İnfliksimab testiküler torsiyon/detorsiyon hasarında koruyucu mu? NF-ĸB ve mapk yolakları (ERK1/2, JNK, p38 MAPK)

Öz

Amaç: Bu çalışmada, TNF-α inhibitörü infliksimab (INF) kullanarak, testis torsiyonu/detorsiyonu (TT/D) yaralanmasında tümör nekroz faktörü-alfa (TNF-α) etkilerini ve nükleer faktör kappa B (NF-κB) ve mitojenle aktive olan protein kinaz (MAPK) yolaklarının bu etkilere katkıda bulunup bulunmadığını araştırmayı amaçladık. Gereç ve Yöntemler: Sıçanlar 3 gruba ayrıldı: Sham, T/D ve INF+T/D. Torsiyon, sağ testisin saat yönünde 720° döndürülmesiyle oluşturuldu ve testis 3 saat boyunca sabitlendi ve korundu. Sıçanlar, detorsiyondan 10 dakika önce 5 mg/kg infliksimab (i.p.) veya çözücüsü ile tedavi edildi. Torsiyonun giderilmesinden 2 ve 24 saat sonra testis ağırlıkları ölçüldü. Torsiyonun giderilmesinden 24 saat sonra, testislere verilen histopatolojik hasarın derecesi Cosentino skoru ve MAPK yolakları (JNK, p38 MAPK ve ERK1/2) ve NF-κB için immüno boyama kullanılarak değerlendirildi. Bulgular: T/D grubunda testis ağırlığında artış ve histopatolojik hasar gözlendi. Bu grupta, spermatogenez hücrelerinde NF-κB ve ERK1/2 için orta yoğunlukta immüno boyama, spermatidlerde JNK için şiddetli yoğunlukta immüno boyama ve spermatositlerde ve spermlerde p38 MAPK için şiddetli yoğunlukta immüno boyama gözlendi. INF tedavisinin testis ağırlıklarını, histopatolojik hasarı ve immüno boyama yoğunluğunu azalttığı gözlenmedi. Bu çalışma, NF-κB ve MAPK yolaklarının, özellikle JNK ve p38 MAPK'nin, TT/D hasarında germ hücrelerinde aktif olduğunu göstermiştir. TNF-α inhibitörü INF tedavisi, TT/D hasarında koruyucu bir etki göstermemiştir. Sonuç: TNF-α inhibisyonu tek başına TT/D hasarını azaltmak ve NF-κB ve MAPK'lerin aktivasyonunu inhibe etmek için yetersiz olabilir.

Anahtar Kelimeler

• Testiküler torsiyon/detorsiyon
• TNF-α inhibitörü
• infliksimab
• MAP kinaz yolağı

Destekleyen Kurum

Bu çalışma, Zonguldak Bülent Ecevit Üniversitesi Bilimsel Araştırma Projeleri (BEU BAP-2013-43341027-01) tarafından finansal olarak desteklenmiştir.

Proje Numarası

BEU BAP-2013-43341027-01

Etik Beyan

Bu çalışma, Zonguldak Bülent Ecevit Üniversitesi Deney Hayvanları Etik Kurulu tarafından onaylanmıştır (No:2014.02.15/02).

Teşekkür

Yazarlar, çalışmanın tamamlanması için gerekli deney ekipmanını ve teknik rehberliği sağlayan Zonguldak Bülent Ecevit Üniversitesi Deneysel Araştırma Uygulama ve Araştırma Merkezi'ne teşekkür ederler.

Is infliximab protective in testicular torsion/detorsion injury? NF-κB and MAPKs pathway (ERK1/2, JNK, p38 MAPK)

Öz

Aim: In this study, we aimed to investigate the effects of tumor necrosis factor-alpha (TNF-α) in testicular torsion/detorsion (TT/D) injury and whether the nuclear factor kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) pathways contribute to these effects, using the TNF-α inhibitor infliximab (INF). Material and Methods: Rats were randomly divided into three groups: Sham, T/D, and INF+T/D. Torsion was induced by rotating the right testis 720° clockwise, than the testis was fixed and maintained in that position for 3 hours. Ten minutes before detorsion, the rats were administered infliximab (5 mg/kg, i.p.) or the vehicle. Testicular weights were measured 2 and 24 hours after detorsion. Twenty-four hours after the detorsion procedure, the degree of histopathological damage to the testes was evaluated using the Cosentino score and immunostaining for MAPK pathways (JNK, p38 MAPK, and ERK1/2) and NF-κB. Results: In theT/D group, an increase in weight of testes and histopathological damage were observed. In this group, moderate intensity immunostaining was observed for NF-κB and ERK1/2 in spermatogenic cells, and strong immunostaining for JNK in spermatids, as well as for p38 MAPK in spermatocytes and sperm. INF treatment was not observed to reduce testicular weights, histopathological damage, and the intensity of immunostaining. Conclusion: This study presented that NF-κB and MAPK pathways, particularly JNK and p38 MAPK, are active in germ cells in TT/D injury. TNF-α inhibitor INF treatment did not show a protective effect in TT/D damage. Consequently, TNF-α inhibition alone may be insufficient to reduce TT/D damage and supress the activation of NF-κB and MAPKs.

Anahtar Kelimeler

• Testicular torsion/detorsion
• TNF-α inhibitor
• infliximab
• MAP kinase pathway.

Destekleyen Kurum

This study was financially supported by the Scientific Research Projects of Zonguldak Bülent Ecevit University (BEU BAP-2013-43341027-01).

Proje Numarası

BEU BAP-2013-43341027-01

Etik Beyan

This study has been approved by the Animal Ethics Committee of Zonguldak Bülent Ecevit University (No: 2014.02.15/02).

Teşekkür

The authors would like to thank the Experimental Research Application and Research Centre at Zonguldak Bülent Ecevit University for providing the experimental equipment and technical guidance necessary to complete the work.

Kaynakça

1. Schick MA, Sternard BT. Testicular Torsion. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2025 [cited 2025 Dec 25]. Available from: http://www.ncbi.nlm.nih.gov/books/NBK448199/
2. Moore SL, Chebbout R, Cumberbatch M, Bondad J, Forster L, Hendry J, et al. Orchidopexy for testicular torsion: a systematic review of surgical technique. Eur Urol Focus. 2021;7(6):1493-1503. https://doi.org/10.1016/j.euf.2020.07.006
3. Zvizdic Z, Aganovic A, Milisic E, Jonuzi A, Zvizdic D, Vranic S. Duration of symptoms is the only predictor of testicular salvage following testicular torsion in children: A case-control study. Am J Emerg Med. 2021;41:197-200. https://doi.org/10.1016/j.ajem.2020.11.023
4. Abdelzaher WY, Mostafa-Hedeab G, Sayed AboBakr Ali AH, Fawzy MA, Ahmed AF, Bahaa El-Deen MA, et al. Idebenone regulates sirt1/Nrf2/TNF-α pathway with inhibition of oxidative stress, inflammation, and apoptosis in testicular torsion/detorsion in juvenile rats. Hum Exp Toxicol. 2022;41:9603271221102515. https://doi.org/10.1177/09603271221102515
5. Wei SM, Huang YM. Baicalein alleviates testicular ischemia-reperfusion injury in a rat model of testicular torsion-detorsion. Oxid Med Cell Longev. 2022;2022:1603469. https://doi.org/10.1155/2022/1603469
6. Yakut S, Karabulut M, Koca RH, Erbaş E, Özkanlar S, Gençer BT, et al. Protective effects of bromelain in testicular torsion-detorsion: reducing inflammation, oxidative stress, and apoptosis while enhancing sperm quality. Biomolecules. 2025;15(2):292. https://doi.org/10.3390/biom15020292
7. Abdel-Kader MS, Abdel-Rahman RF, Althurwi HN, Soliman GA, Ogaly HA, Albaqami FF. Samarcandin protects against testicular ischemia/reperfusion injury in rats via activation of Nrf2/HO-1-mediated antioxidant responses. Saudi Pharm J. 2023;31(7):1186-1196. https://doi.org/10.1016/j.jsps.2023.05.007
8. Demir S, Kazaz IO, Kerimoglu G, Alemdar NT, Colak F, Arici T, et al. Gallic acid attenuates torsion/detorsion-induced testicular injury in rats through suppressing of HMGB1/NF-κB axis and endoplasmic reticulum stress. Rev Int Androl. 2024;22(1):1-7. https://doi.org/10.22514/j.androl.2024.001

9. Conrad M, Angeli JPF, Vandenabeele P, Stockwell BR. Regulated necrosis: disease relevance and therapeutic opportunities. Nat Rev Drug Discov. 2016;15(5):348-366. https://doi.org/10.1038/nrd.2015.6
10. Afolabi OA, Anyogu DC, Hamed MA, Odetayo AF, Adeyemi DH, Akhigbe RE. Glutamine prevents upregulation of NF-kB signaling and caspase 3 activation in ischaemia/reperfusion-induced testicular damage: An animal model. Biomed Pharmacother. 2022;150:113056. https://doi.org/10.1016/j.biopha.2022.113056
11. Besong EE, Akhigbe RE. Sodium acetate prevents testicular damage in Wistar rats subjected to testicular ischaemia/reperfusion injury. Exp Mol Pathol. 2024;137:104901. https://doi.org/10.1016/j.yexmp.2024.104901
12. Demir EA, Demir S, Kazaz IO, Kucuk H, Alemdar NT, Buyuk A, et al. Arbutin abrogates testicular ischemia/reperfusion injury in rats through repression of inflammation and ER stress. Tissue Cell. 2023;82:102056. https://doi.org/10.1016/j.tice.2023.102056
13. Abdel-Fattah MM, Ahmed AM, Saleh RK, Messiha BAS, Rofaeil RR. The protective effect of pregabalin and xanthenone on testicular ischemia/reperfusion injury in rats. Fundam Clin Pharmacol. 2024;38(6):1080-1093. https://doi.org/10.1111/fcp.13027
14. Minutoli L, Antonuccio P, Polito F, Bitto A, Fiumara T, Squadrito F, et al. Involvement of mitogen-activated protein kinases (MAPKs) during testicular ischemia-reperfusion injury in nuclear factor-kappaB knock-out mice. Life Sci. 2007;81(5):413-422. https://doi.org/10.1016/j.lfs.2023.122077
15. Zhao YG, Zheng XM, Zhou J, Liu XL, Hu XM, Chen DH, et al. Sulfasalazine prevents apoptosis in spermatogenic cells after experimental testicular torsion/detorsion. Acta Pharmacol Sin. 2006;27(5):603-608. https://doi.org/10.1111/j.1745-7254.2006.00287.x
16. Plotnikov A, Zehorai E, Procaccia S, Seger R. The MAPK cascades: signaling components, nuclear roles and mechanisms of nuclear translocation. Biochim Biophys Acta. 2011;1813(9):1619-1633. https://doi.org/10.1016/j.bbamcr.2010.12.012
17. Yang S, Shih HJ, Chow YC, Tsai PS, Huang CJ. Hemin induced heme oxygenase-1 over expression involves nuclear factor-E2 related factor-2, nuclear factor-kappaB and extracellular regulated kinase: an experimental study in a testicular torsion-detorsion rodent model. J Urol. 2008;179(6):2456-2463. https://doi.org/10.1016/j.juro.2008.01.085
18. Ahmed MAE, Ahmed AAE, El Morsy EM. Acetyl-11-keto-β-boswellic acid prevents testicular torsion/detorsion injury in rats by modulating 5-LOX/LTB4 and p38-MAPK/JNK/Bax/Caspase-3 pathways. Life Sci. 2020;260:118472. https://doi.org/10.1016/j.lfs.2020.118472
19. Alotaibi SR, Renno WM, Al-Maghrebi M. c-Jun N-terminal Kinase Supports Autophagy in Testicular Ischemia but Triggers Apoptosis in Ischemia-Reperfusion Injury. Int J Mol Sci. 2024;25(19):10446. https://doi.org/10.3390/ijms251910446
20. Antonuccio P, Minutoli L, Romeo C, Nicòtina PA, Bitto A, Arena S, et al. Lipid peroxidation activates mitogen-activated protein kinases in testicular ischemia-reperfusion injury. J Urol. 2006;176(4 Pt 1):1666-1672. https://doi.org/10.1016/j.juro.2006.06.086
21. Liu H, Shi M, Li X, Lu W, Zhang M, Zhang T, et al. Adipose mesenchymal stromal cell-derived exosomes prevent testicular torsion injury via activating PI3K/AKT and MAPK/ERK1/2 pathways. Oxid Med Cell Longev. 2022;2022:8065771. https://doi.org/10.1155/2022/8065771
22. Fadel F, Al-Kandari N, Khashab F, Al-Saleh F, Al-Maghrebi M. JNK inhibition alleviates oxidative DNA damage, germ cell apoptosis, and mitochondrial dysfunction in testicular ischemia reperfusion injury. Acta Biochim Biophys Sin (Shanghai). 2020;52(8):891-900. https://doi.org/10.1093/abbs/gmaa074
23. Li L, Hao Y, Zhao Y, Wang H, Zhao X, Jiang Y, et al. Ferroptosis is associated with oxygen-glucose deprivation/reoxygenation-induced Sertoli cell death. Int J Mol Med. 2018;41(5):3051-3062. https://doi.org/10.3892/ijmm.2018.3469
24. Mitoma H, Horiuchi T, Tsukamoto H, Ueda N. Molecular mechanisms of action of anti-TNF-α agents - Comparison among therapeutic TNF-α antagonists. Cytokine. 2018;101:56-63. https://doi.org/10.1016/j.cyto.2016.08.014
25. Tasdemir C, Tasdemir S, Vardi N, Ates B, Parlakpinar H, Kati B, et al. Protective effect of infliximab on ischemia/reperfusion-induced damage in rat kidney. Ren Fail. 2012;34(9):1144-9. https://doi.org/10.3109/0886022X.2012.717490
26. Guven C, Borcek AO, Cemil B, Kurt G, Yildirim Z, Ucankus NL, et al. Neuroprotective effects of infliximab in experimental spinal cord ischemic injury. J Clin Neurosci. 2010;17(12):1563-1567. https://doi.org/10.1016/j.jocn.2010.04.027
27. Livia C, Inglis S, Crespo-Diaz R, Rizzo S, Mahlberg R, Bagwell M, et al. Infliximab limits injury in myocardial infarction. J Am Heart Assoc. 2024;13(9):e032172. https://doi.org/10.1161/JAHA.123.032172
28. Yang Q, Zheng FP, Zhan YS, Tao J, Tan SW, Liu HL, et al. Tumor necrosis factor-α mediates JNK activation response to intestinal ischemia-reperfusion injury. World J Gastroenterol. 2013;19(30):4925-4934. https://doi.org/10.3748/wjg.v19.i30.4925
29. Hassan SM, Mohammed MH, Jawad MJ, Abbas AN. Use of infliximab to attenuate cerebral apoptosis induced by cerebral ischemia/reperfusion in male rats. Wiad Lek. 2023;76(2):326-331. https://doi.org/10.36740/WLek202302112
30. Albayrak Semalar S, Yılmaz Z, Özbakış Dengiz G, Gülle K, Akpolat Ferah M, Can M, et al. Effects of infliximab on testicular torsion detorsion injury in rats. Cukurova Medical Journal. 2024;49(4):925-934. https://doi.org/10.17826/cumj.1529709
31. Cosentino MJ, Nishida M, Rabinowitz R, Cockett AT. Histopathology of prepubertal rat testes subjected to various durations of spermatic cord torsion. J Androl. 1986;7(1):23-31. https://doi.org/10.1002/j.1939-4640.1986.tb00862.x
32. Lysiak JJ. The role of tumor necrosis factor-alpha and interleukin-1 in the mammalian testis and their involvement in testicular torsion and autoimmune orchitis. Reprod Biol Endocrinol. 2004;2:9. https://doi.org/10.1186/1477-7827-2-9
33. Souza LF, Horn AP, Gelain DP, Jardim FR, Lenz G, Bernard EA. Extracellular inosine modulates ERK 1/2 and p38 phosphorylation in cultured Sertoli cells: possible participation in TNF-alpha modulation of ERK 1/2. Life Sci. 2005;77(24):3117-3126. https://doi.org/10.1016/j.lfs.2005.05.049
34. Sabio G, Davis RJ. TNF and MAP kinase signaling pathways. Semin Immunol. 2014;26(3):237-245. https://doi.org/10.1016/j.smim.2014.02.009
35. de Oliveira SA, Cerri PS, Sasso-Cerri E. Impaired macrophages and failure of steroidogenesis and spermatogenesis in rat testes with cytokines deficiency induced by diacerein. Histochem Cell Biol. 2021;156(6):561-581. https://doi.org/10.1007/s00418-021-02023-7
36. Suh JH, Gong EY, Hong CY, Park E, Ahn RS, Park KS, et al. Reduced testicular steroidogenesis in tumor necrosis factor-alpha knockout mice. J Steroid Biochem Mol Biol. 2008;112(1-3):117-121. https://doi.org/10.1016/j.jsbmb.2008.09.003
37. Lysiak JJ, Nguyen QAT, Kirby JL, Turner TT. Ischemia-reperfusion of the murine testis stimulates the expression of proinflammatory cytokines and activation of c-jun N-terminal kinase in a pathway to E-selectin expression. Biol Reprod. 2003;69(1):202-210. https://doi.org/10.1095/biolreprod.102.013318
38. Minutoli L, Antonuccio P, Romeo C, Nicòtina PA, Bitto A, Arena S, et al. Evidence for a role of mitogen-activated protein kinase 3/mitogen-activated protein kinase in the development of testicular ischemia-reperfusion injury. Biol Reprod. 2005;73(4):730-736. https://doi.org/10.1095/biolreprod.105.040741
39. Al-Maghrebi M, Renno WM, Al-Ajmi N. Epigallocatechin-3-gallate inhibits apoptosis and protects testicular seminiferous tubules from ischemia/reperfusion-induced inflammation. Biochem Biophys Res Commun. 2012;420(2):434-439. https://doi.org/10.1016/j.bbrc.2012.03.013
40. Zhang Y, Lv Y, Liu YJ, Yang C, Hu HJ, Meng XE, et al. Hyperbaric oxygen therapy in rats attenuates ischemia-reperfusion testicular injury through blockade of oxidative stress, suppression of inflammation, and reduction of nitric oxide formation. Urology. 2013;82(2):489.e9-489.e15. https://doi.org/10.1016/j.urology.2013.04.016
41. Seker U, Gökçe Y, Kati B, Yuksel M, Guzel BC, Shokoohi M. Regulation of STAT3 and NF-κB signaling pathways by trans-Anethole in testicular ischemia-reperfusion injury and its gonadoprotective effect. Rev Int Androl. 2024;22(3):57-67. https://doi.org/10.22514/j.androl.2024.015
42. Guo Q, Jin Y, Chen X, Ye X, Shen X, Lin M, et al. NF-κB in biology and targeted therapy: new insights and translational implications. Sig Transduct Target Ther. 2024;9(1):53. https://doi.org/10.1038/s41392-024-01757-9
43. Ozcan L, Otunctemur A, Polat EC, Ozbek E, Kirecci SL, Somay A. Selective nuclear factor kappa b (NFkB) inhibitor, pyrrolidium dithiocarbamate prevents, long-term histologic damage in ischemia-reperfusion injuries after delayed testicular torsion. Urol J. 2016;13(3):2702-2706. https://doi.org/10.22037/uj.v13i3.3296.
44. Hashmi SF, Rathore HA, Sattar MA, Johns EJ, Gan CY, Chia TY, et al. Hydrogen sulphide treatment prevents renal ischemia-reperfusion injury by inhibiting the expression of ICAM-1 and NF-kB concentration in normotensive and hypertensive rats. Biomolecules. 2021;11(10):1549. https://doi.org/10.3390/biom11101549
45. Lysiak JJ, Bang HJ, Nguyen QAT, Turner TT. Activation of the nuclear factor kappa B pathway following ischemia-reperfusion of the murine testis. J Androl. 2005;26(1):129-135. PMID: 15611577. https://doi.org/10.1002/j.1939-4640.2005.tb02882.x
46. Zhou XL, Yang QS, Ni SZ, Tu XP, Zhao Y, Xu B, et al. Protective effects of lipoxin A4 in testis injury following testicular torsion and detorsion in rats. Mediators Inflamm. 2014;2014:898056. https://doi.org/10.1155/2014/898056
47. Ghasemi M, Basiri A, Kazemzadeh H, Manavi MA, Tavangar SM, Dehpour AR, et al. Linezolid mitigates tissue injury in experimental model of pediatric testicular torsion: TLR-4/MAPK/NF-κB involvement. Clin Exp Pediatr. 2025;68(9):700-711. https://doi.org/10.3345/cep.2025.00080
48. Kabay S, Ozden H, Guven G, Burukoglu D, Ustuner MC, Topal F, et al. Protective effects of the nuclear factor kappa B inhibitor pyrrolidine dithiocarbamate on experimental testicular torsion and detorsion injury. Korean J Physiol Pharmacol. 2014;18(4):321-326. https://doi.org/10.4196/kjpp.2014.18.4.321
49. Cargnello M, Roux PP. Activation and function of the MAPKs and their substrates, the MAPK-activated protein kinases. Microbiol Mol Biol Rev. 2011;75(1):50-83. https://doi.org/10.1128/MMBR.00031-10
50. Roux PP, Blenis J. ERK and p38 MAPK-activated protein kinases: a family of protein kinases with diverse biological functions. Microbiol Mol Biol Rev. 2004;68(2):320-344. https://doi.org/10.1128/MMBR.68.2.320-344.2004
51. Xia Z, Dickens M, Raingeaud J, Davis RJ, Greenberg ME. Opposing effects of ERK and JNK-p38 MAP kinases on apoptosis. Science. 1995;270(5240):1326-1331. https://doi.org/10.1126/science.270.5240.1326
52. Chang L, Karin M. Mammalian MAP kinase signalling cascades. Nature. 2001;410(6824):37-40. https://doi.org/10.1038/35065000
53. Wong CH, Cheng CY. Mitogen-activated protein kinases, adherens junction dynamics, and spermatogenesis: a review of recent data. Dev Biol. 2005;286(1):1-15. https://doi.org/10.1016/j.ydbio.2005.08.001
54. Moon C, Kim JS, Jang H, Lee HJ, Kim SH, Kang SS, et al. Activation of Akt/protein kinase B and extracellular signal-regulated kinase in rats with acute experimental testicular torsion. J Vet Med Sci. 2008;70(4):337-341. https://doi.org/10.1292/jvms.70.337
55. Sinha Hikim AP, Lue Y, Diaz-Romero M, Yen PH, Wang C, Swerdloff RS. Deciphering the pathways of germ cell apoptosis in the testis. J Steroid Biochem Mol Biol. 2003;85(2-5):175-182. https://doi.org/10.1016/S0960-0760(03)00193-6