIUM TOXICITY / SELENOSIS Changes to hair & nails Skin lesions Numbness Convulsions Paralysis Clinical Neurological effects

Peripheral hypoaesthesia Acroparesthesiae Pain Hyperreflexia

As a consequence of inadequate human cases of Selenosis, much of our knowledge is based on animal models. As per that, selenium toxicity is a dose dependent process.

No fatalities from Se used as a nutritional supplement. Fatalities – Assoc. with Industrial & environmental exposures

to Se.- Selenous acid (Gun-bluing solution), selenium dioxide Brazil Nut grown in the Andes mountain range of

Venezuela has the highest concentration of selenium measured in the food.

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TREATMENT CHELATION THERAPY

To identify a compound that will bind to the toxic substance and enhance elimination without redistributing the substance to other vital areas in the body (like CNS).

No proven chelators for selenium so far. Animal data with BAL, edentate calcium disodium & succimer –

demonstrate the formation of nephrotoxic complexes with selenium that likely worsen toxicity.

Proposed that Vit-C will compete with selenium absorption and uptake into vital enzymatic processes, thereby limiting selenium toxicity.

PAIN MANAGEMENT Skin pain –(Burning sensation) & Painful nails –(Throbbing pressure) Cornerstone therapy – removal of patient from exposure. For exposures to dietary supplementation, Analgesics – Main Treatment If Dermal pain is from a topical occupational exposure to selenous acid

or selenium dioxide, then sodium thiosulphate solution or ointment – provides symptomatic relief

Sodium thiosulphate – Act as a Reducing agent, forming Se byproducts which are less irritant to skin.

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RISK ASSESSMENT

ENDEMIC SELENOSIS In parts of Venezuala & China Symptoms – Typical Integumentary Findings

Hair Loss Nail Changes

Selenium has a variety of toxic endpoints in both animals & humans

In man, 1st sign of chronic toxicity appear to be pathological changes in hair and nails,

Followed by adverse effects on the nervous system Changes in biochemical parameters also found Available studies indicate the development of selenosis is

associated with selenium intakes in excess of 0.85 mg/day (0.014 mg/kg body wt for a 60 kg adult)

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CARCINOGENECITY & GENOTOXICITY Selenium sulphide – carcinogenic in rats & mice. Other selenium compounds are not carcinogenic Increase in chromosomal aberrations in Hamster bone

marrow occurred, but at lethal doses of sodium selenite. Selenium compounds are not mutagenic in vivo

MECHANISM OF TOXICITY No specific mechanisms yet identified Redox cycling of auto-oxidisable metabolites Glutathione depletion Inhibition of protein synthesis Depletion of SAM ( S-adenosyl methionine) Replacement of sulphur by selenium in critical

sulphydryl groups17

ANALYSIS OF TISSUE LEVELS & SELENIUM STATUS Can be measured directly from plasma, serum, red

cells, nails and hair. Can also be determined indirectly by measuring

tissue glutathione peroxidase activity, although a plateau activity may occur with higher levels of selenium intake not producing a corresponding increase in activity.

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Selenium metabolism in plantsSe-accumulating plants

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normal: < 100 μg/g Se (dry weight)

Se-accumulators: > 1000 μg/g Se (dry weight)

garlic(allium sativum)

broccoli (brassica oleracea)

common mushroom(agaricus bisporus)

Selenium metabolism in humans

SeO32-

SeO42-

Se-Met

SeO32-

human body

H2Se

Se-CysSe-Met

CH3SeH

GSH (glutathione)

-lyase

-lyase

Se-Cys methyl-

transferase

dem

ethy

las

e

GSH

A) low Se intake

Seleno-proteins (Se-Cys in active site)

urine22

Selenium metabolism in humans

SeO32-

SeO42-

Se-Met

SeO32-

human body

H2Se

Se-CysSe-Met

CH3SeH

(CH3)2Se(CH3)3Se+

breath urine

GSH (glutathione)

Seleno-proteins

-lyase

-lyase

Se-Met-containingproteins (Se stores)

Se-sugarsSe-Cys methyl-

transferase

dem

ethy

las

e

GSH

B) high Se intake

(Se-Cys in active site)

(Se-Met incorporated instead of Met)

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Immune function

Viral infection

Thyroid function

Cardiovascular disease

Cancer and chemoprevention

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The importance of selenium to human health

Role of selenium in immune function

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Innate “nonadaptive” immunity

Acquired “adaptive” immunity

Immune system

Humoral Cellular Humoral CellularCytokines

- Interferon- Interleukins (eg. IL-3, IL-2, etc)

MacrophagesGranulocytes

- Neutrophils- Basophils-Eosinophils-Natural-Killer (NK) cells

Antibodies- Immunoglobulin (eg: IgG, IgA, etc)

Lymphocytes- T-cell- B-cell

Involvement of Se in immune system:

• Increased activity of NK cells

• Proliferation of T-cells• Increased interferon

production• Increased interleukin

receptors

• Stimulation of vaccine-induced immunity

• More antibody-producing B-cells• Macrophage and neutrophil activity

….proposed mechanismsSelenoproteins influence three broad areas of cell function an therefore affect immune function through:

•Antioxidant activities•Thyroid hormone metabolism•Regulation of the activity of redox-active proteins

Example: Neutrophil function

-Neutrophils produce peroxi-derived radicals to kill invading microbes, but also the neutrophils themselves need to be protected from the radicals by the system.-Radical production and protection depends on GPx activity in neutrophils, which is impaired in Se-deficiency. 28

Role of selenium in viral infection In Se-deficient mice harmless viruses can become virulent. ( e.g. coxsackie virus causing cardiomyopathy in Keshan disease)

Subjects supplemented with Se showed less mutations in poliovirus compared to placebo group.

HIV patients are nearly 20 times more likely to die from HIV-related causes than those with adequate levels. And low Se levels in HIV-infected children are related to faster disease progression.

Infection with virus

GPx Adequate

↑ Oxidative stress

Normal immune response

Mild to no tissue damage

Se + Se -

GPx deficient

↑↑↑ Oxidative stress

Altered immune response Viral mutation

Moderate to severe tissue damage

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Role of selenium in thyroid function Deiodinases are Se-containing enzymes playing important roles in thyroid hormone metabolism.

Low plasma T3:T4 ratios found in people with low Se intake.

A combined deficiency of iodine and selenium is associated with severe endemic myxedematous cretinism (in Democratic Republic of Congo).

Se supplementation in elderly subjects decreased plasma thyroxine (T4) concentrations , increased deiodinase activity and improved conversion of the active hormone T3.

Impaired thyroid hormone metabolism has adverse effects on immune function.

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Myxedematous endemic cretinism in

the Democratic Republic of Congo

Role of selenium in cardiovascular disease (CVD)Increased production of ROS can cause oxidative stress and cause damage to cellular lipids, proteins and DNA, leading to CVD.

Selenoproteins may help to combat oxidative modification of lipids and to reduce platelet aggregation.

Epidemiological studies produced mixed findings (examples):

2 to 3-fold increase in CV morbidity and mortality in subjects with serum Se conc. below 45 µg/L compared to subjects above at baseline.

Increased risk of ischaemic heart disease in Danish men with serum Se below 79 μg/L.

Inverse association between toenail Se and risk of myocaridal infarction only in areas with the lowest Se status. Thus, effect maybe only apparent in populations of low Se status.

Ebselen, a synthetic mimic of glutathione reductase showed cardio- protective effects in mice.

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Role of selenium in cancer preventionPossible mechanisms of cancer Possible mechanisms of cancer

prevention by seleniumprevention by selenium

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…history of selenium in cancer prevention• In 1969 Shamberger found that his cancer patients had

selenium blood levels only 60 to 80 % as high as non-cancer patients.

• In 1977, Schrauzer reported that selenium intake in 27 countries was inversely correlated with breast cancer death rates.

• Several human epidemiological studies found a statistically significant inverse relationship between Se level and risk of cancer overall, particularly in men.

• Also early animal studies have shown that selenium added to the diet reduced cancer incidence.

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Se in cancer prevention • In the first RCT, supplementation with selenized yeast (200

µg/day), predominantly in the form of selenomethionine (Se-Met) for 4 years, led to a reduction of nearly 50% in overall cancer morbidity (Clark, 1996).

Average selenium intake of the study subjects 90 µg/day, well above levels needed for optimal selenoprotein activity.

This suggests additional cancer preventive mechanisms.

• Inorganic forms, such as selenite/selenate, were more effective than commonly used organic form Se-Met in fighting cancer.

• Because cells can not distinguish Se-Met from essential amino acid methionine, it is incorporated into general body proteins.

• However, inorganic Se is more toxic compared to organic Se forms.

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Methylselenocystein (Met-Se-Cys): Less toxic alternative • Is formed naturally in various plants grown on high selenium

soil.

• Met-Se-Cys rich foods have shown good anticancer activity, without

excess tissue accumulation or toxicity.

• Met-Se-Cys is converted into methylselenol (CH3SeH), which has been shown to be an active anticancer form of Se.

Mechanism of action: Apoptosis and inhibition of angiogenesis.

H2Se CH3SeH

(CH3)2Se

breath urine

methyl-transferase

demethylase

SeMC

MSA

β-lyase

GSH

Monomethylated Se compounds, SeMC and methylselenic acid (MSA), were shown to be more effective than other Se compounds in chemoprevention.

And it is less toxic!

toxic

MSA

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Selenium in human health – A double-edged sword•Several organic and inorganic Se compounds have been investigated as Se supplements. Their safety and efficacy differ markedly because of their differential metabolic processing by the body.

•The inorganic Se compound, sodium selenite, due to its prooxidant character, represents a promising alternative for cancer therapy.

•However, this Se compound is highly toxic compared to organic Se forms.

•To achieve chemopreventive effects, Se had to be given at Se levels 5-10 fold higher than what would be required for normal nutritional need.

•Due to a broad interest in the positive effects of Se on human health and cancer therapy, studies investigating the negative effects such as toxicity and DNA damage are highly required.

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Literature

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Rayman M.P. (2000); The importance of selenium to human health. Lancet 356: 233-41 Dumont E., Vanhaecke F., Cornelis R. (2006); Selenium speciation from food source to

metabolites: a critical review. Anal. Bioanal. Chem. 385: 1304-1323 www.sge-ssn.ch Brozmanova J., Manikova D., Vlckova V., Chovanec M. (2010); Selenium: a double-

edged sword for defense and offence in cancer. Arch. Toxicol. 84: 919-938 Ip, C. (1998); Lessons from Basic Research in Selenium and Cancer Prevention. J. Nutr.

128(11): 1845-1854 Janghorbani M., Xia Y., et. al. (1999); Metabolism of Selenite in Men with Widely

Varying Selenium Status. J. Am. Coll. Nutr. 18(5), 462-469 Tinngi U (2008); Selenium: its role as antioxidant in human health. Environ Health Prev

Med. 13:102-108 Arthur J.R, McKenzie RC, Beckett J (2003); Selenium in the Immune System. J. Nutr 133:

1457S-1459S Beck MA (2001); Antioxidants and Viral Infections: Host Immune Response and Viral

Pathogenicity. J. Am College of Nutrition. 20(5), 384S-388S http://www.dkfz.de/en/tox/c010-2_projects/list_assays.html Reilly, C. 1996. Selenium in food and health. London, Blackie Academic and

Professional. Levander, O.A. 1987. A global view of human selenium nutrition. Annu. Rev. Nutr.,

7:227-250.