Güleser GÖKTAŞ, İskender KAPLANOĞLU, Deniz ERDOĞAN, Celal ILGAZ, Gülnur KAPLANOĞLU TAKE, İrem İNANÇ, Arzu YURTCU, Fatma HELVACIOĞLU

Hipertermide duktus epididimis yapısı

Amaç: Hipertermi; vücut ısısının 41°Cye ya da daha çok yükselmesi durumudur. Sıcak stresi duktus epididimis üzerinde çeşitli anomalilere neden olur. Super oksit dismutaz (SOD) ise oksidatif stresi engelleyebilen ve apopitotik süreci geri döndürebileceği düşünülen antioksidan bir maddedir. Bu nedenle çalışmamızda; hipertermi ile oluşturulan ısı stresinin ve stres öncesinde kullanılan SODun, duktus epididimis üzerindeki etkilerinin immünohistokimyasal olarak belirlenmesi amaçlanmıştır. Yöntemler: Çalışmada Wistar-Albino cinsi erkek sıçanlar, her grupta 6 denek olacak şekilde 7 gruba ayrıldı. Hipertermi deneklerin 42°Cde sıcak su banyosuna 20 dakika bırakılmasıyla gerçekleştirildi. Kontrol grubu olarak belirlenen 1. grup, 22°Cde 20 dakika bekletilerek 24 saat sonra duktus epididimis dokuları alındı. 2, 4, 6. gruplara hipertermiden bir saat önce NaCl+Katalaz uygulandı, 2. grup hipertermiden 6 saat, 4. grup 24 saat ve 6. grup 72 saat sonra dokuları alındı. 3, 5, 7. gruplara hipertermiden bir saat önce NaCl+Katalaz+SOD uygulaması yapıldı. 3. grup hipertermiden 6 saat, 5. grubun 24 saat, 7. grubun 72 saat sonra dokuları alındı. Alınan dokulara, hiperterminin neden olduğu apoptozisi belirlemek için Kaspaz 9, Kaspaz 8 ile Kaspaz 3 protein yapılarına etkisinin belirlenebilmesi amacıyla HSP 70 primer antikorlarıyla indirekt immünohistokimyasal yöntem uygulanmıştır. Bulgular: Çalışmamızda morfolojik olarak bozuk spermiyum içeren lümenlerde Kaspaz 3 immunreaktivitesi belirlendi. Bu bulgu hipertermi etkisiyle testiste üretilen anormal spermiyumların, epididimise ulaşabildiklerini gösterdi. Bazı bölgelerde yuvarlak spermatit başlarının epitele yöneldiği ve sterosilyalar arasında sokulduğu saptandı. Bu yönelmenin Sertoli hücrelerince ortadan kaldırılamamış artık cisimleri ve bozulan spermiyumları fagosite eden esas hücrelerce düzenlenen bir mekanizma sonucu gerçekleştiği düşünüldü. Tüm Kaspazlar için SOD uygulamasının tutulumu azalttığı gözlendi. Sonuç: Skrotal ısı yükselmesinin duktus epididimiste olumsuz etkisini gösterdiği ve SOD un süreye koşut artan HSP70 senteziyle apoptozisi baskılayabileceği sonucuna varıldı.

The structure of ductus epididymis in hyperthermia

Objective: Hyperthermia is the condition of the body temperature rising at 41°C or more. Heat stres on ductus epididymis leads to abnormal spermium production and spermatogenesis. Hyperthermia induced oxidative stres and apopitosis can be blocked by superoxide dismutase (SOD). In this study we aimed to determine the protective effects of SOD on ductus epididymis at hyperthermic conditions by using apoptotic and oxidative stress markers Methods: Wistar Albino male rats were divided into seven groups. Every group has 6 rats. Control group (group 1) had set for 20 minutes in a pool which was 22° C temperature. On the 2nd, 4th, and 6th groups NaCl + catalase were applied one hour before hyperthermia, and then the rats were abondoned in 42° C water bath for 20 minutes. After that tissue were taken th th th nd th that 6 , 24 , 72 hours regarding 2 , 4 , and 6 groups respectively. On the 3rd, 5th, and 7th groups NaCl + catalase + SOD was applied one hour before hyperthermia and then the rats were abandoned in 42° C water bath for 20 th th minutes. After hyperthermia for 20 minutes tissues were taken at 6 , 24and 72th hours regarding 3rd, 5th and 7th groups respectively. In order to determine apoptosis caused by hyperthermia, removed tissues were subjected to indirect immunohistochemical method with HSP-70 primary antibodies to detect impact on Caspase 9, Caspase 8 and Caspase 3 protein structures. Results: Morphologically abnormal spermium including lumens show positive Caspase-3 reaction. This was accepted as an indicator of abnormal spermiums, which were caused by effects of hyperthermia, can reach ductus epididymis. It was also remarkable that some areas, round spermatide heads directed into epithelium an even intruded into stereocilias. It was thought that, this direction can be caused by a mechanism which is organized by principle cells, which were not destroyed in Sertoli cells. It was observed that, for all caspases, SOD application is reducing the immunreactivity in parallel to time. Conclusion: The scrotal temperature elevation shows its harmfull effects on epididymis, but the SOD application may suppress apoptosis in parallel to time by increasing HSP70 level.

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1.Sund-Levander M, Forsberg C, Wahren LK. Normal oral, rectal, tympanic and axillary body temperature in adult men and women. Scand J Caring Sci 2002; 6): 122-8.
2. Çelik Ömür S. Yoğun Bakım Hastalarında Vital Bulguların Takibi ve Önemi. Güncel Gastroenteroloji Derg 2004; 8: 146-50.
3. Kierszerbaum AL. Histoloji ve Hücre Biyolojisi Patolojiye Giriş. Demir R (Çeviri Editörü), Ankara: Palme Yayıncılık; 2006.
4. Knobil and Neills Physiology of Reproduction. Jimmy D. Neill, editör, Elsevier Academic Press Publications St. Louis USA; 2006.
5. Berne RM, Levy MN, Koeppen BM, Stanton BA. Fizyoloji. Türk Fizyolojik Bilimler Derneği (Çev), 5. Baskı. Ankara: Güneş Tıp Kitabevleri; 2008.
6. Guyton AC, Hall JE. Tıbbi Fizyoloji. Çavuşoğlu H., Çağlayan Yeğen B. (Çeviri Editörleri), 11. Baskı. İstanbul: Nobel Tıp Kitabevleri; 2007.
7. Bozdoğan Ö. Fizyoloji. 2. Baskı. Ankara: Palme Yayıncılık; 2004.
8. Andonian S, Hermo L. Cell-and region-specific localization of lysosomal and secretory proteins and endocytic receptors in epithelial cells of the cauda epididymidis and vas deferens of the adult rat. Journal of Andrology 1999; 20: 415.
9. Chinoy NJ. Structure and physiology of mammalian vas deferens in relation to fertility regulation. Journal of Biosciences 1985; 7: 215-21.
10. Sheynkin Y, Jung M, Yoo P, Schulsinger D, Komaroff E. Increase in scrotal temperature in laptop computer users. Human Reproduction 2005; 20: 452-5.
11. Dada R, Gupta NP, Kucheria K. Spermatogenic Altertions in Men with High Testiculo Epididymal Temperatures. Indian J Hum Genet 2002; 8: 20-5.
12. Rockett JC, Mapp FL, Garges JB, Luft JC, Mori C, Dix DJ. Effects of Hyperthermia on Spermatogenesis, Apoptosis, Gene Expression and Fertility in Adult Male Mice. Biol Reprod 2001; 65: 229-39.
13. Jannes P, Spiessens C, Auwera IV, DHooghe T, Verhoeven G, Vanderschueren D. Male subfertility induced by acute scrotal heating affects embryo quality in normal female mice. Human Reproduction 1998; 13: 372-5.
14. Vydra N, Malusecka E, Jarzab M, Lisowska K, Glowala-Kosinska M, Benedyk K, Widlak P, Krawczyk Z, Widlak W. Spermatocyte-specific expression of constitutively active heat shock factor 1 induces HSP70 i- resistant apoptosis in male germ cells. Cell Death and Differentiation 2006; 13: 212-22.
15. Fiorenza MT, Mangia F. Hyperthermia Specifically Inhibits Bivalent Chromosome Disjunction in Maturing Mouse Oocytes. Biol Reprod 1992; 46: 658-64.
16. Ranawat P, Bansal MP. Decreased glutathione levels potentiate the apoptotic efficacy of selenium: Possible involvement of p38 and JNK MAPKs-in vitro studies. Mol Cell Biochem 2008; 309: 2132.
17. Yılmaz F. Desfluranın Antioksidan Etkinliğinin Propofol İle Karşılaştırılması. Uzmanlık Tezi. Isparta: Süleyman Demirel Üniversitesi Anesteziyoloji ve Reanimasyon Anabilim Dalı; 2006.
18. Kehrer JP. Free radicals as mediators of tissue injury and disease. Crit Rev Toxicol 1993; 23: 21-48.
19. Southorn PA, Powis G. Free radicals in medicine. I. Chemical nature and biologic reactions. Mayo Clin Proc 1988; 63: 381-9.
20. Ames BN, Shigenaga MK, Hagen MT. Oxidants, antioxidants and the degenerative diseases of aging. Proc Natl Acad Sci 1993; 90: 7915-22.
21. Gutteridge JMC. Lipid peroxidation and antioxidants as biomarkers of tissue damage. Clin Chem 1995; 41: 1819-28.
22. Akkuş İ. Serbest radikaller ve fizyopatolojik etkileri. 1. Baskı. İstanbul: Mimoza Yayınları; 2000.
23. Hileman EA, Achanta G, Huang P. Superoxide dismutase: an emerging target for cancer therapeutics. Expert Opin Ther Targets 2001; 5: 697- 710.
24. Ross MH, Kaye IG, Pawlina W. Histology A Text and Atlas with Correlated Cell and Molecular Biology. 5. Baskı. Philadelphia: Lippincott Williams & Wilkins; 2006.
25. Sheynkin Y, Jung M, Yoo P, Schulsinger D, Komaroff E. Increase in scrotal temperature in laptop computer users. Hum. Reprod. 2005;20:452-5.
26. Henle KJ, Leeper DB.Effects of hyperthermia (45 degrees) on macromolecular synthesis in Chinese hamster ovary cells.Cancer Res. 1979;39:2665-74.
27. Lysıak JJ, Nguyen QAT, Turner TT. Peptide and nonpeptide reactive oxygen scavengers provide partial rescue of the testis after torsion. Journal of Andrology. 2002; 23: 400-9.
28. Legare C, Thabet M, Sullivan R. Expression of heat shock protein 70 in normal and cryptorchid human excurrent duct. Molecular Human Reproduction 2004; 10: 197-202.
29. Setchell BP. The Parkes Lecture. Heat and the testis. J Reprod Fertil 1998;114: 179-94.
30. Jara M, Esponda P, Carballada R. Abdominal temperature induces region-specific p53-independent apoptosis in the cauda epididymidis of the mouse. Biol Reprod 2002; 67: 1189-96.
31. Hjollund NH, Bonde JP, Jensen TK, Olsen J. Diurnal scrotal skin temperature and semen quality. The Danish First Pregnancy Planner Study Team Int J Androl 2000; 23: 309-18.
32. Perez-Crespo M, Pindato B, Gutierrez-Adan A. Scrotal Heat Stress Effects on Sperm Viability, Sperm DNA Integrity, and the Offspring Sex Ratio in Mice. Molecular Reproduction and Dev 2008; 75: 40-7.
33. Banks S, King SA, Irvine DS, Saunders PT. Impact of a mild scrotal heat stress on DNA integrity in murine spermatozoa. Reproduction 2005; 129: 505-14.
34. Salo DC, Donovan CM, Davies KJ. HSP70 and other possible heat shock or oxidative stress proteins are induced in skeletal muscle, heart and liver during exercise. Free Radic Biol Med 1991; 11: 239-46.
35. Zuo L, Christofi FL, Wright VP, Merola AJ, Berliner LJ, Clanton TL. Intra- and extracellular measurement of reactive oxygen species produced during heat stress in diaphragm muscle. Am J Cell Physiol 2000; 279: 105866.
36. Zhang P, Omaye ST. Antioxidant and prooxidant roles for β-carotene, α-tocopherol and ascorbic acid in human lung cells. Toxicol In Vitro 2001; 15: 13-24.
37. Venkatakrishnan CD, Tewari AK, Moldovan L, Cardounel AJ, Zweier JL, Kuppusamy P, Ilangovan G. Heat shock protects cardiac cells from doxorubicin-induced toxicity by activating p38 MAPK and phosphorylation of small heat shock protein 27. Am J Physiol Heart Circ Physiol 2006; 291: 2680-91.
38. Du J, Di HS, Wang GL. Establishment of a bovine epithelial mammary cell line and its ultrastructural changes when exposed to heat stres. Sheng Wu Gong Cheng Xue Bao 2007; 23: 471-6.