The aim of this study was designed to evaluate the possible protective effects of thymoquinone (TQ) on the spermatogenesis after testicular injury caused by chronic toluene exposure in rats. The rats were randomly allotted into one of three experimental groups: control, toluene-treated and toluene treated with TQ; each group contained 10 animals. Control group received 1 mL serum physiologic and toluene treatment was performed by inhalation of 3000 ppm toluene, in an 8-hour/day and 6-day/week order for 12 weeks. The rats in TQ-treated group was given TQ (50 mg/kg body weight) once a day orally for 12 weeks starting just after toluene exposure. Tissue samples were obtained for histopathological investigation. To date, no histopathological changes of testis in rats after chronic toluene exposure by TQ treatment have been reported. Spermatogenesis and mean seminiferous tubule diameter (MSTD) were significantly decreased in toluene treated groups when compared to the control group. Furthermore, the TQ-treated animals showed an improved histological appearance in toluene-treated group. Our data indicate a significant reduction in the activity of in situ identification of apoptosis using terminal dUTP nick end-labeling (TUNEL), endothelial nitric oxide synthase (eNOS), inducible nitric oxide synthase (iNOS) and there was a rise in the expression of proliferating cell nuclear antigen (PCNA) in testis tissues of the toluene-treated group with TQ therapy. Electron microscopy of the testes of the rats demonstrated that pretreatment with TQ was particularly effective in preventing the mitochondrial degeneration, dilatation of smooth endoplasmic reticulum (SER) and enlarged intercellular spaces in both Sertoli and spermatid cells in the toluene-treated animals. We believe that further preclinical research into the utility of TQ may indicate its usefulness as a potential treatment on the spermatogenesis after testicular injury caused by chronic toluene exposure in rats.
Aboutabl EA, El-Azzouny AA, Hammerschmidt FJ, et al. (1986) Aroma volatiles of Nigella sativa L. seeds . In: Brunke EJ (eds) Progress in Essential Oil Research. Proceedings of the International Symposium on Essential Oils, Berlin de Gruyter, 44-55.
2.
Al-Gharably M., Badary OA, Nagi MN, et al. (1997) Protective effect of thymoquinone against carbon tetrachloride-induced hepatotoxicity in mice. Research Communications in Pharmacology and Toxicology2: 41-50.
3.
Al-Shabanah OA, Badary OA, Naagi MN, al-Gharably NM, al-Rikabi AC, and al-Bekairi AM ( 1998) Thymoquinone protects against doxorubicin-induced cardio-toxicity without compromising its antitumor activity. Journal of Experimental and Clinical Cancer Research17: 193-198.
4.
Badary OA ( 1999) Thymoquinone attenuates ifosfamideinduced Fanconi syndrome in rats and enhances its antitumor activity in mice. Journal of Ethnopharmacology67: 135-142.
5.
Badary OA, Nagi MN, al-Shabanah OA, al-Sawaf HA, al-Sohaibani MO, and al-Bekairi AM ( 1997) Thymoquinone ameliorates the nephrotoxicity induced by cisplatin in rodents and potentiates its antitumor activity. Canadian Journal of Physiology and Pharmacology75: 1356-1361.
6.
Bartoov B., Ben Barak J., Mayevsky A., Sneider M., Yogev L., and Lightman A. ( 1991) Sperm motility index: a new parameter for human sperm evaluation. Fertility Sterility56: 108-112.
7.
Celis JE, Madsen P., Celis A., Nielsen HV, and Gesser B. ( 1987) Cyclin (PCNA, auxiliary protein of DNA polymerase delta) is a central component of the pathway(s) leading to DNA replication and cell division. FEBS Letters220: 1-7.
8.
Chakravarty N. ( 1993) Inhibition of histamine release from mast cells by Nigellone . Annals of Allergy70: 237-42.
9.
Clifton DR, Gross RA, Sahni SK, et al. (1998) NF-kappaB-dependent inhibition of apoptosis is essential for host cell survival during Richettsia rickettsii infection . Proceedings of National Academy of Sciences USA95: 4646-4651.
10.
De Celis R., Feria-Velasco A., Gonzalez-Unzaga M., Torres-Calleja J., and Pedrón-Nuevo N. (2000) Semen quality of workers occupationally exposed to hydrocarbons. Fertility Sterility73: 221-228.
11.
Donald JM, Hooper K., and Hopenhayn-Rich C. (1991) Reproductive and developmental toxicity of toluene: a review. Environmental Health Perspective94: 237-244.
12.
El-Alfy TS, El-Fatatry HM, Tooma MA, et al. (1975) Isolation and structure assignment of an antimicrobial principle from the volatile oil of Nigella sativa L.seeds. Pharmazie30: 109-111.
13.
El-Kadi A., Kandil O., and Tabuni AM ( 1987) Nigella cell mediated immunity. Archives of AIDS Research1: 232-233.
14.
El-Tahir KE, Ashour MM, and Al-Harbi MM ( 1993) The cardiovascular actions of the volatile oil of the black seed (Nigella sativa) in rats: elucidation of the mechanism(s) of action . General Pharmacology24: 1123-1131.
15.
Fararh KM, Shimizu Y., Shiina T., Nikami H., Ghanem MM, and Takewaki T. ( 2005) Thymoquinone reduces hepatic glucose production in diabetic hamsters. Research in Veterinary Science79: 219-223.
16.
Gotohda T., Tokunaga I., Kubo S., Kitamura O., and Ishigami A. ( 2002) Toluene inhalation induces glial cell line-derived neurotrophic factor, transforming growth factor and tumor necrosis factor in rat cerebellum . Legal Medicine4: 21-28.
17.
Hansson T., Pettersson BM, Eneroth P., and Gustafsson JA ( 1985) Neonatal exposure to toluene: effects on the development of liver microsomal cytochrome P-450 and serum hormone levels in the rat. Toxicology37: 39-50.
18.
Houghton PJ, Zarka R., Heras B., and Hoult JR ( 1995) Fixed oil of Nigella sativa and derived thymoquinone inhibit eicosanoid generation in leucocytes and membrane lipid peroxidation . Planta Medica61: 33-36.
19.
Hsu SM, Raine L., and Fanger H. ( 1981) Use of avidin- biotin-peroxidase complex (ABC) in immunperoxidase techniques: a comparison between ABC and unlabeled antibody (PAP) procedures . Journal of Histochemistry and Cytochemistry29: 577-580.
20.
Hu JH, Jiang J., Ma YH, Yang N., Zhang MH, Wu M., et al. (2003) Enhancement of germ cell apoptosis induced by ethanol in transgenic mice overexpressing Fas Ligand. Cell Research13: 361-367.
21.
Ishigami A., Tokunaga I., Kubo S., and Gotohda T. ( 2005) Immunohistochemical study of rat spermatogenesis after toluene-inhalation. Legal Medicine7: 42-46.
22.
Jaskulski D., deRiel JK, Mercer WE, Calabretta B., and Baserga R. ( 1988) Inhibition of cellular proliferation by antisense oligodeoxynucleotides to PCNA cyclin. Science240: 1544-1546.
23.
Kang MJ, Kim MK, Terhune A., Park JK, Kim YH, and Koh GY ( 1997) Cytoplasmic localization of cyclin D3 in seminiferous tubules during testicular development. Experimental Cell Research234: 27-36.
24.
Lemasters GK, Olsen DM, Yiin JH, et al (1999) Male reproductive effects of solvent and fuel exposure during aircraft maintenance. Reproductive Toxicology13: 155-166.
25.
Li Y., Jiang QG, Yao SQ, Liu W., Tian GJ, and Cui JW ( 1993) Effects of exposure to trinitrotoluene on male reproduction . Biomedical and Environmental Sciences6: 154-160.
26.
Luderer U., Morgan MS, Brodkin CA, Kalman DA, and Faustman EM ( 1999) Reproductive endocrine effects of acute exposure to toluene in men and women. Occupational and Environmental Medicine56: 657-666.
27.
Lue Y., Sinha Hikim AP, Wang C., Leung A., and Swerdloff RS ( 2003) Functional role of inducible nitric oxide synthase in the induction of male germ cell apoptosis, regulation of sperm number, and determination of testes size: evidence from null mutant mice. Endocrinology144: 3092-3100.
28.
Mahfouz M., El-Dakhakhny M. ( 1960) The isolation of a crystalline active principle from Nigella sativa L. seeds. Journal of Pharmaceutical Science UAR1: 1-19.
29.
Nagi MN, Alam K., Badary OA, al-Shabanah OA, al-Sawaf HA, and al-Bekairi AM ( 1999) Thymoquinone protects against carbon tetrachloride hepatotoxicity in mice via an antioxidant mechanism. Biochemistry and Molecular Biology International47: 153-159.
30.
NTP (1990) National Toxicology Program technical report series toxicology and carcinogenesis studies of toluene (CAS No. 108-88-3) in F344: N rats and B6C3F1 mice (inhalation studies). Research Triangle Park, NC: U.S. Enviromental Protection Agency. Department of Health and Human Services . No. 371. PB90-256371. Springfield, VA, 253.
31.
Ono A., Sekita K., Ogawa Y., et al. (1996) Reproductive and developmental toxicity studies of toluene. II. Effects of inhalation exposure on fertility in rats . Journal of Environmental Pathology Toxicology and Oncology15: 9-20.
32.
Pagliaro P. ( 2003) Differential biological effects of products of nitric oxide (NO) synthase: it is not enough to say NO. Life Science73: 2137-2149.
33.
Rendon A., Rojas A., Fernandez SI, and Pineda I. ( 1994) Increases in chromosome aberrations and in abnormal sperm morphology in rubber factory workers. Mutation Research323: 151-157.
34.
Riedl SJ, Shi Y. ( 2004) Molecular mechanisms of caspase regulation during apoptosis . Nature Reviews Molecular Cell Biology5: 897-907.
35.
Said TM, Paasch U., Glander HJ, and Agarwal A. ( 2004) Role of caspases in male infertility. Human Reproduction10: 39-51.
36.
Shiraishi K., Yoshida K., and Naito K. ( 2003) Activation of endothelial nitric oxide synthase in contralateral testis during unilateral testicular torsion in rats. Archives of Andrology49: 179-190.
37.
Sinha Hikim AP, Swerdloff RS ( 1999) Hormonal and genetic control of germ cell apoptosis in the testis. Reviews of Reproduction4: 38-47.
38.
Suzuki T., Kashimura S., and Umetsu K. ( 1983) Thinner abuse and aspermia. Medical Science and Law23: 199-202.
39.
Svensson BG, Nise G., Erfurth EM, et al. (1992a) Hormonestatus in occupational toluene exposure . American Journal of Industrial Medicine22: 99-107.
40.
Svensson BG, Nise G., Erfurth EM, et al. (1992b) Neuroendocrine effects in printing workers exposed to toluene. British Journal of Industrial Medicine49: 402-408.
41.
Taneli F., Vatansever S., Ulman C., et al. (2005) The effect of spermatic vessel ligation on testicular nitric oxide levels and germ cell-specific apoptosis in rat testis. Acta Histochemica106: 459-466.
42.
Taskinen H., Anttila A., Lindbohm ML, Sallmén M., and Hemminki K. ( 1989) Spontaneous abortions and congenital malformations among the wives of men occupationally exposed to organic solvents. Scandinavian Journal of Work Environment and Health15: 345-352.
43.
Yamada K. ( 1993) Influence of lacquer thinner and some organic solvents on reproductive and accessory reproductive organs in the male rat. Biological and Pharmaceutical Bulletin16: 425-427.
44.
Yilmaz B., Canpolat S., Sandal S., et al. (2006) Paint thinner exposure inhibits testosterone synthesis and secretion in a reversible manner in the rat. Reproductive Toxicology22: 791-796.
45.
Yilmaz B., Kutlu S., Canpolat S., et al. (2001) Effects of paint thinner exposure on serum LH, FSH and testosterone levels and hypothalamic catecholamine contents in the male rat. Biological and Pharmaceutical Bulletin24: 163-166.
46.
Zini A., Abitbol J., Girardi SK, Schulsinger D., Goldstein M., and Schlegel PN ( 1998) Germ cell apoptosis and endothelial nitric oxide synthase (eNOS) expresion following ischemia- reperfusion injury to testis . Archives of Andrology41: 57-55.
