Anti-Apoptotic Actions of Omega-3 Fatty Acids on Testes in Experimental Formaldehyde Intoxication: An Immunohistochemical Study

Abstract

Objective: In this study, which we have performed immunohistochemically, it was aimed to investigate effects of formaldehyde toxicity on the testes, especially on apoptotic changes. Potential antioxidant effects of omega-3 fatty acids against these toxic effects were also evaluated. Materials and Methods: A total of 21 adult male Wistar-albino rats used in our study were divided into three equal groups. Rats in group I were used as control. Rats in group II were injected intraperitoneally with formaldehyde every other day while the other ones received omega-3 fatty acids daily via intragastric gavage (Marincap capsule®) with injections of formaldehyde. At the end of two-week experimental period, all animals were killed by decapitation and their testes were removed. For light microscopic examination, tissue specimens were embedded in paraffin blocks following routine histological procedures. Section obtained from paraffin blocks stained with Bax protein immunohistochemically for the determination of apoptosis. The intensity of immunohistochemical Bax stain within cell cytoplasm was scored semi-quantitatively. Results: it was found an intensive immunohistochemical Bax staining in tissue specimens of formaldehyde-exposed rats, while no staining was seen in tissue sections of control testes. On the other hand, immunohistochemical Bax staining was observed minimally in tissue sections of testes of rats administered with formaldehyde plus omega-3 fatty acids. Conclusion: According to results of this study performed immunohistochemically, omega-3 fatty acids prevent apoptosis caused by formaldehyde in the testes. ©2008, Firat University, Medical Faculty

Keywords

• -

Deneysel Formaldehit Zehirlenmesinde Omega-3 Yağ Asitlerinin Testislerdeki Antiapopitotik Etkileri: İmmunohistokimyasal Bir Çalışma

Öz

Amaç: İmmunohistokimyasal düzeyde yapmış olduğumuz bu araştırmada, testiküler doku üzerindeki formaldehit toksisitesinin özellikle apoptozis yönüyle incelenmesi amaçlandı. Ayrıca, bu toksisiteye karşı omega-3 yağ asitlerinin muhtemel koruyucu etkileri de değerlendirildi. Gereç ve Yöntem: Araştırmamızda kullanılan toplam 21 adet Wistar-Albino cinsi erkek sıçanlar üç eşit gruba ayrıldı. Grup I (n=7)'deki sıçanlar kontrol olarak kullanıldı. Grup II (n=7)'deki hayvanlara gün aşırı olarak ve de intraperitoneal yolla formaldehit enjekte edildi. Grup III (n=7)'deki sıçanlara ise formaldehit enjeksiyonu ile birlikte intragastrik gavaj yoluyla ve günlük olarak omega-3 yağ asiti (Marincap kapsül®) verildi. İki haftalık deney süresi sonunda bütün sıçanlar dekapite edilerek testisleri çıkartıldı. Mikroskobik incelemeler için, testis doku örnekleri rutin histolojik prosedürlerden geçirilerek parafine gömüldü. Elde edilen parafin bloklardan alınan kesitler apoptozis varlığının belirlenmesi amacıyla immunohistokimyasal olarak Bax proteini ile boyandı. Hücre sitoplazmalarında gözlenen immunohistokimyasal Bax boyanmasının şiddeti semi- kantitatif olarak derecelendirildi. Bulgular: Kontrol grubu sıçanlarına ait testis doku kesitlerinde herhangi bir Bax boyanması gözlenmezken, formaldehite maruz kalan gruba ait doku preparatlarında şiddetli derecede immunohistokimyasal Bax boyanmasının olduğu tespit edildi. Formaldehit enjeksiyonu ile birlikte omega-3 yağ asitleri verilen sıçanlara ait testis doku kesitlerinde ise immunohistokimyasal Bax boyanmasının minimal derecede olduğu görüldü. Sonuç: İmmunohistokimyasal olarak gerçekleştirilen bu çalışma bulgularına göre, omega-3 yağ asitleri formaldehitin testislerde neden olduğu apoptozisi önlemektedir. ©2008, Fırat Üniversitesi, Tıp Fakültesi

Anahtar Kelimeler

• Omega-3 Yağ Asitleri, Formaldehit, Testis, Apoptozis, İmmunohistokimya

References

1. Smith AE. Formaldehyde. Occup Med 1992; 42: 83-88.
2. Usanmaz SE, Akarsu ES, Vural N. Neurotoxic effects of acute and subacute formaldehyde exposures in mice. Envir Toxicol Pharmacol 2002; 11: 93-100.
3. Monteiro Riviere NA, Popp JA. Ultrastructural evaluation of acute nasal toxicity in the rat respiratory epitelium in response to formaldehyde gas. Fundam Appl Toxicol 1986; 6: 251-262.
4. Morgan KT, Patterson DL, Gross EA. Responses of the nasal mucociliary apparatus of F-344 rats to formaldehyde gas. Toxicol Appl Pharmacol 1986; 82: 1-13.
5. Kilburn KH, Warshaw R, Thornton JC. Formaldehyde impairs memory, equilibrium, and dexterity in histology technicians: effects which persist for days after exposure. Arch Environ Health 1987; 42: 117-120.
6. Zararsiz I, Kus I, Akpolat N, et al. Protective effects of omega-3 essential fatty acids against formaldehyde-induced neuronal damage in prefrontal cortex of rats. Cell Biochem Funct 2006; 24: 237-244.
7. Songur A, Akpolat N, Kus I, et al. The effects of the inhaled formaldehyde during the early postnatal period in the hippocampus of rats: a morphological and immunohistochemical study. Neurosci Res Commun 2003; 33: 168-178.
8. Thrasher JD, Kilburn KH. Embryo toxicity and teratogenicity of formaldehyde. Arch Env Health 2001; 56: 300-311.

9. Chowdhury AR, Gautam AK, Patel KG, Trivedi HS. Steroidogenic inhibition in testicular tissue of formaldehyde exposed rats. Indian J Physiol Pharmacol 1992; 36: 162-168.
10. Sarsılmaz M, Özen OA, Akpolat N, Kuş İ, Songur A. Subakut dönemde solunan formaldehitin sıçanların Leydig hücreleri üzerindeki histopatolojik etkileri. Fırat Üniversitesi Sağlık Bilimleri Dergisi 1999; 13: 37-40.
11. Ozen OA, Akpolat N, Songur A, et al. Effect of formaldehyde inhalation on Hsp70 in seminiferous tubules of rat testes: an immunohistochemical study. Toxicol Ind Health 2005; 21: 249- 254.
12. Kuş MA. Sıçanlarda formaldehit maruziyetiyle testislerde oluşan morfolojik değişiklikler üzerine melatonin hormonunun koruyucu etkisi. Yüksek Lisans Tezi, Afyon: Afyon Kocatepe Üniversitesi Tıp Fakültesi, Anatomi Bölümü, Afyon 2007.
13. Nordoy A. Is there a rational use for n-3 fatty acids (fish oils) in clinical medicine? Drugs 1991; 42: 331-342.
14. Bourre JM, Bonneil M, Chaudiere J, et al. Structural and functional importance of dietary polyunsaturated fatty acids in the nervous system. Adv Exp Med Bio 1992; 318: 211-229.
15. Masters C. Omega-3 fatty acids and the peroxisome. Mol Cell Biochem 1996; 165: 83-93.
16. Stone NJ. Fish consumption, fish oil, lipids and coronary heart disease. Am J Clin Nutr 1997; 65: 1083-1086.
17. Miyasaka CK, Alves de Souza JA, Torres RP, et al. Effects of the administration of fish oil by gavage on activities of antioxidant enzymes of rat lymphoid organs. Gen Pharmacol 1998; 30: 759- 762.
18. Zararsız İ, Sönmez MF, Yılmaz HR, ve ark. Sıçanlarda formaldehit uygulamasıyla akciğerlerde oluşan hasar üzerine omega-3 yağ asitlerinin koruyucu etkisi. Selçuk Tıp Dergisi 2004; 20: 93-98.
19. Zararsiz I, Sonmez MF, Yilmaz HR, et al. Effects of -3 essential fatty acids against formaldehyde-induced nephropathy in rats. Toxicol Ind Health 2006; 22: 223-229.
20. Halperin WE, Goodman M, Stayner L, et al. Nasal cancer in a worker exposed to formaldehyde. JAMA 1983; 249: 510-516.
21. Taskinen H, Kyyronen P, Hemminki K, et al. Laboratory work and pregnancy outcome. J Occup Med 1994; 36: 311-319.
22. Zhou DX, Qui SD, Zhang J, Tian H, Wang HX. The protective effect of vitamin E against oxidative damage caused by formaldehyde in the testes of adult rats. Asian J Androl 2006; 8: 584-588.
23. Tang M, Xie Y, Yi Y, Wang W. Effects of formaldehyde on germ cells of male mice. Wei Sheng Yan Jiu 2003; 32: 544-548.
24. Majumder PK, Kumar VL. Inhibitory effects of formaldehyde on the reproductive system of male rats. Indian J Physiol Pharmacol 1995; 39: 80-82.
25. Öztürk F. Apopitoz. İnönü Üniversitesi Tıp Fakültesi Dergisi 2002; 9: 143-148.
26. Ergin M. Apoptosis. Arşiv 2002; 11: 495-504.
27. Lawen A, Grutter MG. Caspases: key players in programmed cell death. Curr Opin Struct Biol 2000; 10: 649-655.
28. Deveraux QL, Schendel SL, Reed JC. Antiapoptotic proteins. The bcl-2 and inhibitor of apoptosis protein families. Cardiol Clin 2001; 19: 57-74.
29. Friedlander RM, Hengartner MO. The biochemistry of apoptosis. Nature 2000; 1407: 770-776.
30. Gross A, McDonnell JM, Korsmeyer SJ. Bcl-2 family members and the mitochondria in apoptosis. Genes Dev 1999; 13: 1899- 1911.
31. Sarsilmaz M, Songur A, Ozyurt H, et al. Potential role of dietary omega-3 essential fatty acids on some oxidant/antioxidant parameters in rats’ corpus striatum. Prostag Leukotr Ess 2003; 69: 253-259.
32. Lonergan PE, Martin DS, Horrobin DF, Lynch MA. Neuroprotective effect of eicosapentaenoic acid in hippocampus of rats exposed to gamma-irradiation. J Biol Chem 2002; 277: 20804-20811.
33. Martin DS, Lonergan PE, Boland B, Fogarty MP, Brady M. Apoptotic changes in the aged brain are triggered by interleukin1- induced activation of p38 and reversed by treatment with eicosapentaenoic acid. J Biol Chem 2002; 277: 34239-34246.
34. Takezaki T, Inoue M, Kataoka H, et al. Diet and lung cancer risk from a 14-year population-based prospective study in Japan: with special reference to fish consumption. Nutr Cancer 2003; 45: 160-167.
35. Kamal KK, Gomaa A, El Khafif M, Hammad AS. Plasma lipid peroxides among workers exposed to slica or asbestos dust. Environ Res 1989; 49: 173-180.
36. Halliwell B. Free radicals, antioxidants and human disease; curiosity, course or consequence? Lancet 1994; 344: 721-724.
37. Kabul Tarihi:23.07.2008