Protective Effect of Mesenchymal Stem Cells and Melatonin on Testicular Torsion-Induced Infertility

Öz

We aimed to explore the effects of systemic melatonin and intratesticular Adipose tissue-derived mesenchymal stem cells (AdMSCs) administration on rats with acute unilateral testicular torsion. Rats were randomized into Sham group (S) (n=8), torsion/detorsion group (T/D by torsion of right testis with rotated 720° counter clockwise for 3 h, then detorsion) (n=8), Melatonin group given 25 mg/kg after torsion/detorsion (M) (n=8), Adipose tissue-derived mesenchymal stem cell-treated group after torsion/detorsion (MSC) (n=8), Adipose tissue-derived mesenchymal stem cell-treated group with melatonin after torsion/detorsion (MSC+M) (n=8). We measured MDA, Testosterone, FSH and LH levels, performed histopathological analyses in testicles, and identified SOX, VASA and Caspas-3 reactions immunohistochemically. Testosterone, FSH, LH values did not yield any significant difference between the groups. While the Johnson score in the right testis remained the lowest in T/D, the highest score was noted in the S. The T/D manifested some degenerative seminiferous tubules, abnormal spermatogenesis and maturation arrest. The degenerative appearance monitored in M, MSC and MSC+M groups persisted in some tubules, while markedly reduced degeneration was observed in some other tubules. The highest Caspase-3 expression in T/D, whereas SOX-9 expression remained significantly higher in the treatment groups. Another aspect deserving attention is that MSC were characterized by low VASA expression. Our experimental trial suggests that the torsion-induced degeneration in testicular tissue was ameliorated in all the treatment groups. Although MSC, MSC+M and M administrations decreased the torsion-induced degeneration in the testicular tissue, these treatments did not prove to be superior to each other.

Anahtar Kelimeler

• Testicular Torsion-Detorsion
• Stem cell
• Melatonin
• Apoptosis

Testiküler Torsiyonun İndüklediği Kısırlığa Mezenkimal Kök Hücrenin ve Melatoninin Koruyucu Etkisi

Öz

Sistemik melatonin ve intratestiküler adipoz doku kaynaklı mezenkimal kök hücrelerin (AdMSCs) uygulamasının akut tek taraflı testis torsiyonu olan sıçanlar üzerindeki etkilerini araştırmayı amaçladık. Sıçanlar, Sham grubu (S) (n=8), torsiyon/detorsiyon grubu (sağ testisin 3 saat boyunca saat yönünün tersine 720° döndürülerek torsiyonu ve ardından detorsiyonu ile T/D) (n=8), torsiyon/detorsiyon sonrası 25 mg/kg melatonin verilen grup (M) (n=8), torsiyon/detorsiyon sonrası adipoz doku kaynaklı mezenkimal kök hücre ile tedavi edilen grup (MSC) (n=8), torsiyon /detorsiyon sonrası melatonin ile adipoz dokusu kaynaklı mezenkimal kök hücre ile tedavi edilen grup (MSC+M) (n=8) olarak randomize edildi. MDA, Testosteron, FSH ve LH düzeyleri ölçüldü. Testislerde histopatolojik analizler yapıldı ve SOX, VASA ve Caspas-3 reaksiyonları immünohistokimyasal olarak incelendi. Testosteron, FSH, LH değerleri gruplar arasında anlamlı bir fark göstermedi. Sağ testiste Johnson skoru T/D'de en düşük seviyede kalırken, en yüksek skor S grubunda kaydedildi. T/D grubunda bazı dejeneratif seminifer tübüller, anormal spermatogenez ve olgunlaşma durması görüldü. M, MSC ve MSC+M gruplarında izlenen dejeneratif görünüm bazı tübüllerde devam ederken, diğer bazı tübüllerde belirgin şekilde azalmış olduğu gözlendi. En yüksek Kaspaz-3 ekspresyonu T/D'de görülürken, SOX-9 ekspresyonu tedavi gruplarında anlamlı derecede yüksek kalmıştır. Dikkat edilmesi gereken bir diğer husus da MSC'nin düşük VASA ekspresyonu ile karakterize olmasıdır. Deneysel çalışmamız, testis dokusunda torsiyona bağlı dejenerasyonun tüm tedavi gruplarında iyileştiğini göstermektedir. MSC, MSC+M ve M uygulamaları testis dokusunda torsiyona bağlı dejenerasyonu azaltmış olsa da, bu tedavilerin birbirlerine üstünlüğü kanıtlanmamıştır.

Anahtar Kelimeler

• Testiküler Torsiyon-Detorsiyon
• Kök hücre
• Melatonin
• Apoptozis

Kaynakça

1. 1. Gul SS, Gurgul S, Uysal M, Erdemir F. The Protective Effects of Pulsed Magnetic Field and Melatonin on Testis Torsion and Detorsion Induced Rats Indicated by Scintigraphy, Positron Emission Tomography/Computed Tomography and Histopathological Methods. Urol J. 2018;15(6):387-96.
2. 2. Abasiyanik A, Dağdönderen L. Beneficial effects of melatonin compared with allopurinol in experimental testicular torsion. J Pediatr Surg. 2004;39(8):1238-1241. h
3. 3. Sekmenli T, Gunduz M, Öztürk B, Karabağlı P, Ciftci I, Tekin G, et al. The effects of melatonin and colchicine on ischemia-reperfusion injury in experimental rat testicular torsion model. J Pediatr Surg. 2017;52(4):582-6.
4. 4. Morielli T, O'Flaherty C. Oxidative stress impairs function and increases redox protein modifications in human spermatozoa. Reproduction 2015;149(1):113-23.
5. 5. Karimaghai N, Tamadon A, Rahmanifar F, Mehrabani D, Raayat Jahromi A, Zare S, et al. Spermatogenesis after transplantation of adipose tissue-derived mesenchymal stem cells in busulfan-induced azoospermic hamster. Iran J Basic Med Sci. 2018;21(7):660-7.
6. 6. Hsiao CH, Ji AT, Chang CC, Cheng CJ, Lee LM, Ho JH. Local injection of mesenchymal stem cells protects testicular torsion-induced germ cell injury. Stem Cell Res Ther. 2015;6(1):113.
7. 7. Cakici C, Buyrukcu B, Duruksu G, Haliloglu AH, Aksoy A, Isık A, et al. Recovery of fertility in azoospermia rats after injection of adipose-tissue-derived mesenchymal stem cells: the sperm generation. Biomed Res Int. 2013;529589.
8. 8. Reiter RJ, Tan DX, Rosales-Corral S, Manchester LC. The universal nature, unequal distribution and antioxidant functions of melatonin and its derivatives. Mini Rev Med Chem. 2013;13(3):373-84.

9. 9. Zhou L, Song K, Xu L, Zhao F, Tian H, Zhou C, et al. Protective Effects of Uncultured Adipose-Derived Stromal Vascular Fraction on Testicular Injury Induced by Torsion-Detorsion in Rats. Stem Cells Transl Med. 2019;8(4):383-91.
10. 10. Yurtçu M, Abasiyanik A, Avunduk MC, Muhtaroğlu S. Effects of melatonin on spermatogenesis and testicular ischemia-reperfusion injury after unilateral testicular torsion-detorsion. Journal of pediatric surgery 2008;43:1873–1878.
11. 11. Jeong SJ, Choi WS, Chung JS, Baek M, Hong SK, Choi H. Preventive effects of cyclosporine combined with prednisolone and melatonin on contralateral testicular damage after ipsilateral torsion-detorsion in pubertal and adult rats. The Journal of urology 2010;184:790–796. https://doi.org/10.1016/j.juro.2010.03.109
12. 12. Ekici S, Doğan Ekici AI, Öztürk G, Benli Aksungar F, Sinanoğlu O, Turan G, Lüleci N. Comparison of melatonin and ozone in the prevention of reperfusion injury following unilateral testicular torsion in rats. Urology 2012;80(4):899-6.
13. 13. Hu C, Li L. Melatonin plays critical role in mesenchymal stem cell-based regenerative medicine in vitro and in vivo. Stem Cell Res Ther. 2019;10(1):13.
14. 14. Mirhoseini M, Talebpour Amiri F, Karimpour Malekshah AA, Rezanejad Gatabi Z, Ghaffari E. Protective effects of melatonin on testis histology following acute torsion-detorsion in rats. Int J Reprod Biomed. 2017;15(3):141-6.
15. 15. Mouro VGS, Menezes TP, Lima GDA, Domingues RR, Souza ACF, Oliveira JA, et al. How Bad Is Aluminum Exposure to Reproductive Parameters in Rats? Biol Trace Elem Res. 2018;183(2):314-324.
16. 16. Johnsen SG. Testicular biopsy score count--a method for registration of spermatogenesis in human testes: normal values and results in 335 hypogonadal males. Hormones 1970;1(1):2-25.
17. 17. Hsiao CH, Ji AT, Chang CC, Chien MH, Lee LM, Ho JH. Mesenchymal stem cells restore the sperm motility from testicular torsion-detorsion injury by regulation of glucose metabolism in sperm. Stem Cell Res Ther. 2019;10(1):270.
18. 18. Siregar S, Sasongko Noegroho B, Adriansjah R, Mustafa A, Wijayanti Z. Intratesticular Human Adipose-Derived Stem Cell (hADSC) Transplantation Decreased Oxidative Stress in Testicular Torsion Model of Wistar Rat. Res Rep Urol. 2021;13:1-8.
19. 19. Karabulut D, Akın A, Sayan M, Kaymak E, Öztürk E, and Yakan B. Effects of Melatonin Against Thioacetamide-Induced Testicular Toxicity in Rats. International Journal of Morphology 2020;38:1455-1462.
20. 20. Yu K, Deng SL, Sun TC, Li YY, Liu YX. Melatonin Regulates the Synthesis of Steroid Hormones on Male Reproduction: A Review. Molecules. 2018;23(2):447.
21. 21. Semercioz A, Baltaci AK, Mogulkoc R, Avunduk MC. Effect of Zinc and Melatonin on Oxidative Stress and Serum Inhibin-B Levels in a Rat Testicular Torsion-Detorsion Model. Biochem Genet. 2017;55(5-6):395-409.
22. 22. Muratoğlu S, Akarca Dizakar OS, Keskin Aktan A, Ömeroğlu S, Akbulut KG. The protective role of melatonin and curcumin in the testis of young and aged rats. Andrologia. 2019;51(3).
23. 23. Parlaktas BS, Atilgan D, Ozyurt H, Gencten Y, Akbas A, Erdemir F, Uluocak N. The biochemical effects of ischemia-reperfusion injury in the ipsilateral and contralateral testes of rats and the protective role of melatonin. Asian J Androl. 2014;16(2):314-8.
24. 24. Chabra A, Shokrzadeh M, Naghshvar F, Salehi F, Ahmadi A. Melatonin ameliorates oxidative stress and reproductive toxicity induced by cyclophosphamide in male mice. Hum Exp Toxicol. 2014;33(2):185-5.
25. 25. Chen YT, Chuang FC, Yang CC, Chiang JY, Sung PH, Chu YC, Huang CR, Huang KH, Yip HK. Combined melatonin-adipose derived mesenchymal stem cells therapy effectively protected the testis from testicular torsion-induced ischemia-reperfusion injury. Stem cell research & therapy 2021;12:370.
26. 26. Zhong L, Yang M, Zou X, Du T, Xu H, Sun J. Human umbilical cord multipotent mesenchymal stromal cells alleviate acute ischemia-reperfusion injury of spermatogenic cells via reducing inflammatory response and oxidative stress. Stem Cell Res Ther. 2020;11(1):294.
27. 27. Liu ZM, Zhu WP, Huang DP, Bai PM.Effects of melatonin on oxidative stress and apoptosis-related gene signaling pathways following testicular torsion in rats. Zhonghua Nan Ke Xue. 2019;25(5):309-4.
28. 28. Cebi Sen C, Yumusak N, Atilgan HI, Sadic M, Koca G, Korkmaz M. The protective effect of melatonin on sperm quality in rat after radioiodine treatment. Andrologia. 2018. https://doi.org/10.1111/and.12962