Apoptosis Induction Through Increased TRPV1 Activation by Synergic Effect of Melatonin and Doxorubicin in Human Osteosarcoma and Chondrosarcoma Cell Lines

Year 2023, , 145 - 152, 29.10.2023

Ahmet Koçak , Anıl Gülcü , İshak Suat Övey

https://doi.org/10.30565/medalanya.1313745

Abstract

Aim: We aimed to reveal the role of doxorubicin (Dox), melatonin (Mel) and transient receptor potential Vanilloid 1 (TRPV1) channels in bone and cartilage cancer cells during the treatment process. Human Bone Osteosarcoma (Saos-2/An1) and Human Chondrosarcoma (Hs 819.T) cell lines were used to prepare in-vitro experiment models.

Methods: Both cell lines were cultured at 37°C. We have separated each cell line into five groups as follows: Controls, Dox, Dox+Capsazepine (Cpz), Dox+Melatonin (Mel), and combined Dox+Mel+Cpz given group. Capsaicin and capsazepine were added to cell culture mediums to activate or inactivate the TRPV1 channels, respectively. Cytosolic calcium, apoptosis, intracellular reactive oxygen, mitochondrial depolarization, caspase-3 and caspase-9 levels were measured.

Results: Increased apoptotic activity was detected in doxorubicin given cell lines (Group II) when compared with the controls (p˂0.001). There was also a significantly higher apoptotic level in Dox+Mel group (Group IV), when compared with only Dox given group (p˂0.001). TRPV1 inhibition applied groups (Group III and V) have had lower apoptotic levels than other drug administered groups (p˂0.001).

Conclusion: This study has indicated that apoptotic effects of Dox and Mel on both osteosarcoma and chondrosarcoma were strictly associated to TRPV1 channels, and that TRPV1 channels played an important role in whole mitochondria dependent pathways of apoptosis, which in turn may lead to increased intracellular Ca+2 levels and mitochondrial depolarization.

Keywords

Osteosarcoma, chondrosarcoma, TRPV1, doxorubicin, melatonin

İnsan Osteosarkoma ve Kondrosarkoma Hücre Hatlarında Melatonin ve Doksorubisinin Sinerjik Etkisi Yoluyla Artan TRPV1 Etkinliği Üzerinden Apopitoz Uyarımı

Abstract

Amaç: Kemik ve kıkırdak kanseri hücrelerinde doksorubisin (Dox) ve melatonin (Mel) ile birlikte geçici reseptör potansiyeli olan Vanilloid 1 (TRPV1) kanallarının tedavi sürecindeki rolünü ortaya çıkarmayı amaçladık. İn-vitro deney modellerinin hazırlanmasında İnsan Kemik Osteosarkomu (Saos-2/An1) ve İnsan Kondrosarkomu (Hs 819.T) hücre hatları kullanıldı.

Gereç ve Yöntem: Her iki hücre hattı da 37°C'de kültürlendi. Her hücre hattını aşağıdaki gibi beş gruba ayırdık: Kontroller, Dox, Dox+Kapsazepin (Cpz), Dox+Mel ve kombine Dox+Mel+Cpz verilen grup. Kapsaisin ve kapsazepin, sırasıyla TRPV1 kanallarını etkinleştirmek veya inaktive etmek için hücre kültürü ortamlarına eklendi. Sitozolik kalsiyum, apoptoz, hücre içi reaktif oksijen, mitokondriyal depolarizasyon, kaspaz 3 ve kaspaz 9 seviyeleri ölçüldü.

Bulgular: Doksorubisin (Dox) verilen hücre hatlarında (Grup II) kontrollere göre artmış apoptotik aktivite saptandı (p˂0.001). Ayrıca Dox+Mel grubunda (Grup IV), sadece Dox verilen grupla (p˂0.001) karşılaştırıldığında anlamlı derecede daha yüksek bir apoptotik seviye vardı. TRPV1 inhibisyonu uygulanan gruplar (Grup III ve V) diğer ilaç uygulanan gruplara göre daha düşük apoptotik düzeylere sahiptir (p˂0.001).

Sonuç: Bu çalışma, Dox ve Mel'in hem osteosarkom hem de kondrosarkom üzerindeki apoptotik etkilerinin TRPV1 kanalları ile kesin olarak ilişkili olduğunu ve TRPV1 kanallarının apoptozun tüm mitokondriye bağımlı yolaklarında önemli bir rol oynadığını ve bunun da hücre içi Ca+2 düzeylerinde artışa ve mitokondriyal depolarizasyona yol açabileceğini göstermiştir.

Keywords

Osteosarkom, kondrosarkom, TRPV1, doksorubisin, melatonin

References

• 1. Gelderblom H, Hogendoorn PC, Dijkstra SD, Rijswijk CS van, Krol AD, Taminiau AH, et al. The clinical approach towards chondrosarcoma. Oncologist. 2008;13(3):320-9 (2008). doi: 10.1634/theoncologist.2007-0237.
• 2. MacDonald LJ, Lin CY, Kuo SJ, Su CM, Tang CH. An update on current and future treatment options for chondrosarcoma, Expert Rev Anticancer Ther. 2019;19(9):773-86. doi: 10.1080/14737140.2019.1659731.
• 3. Dai X, Ma W, He X, Jha RK. Review of therapeutic strategies for osteosarcoma, chondrosarcoma and Ewing's sarcoma. Med Sci Monit. 2011;17(8):RA177-90. doi: 10.12659/msm.881893.
• 4. Huang YW, Lin CY, Tsai HC, Fong YC, Han CK, Huang YL, et al. Amphiregulin promotes cisplatin chemoresistance by upregulating ABCB1 expression in human chondrosarcoma. Aging (Albany NY). 2020;12(10):9475-88. doi: 10.18632/aging.103220.
• 5. Li Y, Li S, Zhou Y, Meng X, Zhang JJ, Xu DP, et al. Melatonin for the prevention and treatment of cancer. Oncotarget. 2017;8(24):39896-921. doi: 10.18632/oncotarget.16379.
• 6. Moran MM. TRP Channels as Potential Drug Targets. Annu Rev Pharmacol Toxicol. 2018;58:309-30. doi: 10.1146/annurev-pharmtox-010617-052832.
• 7. Li H. TRP Channel Classification. Adv Exp Med Biol. 2017;976:1-8. doi: 10.1007/978-94-024-1088-4_1.
• 8. Takahashi N, Chen HY, Harri IS, Stover DG, Selfors LM, Bronson RT, et al. Cancer Cells Co-opt the Neuronal Redox-Sensing Channel TRPA1 to Promote Oxidative-Stress Tolerance. Cancer Cell. 2018;33(6):985-1003.e7. doi: 10.1016/j.ccell.2018.05.001.
• 9. Jin T, Wu H, Wang Y, Peng H. Capsaicin induces immunogenic cell death in human osteosarcoma cells. Exp Ther Med. 2016;12(2):765-70. doi: 10.3892/etm.2016.3368.
• 10. Jadid MFS, Aghaei E, Taheri E, Seyyedsani N, Chavoshi R, Abbasi S, et al. Melatonin increases the anticancer potential of doxorubicin in Caco-2 colorectal cancer cells. Environ Toxicol. 2021;36(6):1061-69. doi: 10.1002/tox.23105.
• 11. Li W, Wang Z, Chen Y, Wang K, Lu T, Ying F, et al. Melatonin treatment induces apoptosis through regulating the nuclear factor-κB and mitogen-activated protein kinase signaling pathways in human gastric cancer SGC7901 cells. Oncol Lett. 2017;13(4):2737-44. doi: 10.3892/ol.2017.5785.
• 12. Martinez NA, Ayala AM, Martinez M, Martinez-Rivera FJ, Miranda JD, Silva WI. Caveolin-1 Regulates the P2Y2 Receptor Signaling in Human 1321N1 Astrocytoma Cells. J Biol Chem. 2016;291(23):12208-22. doi: 10.1074/jbc.M116.730226.
• 13. Övey İS, Güler Y. Apoptotic efficiency of capecitabine and 5-fluorouracil on human cancer cells through TRPV1 channels. Indian J Biochem Biophys. 2020;57(1):64-72.
• 14. Öz A, Çelik Ö. Curcumin inhibits oxidative stress-induced TRPM2 channel activation, calcium ion entry and apoptosis values in SH-SY5Y neuroblastoma cells: Involvement of transfection procedure. Mol Membr Biol. 2016;33(3-5):76-88. doi: 10.1080/09687688.2017.1318224.
• 15. Hattinger CM, Patrizio MP, Luppi S, Serra M. Pharmacogenomics and Pharmacogenetics in Osteosarcoma: Translational Studies and Clinical Impact. Int J Mol Sci. 2020;21(13):4659. doi: 10.3390/ijms21134659.
• 16. J.Z. Tan, S.M. Schlicht, G.J. Powell, et al., Multidisciplinary approach to diagnosis and management of osteosarcoma - a review of the St Vincent's Hospital experience. Int. Semin. Surg. Oncol. 2006;3:38. doi:10.1186/1477-7800-3-38.
• 17. Zheng J. Molecular mechanism of TRP channels. Compr Physiol. 2013;3(1):221-42. doi: 10.1002/cphy.c120001.
• 18. Zorov DB, Juhaszova M, Sollott SJ. Mitochondrial reactive oxygen species (ROS) and ROS-induced ROS release. Physiol Rev. 2014;94(3):909-50. doi: 10.1152/physrev.00026.2013.
• 19. Bao Z, Dai X, Wang P, Tao Y, Chai D. Capsaicin induces cytotoxicity in human osteosarcoma MG63 cells through TRPV1-dependent and -independent pathways. Cell Cycle. 2019;18(12):1379-92. doi: 10.1080/15384101.2019.1618119.
• 20. Gees M, Colsoul B, Nilius B. The role of transient receptor potential cation channels in Ca2+ signaling. Cold Spring Harb Perspect Biol. 2010;2(10):a003962. doi: 10.1101/cshperspect.a003962.
• 21. Niu G, Yousefi B, Qujeq D, Marjani A, Asadi J, Wang Z, et al. Melatonin and doxorubicin co-delivered via a functionalized graphene-dendrimeric system enhances apoptosis of osteo sarcoma cells. Mater Sci Eng C. Mater Biol Appl. 2021;119:111554. doi: 10.1016/j.msec.2020.111554.
• 22. Kumari R, H. DR Li, Haudenschild, F. Fierro, C.S. Carlson, P. Overn, et al., The oncogene LRF is a survival factor in chondrosarcoma and contributes to tumor malignancy and drug resistance. Carcinogenesis. 2012;33(11):2076-83. doi: 10.1093/carcin/bgs254.
• 23. Mukherjee SB, Das M, Sudhandiran G, Shaha C. Increase in cytosolic Ca2+ levels through the activation of non-selective cation channels induced by oxidative stress causes mitochondrial depolarization leading to apoptosis-like death in Leishmania Donovani promastigotes. J Biol Chem. 2002;277(27):24717-27. doi: 10.1074/jbc.M201961200.
• 24. Hsu YN, Shyu HW, Hu TW, Yeh JP, Lin YW, Lee LY, et al. Anti-proliferative activity of biochanin A in human osteosarcoma cells via mitochondrial-involved apoptosis. Food Chem Toxicol. 2018;112:194-204. doi: 10.1016/j.fct.2017.12.062.
• 25. Koşar PA, Nazıroğlu M, Övey İS, Çiğ B. Synergic Effects of Doxorubicin and Melatonin on Apoptosis and Mitochondrial Oxidative Stress in MCF-7 Breast Cancer Cells: Involvement of TRPV1 Channels. J Membr Biol. 2016;249(1-2):129-40. doi: 10.1007/s00232-015-9855-0.