Skip to main content
SpringerLink
Log in

α-lipoic acid protects testis and epididymis against linuron-induced oxidative toxicity in adult rats

  • Original Article
  • Published:
Toxicological Research Aims and scope Submit manuscript

Abstract

Linuron is well known for its antiandrogenic property. However, the effects of linuron on testicular and epididymal pro- and antioxidant status are not well defined. On the other hand, α-lipoic acid is well known as universal antioxidant. Therefore, the purpose of this study was twofold: firstly to investigate whether linuron exposure alters antioxidant status in the testis and epididymis of rats and if so, whether the supplementation of α-lipoic acid mitigates linuron-induced oxidative toxicity in rats. To address this question, α-lipoic acid at a dose of 70 mg/Kg body weight (three times a week) was administered to linuron exposed rats (10 or 50 mg/Kg body weight, every alternate day over a period of 60 days), and the selected reproductive endpoints were analyzed after 60 days. Respective controls were maintained in parallel. Linuron at selected doses reduced testicular daily sperm count, and epididymal sperm count, sperm motility, sperm viability, and number of tail coiled sperm, reduced activity levels of 3β- and 17β-hydroxysteroid dehydrogenases, decreased expression levels of StAR mRNA, inhibition of testosterone levels, and elevated levels of testicular cholesterol in rats over controls. Linuron intoxication deteriorated the structural integrity of testis and epididymis associated with reduced the reproductive performance over controls. Conversely, α-lipoic acid supplementation enhanced sperm quality and improved the testosterone synthesis pathway in linuron exposed rats over its respective control. Administration of α-lipoic acid restored inhibition of testicular and epididymal enzymatic (superoxide dismutase, catalase, glutathione reductase, glutathione peroxidise) and non-enzymatic (glutathione content), increased lipid peroxidation and protein carbonyl content produced by linuron in rats. α-lipoic acid supplementation inhibited the expression levels of testicular caspase-3 mRNA levels and also its activity in linuron treated rats. To summate, α-lipoic acid-induced protection of reproductive health in linuron treated rats could be attributed to its antioxidant, and steroidogenic properties.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9

Similar content being viewed by others

References

  1. EFSA (European Food Safety Authority) (2016) Conclusion on the peer review of the pesticide risk assessment of the active substance linuron. EFSA J 14:4518

    Google Scholar 

  2. Gupta PK (2018) Herbicides and fungicides. In: Veterinary toxicology, basic and clinical principle, 3rd edn. pp 553–567

    Google Scholar 

  3. Gatidou G, Thomaidis NS, Stasinakis AS, Lekkas TD (2007) Simultaneous determination of the endocrine disrupting compounds nonylphenol, nonylphenol ethoxylates, triclosan and bisphenol A in wastewater and sewage sludge by gas chromatography–mass spectrometry. J Chromatogr A 1138:32–41

    Article  CAS  PubMed  Google Scholar 

  4. Pest management regulatory agency (2012) Proposed re-evaluation decision PRVD2012-02 linuron. Tecnical report prepared for pest management regulatory agency. Health Canada: Ottawa Ontario Canada

  5. Kojima H, Katsura E, Takeuchi S, Niiyama K, Kobayashi K (2004) Screening for estrogen and androgen receptor activities in 200 pesticides by in vitro reporter gene assays using Chinese hamster ovary cells. Environ Health Perspect 112:524–531

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Orton F, Lutz I, Kloas W, Routledge EJ (2009) Endocrine disrupting effects of herbicides and pentachlorophenol: in vitro and in vivo evidence. Environ Sci Technol 43:2144–2150

    Article  CAS  PubMed  Google Scholar 

  7. Freyberger A, Witters H, Weimer M, Ahr HJ (2010) Screening for (anti) androgenic properties using a standard operation protocol based on the human stably transfected androgen sensitive PALM cell line first steps towards validation. Reprod Toxicol 30:9–17

    Article  CAS  PubMed  Google Scholar 

  8. Wilson VS, Lambright CR, Furr JR, Howdeshell KL, Earl Gray L (2009) The herbicide linuron reduces testosterone production from the fetal rat testis during both in utero and in vitro exposures. Toxicol Lett 186:73–77

    Article  CAS  PubMed  Google Scholar 

  9. Uren TM, Perry MH, Santos EM (2015) The herbicide linuron inhibits cholesterol biosynthesis and induces cellular stress responses in brown trout. Environ Sci Technol 49:3110–3118

    Article  CAS  Google Scholar 

  10. Lambright C, Ostby J, Bobseine K, Wilson V, Hotchkiss AK, Mann PC, Gray LE (2000) Cellular and molecular mechanisms of action of linuron: an antiandrogenic herbicide that produces reproductive malformations in male rats. Toxicol Sci 56:389–399

    Article  CAS  PubMed  Google Scholar 

  11. McIntyre BS, Barlow NJ, Wallace DG, Maness SC, Gaido KW, Foster PM (2000) Effects of in utero exposure to linuron on androgen-dependent reproductive development in the male Crl:CD(SD)BR rat. Toxicol Appl Pharmacol 167:87–99

    Article  CAS  PubMed  Google Scholar 

  12. Waller CL, Uma BW, Gray LE Jr, Kelce WR (1996) Three dimensional quantitative structure-activity relationships for androgen receptor ligands. Toxicol Appl Pharmacol 137:219–227

    Article  CAS  PubMed  Google Scholar 

  13. Bauer ER, Meyer HH, Stahischmidt-Allner P, Sauerwein H (1998) Application of an androgen receptor assay for the characterisation of the androgenic or antiandrogenic activity of various phenylurea herbicides and their derivatives. Analyst 123:2485–2487

    Article  CAS  PubMed  Google Scholar 

  14. Bai J, Han H, Wang F, Ding H, Hu X, Hu B, Li H, Zheng W, Li Y (2017) Maternal linuron exposure alters testicular development in male offspring rats at the whole genome level. Toxicology 389:13–20

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Ding HW, Zheng W, Han H, Hu X, Hu B, Wang F, Su L, Li H, Li Y (2017) Reproductive toxicity of linuron following gestational exposure in rats and underlying mechanisms. Toxicol Lett 266:49–55

    Article  CAS  PubMed  Google Scholar 

  16. Hotchkiss AK, Parks-Saldutti LG, Ostby JS, Lambright C, Furr J, Vandenbergh JG, Gray LE (2004) A mixture of the antiandrogens linuron and butyl benzyl phthalate alters sexual differentiation of the male rat in a cumulative fashion. Biol Reprod 71:1852–1861

    Article  CAS  PubMed  Google Scholar 

  17. Santos SM, Videira RA, Fernandes MA, Santos MS, Moreno AJ, Vicente JA, Jurado AS (2014) Toxicity of the herbicide linuron as assessed by bacterial and mitochondrial model systems. Toxicol In Vitro 28:932–939

    Article  CAS  PubMed  Google Scholar 

  18. Nam RK, Hang W, Siminovitch K, Shlien A, Kattan MW, Klotz LH, Trachtenberg J, Lu Y, Zhang J, Yu C, Toi A, Loblaw DA, Venkateswaran V, Stanimirovic A, Sugar L, Malkin D, Seth A, Narod SA (2011) New variants at 10q26 and 15q21 are associated with aggressive prostate cancer in a genome-wide association study from a prostate biopsy screening cohort. Cancer Biol Ther 12:997–1004

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Bilska A, Wodek L (2005) Lipoic acid – the drug of the future? Pharmacol Rep 57:570–577

    CAS  PubMed  Google Scholar 

  20. El-Bishbishy HA, Aly HA, El-Shafey M (2014) Lipoic acid mitigates bisphenol A induced testicular mitochondrial toxicity in rats. Environ Occup Health 29:875–887

    Google Scholar 

  21. Gules O, Eren U (2016) Protective role of alpha lipoic acid against polychlorobiphenyl (Aroclor 1254)-induced testicular toxicity in rats. Rom J Morph Embryol 57:451–459

    Google Scholar 

  22. Prathima P, Venkaiah K, Pavani R, Daveedu T, Munikumar M, Gobinath M, Valli M, Sainath SB (2017) α-lipoic acid inhibits oxidative stress in testis and attenuates testicular toxicity in rats exposed to carbimazole during embryonic period. Toxicol Rep 4:373–381

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Prathima P, Pavani R, Sukeerthi S, Sainath SB (2018) α-Lipoic acid inhibits testicular and epididymal oxidative damage and improves fertility efficacy in arsenic-intoxicated rats. J Biochem Mol Toxicol. https://doi.org/10.1002/jbt.22016

    Article  PubMed  Google Scholar 

  24. Koga T, Ishida T, Takeda T, Ishii Y, Uchi H, Tsukimori K, Yamamot M, Himeno M, Furue M, Yamada H (2012) Restoration of dioxin-induced damage to fetal steroidogenesis and gonadotropin formation by maternal co-treatment with α-lipoic acid. PLoS ONE 7:e40322

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. CPCSEA (2003) Committee for the Purpose of control and supervision on experiments on animals PCSEA guidelines for laboratory animal facility. Indian J Pharmacol 35:257–274

    Google Scholar 

  26. Dixit S, Dhar P, Mehra RD (2015) Alpha lipoic acid (ALA) modulates expression of apoptosis associated proteins in hippocampus of rats exposed during postnatal period to sodium arsenite (NaAsO2). Toxicol Rep 2:78–87

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. O’Connor TG, Heron J, Golding J, Beveridge M, Glover V (2002) Maternal antenatal anxiety and children’s behavioural emotional problems at 4 years. Br J Psychiatry 180:502–508

    Article  PubMed  Google Scholar 

  28. Hayes W, Kruger CL (2014) Hayes’ principles and methods of toxicology, 6th edn. CRC Press, Taylor and Francis Group, Boca Raton, Florida

    Book  Google Scholar 

  29. Belsey MA, Moghissi KS, Eliasson R, Paulsen CA, Gallegos AJ, Prasad MR (1980) Laboratory manual for the examination of human semen and semen cervical mucus interaction WHO 1211 Geneva-27 Switzerland

  30. Talbot P, Chacon RS (1981) A triple-stain technique for evaluating normal acrosome reactions of human sperm. J Exp Zool 215:201–208

    Article  CAS  PubMed  Google Scholar 

  31. Jeyendran RS, Van der Ven HH, Zaneveld LJD (1992) The hypoosmotic swelling test: an update. Arch Androl 29:105–116

    Article  CAS  PubMed  Google Scholar 

  32. Ramu S, Jeyendran RS (2013) The hypo-osmotic swelling test for evaluation of sperm membrane integrity. Methods Mol Biol 927:21–25

    Article  CAS  PubMed  Google Scholar 

  33. Robb GW, Amann RP, Killian GJ (1978) Daily sperm production and epididymal sperm reserves of pubertal and adult rats. J Reprod Fertil 54:103–107

    Article  CAS  PubMed  Google Scholar 

  34. Linder RE, Strader LF, Slott VL, Suarez JD (1992) Endpoints of spermatotoxicity in the rat after short duration exposures to fourteen reproductive toxicants. Reprod Toxicol 6:491–505

    Article  CAS  PubMed  Google Scholar 

  35. Miranda-Spooner M, Paccola CC, Neves FM, de Oliva SU, Miraglia SM (2016) Late reproductive analysis in rat male offspring exposed to nicotine during pregnancy and lactation. Andrology 4:218–231

    Article  CAS  PubMed  Google Scholar 

  36. Zlatkis A, Zak B, Boyle AJ (1953) A new method for the direct determination of serum cholesterol. J Lab Clin Med 41:486–492

    CAS  PubMed  Google Scholar 

  37. Bergmeyer HU (1974) Beta-hydroxysteroid dehydrogenase. In: Bergmeyer HU (ed.) Methods Enzy Analy Academic Press, New York, pp 447–489

  38. Misra HP, Fridovich I (1972) The role of superoxide anion in the autoxidation of epinephrine and a simple assay for superoxide dismutase. J Biol Chem 247:3170–3175

    CAS  PubMed  Google Scholar 

  39. Chance B, Maehly AC (1955) Assay of catalase and peroxidises. Methods Enzymol 2:764–775

    Article  Google Scholar 

  40. Flohe L, Gunzler WA (1984) Assays of glutathione peroxidise. Methods Enzymol 105:114–120

    Article  CAS  PubMed  Google Scholar 

  41. Carlberg I, Mannervik B (1985) Glutathione reductase assay. Methods Enzymol 113:484–495

    Article  CAS  PubMed  Google Scholar 

  42. Beutler E (1975) The preparation of red cells for assay. In: Beutler E (ed) Red cell metabolism: a manual of biochemical methods. Grune and Straton Company, New York, pp 8–18

    Google Scholar 

  43. Ohkawa H, Ohishi N, Yagi K (1979) Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction. Anal Biochem 95:351–358

    Article  CAS  PubMed  Google Scholar 

  44. Levine RL, Garland D, Oliver CN, Amici A, Climent I, Lenz AG, Ahn BW, Shaltiel S, Stadtman ER (1990) Determination of carbonyl content in oxidatively modified proteins. Methods Enzymol 186:464–478

    Article  CAS  PubMed  Google Scholar 

  45. Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the folin phenol reagent. J Biol Chem 193:265–275

    CAS  PubMed  Google Scholar 

  46. Cid C, Alvarez-Cermeno JC, Regidor I, Plaza J, Salinas M, Alcazar A (2003) Caspase inhibitors protect against neuronal apoptosis induced by cerebrospinal fluid from multiple sclerosis patients. J Neuroimmunol 136:119–124

    Article  CAS  PubMed  Google Scholar 

  47. Brancraft JD, Stevens A (1982) Theory and practice of histological techniques, 2nd edn. Chruchill Livingstone, New York

    Google Scholar 

  48. Turner TT, Lysiak JJ (2008) Oxidative stress: a common factor in testicular dysfunction. J Androl 29:488–498

    Article  CAS  PubMed  Google Scholar 

  49. Stocco DM, Clark BJ (2000) Role of the steroidogenic acute regulatory protein (StAR) in steroidogenesis. Biochem Pharmacol 51:197–205

    Article  Google Scholar 

  50. Clair E, Mesnage R, Travert C, Seralini GE (2012) A glyphosate-based herbicide induces necrosis and apoptosis in mature rat testicular cells in vitro and testosterone decrease at lower levels. Toxicol In Vitro 26:269–279

    Article  CAS  PubMed  Google Scholar 

  51. Victor-Costa AB, Bandeira SM, Oliveira AG, Mahecha GA, Oliveira CA (2010) Changes in testicular morphology and steroidogenesis in adult rats exposed to atrazine. Reprod Toxicol 29:323–331

    Article  CAS  PubMed  Google Scholar 

  52. Knez J (2013) Endocrine-disrupting chemicals and male reproductive health. Reprod Biomed Online 26:440–448

    Article  CAS  PubMed  Google Scholar 

  53. Mehrpour O, Karrari P, Zamani N, Tsatsakis AM, Abdollahi M (2014) Occupational exposure to pesticides and consequences on male semen and fertility: a review. Toxicol Lett 230:146–156

    Article  CAS  PubMed  Google Scholar 

  54. Espinoza Navarro O, Bustos Obregon E (2014) Effect of Malathion on cellularity and sperm differentiation in testis and eididymis of adult rats. Int J Morphol 32:119–124

    Article  Google Scholar 

  55. Nakai M, Hess RA, Moore BJ, Guttroff RF, Strader LF, Linder RE (1992) Acute and long-term effects of a single dose of the fungicide carbenduim (metyl-2-benzemidazole carbamate) on the male reproductive system in the rat. J Androl 13:507–518

    CAS  PubMed  Google Scholar 

  56. Hess RA, Gist DH, Bunick D, Lubahn DB, Farrell A, Bahr J, Cooke PS, Greene GL (1997) Estrogen receptor (alpha and beta) expression in the excurrent ducts of the adult male rat reproductive tract. J Androl 18:602–611

    CAS  PubMed  Google Scholar 

  57. Spirhanzlova P, De Groef B, Nicholson FE, Grommen SVH, Marras G, Sébillot A, Demeneix BA, Pallud-Mothré S, Lemkine GF, Tindall AJ, Du Pasquier D (2017) Using short-term bioassays to evaluate the endocrine disrupting capacity of the pesticides linuron and fenoxycarb. Comp Biochem Physiol C Toxicol Pharmacol 200:52–58

    Article  CAS  PubMed  Google Scholar 

  58. U S EPA (1995) Reregistration Eligibility Decision (RED): Linuron. US Environmental Protection Agency, Office of Prevention, Pesticides and Toxic Substances, Office of Pesticide. Programs US Government Printing Office, Washington DC, EPA 738-R-95-003

  59. Pratap Reddy K, Girish BP, Sreenivasula Reddy P (2014) Reproductive and paternal mediated developmental toxicity of Benzo(a)Pyrene in adult male Wistar rats. Toxicol Res (Camb). https://doi.org/10.1039/c4tx00121d

    Article  Google Scholar 

  60. Hamdy BG, Nabil T, Saad NN, Abdelwahab M, Mohamed L (2016) Protective role of alpha lipoic acid against the deleterious effects of both natural and artificial sweetener (Sucrose and Aspartame) in albino rats. Alex J Vet Sci 49:105–115

    Google Scholar 

  61. Lebda M, Gad S, Gaafar H (2014) Effects of lipoic acid on acrylamide induced testicular damage. Mater Soc Med 26:208–212

    Article  Google Scholar 

  62. Holmquist L, Stuchbury G, Berbaum K, Muscat S, Young S, Hager K, Engel J, Munch G (2007) Lipoic acid as a novel treatment for alzheimer’s disease and related dementias. Pharmacol Ther 113:154–164

    Article  CAS  PubMed  Google Scholar 

  63. Biewenga GP, Haenen GR, Bast A (1997) The pharmacology of the antioxidant lipoic acid. Gen Pharmacol 29:315–331

    Article  CAS  PubMed  Google Scholar 

  64. Shila S, Kokilavani V, Subathra M, Panneerselvam C (2005) Brain regional responses in antioxidant system to alpha-lipoic acid in arsenic intoxicated rat. Toxicology 210:25–36

    Article  CAS  PubMed  Google Scholar 

  65. McIlwain DR, Berger T, Mak TW (2013) Caspase functions in cell death and disease. Cold Spring Harb Perspect Biol 5:a008656

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  66. Santos PS, Campelo LM, Freitas CM, Saldanha GB, Freitas RM (2011) Lipoic acid effects on glutamate and taurine concentrations in rat hippocampus after pilocarpine-induced seizures. Arq Neuropsiquiatr 69:360–364

    Article  PubMed  Google Scholar 

  67. Astiz M, de Alaniz MJT, Marra CA (2012) The oxidative damage and inflammation caused by pesticides are reverted by lipoic acid in rat brain. Neurochem Int 61:1231–1241

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank the Head, Dept. of Biotechnology, VSU, Nellore, AP, India for providing laboratory space and allowed us to utilize the equipments purchased under DST-FIST programme, New Delhi. We thank the Head, Department of Genetics, Narayana Medical College, Nellore for providing animals and the Dr. M. Gobinath, Ratnam Pharmacy College, Muthurkur for providing the animal house facilities. Our special thanks to the Head, Department of Marine Biology for allowing us to utilize ELISA Microplate Reader.

Funding

This research work was not supported by any funding agency.

Author information

Authors and Affiliations

  1. Department of Biotechnology, Vikrama Simhapuri University, Nellore, AP, 524 320, India

    P. Prathima, K. Venkaiah, T. Daveedu, R. Pavani, S. Sukeerthi & Sri Bhashaym Sainath

  2. Department of Pharmacy, Ratnam Pharmacy College, Muthukur, Nellore, AP, India

    M. Gopinath

Authors
  1. P. Prathima
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. K. Venkaiah
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. T. Daveedu
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. R. Pavani
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. S. Sukeerthi
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. M. Gopinath
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Sri Bhashaym Sainath
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Sri Bhashaym Sainath.

Ethics declarations

Conflict of interest

Nothing to disclose.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (JPEG 26 kb)

Supplementary material 2 (JPEG 32 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Prathima, P., Venkaiah, K., Daveedu, T. et al. α-lipoic acid protects testis and epididymis against linuron-induced oxidative toxicity in adult rats. Toxicol Res. 36, 343–357 (2020). https://doi.org/10.1007/s43188-019-00036-y

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s43188-019-00036-y

Keywords

Search

Navigation