K562 KRONİK MİYELOİD LÖSEMİ HÜCRE HATTINDA SIRT5 MODÜLATÖRLERİNİN SIRT5 VE SİTOKROM C PROTEİN EKSPRESYONLARI ÜZERİNE ETKİLERİNİN ARAŞTIRILMASI

Yıl 2022, , 805 - 816, 30.09.2022

Tulin Ozkan , Aslı Koc , Arzu Zeynep Karabay , Yalda Hekmatshoar , Asuman Sunguroglu

https://doi.org/10.33483/jfpau.1081893

Öz

Amaç: SIRT5, hedef proteinlerdeki lizin rezidülerinden, asetil, malonil ve süksinil gruplarını uzaklaştıran ve sitokrom c ile etkileşerek, onun deasetilasyonuna neden olan bir mitokondriyal proteindir. SIRT5'in K562 kronik miyeloid lösemi hücrelerindeki etkilerine ilişkin bir çalışma bulunmamaktadır. Resveratrol ve Suramin'in SIRT5'in deasetilaz ve desüksinilaz aktivitelerini modüle etmede rol oynadığı bilinmektedir. Resveratrol'ün K562 hücrelerinin apoptozunu indüklediği bildirilmiştir. Ancak Suramin'in K562 hücrelerinin apoptozu üzerindeki etkileri büyük ölçüde bilinmemektedir. Bu çalışmada, SIRT5 modülatörleri Resveratrol ve Suramin'in K562 hücrelerinin proliferasyonu ve apoptozu ile SIRT5 ve SIRT5'in bilinen bir hedefi olan sitokrom c proteini üzerindeki etkilerinin aydınlatılması amaçlanmıştır.
Gereç ve Yöntem: K562 kronik miyeloid lösemi hücrelerine artan konsantrasyonlarda Suramin ve Resveratrol uygulandı. Hücre proliferasyonu MTT analizi ve BrdU inkoporasyon yöntemi ile belirlendi. Apoptoz, Akım sitometrisi ile Annexin V boyaması ile belirlendi. Resveratrol ve Suramin'in SIRT5 ve Sitokrom c protein ekspresyon seviyeleri üzerindeki etkisini belirlemek için Western Blot analizi yapıldı.
Sonuç ve Tartışma: Sonuçlarımız, Suramin'in SIRT5 ve sitokrom c protein ekspresyonlarını önemli ölçüde etkilemediğini ve Resveratrol'ün SIRT5'i azalttığını ve sitokrom c ekspresyonunu artırdığını göstermiştir. Suramin, K562 hücrelerinin apoptozunda herhangi bir değişikliğe neden olmamıştır. Resveratrol, literatüre uygun olarak hücre proliferasyonunu azaltmış ve K562 hücrelerinin apoptozunu indüklemiştir. Resveratrolün, SIRT5 protein ekspresyonunu azaltıcı etkisi ile apoptotik etkilerine aracılık etmiş olabileceği düşünülmektedir

Anahtar Kelimeler

Apoptoz, K562, Resveratrol, SIRT5, Suramin

Destekleyen Kurum

Ankara Üniversitesi Bilimsel araştırma Projeleri (BAP)

Proje Numarası

18H0237008.

AN INVESTIGATION ON THE EFFECTS OF SIRT5 MODULATORS ON SIRT5 AND CYTOCHROME C PROTEIN EXPRESSIONS IN K562 CHRONIC MYELOID LEUKEMIA CELL LINE

Öz

Objective: SIRT5 is a mitochondrial protein that removes acetyl, malonyl and succinyl groups from lysine moieties in target proteins and interacts with cytochrome c and causes its deacetylation. There is no study on the effects of SIRT5 in K562 chronic myeloid leukemia cells. Resveratrol and Suramin are known to play a role in modulating the deacetylase and desuccinylase activities of SIRT5. It has been reported that Resveratrol induces apoptosis of K562 cells but effects of Suramin on the apoptosis of K562 cells are largely unknown. In this study, it was aimed to elucidate the effects of SIRT5 modulators Resveratrol and Suramin on proliferation and apoptosis of K562 cells and on SIRT5 and cytochrome c protein, a known target of SIRT5.
Material and Method: K562 chronic myeloid leukemia cells were treated with increasing concentrations of suramin and resveratrol, cell proliferation was determined by MTT assay and BrdU incorporation. Apoptosis was determined with Annexin V staining by Flow cytometry. Western Blot analysis was performed to determine the effect of resveratrol and suramin on SIRT5 and Cytochrome c protein expression levels.
Result and Discussion: Our results showed that suramin did not affect SIRT5 and cytochrome c protein expressions significantly and resveratrol decreased SIRT5 and increased cytochrome c expression. Suramin did not cause any changes on the apoptosis of K562 cells. Resveratrol decreased cell proliferation and induced apoptosis of K562 cells in accordance with the literature. The SIRT5-lowering effect of Resveratrol may have mediated its apoptotic effects.

Anahtar Kelimeler

Apoptosis, K562, Resveratrol, SIRT5, Suramin

Proje Numarası

18H0237008.

Kaynakça

• Deininger, M.W., Goldman, J.M., Melo, J.V. (2000). The molecular biology of chronic myeloid leukemia. Blood, 96(10), 3343-3356. [CrossRef]
• 2. Pophali, P.A., Patnaik, M.M. (2016). The Role of New Tyrosine Kinase Inhibitors in Chronic Myeloid Leukemia. The Cancer Journal, 22(1), 40-50. [CrossRef]
• 3. Chen, X., Xu, Z., Zeng, S.,Wang, X., Liu,W.,Qian, L.,Wei, J., Yang, X., Shen, Q., Gong, Z., Yan, Y. (2019). SIRT5 downregulation is associated with poor prognosis in glioblastoma. Cancer Biomarkers, 24(4), 449-459. [CrossRef]
• 4. Bringman-Rodenbarger, L.R., Guo, A.H., Lyssiotis, C. A., Lombard, D.B. (2018). Emerging roles for SIRT5 in metabolism and cancer. Antioxidants & Redox Signaling, 28(8), 677-690. [CrossRef]
• 5. Kumar, S., Lombard, D. B. (2018). Functions of the sirtuin deacylase SIRT5 in normal physiology and pathobiology. Critical Reviews in Biochemistry and Molecular Biology, 53(3), 311-334. [CrossRef]
• 6. Wang, C. H., Wei, Y.H. (2020). Roles of mitochondrial sirtuins in mitochondrial function, redox homeostasis, insulin resistance and type 2 diabetes. International Journal of Molecular Sciences, 21(15), 5266. [CrossRef]
• 7. Greene, K. S., Lukey, M. J., Wang, X., Blank, B., Druso, J. E., Lin, M. J., Stalnecker, C.A., Zhang, C., Abril, Y. N., Erickson, J.W., Wilson, K.F., Lin, H., Weiss, R.S., Cerione, R. A. (2019). SIRT5 stabilizes mitochondrial glutaminase and supports breast cancer tumorigenesis. Proceedings of the National Academy of Sciences of the United States of America, 116(52), 26625-26632. [CrossRef]
• 8. Guan, J., Jiang, X., Gai, J., Sun, X., Zhao, J., Li, J., Li, Y., Cheng, M., Du, T., Fu, L., Li, Q. (2020). Sirtuin 5 regulates the proliferation, invasion and migration of prostate cancer cells through acetyl-CoA acetyltransferase 1. Journal of Cellular and Molecular Medicine, 24(23), 14039-14049. [CrossRef]
• 9. Wang, K., Hu, Z., Zhang, C., Yang, L., Feng, L., Yang, P., Hu, H. (2020). SIRT5 Contributes to Colorectal Cancer Growth by Regulating T Cell Activity. Journal of Immunology Research, 2020, 3792409. [CrossRef]
• 10. Sun, X., Wang, S., Gai, J., Guan, J., Li, J., Li, Y., Zhao, J., Zhao, C., Fu, L., Li, Q. (2019). SIRT5 Promotes Cisplatin Resistance in Ovarian Cancer by Suppressing DNA Damage in a ROS-Dependent Manner via Regulation of the Nrf2/HO-1 Pathway. Frontiers in Oncology, 9, 754. [CrossRef]
• 11. Guo, D., Song, X., Guo, T., Gu, S., Chang, X., Su, T., Yang, X., Liang, B., Huang, D. (2018). Vimentin acetylation is involved in SIRT5-mediated hepatocellular carcinoma migration. American Journal of Cancer Research, 8(12), 2453-2466.
• 12. Can, G., Cakir, Z., Kartal, M., Gunduz, U., Baran, Y. (2012). Apoptotic effects of resveratrol, a grape polyphenol, on imatinib-sensitive and resistant K562 chronic myeloid leukemia cells. Anticancer Research, 32(7), 2673-2678.
• 13. Chakraborty, P.K., Mustafi, S.B., Ganguly, S., Chatterjee, M., Raha, S. (2008). Resveratrol induces apoptosis in K562 (chronic myelogenous leukemia) cells by targeting a key survival protein, heat shock protein 70. Cancer Science, 99 (6), 1109-1116. [CrossRef]
• 14. Kartal, M., Saydam, G., Sahin, F., Baran, Y. (2011). Resveratrol triggers apoptosis through regulating ceramide metabolizing genes in human K562 chronic myeloid leukemia cells. Nutrition and Cancer, 63(4), 637-644. [CrossRef]
• 15. Wang, B., Liu, J., Gong, Z. (2015). Resveratrol induces apoptosis in K562 cells via the regulation of mitochondrial signaling pathways. International Journal of Clinical and Experimental Medicine, 8(9), 16926-16933.
• 16. Yan, H.W., Hu, W.X., Zhang, J.Y., Wang, Y., Xia, K., Peng, M. Y., Liu, J. (2014). Resveratrol induces human K562 cell apoptosis, erythroid differentiation, and autophagy. Tumour Biology, 35, 5381-5388. [CrossRef]
• 17. Bai, L., Naomoto, Y., Miyazaki, M., Orita, K., Namba, M. (1992). Antiproliferative effects of suramin on human cancer cells in vitro and in vivo. Acta Medica Okayama, 46(6), 457-463. [CrossRef]
• 18. Carafa, V., Rotili, D., Forgione, M., Cuomo, F., Serretiello, E., Hailu, G. S., Jarho, E., Lahtela-Kakkonen, M., Mai, A., Altucci, L. (2016). Sirtuin functions and modulation: from chemistry to the clinic. Clinical Epigenetics, 8, 61. [CrossRef]
• 19. Costa-Machado, L. F., Fernandez-Marcos, P. J. (2019). The sirtuin family in cancer. Cell Cycle, 18(18), 2164-2196. [CrossRef]
• 20. O'Callaghan, C., Vassilopoulos, A. (2017). Sirtuins at the crossroads of stemness, aging, and cancer. Aging Cell, 16, 1208-1218. [CrossRef]
• 21. Lim, M.L., Lum, M.G., Hansen, T.M., Roucou, X., Nagley, P. (2002). On the release of cytochrome c from mitochondria during cell death signaling. Journal of Biomedical Science, 9, 488-506. [CrossRef]
• 22. Guan, D., Lim, J.H., Peng, L., Liu, Y., Lam, M., Seto, E., Kao, H.Y. (2014). Deacetylation of the tumor suppressor protein PML regulates hydrogen peroxide-induced cell death. Cell Death & Disease, 5, e1340. [CrossRef]
• 23. Zhang, R., Wang, C., Tian, Y., Yao, Y., Mao, J., Wang, H., Li, Z., Xu, Y., Ye, M., Wang, L. (2019). SIRT5 Promotes Hepatocellular Carcinoma Progression by Regulating Mitochondrial Apoptosis. Journal of Cancer, 10, 3871-3782. [CrossRef]
• 24. Chao, S. C., Chen, Y. J., Huang, K. H., Kuo, K. L., Yang, T. H., Huang, K. Y., Wang, C.C., Tang, C.H., Yang, R.S., Liu, S.H. (2017). Induction of sirtuin-1 signaling by resveratrol induces human chondrosarcoma cell apoptosis and exhibits antitumor activity. Scientific Reports, 7, 3180. [CrossRef]
• 25. Luzi, C., Brisdelli, F., Cinque, B., Cifone, G., Bozzi, A. (2004). Differential sensitivity to resveratrol-induced apoptosis of human chronic myeloid (K562) and acute lymphoblastic (HSB-2) leukemia cells. Biochemical Pharmacology, 68(10), 2019-2030. [CrossRef]
• 26. Yu, H., Pan, W., Huang, H., Chen, J., Sun, B., Yang, L., Zhu, P. (2019). Screening Analysis of Sirtuins Family Expression on Anti-Inflammation of Resveratrol in Endothelial Cells. Medical Science Monitor, 25, 4137-4148. [CrossRef]
• 27. Gertz, M., Nguyen, G.T.T., Fischer, F., Suenkel, B., Schlicker, C., Fränzel, B., Tomaschewski, J., Aladini, F., Becker, C., Wolters, D., Steegborn, C. (2012). A molecular mechanism for direct sirtuin activation by resveratrol. PLoS One, 7(11), e49761. [CrossRef]
• 28. Marti, N., Bouchoucha, N., Sauter, K. S., Flück, C. E. (2017). Resveratrol inhibits androgen production of human adrenocortical H295R cells by lowering CYP17 and CYP21 expression and activities. PLoS One, 12, e0174224. [CrossRef]
• 29. Perabo, F. G., Müller, S. C. (2005). New agents in intravesical chemotherapy of superficial bladder cancer. Scandinavian Journal of Urology and Nephrology, 39(2), 108-116. [CrossRef]
• 30. Gertz, M., Steegborn, C. (2010). Function and regulation of the mitochondrial sirtuin isoform Sirt5 in Mammalia. Biochimica et Biophysica Acta (BBP)- Proteins and Proteomics, 1804(8), 1658-1665. [CrossRef]
• 31. Schuetz, A., Min, J., Antoshenko, T., Wang, C. L., Allali-Hassani, A., Dong, A., Loppnau, P., Vedadi, M., Bochkarev, A., Sternglaz, R., Plotnikov, A.N. (2007). Structural basis of inhibition of the human NAD+-dependent deacetylase SIRT5 by suramin. Structure, 15(3),377-389. [CrossRef]
• 32. Wiedemar, N., Hauser, D. A., Mäser, P. (2020). 100 years of suramin. Antimicrobial Agents and Chemotherapy, 64(3), e01168-19. [CrossRef]
• 33. Choe, G., Kim, W. H., Park, J. G., Kim, Y. I. (1997). Effect of suramin on differentiation of human stomach cancer cell lines. Journal of Korean Medical Science, 12(5), 433-442. [CrossRef]
• 34. Bhargava, S., Hotz, B., Hines, O.J., Reber , H.A., Buhr, H. J., Hotz, H. G. (2007). Suramin inhibits not only tumor growth and metastasis but also angiogenesis in experimental pancreatic cancer. Journal of Gastrointest Surgery, 11, 171-178. [CrossRef]
• 35. Gogada, R., Prabhu, V., Amadori, M., Scott, R., Hashmi, S., Chandra, D. (2011). Resveratrol induces p53-independent, X-linked inhibitor of apoptosis protein (XIAP)-mediated Bax protein oligomerization on mitochondria to initiate cytochrome c release and caspase activation. Journal of Biological Chemistry, 286(33), 28749-28760. [CrossRef]