As 2

Return to top The rhizome (root) of turmeric ( Curcuma longa Linn.) has long been used in traditional Asian medicine to treat gastrointestinal upset, arthritic pain, and "low energy." Laboratory and animal research has demonstrated anti-inflammatory, antioxidant, and anti-cancer properties of turmeric and its constituent curcumin. Preliminary human evidence, albeit poor quality, suggests possible efficacy in the management of dyspepsia (heartburn), hyperlipidemia (high cholesterol), and scabies (when used on the skin).

SynonymsReturn to top Amomoum curcuma, anlatone (constituent), ar-tumerone, CUR, Curcuma , Curcuma aromatica , Curcuma aromatica salisbury, Curcuma domestica , Curcuma domestica valet, Curcuma longa , Curcuma longa Linn, Curcuma longa rhizoma, curcuma oil, curcumin, diferuloylmethane, E zhu, Gelbwurzel, gurkemeje, haldi, Haridra, Indian saffron, Indian yellow root, jiang huang, kunir, kunyit, Kurkumawurzelstock, kyoo, NT, number ten, Olena, radix zedoaria longa, rhizome de curcuma, safran des Indes, sesquiterpenoids, shati, tumeric, turmeric oil, turmeric root, tumerone (constituent), Ukon, yellowroot, zedoary, Zingiberaceae (family), zingiberene, Zitterwurzel.

EvidenceReturn to top These uses have been tested in humans or animals. Safety and effectiveness have not always been proven. Some of these conditions are potentially serious, and should be evaluated by a qualified healthcare provider. Uses based on scientific evidence

Grade*

Blood clot prevention Early research suggests that turmeric may prevent the formation of blood clots. However, more research is needed before turmeric can be recommended for these conditions.

C

Cancer Several early animal and laboratory studies report anti-cancer (colon, skin, breast) properties of curcumin. Many mechanisms have been considered, including antioxidant activity, antiangiogenesis (prevention of new blood vessel growth), and direct effects on cancer cells. Currently it remains unclear if turmeric or curcumin has a role in preventing or treating human cancers. There are several ongoing studies in this area.

C

Cognitive function Curcumin has been shown to have antioxidant and antiinflammatory properties and to reduce beta-amyloid and plaque C burden in lab studies. However, there is currently not enough evidence to suggest the use of curcumin for cognitive performance. Dyspepsia (heartburn) Turmeric has been traditionally used to treat stomach problems (such as indigestion from a fatty meal). There is preliminary evidence that turmeric may offer some relief from these stomach problems. However, at high doses or with prolonged

C

use, turmeric may actually irritate or upset the stomach. Reliable human research is necessary before a recommendation can be made. Gallstone prevention/bile flow stimulant It has been said that there are fewer people with gallstones in India, which is sometimes credited to turmeric in the diet. Early studies report that curcumin, a chemical in turmeric, may C decrease the occurrence of gallstones. However, reliable human studies are lacking in this area. The use of turmeric may be inadvisable in patients with active gallstones. High cholesterol Early studies suggest that turmeric may lower levels of lowdensity lipoprotein ("bad cholesterol") and total cholesterol in the blood. Better human studies are needed before a recommendation can be made.

C

HIV/AIDS Several laboratory studies suggest that curcumin, a component of turmeric, may have activity against HIV. However, reliable human studies are lacking in this area.

C

Inflammation Laboratory and animal studies show anti-inflammatory activity of turmeric and its constituent curcumin. Reliable human research is lacking.

C

Irritable bowel syndrome (IBS) Irritable bowel syndrome (IBS) is a common functional disorder for which there are limited reliable medical treatments. One study investigated the effects of Curcuma xanthorriza on IBS and found that treatment did not show any therapeutic benefit over placebo. More studies are needed to verify these findings.

C

Liver protection In traditional Indian Ayurvedic medicine, turmeric has been used to tone the liver. Early research suggests that turmeric may have a protective effect on the liver, but more research is needed before any recommendations can be made.

C

Oral leukoplakia Results from lab and animal studies suggest turmeric may have C anticancer effects. Large, well-designed human studies are needed before a recommendation can be made. Osteoarthritis Turmeric has been used historically to treat rheumatic conditions. Laboratory and animal studies show antiinflammatory activity of turmeric and its constituent curcumin, which may be beneficial in people with osteoarthritis. Reliable human research is lacking.

C

Peptic ulcer disease (stomach ulcer) C Turmeric has been used historically to treat stomach and duodenal ulcers. However, at high doses or with prolonged use,

turmeric may actually further irritate or upset the stomach. Currently, there is not enough human evidence to make a firm recommendation. Rheumatoid arthritis Turmeric has been used historically to treat rheumatic conditions and based on animal research may reduce inflammation. Reliable human studies are necessary before a recommendation can be made in this area.

C

Scabies Historically, turmeric has been used on the skin to treat chronic skin ulcers and scabies. It has also been used in combination with the leaves of the herb Azadirachta indica ADR or "neem." More research is necessary before a firm recommendation can be made.

C

Uveitis (eye inflammation) Laboratory and animal studies show anti-inflammatory activity of turmeric and its constituent curcumin. A poorly designed human study suggests a possible benefit of curcumin in the treatment of uveitis. Reliable human research is necessary before a firm conclusion can be drawn.

C

Viral infection Evidence suggests that turmeric may help treat viral infections. However, there is not enough human evidence in this area. C Well-designed trials are needed to determine if these claims are true. *Key to grades A: Strong scientific evidence for this use; B: Good scientific evidence for this use; C: Unclear scientific evidence for this use; D: Fair scientific evidence against this use; F: Strong scientific evidence against this use. Grading rationale Uses based on tradition or theory The below uses are based on tradition or scientific theories. They often have not been thoroughly tested in humans, and safety and effectiveness have not always been proven. Some of these conditions are potentially serious, and should be evaluated by a qualified healthcare provider. Abdominal bloating, Alzheimer's disease, antibacterial, antifungal, antimicrobial, antispasmodic, anti-venom, appetite stimulant, asthma, boils, breast milk stimulant, bruises, cataracts, chemoprotective, contraception, cough, cystic fibrosis, diabetes, diarrhea, dizziness, epilepsy, flavoring agent, gas, gonorrhea, heart damage from doxorubicin (Adriamycin®, Doxil®), Helicobacter pylori infection, hepatitis, high blood pressure, histological dye, human papillomavirus (HPV), hypoglycemic agent (blood sugar lowering), infections (methicillin-resistant Staphylococcus aureus), insect bites, insect repellent, jaundice, kidney disease, kidney stones, leprosy, liver damage from toxins/drugs, male fertility, menstrual pain, menstrual period problems/lack of menstrual period, multidrug resistance, neurodegenerative disorders, pain, parasites, ringworm, scarring, scleroderma, weight reduction.

DosingReturn to top The below doses are based on scientific research, publications, traditional use, or expert opinion. Many herbs and supplements have not been thoroughly tested, and safety and effectiveness may not be proven. Brands may be made differently, with variable ingredients, even within the same brand. The below doses may not apply to all products. You should read product labels, and discuss doses with a qualified healthcare provider before starting therapy. Adults (over 18 years old) Doses used range from 450 milligrams of curcumin capsules to 3 grams of turmeric root daily, divided into several doses, taken by mouth. As a tea, 1 to 1.5 grams of dried root may be steeped in 150 milliliters of water for 15 minutes and taken twice daily. Average dietary intake of turmeric in the Indian population may range between 2 to 2.5 grams, corresponding to 60 to 200 milligrams of curcumin daily. A dose of 0.6 milliliters of turmeric oil has been taken three times daily for one month and a dose of 1 milliliter in three divided doses has been taken for two months. One reported method for treating scabies is to cover affected areas once daily with a paste consisting of a 4:1 mixture of Azadirachta indica ADR ("neem") to turmeric, for up to 15 days. Scabies should be treated under the supervision of a qualified healthcare professional. Children (under 18 years old) There is no proven or safe medicinal dose of turmeric in children.

SafetyReturn to top The U.S. Food and Drug Administration does not strictly regulate herbs and supplements. There is no guarantee of strength, purity or safety of products, and effects may vary. You should always read product labels. If you have a medical condition, or are taking other drugs, herbs, or supplements, you should speak with a qualified healthcare provider before starting a new therapy. Consult a healthcare provider immediately if you experience side effects. Allergies Allergic reactions to turmeric may occur, including contact dermatitis (an itchy rash) after skin or scalp exposure. People with allergies to plants in the Curcuma genus are more likely to have an allergic reaction to turmeric. Use cautiously in patients allergic to turmeric or any of its constituents (including curcumin), to yellow food colorings, or to plants in the Zingiberaceae (ginger) family. Side Effects and Warnings Turmeric may cause an upset stomach, especially in high doses or if given over a long period of time. Heartburn has been reported in patients being treated for stomach ulcers. Since turmeric is sometimes used for the treatment of heartburn or ulcers, caution may be necessary in some patients. Nausea and diarrhea have also been reported. Based on laboratory and animal studies, turmeric may increase the risk of bleeding. Caution is advised in patients with bleeding disorders or taking drugs that may increase the risk of bleeding. Dosing adjustments may be necessary. Turmeric should be stopped prior to scheduled surgery. Limited animal studies show that a component of turmeric, curcumin, may increase liver function tests. However, one human study reports that turmeric has no effect on these tests. Turmeric or curcumin may cause gallbladder squeezing (contraction) and may not be advised in patients with gallstones. In animal studies, hair loss (alopecia) and lowering of blood pressure have been reported. In theory, turmeric may weaken the immune system and should be used cautiously in patients with immune system deficiencies. Turmeric should be used with caution in people with diabetes or hypoglycemia or people taking drugs or supplements that lower blood sugar. Pregnancy and Breastfeeding

Historically, turmeric has been considered safe when used as a spice in foods during pregnancy and breastfeeding. However, turmeric has been found to cause uterine stimulation and to stimulate menstrual flow and caution is therefore warranted during pregnancy. Animal studies have not found turmeric taken by mouth to cause abnormal fetal development.

InteractionsReturn to top Most herbs and supplements have not been thoroughly tested for interactions with other herbs, supplements, drugs, or foods. The interactions listed below are based on reports in scientific publications, laboratory experiments, or traditional use. You should always read product labels. If you have a medical condition, or are taking other drugs, herbs, or supplements, you should speak with a qualified healthcare provider before starting a new therapy. Interactions with Drugs Based on laboratory and animal studies, turmeric may inhibit platelets in the blood and increase the risk of bleeding caused by other drugs. Some examples include aspirin, anticoagulants ("blood thinners") such as warfarin (Coumadin®) or heparin, anti-platelet drugs such as clopidogrel (Plavix®), and non-steroidal anti-inflammatory drugs such as ibuprofen (Motrin®, Advil®) or naproxen (Naprosyn®, Aleve® ). Based on animal data, turmeric may lower blood sugar and therefore may have additive effects with diabetes medications. In animals, turmeric protects against stomach ulcers caused by non-steroidal anti-inflammatory drugs (NSAIDs) such as indomethacin (Indocin®) and against heart damage caused by the chemotherapy drug doxorubicin (Adriamycin®). Turmeric may lower blood pressure levels and may have an additive effect if taken with drugs that also lower blood pressure. Turmeric may lower blood levels of low-density lipoprotein (LDL or "bad" cholesterol) and increase high-density lipoprotein (HDL or "good" cholesterol). Thus, turmeric may increase the effects of cholesterol-lowering drugs such as lovastatin (Mevacor®) or atorvastatin (Lipitor®). Based on animal studies, turmeric may interfere with the way the body processes certain drugs using the liver's "cytochrome P450" enzyme system. As a result, the levels of these drugs may be increased in the blood and may cause increased effects or potentially serious adverse reactions. Patients using any medications should check the package insert and speak with a healthcare professional or pharmacist about possible interactions. When taken with indomethacin or reserpine, turmeric may help reduce the number of stomach and intestinal ulcers normally caused by these drugs. However, when taken in larger doses or when used for long periods of time, turmeric itself can cause ulcers. Interactions with Herbs and Dietary Supplements Based on animal studies, turmeric may increase the risk of bleeding when taken with herbs and supplements that are believed to increase the risk of bleeding. Multiple cases of bleeding have been reported with the use of Ginkgo biloba , some cases with garlic, and fewer cases with saw palmetto. Based on animal data, turmeric may lower blood sugar. Individuals taking other herbs or supplements or diabetes medications should speak with a healthcare professional before starting turmeric. Turmeric may lower blood levels of low-density lipoprotein (LDL or "bad" cholesterol) and increase high-density lipoprotein (HDL or "good" cholesterol). Thus, turmeric may increase the effects of cholesterol-lowering herbs or supplements such as fish oil, garlic, guggul, or niacin. Based on animal studies, turmeric may interfere with the way the body processes certain herbs or supplements using the liver's "cytochrome P450" enzyme system. As a result, the levels of other herbs or supplements may become too high in the blood. It may also alter the effects that other herbs or supplements possibly have on the P450 system. Turmeric may lower blood pressure and may therefore have an additive effect if taken with herbs that also lower blood pressure.

Methodology Return to top This information is based on a professional level monograph edited and peer-reviewed by contributors to the Natural Standard Research Collaboration (www.naturalstandard.com): Winnie Abramson, ND (Natural Standard Research Collaboration); E-P Barrette, MD (Case Western Reserve University School of Medicine); Ethan Basch, MD (Memorial Sloan-Kettering Cancer Center); Michael Bodock, RPh (Massachusetts General Hospital); Heather Boon, B.Sc.Phm, PhD (University of Toronto); Dawn Costa, BA, BS (Natural Standard Research Collaboration); Sadaf Hashmi, MD (Johns Hopkins School of Hygiene and Public Health); Jenna Hollenstein, MS, RD (Natural Standard Research Collaboration); George Papaliodis, MD (Massachusetts Eye and Ear Infirmary); Michael Smith, MRPharmS, ND (Canadian College of Naturopathic Medicine); Shaina Tanguay-Colucci, BS (Natural Standard Research Collaboration); Catherine Ulbricht, PharmD (Massachusetts General Hospital); Mamta Vora (Northeastern University); Wendy Weissner, BA (Natural Standard Research Collaboration). Methodology details

Selected references Return to top 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15.

Aggarwal BB, Kumar A, Bharti AC. Anticancer potential of curcumin: preclinical and clinical studies. Anticancer Res 2003;23(1A):363-398. Brinkhaus B, Hentschel C, Von Keudell C, et al. Herbal medicine with curcuma and fumitory in the treatment of irritable bowel syndrome: a randomized, placebo-controlled, double-blind clinical trial. Scand J Gastroenterol 2005 Aug;40(8):936-43. Deodhar SD, Sethi R, Srimal RC. Preliminary study on antirheumatic activity of curcumin (diferuloyl methane). Indian J Med Res 1980;71:632-634. Egan ME, Pearson M, Weiner S, et al. Curcumin, a major constituent of turmeric, corrects cystic fibrosis defects. Science 4-23-2004;304(5670):600-602. Kositchaiwat C, Kositchaiwat S, Havanondha J. Curcuma longa Linn. in the treatment of gastric ulcer comparison to liquid antacid: a controlled clinical trial. J Med Assoc Thai 1993;76(11):601-605. Kulkarni RR, Patki PS, Jog VP, et al. Treatment of osteoarthritis with a herbomineral formulation: a double-blind, placebo-controlled, cross-over study. J Ethnopharmacol 1991;33(1-2):91-95. Limtrakul P, Anuchapreeda S, Buddhasukh D. Modulation of human multidrug-resistance MDR-1 gene by natural curcuminoids. BMC Cancer 4-17-2004;4(1):13. Ng TP, Chiam PC, Lee T, et al. Curry consumption and cognitive function in the elderly. Am J Epidemiol 2006 1;164(9):898-906. Nishiyama T, Mae T, Kishida H, et al. Curcuminoids and sesquiterpenoids in turmeric (Curcuma longa L.) suppress an increase in blood glucose level in type 2 diabetic KK-Ay mice. J Agric Food Chem 2-232005;53(4):959-963. Prusty BK, Das BC. Constitutive activation of transcription factor AP-1 in cervical cancer and suppression of human papillomavirus (HPV) transcription and AP-1 activity in HeLa cells by curcumin. Int J Cancer 3-12005;113(6):951-960. Rithaporn T, Monga M, Rajasekaran M. Curcumin: a potential vaginal contraceptive. Contraception 2003;68(3):219-223. Taher MM, Lammering G, Hershey C, et al. Curcumin inhibits ultraviolet light induced human immunodeficiency virus gene expression. Mol Cell Biochem 2003;254(1-2):289-297. Thamlikitkul V, Bunyapraphatsara N, Dechatiwongse T, et al. Randomized double blind study of Curcuma domestica Val. for dyspepsia. J Med Assoc Thai 1989;72(11):613-620. Tilak JC, Banerjee M, Mohan H, et al. Antioxidant availability of turmeric in relation to its medicinal and culinary uses. Phytother.Res 2004;18(10):798-804. Van Dau N, Ngoc Ham N, Huy Khac D, et al. The effects of a traditional drug, turmeric (Curcuma longa), and placebo on the healing of duodenal ulcer. Phytomed 1998;5(1):29-34.

Elemental speciation in human health risk assessment, Environmental Health Criteria 234 (World Health Organization, Geneva) 2007. 238 pages. Price: CHF 30.00/US $ 30.00; in developing countries: CHF 21.00/US$ 18.90 ISBN 92-4-157234-5 This book is based on a draft prepared by WHO

Task Group on Environmental Health Criteria for Elemental Speciation in Human Health Risk Assessment under the supervision of Dr Inge Mangelsdorf of Fraunhofer Institute, Hanover, Germany and Dr A. Aitio, International Programme on Chemical Safety, WHO, Geneva, Switzerland, held on November 15-18, 2005 at Fraunhofer Institute of Toxicology and Experimental Medicine, Hanover, Germany. The book is directed at risk assessors and regulators, to emphasize the importance of consideration of speciation which is hardly taken into consideration as a part of most hazard and risk assessments. Further, the book points at the significance of analysis of speciation of elements to increase knowledge on the effect of speciation on mode of action and also to increase understanding of health effects. Speciation is the evaluation of chemical form and distribution of the elements, which play an important role in their toxicity and bioavailability. The areas of speciation analysis have undergone a continued evolution and development for the last two decades. The rapid development of inorganic instrumental analysis mainly driven by the development of atomic spectrometry enabled the analysts to look into the role of trace elements in different areas, such as health and environment, geochemistry and material sciences, to name some. This book provides a comprehensive description of the major areas involved in elemental speciation with the

Book Reviews Indian J Med Res 127, April 2008, pp 417-420 417

following chapters: structural aspects of speciation; analytical techniques and methodology; bioaccessibility and bioavailability; toxicokinetics and biological monitoring; molecular and cellular mechanisms of metal toxicity; and health effects. In addition, individual chapters are devoted to elemental speciation and mechanism of actions of most of the elements (chromium, manganese, iron, zinc, cobalt, nickel, copper, arsenic, selenium, cadmium, mercury and lead) in detail. Apart from this, role of tin, barium, palladium, platinum, thallium is also discussed in health effects chapter. The book contains an important chapter entitled, “Structural aspects of speciation” where it clearly explained electronic and oxidation states, nuclear (isotopic) composition, inorganic compounds and complexes, organometallic compounds, organic and macromolecular complexes of various elemental species. The book successfully pointed out that the

distribution, mobility and biological availability of chemical elements depends not simply on their concentration but, critically, on the chemical and physical association which they undergo in natural systems. However, the book does not point out some of the error sources in analysis of speciation, e.g., incomplete extraction from solid samples, degradation reactions, matrix-dependent recovery of extraction, lack of adequate internal standards, matrix effects, poor sensitivity of analytical techniques, etc. Actually the most difficult area of speciation remains the provision of a truly representative sample and the ability to transfer this to a suitable measurement technique whilst ensuring that the speciation profile remains intact. Fundamental understanding of trace element behaviour, the realistic formulation of historical perspectives of trace element contamination, an assessment of environmental transformation processes and a thorough appraisal of environment-related health problems and diseases are well documented in this book. 418 INDIAN J MED RES, APRIL 2008

The book explains the toxicokinetics and toxicodynamics of elements influenced by carriers, valence state and isotopes, particle size, element ligands, organic versus inorganic element species and biotransformations with resultant health effects. The book also reflects on understanding of some of the most important forms of an element, the transformation between forms and various consequences in terms of risk assessment, toxicity or biological activity. Different elemental species might not only differ quantitatively in their characteristics (e.g., toxicity) but also qualitatively (toxic versus essential). While Cr (III) compounds do have some positive biological activity and are therefore considered to be essential, Cr (VI) compounds are carcinogenic. Inorganic As (III) trioxide (As2O3) is also a carcinogenic but it was found to be effective in the treatment of certain forms of leukaemia. The methylated forms of arsenic are found to be far less toxic than the inorganic arsenic compounds. Methylated mercury, by contrast, is much more toxic than inorganic mercury. The book successfully highlighted molecular mechanisms of metal toxicity and carcinogenesis in respective chapters. Chronic exposure of many heavy metals and metalderivatives is associated with an increased risk of cancer, although the mechanisms of tumour genesis are largely unknown. Major areas of focus in this book include exposure assessment and biomarker identification, role of ROS or oxidative stress in carcinogenesis, mechanisms of metal-induced DNA damage, metal

signaling, metal-protein interactions and the effects of different elemental species on immune system. In conclusion, this is an excellent reference book and presents a comprehensive insight on the toxicological evaluation of elemental species and their intended usage pattern and possible human exposure, and provides an extensive bibliography. The text is clearly written and presents a vast amount of sound technical information. This book should be useful to all toxicologists, scientists, medical practitioners involved in regulatory agencies and is a good guide to those who are engaged in basic medical research on heavy metals or trace elements. Kusal K. Das Environmental Health Research Unit Department of Physiology Al Ameen Medical College Bijapur 586 108, India e-mail: [email protected] Turmeric: The salt of the orients is the spice of life, Kamala Krishnaswamy (Allied Publishers Private Limited, New Delhi) 2007. 248 pages. Price: Rs.350/ISBN 81-8424-126-7 Spices have been used since times immemorial for culinary purposes but the discovery of their physiological properties and therapeutic potential is much more recent. Several spices had been used in traditional systems of medicine, mainly for preventive purposes. The most important among them undoubtedly is turmeric (rhizome of Curcuma longa Linn). About 30 species of Curcuma have been described and several of these are used for food colouring and flavouring but C. longa is most extensively used. Turmeric occupies an unique place among the spices since besides being used for culinary and medicinal use it is also employed extensively as a cosmetic, colouring agent and preservative. The medicinal use of turmeric is mentioned even among the Vedas and elaborated in classical Ayurvedic texts like the Charaka Samhita. A systematic experimental and clinical evaluation of its medicinal properties, however, has been done largely from the middle of the last century. Major attention has centered round curcumin and related curcuminoids which constitute 2-5 per cent of the biomass of the rhizome and are responsible for its characteristic yellow colour. Curcumin free extracts also exhibit biological activity. There have been several reviews on pharmacology and chemistry of C. longa including a monograph in WHO Monographs on Selected Medicinal Plants, Volume I (1999). The present book is perhaps the first

book to comprehensively cover all aspects of the plant, including the historical development, botany, chemistry, pharmacology and uses in medical and other fields. The opening chapter has reviewed the traditional uses vis-à-vis modern concepts of functional foods and nutraceuticals. It has highlighted the need to provide evidence information to support its traditional uses. It provides justification for turmeric being considered the world’s ‘most important herb’ today and lists about 20 medicinal uses in countries from all parts of the world even though the plant is a native of tropical and subtropical regions. The chapter also illustrates the wide variation in chemical composition of various varieties of turmeric. Thus, the curcumin content varies from 2.8 (Krishna) to 9.3 per cent (Roma, Suroma), oleoresins from 3.8 (Krishna) to 16.2 per cent (IISR Pratibha) and essential oil from 2 (Krishna) to 7 per cent (Suvarna, Sudarsana). These may be responsible partly for the varying results obtained with the crude extracts by different investigators. The next chapter describes the chemical and nutritional composition of turmeric. The chemical description mainly covers curcumin and some other curcuminoids. An important omission is any mention of the pharmacologically active peptide turmerin. The yield of this 5-KDa water soluble cyclic peptide is 0.1 per cent and it contains 40 amino acid residues. The remaining chapters of the book review the major pharmacological activities, mainly of curcuminoids , and related clinical and pharmacokinetic studies. The activities reviewed include antiinflammatory, anti-carcinogenic anti-mutagenic, antioxidant, anti-atherosclerotic and some other minor effects. The experimental data have been reviewed in chapters 3-11. The basic mechanisms underlying the pathological processes like inflammation, carcinogenesis, atherosclerosis, etc., have been reviewed in adequate detail thereby helping to locate the possible sites of action of curcumin, its analogues and metabolites. Curcuminoids act on multiple sites in each of these conditions and some of these sites are common. These include kinins like PKC and tyrosine kinase, TNF, NFkappa B, cyclic D, protein API, STAT family agents, etc. They play important role in mutagenesis, tumour initiation, transformation, progression and metastasis. Pathogenic changes in vasculature are involved in different ways in growth and differentiation of tumours, wound healing, immune responses and thrombotic episodes. Curcumin suppresses proliferation of normal, transformed and malignant cells by modulating growth

factors and induces apoptosis in a variety of tumour and other cells. The mechanisms include mitochondrial dependent and independent pathways. An important observation evident from the review is the potentiation of effects of anti-cancer drugs like cis-platinum by curcumin. The prostaglandins are growth factors for inflammation and angiogenesis and perform a house keeping function. Curcumin modulates their activity by acting on the arachidonic acid metabolism. Other mechanisms involved in its anti-inflammatory activity include effect on lysosomal enzymes and membranes, inhibition of adhesion molecules and cytokine production by inhibition of NF-Kappa B target genes. The mechanism of other effects like hypolipidaemic, wound healing, anti-infective activity, etc., has also been similarly reviewed. The author has reviewed the extensive data exhaustively but has not commented on the procedures used or the results obtained. Some of the studies have employed very high amounts of the test substances like 5 per cent in the diet and up to 5 mg/ml in some in vitro studies. Results of such studies must be interpreted with caution. The clinical studies have been reviewed in a separate chapter and their inadequacies, and in many cases poor design of the trial, have been clearly brought out. It may have been better if the clinical studies had been given along with experimental studies on the condition and permitted better correlation. Still the clinical data provide enough evidence for chemopreventive and therapeutic potential of curcuminoids in malignancy and several chronic conditions. Chronic disorders including degenerative (atherosclerosis, Alzheimer’s disease), proliferative (cancer) or inflammatory (rheumatoid arthritis, chronic infections) conditions share cellular/ subcellular biochemical, molecular regulatory and pathological events including abnormal redox sites, cytokines, apoptotic mechanisms, etc. The book elegantly brings out how the effect of curcuminoids on basic modulators of these processes is responsible for their beneficial effects in such diverse conditions. It helps to identify potential targets on which future work may help in developing novel chemical entities for the management of these chronic conditions. It also highlights the therapeutic potential of the polypeptide tumerin which has not received adequate attention. Similarly data on bioavailability studies suggest the possibility of improving it by combining

with piperine. The studies reviewed in the book also clearly demonstrate that the safety and excellent tolerance of even large doses of curcuminoids and turmeric extracts enhance their utility in preventive health programmes and as nutraceuticals. The author aptly points out (p 237) the paucity of in vivo bioefficacy data in terms of their concentration and synergistic or additive effects with other bioactive compounds. She has also emphasized (p 298) the impact food-based approach for enhancing the intake of such phytochemicals may have on the onset and progression of several chronic diseases. The approach may also provide means of BOOK REVIEW 419

‘improving and prolonging the success of standard therapies. The book is extensively documented and covers references of papers published up to 2004. There are many patents on the curcuminoids and analogues, etc., but they have not been so well covered. The book has been profusely illustrated with black and white photographs, bar diagrams, etc. It would have been better to provide coloured photograph, for example, of the plant (Fig. 1) or the tissues (Fig. 6.1) The legends of some of the figures (e.g. Fig. 7.3, 14.4) could have been more explicit. Similarly the splitting of some of the figures (e.g., 4.3) and tables (e.g., 5.3) should have been avoided. These minor aberrations, however, detract very little from the merits of a timely and authoritative document on biomedical aspects of turmeric. This well produced monograph will be a useful reference book not only for research workers and clinicians but also those involved in economic utilization of turmeric and its value added products in pharmaceuticals, cosmetics, etc. B.N. Dhawan 3, Ramakrishan Marg Faizabad Road Lucknow 226 001, India e-mail: [email protected] Influenza and its global public health significance, T. M. Mathew & T. Mathew, editors (Thejma Publishers, West Orange, NJ, USA) 2006. 203 pages. Price: $ 33.99 ISBN: 0-9727597-5-1 This book published in 2006 is aimed for the public, and government authorities dealing with the influenza epidemic, researchers, scientists working on the treatment and control of influenza during the impending threat of influenza pandemic. It provides a comprehensive overview of epidemic and pandemic influenza. The detailed information about all the facts

related to influenza has been elaborated chapter-wise precisely making comprehensive overview of epidemic and is user friendly and ready referenced. It provides an educational and reference tool for students as well as a practical guide to all those involved in the activities related to influenza. Overall, the book contains 13 chapters. The chapter on history of influenza pandemics describes each pandemic nicely in a tabular form which can be helpful in studying the strain pattern. The structure of the virus has been explained in detail but a diagrammatic representation of the virus would have contributed more, making it more understandable, particularly for the students. Chapter on influenza C highlights its growth properties, which are different from influenza A & B as well as the factors influencing it. Chapter on avian influenza focuses on all the outbreaks and describes the receptor specificity of influenza viruses which helps in human transmission. Also, the different perspectives of avian influenza in canines, equines and swine have been explained in different chapters. Diagnosis of avian influenza and all the significant investigations have been outlined nicely and briefly. The author also describes the experience of infection and isolation of influenza virus in her family during the 1972 - 1973 epidemics in Madras (now Chennai). The interesting fact is that infection was brought to the house by her husband who contracted the disease through his friends who had respiratory illness while the author did not get respiratory infection in spite of working with the respiratory viruses. The steps to be followed for the outbreak investigation starting from descriptive epidemiology, the diagnosis with clinical features of the illness, determining the aetiology, etc. are described. Also included is the case study on influenza like outbreak, which can help in better understanding and analysis of the subject, particularly for the students. In short, it is a handy book but no new information has been added, there is repetition of the material in different chapters. Shashi Khare Department of Microbiology National Institute of Communicable Diseases 22 Sham Nath Marg Delhi 110 054, India 420 INDIAN J MED RES, APRIL 2008 ©2009.Al Ameen Charitable Fund Trust, Bangalore 73

SHORT COMMUNICATION Al Ame en J Med S c i (2 00 9 )2 (1 ) :7 3 -7 7

Studies on Arsenic Toxicity in Male Rat Gonads and its Protection by High Dietary Protein Supplementation Sanjit Mukherjee and Prabir K.Mukhopadhyay*

Department of Physiology, Presidency College, Kolkata-700 073, India Abstract: Arsenic was given orally to rats as arsenic tri oxide, 3mg /kg body wt/day in a single dose for 28 consecutive days. This treatment in male Wistar rats caused increase in seminiferous tubular luminal size coupled with reduced accumulation of spermatozoa, and signs of necrotic changes with disarray in cellular organization. Other significant changes were decrease in sperm count, viability and motility (p<0.001). On high protein diet (containing pea 37gm/100 gm of diet and casein 9gm/100gm of diet) supplementation along with same arsenic exposure caused partial restoration of normalcy. All these sperm physiological changes and altered gonadal features, both histomorphometric and histological observations, were found significantly ameliorated. Results of this study propose that high protein diet supplementation may be effective to recovery from the toxic effect of arsenic on male gonad of rat.

Introduction It has become evident that increasing human activities have modified the global cycle of heavy metals and metalloids, including the toxic non-essential elements like arsenic (As) [1]. Among these metals, arsenic exhibits a complex metabolism and is possibly the most abundant and potential carcinogen [2]. Arsenic is present in the nature in stable form as As 5+ species, and As 3+species. An analysis of 25000 tube wells in West Bengal reveals that the average As concentration reaches to 0.3mg/lt of water, and in some places even the concentration reaches up to 3mg/lt of water, where 0.05mg/lt is the permissible limit for drinking water as per WHO [3]. In the process of arsenic metabolism, inorganic arsenic is methylated to monomethyl arsonic acid (MMA) and finally to dimethyl arsinic acid (DMA) followed by a renal excretion. In this process of biomethylation, constant depletion of methyl causes DNA hypomethylation, and thus generates mutation followed by carcinogenesis [2]. Arsenic affects the mitochondrial enzymes, impairs the cellular respiration and

causes cellular toxicity. It can also substitute phosphate intermediates, which could theoretically slow down the rate of metabolism and interrupt the production of energy [4]. Male infertility is reflected by low sperm count, low sperm motility and bad quality of sperms (4). Sodium arsenite has been found to have an inhibitory effect on the activity of testicular steroidogenic enzyme ∆5­3β­hydroxysteroid dehydrogenase (∆5­3β­HSD) and 17ß-hydroxysteroid dehydrogenase (17ßHSD) and to reduce the weight of testes and accessory sex glands [4] in rat. High Arsenic level may suppress the sensitivity of gonadotroph cells to GnRH as well as gonadotropin secretion by elevating plasma levels of glucocorticoids. These ultimately lead to the development of gonadal toxicity [4-5]. In recent studies, dietary proteins have been found to have antioxidant activities [6-8]. Wheat (Puccinia graminis tritici) and pea (Pisum sativum) are both good sources of dietary plant protein, while casein is an animal protein. The antioxidant activities of pea, wheat, Al Ame en J Med S c i Volume2, No.1, 2009

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and casein has been studied using different liposomal models and the results show a minimization in lipid peroxidation, thus preventing the damage produced by the free radicals [6]. Our present study has been done to test the causal relationship between arsenic generated oxidative stress and testicular cell damage using rat as a model animal and further to examine whether dietary high protein interventions may be an effective strategy of detoxification that may help in preventing the disorders induced by arsenic. Materials and Methods: Animal selection and drug treatment: Normal (Wistar) male rats weighing between 120gm—140gm were selected for our experiment. Animals were maintained in an environmentally controlled animal house (temperature 24 ± 3°C) and in a 12 h light/dark schedule with free access to water. For experiments, rats were randomly selected into three groups consisting six rats in each: group A, control; group B, arsenic-treated; and group C, arsenic + pea + casein-diet supplemented. The animals

of groups A and B were provided with a control diet composed of 71% carbohydrate, 18% protein, 7% fat and 4% salt mixture and vitamins [9]. For chronic oral exposure to arsenic, a dose was selected (3 mg/kg body wt/day), which is within the range of LD50 of a 70-kg body wt human (1–4 mg/kg) and lesser than one-thirteenth of LD50 value of rats (40 mg/kg) [6]. Accordingly, animals of groups B, and C were orally treated with aqueous solution of arsenic trioxide, 3 mg/kg body wt/day for 28 days. The animals of group C, in addition, were supplemented with pea (37 g/100 g of diet), which contributed 8.5% protein, and casein (9 g/100 g of diet) which contributed additional 9% protein in the formulation of a high protein (27%) diet [6]. To overcome the impact of any altered food intake, control (group A) animals were pair-fed with other experimental groups B, and C. 1. Preparation of permanent slide for histological study of testes: Testes from all groups of animals were dissected out and were Bouin’s-fixed. Paraffin blocks were prepared, and 4–5 ∝m thin sections were cut with a high precision microtome (IEC Microtome, USA) and routine microscopic slides were prepared. For staining, standard Haematoxylin/Eosin method was followed to study both the histomorphometric and histological alterations. 2. Sperm viability count: Immediately after the sacrifice the cauda portion of epididymis was cut. The cauda was kept in 1 ml diluent [10]. This was kept for 5 mins at 37oC. It was then taken out, and an incision was given through the cauda and the sperms were dispersed in the fluid. 40µl of this spermatozoal suspension was transferred to an eppendorf. This was stained with EosinY and Nigrosin (40µl each). This was mixed gently and 25µl from the mixture was taken on a grease free slide and a smear was drawn and dried. The number of viable and non-viable sperms was counted under light microscope. 3. Sperm count: From the dispersed spermatozoal suspension 25µl was charged on a Neubauer Haemocytometer [11] and the numbers of sperms were counted and calculated using WBC chambers. 4. Sperm motility count: The cauda epididymis from all three groups was obtained

as mentioned earlier, and each cauda was kept in 1ml PBS 0.2 M, pH 7.4 at 37°C [12]. 25µl of this was taken on a clean slide covered with a cover slip and was Al Ame en J Med S c i Volume2, No.1, 2009

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observed under a microscope. Total numbers of motile sperms per 100 sperms were counted. The readings were taken at the beginning of 1st hour, 2nd hour and 3rd hour of the experiment (the temperature of the spermatozoal suspension was maintained at 37 o C throughout the process). 5. Statistical Analysis: The data were expressed as mean ± SEM (Standard error of mean). For statistical analysis, the quantitative data of each parameter from the different groups were analyzed by Student’s “t” test. The mean ± SEM was calculated for each group and the corresponding level of significance was calculated. Results: Histological analysis: The H/E stained histological sections of arsenic treated testes showed increase in luminal areas associated with reduced accumulation of spermatozoa, signs of necrotic changes with disarray in cellular organization (Fig 2) compared to that of control (Fig 1). Protein supple mented rat testes showed the partial recovery compared to the treated group (Fig 3). Recovery includes an accumulation of increased spermatozoa in the luminal areas along with partial amelioration of arsenic induced changes. Analysis of spermatozoal status : 1. Sperm count: The Table 1 shows that there is a reduction in the number of matured spermatozoa in case of treated group compared to that of the control(p<0.001). The values from high protein supplemented group, reveals an increased (p>0.001) sperm count towards normal. All the values are presented in table1. Results are expressed as mean ±SEM (Standard error of the mean) (n=6). 2. Sperm viability count: The Table1 shows that there is a reduction in the number

of viable sperms in cases of treated group (p<0.001) and this decrease in viability is minimized (p>0.001) in the high protein diet supplemented group. Results are expressed in table 1 following the same statistical analysis as mentioned earlier. 3. Sperm motility count: In case of treated group the decrease in motility was observed in studies done in the 1st, 2nd and 3rd hours compared to control. This decrease was around 10% as observed in three consecutive hours; administration of protein diet minimized the decrease in motility as reflected in the values nearer to that of control. The mean ± SEM (Standard error of mean) was calculated for each Fig: 1:- Section of control testis showing normal features (a) x10 (b) x40 Fig: 2: - Section of treated testis showing an increase in luminal area, reduced spermatozoal mass, disintegrated cell and nuclear membranes and disorganized cellular orientation.(a) x10 (b) x 40

1a 1b 2a 2b Fig 3: - Section of protein diet supplemented testis showing features towards normalcy.(a) x10 (b) x 40

3a 3b

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group and the corresponding level of significance was calculated by using the previous statistical analysis. Table: - 1. Table presenting the effect of protein diet supplementation on the arsenic trioxide induced changes in spermatozoal status in rat: Discussion Increase in the luminal areas of the seminiferous tubules associated with decreased spermatozoal mass might be due to low levels of gonadotropins in arsenic treated rats, and these low levels are responsible for the decreased production of steroidogenic enzymes [4]. It has been established that arsenic administration leads to decrease in ovarian steroidogenic enzymes synthesis [4]. Thus the low levels of gonadotropins and possibly testosterone might be responsible for the decrease in the spermatozoal mass in the lumen. Decrease in epididymal spermatozoal number provides support towards this histological observation. Arsenic causes lipid peroxidation by generation of reactive oxygen species (ROS) [13-14]. This peroxidation may cause rupture of cell as well as nuclear membrane. This might be responsible for the observed necrosis and disarray in cellular organization in histological section (40 X) Fig-2b. Evidence suggests that arsenic induces free

radical formation and thus the generated reactive oxygen species (ROS) react with the polyunsaturatedfattyacid (PUFA) rich spermatozoa, specially the mid spermatozoa and results in peroxidation which finally leads to destruction in spermatozoa causing reduced motility and viability [4]. Pea plus casein diet supplementation along with arsenic treatment reveals that the decrease in sperm count, motility and viability due to toxic effects of arsenic is minimized. As a possible mechanism it could be stated that either pea or casein or both have a recovery role on arsenic tri oxide mediated toxicity by inducing an antioxidant effect against the oxidative stress [15]. The antioxidant properties of milk casein have been established [15]. Studies also indicate that casein phosphopeptides (CPP) and casein hydrolysate bind the peroxidant and thus lipid peroxidation is suppressed [1517]. Studies for finding the antioxidant mechanisms of pea and wheat have also been done [5]. The pea or pea and casein supplement is effective in reducing the Parameters Studied Control groupA Treated groupB Percent decrease % Significance Level between A&B Protein diet supplemented groupC Percentage restored % Significance Level between B&C Mean±SEM Mean±SEM Mean±SEM 1) Sperm count (10^6/cauda) 135.14±5.87 122.24± 5.7 9.5 P<0.001 131.92± 5.31 75 P>0.001 2) Sperm viability (%)

27.16±3.42 14.7± 1.54 46 P<0.001 22.63 ± 3.93 64 P>0.001 3) Sperm motility (%) 1st hr 97.66 ±0.71 89.16 ± 2.8 8.7 P<0.001 94 ± 1.11 57 P>0.001 2nd hr 81 ±1.01 73 ± 3.2 98 P<0.001 74.5 ± 0.71 19 P>0.001 3rd hr 61.33 ±2.7 54.66 ± 4.5 10.3 P<0.001 54.8 ± 2.98 2 P<0.20 Al Ame en J Med S c i Volume2, No.1, 2009 Mukherjee S & Mukhopadhyay PK ©2009. Al Ameen Charitable Fund Trust, Bangalore 77

production of nitric oxide and MDA which could markedly increase the activity of the antioxidant enzymes. This could not only overcome the oxidative stress caused by arsenic but also suppresses the ROS generation from other sources. [6]. Studies on arsenic trioxide induced toxicity on male gonad reveal a good deal of changes in histology of seminiferous tubule, associated with decreased spermatozoal mass. Sperm count, viability, and motility are seen to be affected and the possible mechanisms behind these changes have been discussed. Supplementation of specific proteins with the normal diet causes significant recovery from all these toxic effects. In summary we have demonstrated that the pea and casein by virtue of having antioxidant properties are responsible for restoration of normal gonadal status when supplemented with simultaneous arsenic exposure. Reference 1. Clarkson, T; Envioron. Health Prospect, 1995, 103, 9—12. 2. Roy, P; Saha, A Metabolism and toxicity of arsenic: A human carcinogen Current Sc. Vol 82 No.1, Jan 2002, Pg-38—45. 3. Mandal,BK, ;Roychowdhury,T, Samanta,G , BasuGK, ChowdhuryPP, Chanda,CR (1996) Current science 70, 976—986 4. Sarkar M, Ray Chaudhuri G, Chattopadhyay A, Biswas NM; Effect of sodium arsenite on spermatogenesis, plasma gonadotrophins and testosterone in rats. Indian Asian J Androl 2003 Mar; 5: 27-31 5. Kreiger DT, Porlow MJ, Gibson MJ, Davis TF , Brain grafts reverse hypogonadism of gonadotropin releasing hormone deficiency. Nature (1982) 298,468-471 6. Mukherjee,S, Das,D Darbar D and Mitra*,C Dietary intervention affects arsenic-generated nitric oxide and reactive oxygen intermediate toxicity in islet cells of rats Current Science, vol. 85, no. 6, 25 September 2003 786-793. 7. S.Maiti and A. K. Chatterjee; Effects on levels of glutathione and some related enzymes in tissues after an acute arsenic exposure in rats and their relationship to dietary protein deficiency. Arch Toxicol 2001 Nov;75(9):531-7 8. P. Y. Y. Wong and D. D. Kitts* Chemistry of Buttermilk Solid Antioxidant Activity J. Dairy Sci.

2003, 86:1541-1547. 9. Chatterjee, A. K., Jamdar, S. C. and Ghosh, B. B., Effect of riboflavin deficiency on incorporation in vivo of [14C] amino acid into liver proteins of rats. Br. J. Nutr., 1970, 24, 635–640. 10. Jequier A.M., Male Infertility. A guide for clinician. Blackwell Pg: 64—65. 11. Jequier A.M. Male Infertility. A guide for clinician. Blackwell Pg: 55—61. 12. Jequier A.M. Male Infertility. A guide for clinician. Blackwell Pg: 61—63. 13. Xu Y, Wang Y, Zheng Q, Li X, Li B, Jin Y, Sun X, Sun G. Association of oxidative stress with arsenic methylation in chronic arsenic-exposed children and adults. Toxicol Appl Pharmacol. 2008 Jul 1. (Epub ahead of print). 14. Osbaldo Ramos, Leticia Carrizales, Leticia Yáñez, Jesús Mejía, Lilia Batres, deogracias Ortíz, and Fernando Díaz-Barriga : Arsenic Increased Lipid Peroxidation in Rat Tissues by a Mechanism Independent of Glutathione Levels. Environmental Health Perspectives 103, Supplement 1, February 1995. 15. Cervato G, Cazzola R, Cestaro B. Int. Studies on the antioxidant activity of milk caseins. J Food Sci Nutr. 1999 Jul; 50(4):291-6. 16. Taylor MJ, Richardson T. Antioxidant activity of skim milk: effect of sonication. J Dairy Sci. 1980 Nov; 63(11):1938-42. 17. Diaz M, Decker EA.: Antioxidant mechanisms of caseinophosphopeptides and casein hydrolysates and their application in ground beef. J Agric Food Chem. 2004 Dec 29; 52(26):8208-13. Address for correspondence: Dr. Prabir Kumar Mukhopadhyay, Department of Physiology, Presidency College, 86/1, College street, Kolkata- 700073. West Bengal, India. Comparative toxicity of trivalent and pentavalent inorganic and methylated arsenicals in rat and human cells Styblo, M | Del Razo, LM | Vega, L | Germolec, DR | LeCluyse, EL | Hamilton, GA | Reed, W | Wang, C | Cullen, WR | Thomas, DJ Archives of Toxicology [Arch. Toxicol.]. Vol. 74, no. 6, pp. 289-299. 18 Aug 2000. Biomethylation is considered a major detoxification pathway for inorganic arsenicals (iAs). According to the postulated metabolic scheme, the methylation of iAs yields methylated metabolites in which arsenic is present in both pentavalent and trivalent forms. Pentavalent mono- and dimethylated arsenicals are less acutely toxic than iAs. However, little is known about the toxicity of trivalent methylated species. In the work reported here the toxicities of iAs and trivalent and pentavalent methylated arsenicals were examined in cultured human cells derived from tissues that are considered a major site for iAs methylation (liver) or targets for carcinogenic effects associated with exposure to iAs (skin, urinary bladder, and lung). To characterize the role of methylation in the protection against toxicity of arsenicals, the capacities of cells to produce methylated metabolites were also examined. In addition to human cells, primary rat hepatocytes were used as methylating controls. Among the arsenicals examined, trivalent monomethylated species were the most cytotoxic in all cell types. Trivalent dimethylated arsenicals were at least as cytotoxic as trivalent iAs (arsenite) for most cell types. Pentavalent arsenicals were significantly less cytotoxic than their trivalent analogs. Among the cell types examined, primary rat hepatocytes exhibited the greatest methylation capacity for iAs followed by primary human hepatocytes, epidermal keratinocytes, and bronchial epithelial cells. Cells derived from human bladder did not methylate iAs. There was no apparent correlation between susceptibility of cells to arsenic toxicity and their capacity to methylate iAs. These results suggest that (1) trivalent methylated arsenicals, intermediary products of arsenic methylation, may significantly contribute to the adverse effects associated with exposure to iAs, and (2) high methylation capacity does not protect cells from the acute toxicity of trivalent arsenicals. Descriptors: Article Subject Terms Arsenic | Liver | Lung | Methylation | Skin | Urinary bladder

Abstract The effects of arsenic and ethanol interaction on blood, liver and serum biochemical indices, and arsenic concentration in soft tissues of rats were investigated to determine the influence of these substances in inducing susceptibility to arsenic poisoning. Arsenic, intraperitoneally (100 ppm, once, daily), ethanol in drinking water (10%), or the combination were administered for a period of 6 weeks. Both the chemicals had some additive effects in marginally elevating blood zinc protoporphyrin. Glutathione (GSH) concentrations of blood and liver were reduced by both arsenic and ethanol; however, there was a more pronounced depletion of hepatic GSH concentration in animals coexposed to arsenic and ethanol. Combined arsenic plus ethanol exposure led to significantly more elevated activities of serum transaminases than in animals administered arsenic or ethanol alone. Histopathological alterations in

kidneys and liver occurred following arsenic exposure. Arsenic plus ethanol produced more pronounced liver lesions, whereas kidney changes were the same as with arsenic alone. The concentrations of arsenic in kidney and liver were higher in rats exposed to arsenic plus ethanol. The results suggest that animals exposed to arsenic plus ethanol are more vulnerable to arsenic toxicity.

Subscriptions Performing your original search, arsenic toxicity in rats, within the ACS Publications collection will retrieve 218 results. Go to results Prev. Article Next Article Table of Contents Research Article Insecticide-Rodenticide Toxicology, The Acute Oral Toxicity in Rats of Several DietArsenic Trioxide Mixtures First Page Hi-Res PDF[233 KB] E. W. Packman, D. D. Abbott, J. W. E. Harrison J. Agric. Food Chem., 1961, 9 (4), pp 271–272 DOI: 10.1021/jf60116a008 Publication Date: July 1961 Note: In lieu of an abstract, this is the article's first page.

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Lead-, cadmium-, and arsenic-induced DNA damage in rat germinal cells. Full Abstract Toxic agents can interfere with the male reproductive system at many targets. One of the major unresolved questions concerning male infertility is identification of its molecular origins. Clinical and animal studies indicate that abnormalities of spermatogenesis result from exposure to three toxic metals (lead acetate, cadmium chloride, and arsenic trioxide), but the effects on primary spermatocyte DNA of the male rat after chronic exposure to these metals have not been identified. The aims of this study were to analyze, in three independent experiments, the DNA damage induced by lead (Pb), cadmium (Cd), and arsenic (As) in rat germinal cells during three time periods, and to determine the

relationship between DNA damage and blood Pb, blood Cd, and urine As levels. For lead acetate and cadmium chloride experiments, blood was collected by cardiac puncture, while for arsenic trioxide a 24-h urine sample was collected. Afterward, the animals were sacrificed by decapitation. Pachytene spermatocytes from rat testes were purified by trypsin digestion followed by centrifugal elutriation. After establishment of cell purity and viability, DNA damage (tail length) was measured employing a single cell gel/comet assay. Significant DNA damage was found in primary spermatocytes from rats with chronic exposure (13 weeks) to toxic metals. In conclusion, these findings indicate that exposure to toxic metals affects primary spermatocyte DNA and are suggestive of possible direct testicular toxicity.

27. Blair PC, Thompson MB, Bechtold M, Wilson RE, Moorman MP, Fowler BA. Evidence for oxidative damage to red blood cells in mice induced by arsine gas. Toxicology 63:25-34 (1990). 28. Yamanaka K, Hasegawa A, Sawamura R, Okada S. Dimethylated arsenics induce DNA strand breaks in lung via the production of active oxygen in mice. Biochem Biophys Res Commun 165:43-50 (1989). 29. Keyse SM, Tyrrell RM. Heme oxygenase is the major 32-kDa stress protein induced in human skin fibroblasts by UVA radiation, hydrogen peroxide, and sodium arsenite. Proc Natl Acad Sci USA 86:99-103 (1989). 30. Shannon RL, Strayer, DS. Arsenic-induced skin toxicity. Human Toxicol 8:99104 (1989). Last Update: September 30, 1998 |

ARSENIC Toxicity Significant exposure to arsenic occurs through both anthropogenic and natural sources. Arsenic is released into the air by volcanoes and is a natural contaminant of some deep-water wells. Occupational exposure to arsenic is common in the smelting industry (in which arsenic is a byproduct of ores containing lead, gold, zinc, cobalt, and nickel) and is increasing in the microelectronics industry (in which gallium arsenide is responsible). Low-level arsenic exposure continues to take place in the general population (as do some cases of high-dose poisoning) through the commercial use of inorganic arsenic compounds in common products such as wood preservatives, pesticides, herbicides, fungicides, and paints; through the consumption of foods and the smoking of tobacco treated with arsenic-containing pesticides; and through the burning of fossil fuels in which arsenic is a contaminant. Arsenic was also a major ingredient of Fowler's solution and continues to be found in some folk remedies.

METABOLISM The toxicity of an arsenic-containing compound depends on its valence state (zero-valent, trivalent, or pentavalent), its form (inorganic or organic), and the physical aspects governing its absorption and elimination. In general, inorganic arsenic is more toxic than organic arsenic, and trivalent arsenite is more toxic than pentavalent and zero-valent arsenic. The normal intake of arsenic by adults occurs primarily through ingestion and averages around 50 ug/d (range, 8 to 104 ug/d). Most (around 64 percent) of this amount is accounted for by organic arsenic from fish, seafood, and algae; the specific arsenic compounds obtained from these sources are arsenobentaine and arsenocholine, which are relatively nontoxic and are rapidly excreted in unchanged form in the urine. After absorption, inorganic arsenic accumulates in the liver, spleen, kidneys, lungs, and gastrointestinal tract. It is then rapidly cleared from these sites but leaves a residue in keratin-rich tissues such as skin, hair, and nails. Arsenite (+5) undergoes biomethylation in the liver to the less toxic metabolites methylarsenic acid and dimethylarsenic acid; biomethylation can quickly become saturated, however, and the result is the deposition of increasing doses of inorganic arsenic in soft tissues. Arsenic, particularly in its trivalent form, inhibits critical sulfhydrylcontaining enzymes. In the pentavalent form, the competitive substitution of arsenic for phosphate can lead to rapid hydrolysis of the high-energy bonds in compounds such as ATP. CLINICAL FEATURES Acute arsenic poisoning from ingestion results in increased permeability of small blood vessels and inflammation and necrosis of the intestinal mucosa; these changes manifest as hemorrhagic gastroenteritis, fluid loss, and hypotension. Delayed cardiomyopathy accompanied by electrocardiographic abnormalities may develop. Symptoms include nausea, vomiting, diarrhea, abdominal pain, delirium, coma, and seizures. A garlicky odor may be detectable on the breath. Acute tubular necrosis and hemolysis may develop. The reported lethal dose of arsenic ranges from 120 to 200 mg in adults and is 2 mg/kg in children. Arsine gas causes severe hemolysis within 3 to 4 h of exposure and can lead to acute tubular necrosis and renal failure. In chronic arsenic poisoning, the onset of symptoms comes at 2 to 8 weeks. Typical findings are skin and nail changes, such as hyperkeratosis, hyperpigmentation, exfoliative dermatitis, and Mees' lines (transverse white striae of the fingernails); sensory and motor polyneuritis manifesting as numbness and tingling in a "stocking-glove" distribution, distal weakness, and quadriplegia; and inflammation of the respiratory mucosa. Epidemiologic evidence has linked chronic consumption of water containing arsenic at concentrations in the range of 10 to 1820 ppb with vasospasm and peripheral vascular insufficiency culminating in "blackfoot disease," a gangrenous

condition affecting the extremities. Chronic arsenic exposure has also been associated with a greatly elevated risk of skin cancer and possibly of cancers of the lung, liver (angiosarcoma), bladder, kidney, and colon. LABORATORY FINDINGS When acute arsenic poisoning is suspected, an x-ray of the abdomen may reveal ingested arsenic, which is radiopaque. The serum arsenic level may exceed 0.9 umol/L (7 ug/dL); however, arsenic is rapidly cleared from the blood. Electrocardiographic findings may include QRS complex broadening, QT prolongation, ST-segment depression, T-wave flattening, and multifocal ventricular tachycardia. Urinary arsenic should be measured in 24-h specimens collected after 48 h of abstinence from seafood ingestion; normally, levels of total urinary arsenic excretion are less than 0.67 umol/d (50 ug/d). Arsenic may be detected in the hair and nails for months after exposure. Abnormal liver function, anemia, leukocytosis or leukopenia, proteinuria, and hematuria may be detected. Electromyography may reveal features similar to those of Guillain-Barre syndrome. TREATMENT

Vomiting should be induced with ipecac in the alert patient with acute arsenic ingestion. Gastric lavage may be useful; activated charcoal with a cathartic (such as sorbitol) may be tried. Aggressive therapy with intravenous fluid and electrolyte replacement in an intensive-care setting may be life-saving. Dimercaprol is the chelating agent of choice and is administered intramuscularly at an initial dose of 3 to 5 mg/kg on the following schedule: every 4 h for 2 days, every 6 h on the third day, and every 12 h thereafter for 10 days. (An oral chelating agent may be substituted.) Succimer is sometimes an effective alternative, particularly if adverse reactions to dimercaprol develop (such as nausea, vomiting, headache, increased blood pressure, and convulsions). In cases of renal failure, doses should be adjusted carefully, and hemodialysis may be needed to remove the chelating agent-arsenic complex. Arsine gas poisoning should be treated supportively with the goals of maintaining renal function and circulating red-cell mass.

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Arsenic and many of its compounds are especially potent poisons. Arsenic disrupts ATP production through several mechanisms. At the level of the citric acid cycle, arsenic inhibits pyruvate dehydrogenase and by competing with phosphate it uncouples oxidative phosphorylation, thus inhibiting energy-linked reduction of NAD+, mitochondrial respiration, and ATP synthesis. Hydrogen peroxide production is also increased, which might form reactive oxygen species and oxidative stress. These metabolic interferences lead to death from multisystem organ failure (see arsenic poisoning) probably from necrotic cell death, not apoptosis. A post mortem reveals brick red colored mucosa, due to severe hemorrhage. Although arsenic causes toxicity, it can also play a protective role.[1]

[edit] Exposure Exposure to arsenic can occur from the environment and food consumption.[2] As a component of geologic formations, arsenic can be washed out into the ground water. Human activities such as mining and ore smelting can contribute to this process. Moreover, certain prokaryotes can metabolize arsenic in order to generate energy. This may enhance the process of solubilization from the solid to the aqueous phase. [3]

[edit] Kinetics The two forms of arsenic, reduced (trivalent As (III)) and oxidized (pentavalent As(V)), can be absorbed, and accumulated in tissues and body fluids.[2] In the liver, the metabolism of arsenic involves enzymatic and non-enzymatic methylation, the most frequently excreted metabolite (≥ 90%) in the urine of mammals is dimethylarsinic acid (DMA(V)).[4] In humans inorganic arsenic is reduced nonenzymatically from pentavalent to trivalent state using glutathione (GSH) or its is mediated by enzymes. Methylation occurs through methyltransferase enzymes. S-adenosylmethionine (SAM) may serve as methyl donor. Various pathways are used, the principal route being dependant on the current environment of the cell.[5] Resulting metabolites are monomethylarsonous acid (MMA(III)) and dimethylarsinous acid (DMA(III)). These metabolites are less likely to react with tissue components than inorganic arsenic (especially +3 As) and as they are easly excreted, methylation has been regarded as a detoxification process. While reduction from +5 As to +3 As may be considered as a bioactivation.[6] Another suggestion is that methylation might be a detoxification if "As[III] intermediates are not permitted to accumulate" because the pentavalent organoarsenics have a lower affinity to thiol groups than inorganic pentavalent arsenics.[5] Gebel (2002) stated that methylation is a detoxification through accelerated excretion. With regard to carcinogenicity it has been suggested that methylation should be regarded as a toxification. (Kitchin 2001, Kenyon et al. 2001, Styblo et al. 2002)

Arsenic, especially +3 As, binds to single, but with higher affinity to vicinal sulfhydryl groups, thus reacts with a variety of proteins and inhibits their activity. It was also proposed that binding of arsenite at nonessential sites might contribute to detoxification (Aposhian 1989). Arsenite inhibits members of the disulfide oxidoreductase family like glutathione reductase (Rodríguez et al. 2005) and thioredoxin reductase.[7] The remaining non-excreted arsenic (≤ 10%) accumulates in cells, which over time may lead to skin, bladder, kidney, liver, lung, and prostate cancers.[4] Other forms of arsenic toxicity in humans have been observed in blood, bone marrow, cardiac, central nervous system, gastrointestinal, gonadal, kidney, liver, pancreatic, and skin tissues.[4]

[edit] Mechanism Arsenite inhibits not only the formation of Acetyl-CoA but also the enzyme succinic dehydrogenase. Arsenate can replace phosphate in many reactions. It is able to form Glc-6-Arsenate in vitro; therefore it has been argued that hexokinase could be inhibited (reported by Hughes 2002). (Eventually this may be a mechanism leading to muscle weakness in chronic arsenic poisoning.) In the glyceraldehyde-3-P-dehydrogenase reaction arsenate attacks the enzyme-bound thioester. The formed 1-arseno-3-posphoglycerate is unstable and hydrolyzes spontaneously. Thus, ATP formation in Glycolysis is inhibited while bypassing the phosphoglycerate kinase reaction. (Moreover, the formation of 2,3bisphosphoglycerate in erythrocytes might be affected, followed by a higher oxygen affinity of hemoglobin and subsequently enhanced cyanosis) As shown by Gresser (1981), submitochondrial particles synthesize Adenosine5’-diphosphate-arsenate from ADP and arsenate in presence of succinate. Thus, by a variety of mechanisms arsenate leads to an impairment of cell respiration and subsequently diminished ATP formation. This is consistent with observed ATP depletion of exposed cells and histopathological findings of mitochondrial and cell swelling, glycogen depletion in liver cells and fatty change in liver, heart and kidney. Experiments demonstrated enhanced arterial thrombosis in a rat animal model, elevations of serotonin levels, thromboxane A[2] and adhesion proteins in platelets, while human platelets showed similar responses (Lee et al. 2002). The effect on vascular endothelium may eventually be mediated by the arsenicinduced formation of nitric oxide. It was demonstrated that +3 As concentrations substantially lower than concentrations required for inhibition of the lysosomal protease cathepsin L in B cell line TA3 were sufficient to trigger apoptosis in the same B cell line, while the latter could be a mechanism mediating immunosuppressive effects (Harrison et al. 2001).

[edit] Carcinogenicity It is still a matter of debate whether DNA repair inhibition or alterations in the status of DNA methylation are responsible for the carcinogenic potential of As. As vicinal sulfhydryl groups are frequently found in DNA-binding proteins, transcription factors and DNA-repair proteins, interaction of arsenic with these molecules appears to be likely. However, in vitro, most purified DNA repair enzymes are rather insensitive to As, but in cell culture, As produces a dosedependant decrease of DNA ligase activity. This might indicate that inhibition of DNA repair is an indirect effect due to changes in cellular redox levels or alterated signal transduction and consequent gene expression (Hu et al. 1998). In spite of its carcinogenicity, the potential of arsenic to induce point mutations is weak. If administered with point mutagens it enhances the frequency of mutations in a syngergistic way (Gebel 2001). Its comoutagenic effects may be explained by interference with base and nucleotide excision repair, eventually through interaction with zinc finger structures (Hartwig et al. 2002). DMA showed to effectuate DNA single stand breaks resulting from inhibition of repair enzymes at levels of 5 to 100 mM in human epithelial type II cells (Yamanaka 1997). (Significant DNA strand breaks have also been observed by Bau et al. 2002) +3 MMA and +3 DMA were also shown to be directly genotoxic by effectuating scissions in supercoiled ΦX174 DNA (Mass et al. 2001). Increased arsenic exposure is associated with an increased frequency of chromosomal aberrations (recently shown in an epidemiologic stud by Maki-Paakkanen et al. 1998), mikronuklei (Warner et al. 1994, Gonseblatt et al. 1997) and sister-chromatid exchanges. An explanation for chromosomal aberrations is the sensitivity of the protein tubulin and the mitotic spindle to arsenic. Histological observations confirm effects on cellular integrity, shape and locomotion (reviewed by Bernstam et al. 2000). +3 DMA is able to form reactive oxygen species (ROS) by reaction with molecular oxygen. Resulting metabolites are the dimethylarsenic radical and the dimethylarsenic peroxyl radical (Yamanaka et al. 1990). Both +5 DMA and +3 DMA were shown to release iron from horse spleen as well as from human liver ferritin if ascorbic acid was administered simultaneously. Thus, formation of ROS can be promoted (Ahmad et al. 2000). Moreover, Arsenic could cause oxidative stress by depleting the cell’s antioxidants, especially the ones containing thiol groups. The accumulation of ROS like the cited above and hydroxyl radicals, superoxide radicals and hydrogen peroxides causes aberrant gene expression at low concentrations and lesions of lipids, proteins and DNA in higher concentrations which eventually lead to cellular death. In a rat animal model, urine levels of 8-hydroxy-2’-desoxyguanosine (as a biomarker of ROS DNA damage) were measured after treatment with DMA. In comparison to control levels, they turned out to be significantly increased (Yamanaka et al. 2001). This

theory is further supported by a cross-sectional study which found elevated mean serum lipid peroxides (LPO) in the As exposed individuals which correlated with blood levels of inorganic arsenic and methylated metabolites and inversely correlated with nonprotein sulfhydryl (NPSH) levels in whole blood (Pi et al. 2002). Another study found an association of As levels in whole blood with the level of reactive oxidants in plasma and an inverse relationship with plasma antioxidants (Wu et al. 2001). A finding of the latter study indicates that methylation might in fact be a detoxification pathway with regard to oxidative stress: the results showed that the lower the As methylation capacity was, the lower the level of plasma antioxidant capacity. As reviewed by Kitchin (2001), the oxidative stress theory provides an explanation for the preferred tumor sites connected with arsenic exposure. Considering that a high partial pressure of oxygen is present in lungs and +3 DMA is excreted in gaseous state via the lungs this seems to be a plausible mechanism for special vulnerability. The fact that DMA is produced by methylation in the liver, excreted via the kidneys and latter on stored in the bladder accounts for the other tumor localizations. Regarding DNA methylation, some studies suggest interaction of As with methyltransferases which leads to an inactivation of tumor suppressor genes through hypermethylation, others state that hypomethylation might occur due to a lack of SAM resulting in aberrant gene activation (Goering et al. 1999). An experiment by Zhong et al. (2001) with arsenite-exposed human lung A549, kidney UOK123, UOK109 and UOK121 cells isolated eight different DNA fragments by methylation-sensitive arbitrarily primed PCR. It turned out that six of the fragments were hyper- and two of them were hypomethylated. Higher levels of DNA methltransferase mRNA and enzyme activity were found. Kitchin (2001) proposed a model of altered growth factors which lead to cell proliferation and thus to carcinogenesis. From observations it is known that chronic low-dose arsenic poisoning can lead to increased tolerance to its acute toxicity. MRP1-overexpressing lung tumor GLC4/Sb30 cells poorly accumulate arsenite and arsenate. This is mediated through MRP-1 dependent efflux (Vernhet et al. 2000). The efflux requires GSH, but no As-GSH complex formation (Salerno et al. 2002). Resistant Cell lines have also been studied by Gebel 2001 and Brambila et al. 2002. Although a lot of mechanisms have been proposed, no definite model can be given for the mechanisms of chronic arsenic poisoning. The prevailing events of toxicity and carcinogenicity might be quite tissuespecific. Current consensus on the mode of carcinogenesis is that it acts primarily as a tumor promoter. Its co-carcinogenicity has been demonstrated in several models. However, the finding of several studies that chronically arsenicexposed Andean populations (as most extremely exposed to UV-light) do not develop skin cancer with chronic arsenic exposure, is puzzling (cited by Gebel 2000).[8]

[edit] Heat shock response Another aspect is the similarity of arsenic effects to the heat shock response. Short-term arsenic exposure has effects on signal transduction inducing heat shock proteins with masses of 27,60,70,72,90,110 kDa as well as metallotionein, ubiquitin, mitogen-activated [MAP] kinases, extracellular regulated kinase [ERK], c-jun terminal kinases [JNK] and p38 (Bernstam 2000, Del Razo 2001). Via JNK and p38 it activates c-fos, c-jun and egr-1 which are usually activated by growth factors and cytokines (Cavigelli et al. 1996, Ludwig et al. 1998, Bernstam et al. 2000) The effects are largely dependant on the dosing regime and may be as well inversed. As shown by some experiments reviewed by Del Razo (2001), ROS induced by low levels of inorganic arsenic increase the transcription and the activity of the activator protein 1 (AP-1) and the nuclear factor-kB (NF-kB) (maybe enhanced by elevated MAPK levels), which results in c-fos/c-jun activation, over-secretion of pro-inflammatory and growth promoting cytokines stimulating cell proliferation (reviewed by Simeonova and Luster 2000). Germolec et al. (1996) found an increased cytokine expression and cell proliferation in skin biopsies from individuals chronically exposed to arseniccontaminated drinking water. Increased AP-1 and NF-kB obviously also result in an up-regulation of mdm2 protein, which decreases p53 protein levels (Hamadeh et al. 1999) Thus, taking into account p53’s function, a lack of it could cause a faster accumulation of mutations contributing to carcinogenesis. However, high levels of inorganic arsenic inhibit NF-kB activation and cell proliferation. An experiment of Hu et al. (2002) demonstrated increased binding activity of AP-1 and NF-kB after acute (24 h) exposure to +3 sodium arsenite, whereas long-term exposure (10–12 weeks) yielded the opposite result. The authors conclude that the former may be interpreted as a defense response while the latter could lead to carcinogenesis. As the contradicting findings and connected mechanistic hypotheses indicate, there is a difference in acute and chronic effects of arsenic on signal transduction which is not clearly understood yet.

[edit] Oxidative stress Studies have demonstrated that the oxidative stress generated by arsenic may disrupt the signal transduction pathways of the nuclear transcriptional factors PPAR’s, AP-1, and NFκB,[4][9][10] as well as the pro-inflammatory cytokines IL-8 and TNF-α.[9][4][11][12][10][13][14][15] The interference of oxidative stress with signal transduction pathways may affect physiological processes associated with cell growth, metabolic syndrome X, glucose homeostasis, lipid metabolism, obesity, insulin resistance, inflammation, and diabetes-2.[16][17][18] Recent scientific evidence has elucidated the physiological roles of the PPAR’s in the ωhydroxylation of fatty acids and the inhibition of pro-inflammatory transcription factors (NFκB and AP-1), pro-inflammatory cytokines (IL-1, -6, -8, -12, and TNFα), cell4 adhesion molecules (ICAM-1 and VCAM-1), inducible nitric oxide synthase, proinflammatory nitric oxide (NO), and anti-apoptotic factors.[4][11][16][18][19]

Epidemiological studies have suggested a correlation between chronic consumption of drinking water contaminated with arsenic and the incidence of Type 2-diabetes.[4] The human liver after exposure to therapeutic drugs may exhibit hepatic non-cirrhotic portal hypertension, fibrosis, and cirrhosis.[4] However, the literature provides insufficient scientific evidence to show cause and effect between arsenic and the onset of diabetes mellitus Type 2.[4]

[edit] References 1. ^ Klaassen, Curtis; Watkins, John (2003). Casarett and Doull's Essentials of Toxicology. McGraw-Hill. pp. 512. ISBN 978-0071389143. 2. ^ a b Ueki, K.; Kondo, T.; Tseng, Y.-H.; Kahn, R. C. (2004). "Central role of suppressors of cytokine signaling proteins in hepatic steatosis, insulin resistance, and the metabolic syndrome in the mouse". Proceedings of the National Academy of Sciences of the United States of America 101 (28): 10422–10427. doi:10.1073/pnas.0402511101. 3. ^ "The ecology of arsenic". Science 300: 939–944. 2003. doi:10.1126/science.1081903. http://www.sciencemag.org/cgi/content/abstract/sci;300/5621/939?maxtoshow=& HITS=10&hits=10&RESULTFORMAT=&fulltext=oremland+2003&searchid=1 &FIRSTINDEX=0&resourcetype=HWCIT. 4. ^ a b c d e f g h i Vigo, J. B., and J. T. Ellzey (2006). "Effects of Arsenic Toxicity at the Cellular Level: A Review". Texas Journal of Microscopy 37 (2): 45–49. 5. ^ a b "A chemical hypothesis for arsenic methylation in mammals". Chem Biol Interact 88 (2-3): 89–14. 1993. http://www.ncbi.nlm.nih.gov/pubmed/8403081. 6. ^ "Role of metabolism in arsenic toxicity". Pharmacol Toxicol 89 (1): 1–5. 2001. http://www.ncbi.nlm.nih.gov/pubmed/11484904. 7. ^ Environmental and Occupational Medicine. Environmental and Occupational Medicine. 2006. pp. 1014–1015. http://books.google.com/books?id=H4Sv9XY296oC&pg=RA2PA1014&lpg=RA2-PA1014#PRA2-PA1014,M1. 8. ^ "Confounding variables in the environmental toxicology of arsenic". Toxicology 144: 155–1062. 2000. http://www.ncbi.nlm.nih.gov/pubmed/10781883. 9. ^ a b Hu Y., X. Jin, and E. Snow (2002). "Effect of arsenic on transcription factor AP-1 and NF- K B". Toxicology Letters 133: 33. doi:10.1016/S03784274(02)00083-8. 10. ^ a b Walton FS, Harmon AW, Paul DS, Drobná Z, Patel YM, Styblo M (August 2004). "Inhibition of insulin-dependent glucose uptake by trivalent arsenicals: possible mechanism of arsenic-induced diabetes". Toxicol. Appl. Pharmacol. 198 (3): 424–33. doi:10.1016/j.taap.2003.10.026. PMID 15276423. 11. ^ a b Black PH (October 2003). "The inflammatory response is an integral part of the stress response: Implications for atherosclerosis, insulin resistance, type II diabetes and metabolic syndrome X". Brain Behav. Immun. 17 (5): 350–64. doi:10.1016/S0889-1591(03)00048-5. PMID 12946657. 12. ^ Carey AL, Lamont B, Andrikopoulos S, Koukoulas I, Proietto J, Febbraio MA (March 2003). "Interleukin-6 gene expression is increased in insulin-resistant rat

skeletal muscle following insulin stimulation". Biochem. Biophys. Res. Commun. 302 (4): 837–40. doi:10.1016/S0006-291X(03)00267-5. PMID 12646246. 13. ^ Dandona P, Aljada A, Bandyopadhyay A (January 2004). "Inflammation: the link between insulin resistance, obesity and diabetes". Trends Immunol. 25 (1): 4– 7. doi:10.1016/j.it.2003.10.013. PMID 14698276. http://linkinghub.elsevier.com/retrieve/pii/S1471490603003363. 14. ^ Fischer CP, Perstrup LB, Berntsen A, Eskildsen P, Pedersen BK (November 2005). "Elevated plasma interleukin-18 is a marker of insulin-resistance in type 2 diabetic and non-diabetic humans". Clin. Immunol. 117 (2): 152–60. doi:10.1016/j.clim.2005.07.008. PMID 16112617. 15. ^ Gentry PR, Covington TR, Mann S, Shipp AM, Yager JW, Clewell HJ (January 2004). "Physiologically based pharmacokinetic modeling of arsenic in the mouse". J. Toxicol. Environ. Health Part A 67 (1): 43–71. doi:10.1080/15287390490253660. PMID 14668111. 16. ^ a b Kota BP, Huang TH, Roufogalis BD (February 2005). "An overview on biological mechanisms of PPARs". Pharmacol. Res. 51 (2): 85–94. doi:10.1016/j.phrs.2004.07.012. PMID 15629253. 17. ^ Luquet, S., C. Gaudel, D. Holst, J. Lopez-Soriano, C. Jehl-Pietri, A. Fredenrich (2005). Biochimica et Biophysica Acta 1740: 313–317. 18. ^ a b Moraes, La; Piqueras, L; Bishop-Bailey, D (Jun 2006). "Peroxisome proliferator-activated receptors and inflammation.". Pharmacology & therapeutics 110 (3): 371–85. doi:10.1016/j.pharmthera.2005.08.007. PMID 16168490. 19. ^ Hara K, Okada T, Tobe K, et al. (April 2000). "The Pro12Ala polymorphism in PPAR gamma2 may confer resistance to type 2 diabetes". Biochem. Biophys. Res. Commun. 271 (1): 212–6. doi:10.1006/bbrc.2000.2605. PMID 10777704.

Paul B. Tchounwou1, Jose A. Centeno2 and Anita K. Patlolla1 (1) Molecular Toxicology Research Laboratory, NIH-Center for Environmental Health, School of Science and Technology, Jackson State University, Jackson, MS 39217, USA (2) Environmental and Toxicologic Pathology Laboratory, Armed Forces Institute of Pathology, Washington DC, 20306, USA

Abstract A comprehensive analysis of published data

indicates that arsenic exposure induces cardiovascular diseases, developmental abnormalities, neurologic and neurobehavioral disorders, diabetes, hearing loss, hematologic disorders, and various types of cancer. Although exposure may occur via the dermal, and parenteral routes, the main pathways of exposure include ingestion, and inhalation. The severity of adverse health effects is related to the chemical form of arsenic, and is also time- and dose-dependent. Recent reports have pointed out that arsenic poisoning appears to be one of the major public health problems of pandemic nature. Acute and chronic exposure to

arsenic has been reported in several countries of the world where a large proportion of drinking water (groundwater) is contaminated with high concentrations of arsenic. Research has also pointed significantly higher standardized mortality rates for cancers of the bladder, kidney, skin, liver, and colon in many areas of arsenic pollution. There is therefore a great need for developing a comprehensive health risk assessment (RA) concept that should be used by public health officials and environmental managers for an effective management of the health effects associated with arsenic exposure. With a special emphasis on arsenic toxicity, mutagenesis, and carcinogenesis, this paper is aimed at using the National Academy of Science''s RA framework as a guide, for developing a RA paradigm for arsenic based on a comprehensive analysis of the currently available scientific information on its physical and chemical properties, production and use, fate and transport, toxicokinetics, systemic and carcinogenic health effects, regulatory and health guidelines, analytical guidelines and treatment technologies. arsenic - contamination - health effects - risk assessments and management Arsenic-cadmium interaction in rats: toxic effects Title: in the heart and tissue metal shifts. Yanez, L : Carrizales, L : Zanatta, M T : Mejia, J J : Author: Batres, L : Diaz Barriga, F Citation: Toxicology. 1991 Apr 8; 67(2): 227-34 Abstract: Previously, we had shown that arsenic interacts with cadmium in rats; our results showed that the toxicity of a mixture of arsenic + cadmium cannot be predicted by the toxic mechanisms of the individual components. In this paper, we present further evidence about the interaction of arsenic and cadmium in rats. The results were: arsenic modified the 24 h-LD50 value of cadmium more clearly than cadmium did with the one of arsenic; based on the LD50 values, the mixtures we studied were more toxic than either metal alone. With single doses (As 10 mg/kg, Cd 2.6 mg/kg, and As 10 mg/kg + Cd 2.6 mg/kg) the mixture As + Cd was more toxic than each metal. At these doses, cadmium significantly induces the levels of glutathione, metallothionein, and lipid peroxidation in heart tissue, as compared to

a saline group of rats. Arsenic incremented glutathione and lipid peroxidation at higher values than those obtained with cadmium. The mixture of As + Cd behaved as arsenic in the induction of lipid peroxidation and glutathione and like cadmium in metallothionein induction. Finally, rats treated with As + Cd had less Cd in liver than animals treated only with cadmium, and more As in heart tissue than rats treated only with arsenic. Our results give further evidence about the arsenic-cadmium interaction in rats, demonstrate the utility of employing different biomarkers in the study of chemical mixtures and indicate that heart tissue is affected not only by the mixture of As + Cd, but also by either metal alone. Review None References: Concurrent exposure to lead, cadmium, and arsenic. Effects on toxicity and tissue metal concentrations in the rat. Mahaffey KR, Capar SG, Gladen BC, Fowler BA. Male rats were exposed to dietary Pb (200 ppm), Cd (50 ppm), or As (50 ppm) as arsenate either alone or in combination for 10 weeks using a 2 x 2 x 2 factorial design. Cd and As reduced weight gain even when differences in food intake were taken into account, and administration of both Cd and As depressed weight gain more than did either metal alone. Pb did not adversely affect food consumption or weight gain. Increased RBCs were observed after administration of Pb, Cd, or As, and more cells were observed when two or three metals were concomitantly administered. Despite increased numbers of circulating RBCs, hemoglobin and hematocrit were reduced, especially with the Pb-Cd combination. Analysis of blood chemistries showed normal ranges for blood urea nitrogen, creatinine, cholesterol, calcium, albumin, total protein, and bilirubin. Uric acid was increased by Pb, but not by Cd or As. SGOT activity was reduced by As alone. Serum alkaline phosphatase was reduced by either As or Cd but not Pb. Combinations of As and Cd did not further reduce the activity of this enzyme. Kidney weight and kidney weight/body weight ratios were increased by Pb alone, but Cd or As alone or in combination had no effect. Liver weight/body weight ratios were reduced in animals fed Cd. Kidney histology showed predominantly Pb effects, i.e., intranuclear inclusion bodies and cloudy swelling. Ultrastructural evaluation of kidneys from Pb-treated animals disclosed nuclear inclusion bodies and mitochondrial swelling. Concurrent administration of Cd reduced total mean bone and kidney Pb levels by 50% and 60%, respectively, and this was associated with a decrease in kidney intranuclear inclusions. Cd exposure also reduced renal, femur, and liver concentrations of Fe by

33%, 43%, and 63%, respectively, decreased femur Zn by 27%, but increased renal Zn by 20%. Administration of As produced mild swelling of tubule cell mitochondria, increased mean total renal Cu to 200% of control, and increased liver Fe by 44%. Dietary Pb produced increased urinary excretion of ALA and coproporphyrin. Dietary exposure to As caused increased urinary excretion of uroporphyrin and to a lesser extent coproporphyrin, whereas dietary Cd caused no significant changes in urinary levels of any of the porphyrins measured. Pb plus As produced an additive effect on coproporphyrin excretion but not that of ALA or uroporphyrin. These studies indicate that interactions between common toxic elements do occur and are characterized by alterations in both tissue trace metal levels and toxicity. Humans are exposed to a number of toxic elements in the environment; however, most experiments with laboratory animals investigate only one toxic element. To determine if concomitant exposure to lead (Pb), cadmium (Cd), and/or arsenic (As) modified the changes produced by any one metal in various parameters of toxicity, 168 male, Sprague-Dawley, young adult rats were fed nutritionally adequate diets to which had been added 0 or 200 ppm Pb as Pb acetate, or 50 ppm Cd as Cd chloride, or 50 ppm As as sodium arsenate or arsanilic acid in a factorial design for a period of 10 weeks.At these concentrations, Cd and As reduced weight gain even when differences in food intake were taken into account; administration of both Cd and As depressed weight gain more than did either metal alone. Pb did not adversely affect food consumption or weight gain. Increased numbers of red blood cells (RBCs) were observed following administration of Pb, Cd, or As; usually more cells were observed when two or three metals were administered, compared to individual metals. Despite increasing numbers of circulating RBCs, hemoglobin and hematocrit were reduced, especially with the Pb-Cd combination and the Cd-arsanilic acid combination. Specific effects of Pb on heme synthesis were observed, including increased urinary excretion of delta-aminolevulinic acid; this increase was reduced by the presence of dietary cadmium.Analyses of blood showed values for the laboratory rat within normal ranges for blood urea nitrogen, creatinine, cholesterol, calcium, albumin, total protein, and bilirubin. Uric acid was increased by Pb, with little modification by dietary Cd or As content. Serum glutamate-oxalate transaminase activity was reduced by As. Serum alkaline phosphatase was greatly reduced by either As or Cd but not Pb. Combinations of As and Cd did not further reduce the activity of this enzyme. Kidney weight and kidney weight/body weight ratios were increased by Pb alone, with no effects of Cd or As alone or as interactions. Liver weight/body weight ratios were reduced in animals fed 50 ppm dietary Cd. Kidney histology shows predominantly Pb effects, namely, intranuclear inclusion bodies and cloudy swelling. Ultrastructural evaluation of kidneys from Pb-treated animals disclosed nuclear inclusion bodies of the usual morphology and mitochondrial swelling. Concurrent administration of Cd greatly minimized Pb effects on the kidney under conditions of this experiment. Liver histology suggests an increased rate of cell turnover with either As compound, but few specific changes. PMID: 198203 [PubMed - indexed for MEDLINE]

PMCID: PMC1637428

Effect of sodium arsenite on spermatogenesis, plasma gonadotrophins and testosterone in rats Mahitosh Sarkar, Gargi Ray Chaudhuri, Aloke Chattopadhyay, Narendra Mohan Biswas Reproductive Physiology Unit, Dept. of Physiology, University Colleges of Science and Technology, Calcutta University, Calcutta-700 009, India Asian J Androl 2003 Mar; 5: 27-31

Keywords: arsenite; spermatogenesis; gonadotrophins; testosterone;

testis Abstract Aim: To investigate the effect of arsenic on spermatogenesis. Methods: Mature (4 months old) Wistar rats were intraperitoneally administered sodium arsenite at doses of 4, 5 or 6 mg.kg-1.day1 for 26 days. Different varieties of germ cells at stage VII seminiferous epithelium cycle, namely, type A spermatogonia (ASg), preleptotene spermatocytes (pLSc), midpachytene spermatocytes (mPSc) and step 7 spermatids (7Sd) were quantitatively evaluated, along with radioimmunoassay of plasma follicle-stimulating hormone (FSH), lutuneizing hormone (LH), testosterone and assessment of the epididymal sperm count. Results: In the 5 and 6 mg/kg groups, there were significant dose-dependent decreases in the accessory sex organ weights, epididymal sperm count and plasma concentrations of LH, FSH and testosterone with massive degeneration of all the germ cells at stage VII. The changes were insignificant in the 4 mg/kg group. Conclusion: Arsenite has a suppressive influence on spermatogenesis and gonadotrophin and testosterone release in rats.

1 Introduction Arsenicals are used as herbicides, fungicides and rodenticides and may cause air, soil and water pollution. Arsenical exposure through drinking water is common in many areas of the world [1-3]. Exposure to arsenic is associated with metabolic disorders, hypertrophy of adrenal glands [4] and anemia [5]. A number of sulfhydryl containing proteins and enzyme systems have been found to be altered by exposure to arsenite [6]. Arsenite affects mitochondrial enzymes and impairs tissue respiration, which seem to be related to the cellular toxicity of arsenic [7]. Gonadal effects of arsenic were first evaluated in mice, then in fishes [8-10]. Sodium arsenite has been found to have an inhibitory effect on the activity of testicular steroidogenic enzymes, ∆5-3β-hydroxysteroid dehydrogenase (∆5-3β-HSD) and 17β-hydroxysteroid dehydrogenase (17β-HSD) and to reduce the weight of testes and accessory sex glands [11]. Most of the available data on arsenic reproductive toxicity indicate that the main concern is with the developmental toxicity on the fetus [12]. Till date there is very few evidence on its male reproductively effects [11,13]. The present study was designed to quantitative study the effect of

arsenite on spermatogenesis at stage VII of the seminiferous epithelial cycle and plasma hormone concentrations, which has not been reported so far in the literature.

2 Materials and methods 2.1 Animals and treatment Adult male Wistar rats weighing 160¡À10 g (120~140 days of age) were maintained in a 12 h light and 12 h dark and 26 ¡æ~28 ¡æ animal house and standard laboratory chew and tap water were available ad libitum. Sodium arsenite was purchased from Sigma (USA) and dissolved in sterile distilled water. Thirty-two rats were divided into 4 groups of 8 animals each. Three groups of animals were injected with either 4, 5 or 6 mg.kg-1.day-1 sodium arsenite per 1 mL sterile distilled water for 26 days (Group II, III and IV, respectively). Animals of Group I were injected the same amount of distilled water for 26 days and served as the controls. Treatment for 26 days was selected as the duration of one seminiferous cycle is 13.2 days in Wistar rats. On the 27th day between 08:00 to 10:00, blood samples were collected from the hepatic vein under light ether anesthesia and after that the rats were killed following ethical procedure. Heparinized plasma was prepared and stored at -20 ¡æ until hormone radioimmuno-assay. 2.2 Body and organ weights The body weights were recorded on the first day before injection (initial) and the day of sacrifice (final). The testicles and accessory sex organs were dissected out, trimmed off the attached tissues and weighed. The relative weight of organs was expressed per 100g body weight. The left testis of each rat was used for histological study and the right for estimation of elementary arsenic content. 2.3 Histological study Immediately after removal, the testis was fixed in Bouin's fluid and embedded in paraffin. Sections of 5 µm thickness were taken from the mid portion of each testis and stained with periodic acid Schiff (PAS)-hematoxylin and examined under a light microscope. Quantitative analysis of spermatogenesis was carried out by counting the relative number of each variety of germ-cells at stage VII of the seminiferous epithelium cycle, i.e. type-A spermatogonia (ASg), preleptotene spermatocytes (pLSc), mid pachytene spermatocytes (mPSc) and step 7 spermatids (7Sd), according to the method of Leblond and Clermont [14]. The nuclei of different germ cells (except step 19 spermatids which cannot be enumerated precisely) were counted in 20 round tubules of each rat. All the counts (crude counts) of the germ cells were corrected for differences in the nuclear diameter by the formula of Abercrombie [15]: true count = (crude count ¡Á section thickness)/ (section thickness - nuclear diameter of germ cell). The nuclear diameter of each variety of germ cell was measured with a Leitz micrometer. The possibility of variable tubular shrinkage in the sections of both arsenite and vehicle injected groups were eliminated by the index of tubular shrinkage which was obtained from the average number of Sertoli cell nuclei containing prominent nucleoli in the sections of the treated rats divided by that of the controls [16]. Stage VII spermatogenesis was analyzed because this stage is highly susceptible to testosterone deficiency [17] and also reflects the final stages of spermatid maturation and thus provides evidence of spermatogenesis as a whole [18]. 2.4 Sperm count

The sperm count was determined by counting in a haemocytometer. Sperm samples were collected from the cauda epididymis. To minimize the count was repeated at least five times for each rat. 2.5 Hormone assay Plasma follicle-stimulating hormone (FSH) and lutuneizing hormone (LH) were measured by radioimmunoassay (RIA) as described in the instructions provided with the kits (NIADDK, USA). Carrier free 125I for hormone iodination was obtained from the Bhaba Atomic Research Centre, Bombay, India. Pure rat FSH (NIADDK-r FSH-1-6) and LH (NIADDK-rLH-1-6) were iodinated using the chloramine T (Sigma, USA) method of Greenwood et al [19]. The antisera to FSH and LH were NIADDK anti-r FSH-S-11 and NIADDK-anti-rLH-S-9, and were used at a final dilution of 1:100000 and 1:150 000, respectively. Goat anti-rabbit g-globulin used as the second antibody was obtained from the Indo Medix Inc., USA. The sensitivity of the assay was 2.0 µg/L for FSH and 0.15 mg/L for LH. Each sample was assayed at two concentrations, each in duplicate at a time. The intra assay coefficient of variation in each assay was 7.5 % for FSH and 6.0 % for LH. Hormone concentrations were expressed in terms of NIH reference preparation RP-2. Plasma testosterone was assayed according to Auletta et al [20]. Methodological loss was monitored and accounted for by adding 1000 c. p. m. [1β, 2β3 - H (N)¡¡] testosterone (sp. act. 50.4 Ci/m mol; New England Nuclear, Boston, MA, USA) before extraction with diethyl ether. Samples were assayed at two concentrations, each in duplicate. The antisera to testosterone was purchased from the Endocrine Science, USA, and had a 44 % cross reactivity with dihydrotestosterone. Free and bound testosterone were separated by using dextrancoated charcoal. The recovery of plasma testosterone after ether extraction was estimated to be 88.5 %. The sensitivity of the assay was 69.5 % pmol/L and the intraassay variance was 6.4 %. All samples were measured in a single assay. Since chromatographic purification of the samples was not performed, the testosterone values reported are the sum of testosterone and dihydrotestosterone. 2.6 Testicular arsenic concentration The flameless atomic absorption spectrophotometric technique [21] was used for the determination of arsenic concentration. Animals were scarified 24h after the last arsenite injection and one testis from each animal was collected and digested with a mixture of nitric acid, sulfuric acid and perchloric acid (3:1:1). Values are expressed in µg of arsenic/g of testicular wet tissue. 2.7 Statistical Analysis Data were expressed in mean¡ÀSEM. Statistical analysis was performed by analysis of variance (ANOVA) followed by multiple comparison by two-tailed t-test.

3 Results 3.1 Body and organ weights In all the treated groups, the body weight was not significantly different from that of the controls. The relative weights of the testis, seminal vesicle and ventral prostate were significantly decreased (P<0.05) after 5 mg or 6 mg treatment, but not after 4 mg of sodium arsenite treatment (Table 1). Table 1. Effect of sodium arsenite on body weight (g) and organ weights (mg % body weight) (mean¡ÀSEM, n=8) in rats. bP<0.05, compared with controls. ANOVA followed by multiple comparison two-tailed t-test.

Group Control 4 mg/kg 5 mg/kg 6 mg/kg

Body weight (g) 190.85¡À18.21 185.60¡À16.83 181.81¡À15.39 187.30¡À16.45

Testes (pair) 1478.19¡À32.93 1434.65¡À36.75 1308.39¡À34.26b 1270.89¡À35.19b

Seminal vesicle Ventral prostate 472.62¡À22.81 450.87¡À29.38 349.34¡À20.52b 322.16¡À18.98b

227.29¡À14.78 212.42¡À12.79 174.84¡À12.09b 155.42¡À13.69b

3.2 Histological findings Sodium arsenite treatment at the dose of 5 mg/kg significantly reduced the number of ASg, pLSc, mPSc and 7Sd when compared with those of the controls. Six mg/kg caused a more prominent spermatogenic arrest. No significant change was found in the cellular counts in the 4 mg/kg treated group (Table 2). Theoretically, the ratio of mPSc:7Sd is 1: 4. This ratio was 1:2.76 and 1:2.57 after 5 and 6 mg/kg treatment, respectively; the ratio of the control group was 1:3.41. The percentage spermatid degeneration (35.75 %) as calculated from the above ratio, was highly significant after 6 mg of sodium arsenite treatment (Table 2). Table 2. Effect of sodium arsenite on number of germ cells per tubular cross section at stage VII of the seminiferous epithelial cycle in rats (mean¡ÀSEM, n=8). ASg = spermatogonia A; pLSc = preleptotene spermatocytes; mPSc = mid pachytene spermatocytes; 7Sd = step 7 spermatid. bP<0.05, compared with controls. ANOVA followed by multiple comparison two-tailed ttest.

Spermatogenesis pattern at stage VII Group

ASg

pLSc

mPSc

7Sd

7Sd Effective mPSc: 7Sd degeneration 7Sd (%) degeneration

Control

0.64¡À0.04 19.72¡À0.82 19.07¡À0.69 65.09¡À1.17

1:3.41

14.75

-

4 mg/kg 5 mg/kg

0.62¡À0.06 19.89¡À0.80 19.89¡À0.88 62.92¡À1.07 0.52¡À0.04b 16.37¡À0.68b 14.62¡À0.74b 40.36¡À1.68b

1:3.16 1:2.76

21.00 31.00

+6.25 +16.25

6 mg/kg

0.47¡À0.03b 15.98¡À0.73b 13.63¡À0.39b 35.09¡À1.92b

1:2.57

35.75

+21

3.3 Sperm count The sperm count was significantly reduced in both the 5 and 6 mg/kg, but not in the 4 mg/kg treated groups compared with the controls (Table 3). Table 3. Effect of sodium arsenite on the sperm count and testicular arsenic content in rats (n=8). bP<0.05, compared with controls.

Group

Sperm count (106/cauda epididymis)

Elementary arsenic (mg/g)

Control 4 mg/kg 5 mg/kg 6 mg/kg

135.75¡À8.39 122.87¡À7.28 84.81¡À6.33b 65.09¡À7.32b

0.98¡À0.27 2.68¡À0.42 4.39¡À0.31b 6.58¡À0.29b

3.4 Plasma hormonal levels

At the dose of 5 and 6 but not 4 mg/kg, the plasma levels of FSH and LH were significantly decreased compared with the controls. The changes were more prominent in the 6 mg/kg group (Figures 1 and 2). Figure 1. Effect of sodium arsenite on plasma level of FSH. bP<0.05, compared with controls (n=8). Figure 2. Effect of sodium arsenite on plasma level of LH. bP<0.05, compared with controls (n=8).

Plasma testosterone was significantly decreased (P<0.01) in rats of the 5 and 6 mg/kg group and was more prominent in the latter compared with the controls. It was not significantly changed in the 4 mg/kg group (Figure 3). Figure 3. Effect of sodium arsenite on plasma level of testosterone. bP<0.05, compared with controls (n=8).¡¡

3.5 Testicular arsenic concentration The arsenic concentration was significantly increased in the testes in all the treated animals in a dose-dependent manner (Table 3).

4 Discussion A significant decrease in the plasma FSH, LH and testosterone levels and degenerative changes in testicular histology in arsenite treated rats are in agreement with previous findings of the inhibitory effect of arsenite on the gonadal structure and function in mice [13] and fishes [8-10] and the testicular ∆5-3β-HSD and 17β-HSD activities in rats [11]. As these enzymes are gonadotrophin dependent [22], a decrease in their levels may reflect reduced pituitary gonadotrophin secretion. It was indicated that in parallel with the decrease in 7Sd and increased 7Sd degeneration, the sperm count was markedly reduced. Earlier reports have revealed degenerative changes in the testicular histology in fish and mice treated with arsenite [8, 13]. LH and FSH are required for the initiation and maintenance of spermatogenesis in prepubertal and pubertal rats [23] and for quantitatively normal spermatogenesis in pubertal rats [17]. The reduction of FSH and LH and a consequent reduction in testosterone production may, therefore, be held responsible for this arsenite-induced changes in spermatogenesis. The reduction in the number of ASg in arsenite treated rats is possibly due to the increased rate of degeneration of Asg as FSH inhibits the normal degeneration of ASg and reduced FSH secretion may promote ASg degeneration. We found that higher doses of sodium arsenite increase adrenocortical activity and elevated serum corticosterone level [4], which in turn may reduce the serum gonadotriphin and testosterone levels [24]. Inhibitory effects of glucocorticoids on LH secretion have been reported in cultured pituitary [25]. Glucocorticoids also directly suppress testosterone production and secretion by decreasing the testicular LH receptors [26], resulting in the reduction of spermatogenesis and sperm count. It appears that the primary site of arsenic action may be on the brain or pituitary, however, a direct action on the germ cells cannot be ruled out and further studies are required to clarify these points.

References

[1] Nickson R, McArthur J, Burges W, Ahmed KM, Ravenserof P, Rahman M. Arsenic poisoning of Bangladesh ground water. Nature 1998; 395: 338. [2] Borzsonyl A, Bereczky A, Rudnai P, Csanady M, Horvath A. Epidemiological studies on human subjects exposed to arsenic in drinking Water in Southern Hungary. Arch Toxicol 1992; 66: 77-8. [3] Chatterjee A, Das D, Chatterjee D. The study of ground water contamination by arsenic in the residental area of Behala, Calcutta, due to industrial pollution. Environ Pollution 1993; 80: 57-65. [4] Biswas NM, Roy Chowdhury G, Sarkar M. Effect of sodium arsenite on adrenocortical activities in male rats: dose-duration dependent responses. Med Sci Res 1994; 23: 153-4. [5] Sarkar M, Ghosh D, Biswas HM, Biswas NM. Effect of sodium arsenite on haematology in male albino rats. Ind J Physiol Allied Sc 1992; 46: 116-20. [6] Robert EM, Judd ON. Water and soil pollutants. In : Klassen CD, Ambur MD, J Doull, editors, Toxicology - The basic science of poison. 3rd edition. New York: Macmillan Publishing Company; 1986. p825. [7] Brown MM, Rhyne BC, Boyer RA, Fowler BA. Intracellular effects of chronic arsenite administration on renal proximal tubule cells. J Toxicol Environ Health 1976; 1: 505-14. [8] Shukla JP, Pandey K. Impaired spermatogenesis in arsenic treated fresh water fish. Colisa fasciatus (Bl & Sch). Toxicol Lett 1984; 21: 191-5. [9] Shukla JP, Pandey K. Arsenic induced cellular and biochemical changes during the testicular cycle of a fresh water perch. Colisa fasciatus (Bl & Sch). Cell Mol Biol 1984; 30: 227-31. [10] Shukla J P, Pandey K. Toxicity and long term effect of arsenic on the gonadal protein metabolism in a tropical fresh water fish. Colisa fasciatus (Bl & Sch). Acta Hydrochem Hydrobiol 1985; 13: 127-31. [11] Sarkar M, Biswas NM, Ghosh D. Effect of sodium arsenite on testicular ∆5- 3β and 17βhydroxysteroid dehydrogenase activities in albino rats: dose and duration dependent response. Med Sci Res 1991; 19: 789-90. [12] Golub MS, Macintoch MS, Baumrind N. Development and reproductive toxicity of inorganic arsenic: animal studies and human concerns. J Toxicol Environ Health B Crit Rev 1998; 1: 199241. [13] Pant N, Kumar R, Murthy RC, Srivastava SP. Male reproductive effect of arsenic in mice. Biometals 2001; 14 : 113-7. [14] Leblond PC, Clermont Y. Definition of the stages of the seminiferous epithelium in the rat. Ann N Y Acad Sci 1952; 55: 548-73. [15] Abercrombie M. Estimation of nuclear population from microtome sections. Anat Rec 1946; 94: 239-47. [16] Clermont Y, Morgentalor H. Quantitative study of spermatogenesis in hypophysectomized rats. Endocrinology 1955; 57: 369-82. [17] Russell LD, Alger LF, Naquin LG. Hormonal control of pubertal spermatogenesis. Endocrinology 1987; 120: 1615-32. [18] Clermont Y, Harvey SC. Effect of hormones on spermatogenesis in the rat. Ciba Foundation Coll. Endocrinology 1967; 16: 173-96. [19] Greenwood FC, Hunter WM, Glover J S. The preparation of 131I-labeled human growth hormone of high specific activity. Biochem J 1963; 89: 114-23. [20] Auletta FJ, Caldwell BV, Hamilton G. Androgen: testosterone and dihydrotestosterone. In: Jaffe BM, Behrman HR, editors. Methods of Hormone Radioimmunoassay. New York: Academic Press; 1974. p359-70. [21] N¨¹rnberg HW. Processing Biological Samples for Metal Analysis. In: Brown SS, Savory J, editors. International Union of Pure and Applied Chemistry; New York: Academic Press; 1983. p31-44. [22] Murono EP, Payne AH. Testicular maturation in the rat. In vivo effect of gonadotrophins on steroidogenic enzymes in hypophysectomized immature rats. Biol Reprod 1979; 20: 911-6. [23] Chowdhury AK. Dependence of testicular germ cells on hormones: A quantitative study in hypophysectomized testosterone - treated rats. J Endocrinology 1979; 83: 331-40. [24] Philips DM, Lakshmi V, Monder C. Corticosteroid 11b-dehydrogenase in rat testis. Encocrinology 1989; 125: 209-16. [25] D'Agostino JB, Valadka RJ, Schwartz NB. Differntial effect of in vitro glucocorticoids on LH and FSH secretion: Dependence of sex of pituitary donor. Endocrinology 1990; 127: 891.

[26] Bombino TH, Hsueh AJW. Direct effect of glucocorticoids upon testicular luteinizing hormone receptor and steroidogenesis in vivo and in vitro. Endocrinology 1989; 125 : 209-16.

Niraj Pant1 , Rakesh Kumar1 Satya P. Srivastava1

, Ramesh C. Murthy1

and

(1) Industrial Toxicology Research Centre, Post Box No 80, M.G. Marg, Lucknow-, 226001, India

Abstract Arsenic, a known human carcinogen, was given to mice via drinking water as sodium arsenite at a dose 53.39, 133.47, 266.95 and 533.90 activity of 17

mol l for 35 days. A decrease in the

HSD along with increase in LDH,

GT activity

were observed at 533.90 mol l. The observed sperm count, motility and morphological abnormalities in sperm were similar to control at lower dose levels. However at 533.90 mol l a significant decrease in sperm count and motility along with increase in abnormal sperm were noticed. Significant accumulation of arsenic in testes and accessory sex organs may be attributed to the arsenic binding to the tissues or greater cellular uptake. No effects were observed on indices studied for reproductive effects at 53.39 mol l arsenic close to which human being are exposed through drinking water under the present set of experimental conditions. arsenic - drinking water - sperm - testes