Pleurotus ostreatus (Jacq.) P. Kumm. Ekstraktı Bazı Apoptotik Genlerin İfadesini Değiştirir

Abstract

Mantarlar, eski çağlardan beri yiyecek ve tıbbi amaçlarla kullanılmıştır. Özellikle tedavi edici etkisi olan mantarlar birçok araştırma grubunun ilgisini çekmektedir. Aynı zamanda, mantarlardan türetilen aktif bileşiklerin potansiyel olarak yeni antikanser ajanların değerli bir kaynağı olabileceği düşünülmektedir. Bu çalışmanın amacı, Pleurotus ostreatus (Jacq.) P. Kumm'un metanolik ekstraktının apoptozda intrinsik yolda önemli olan bazı gen bölgelerinin ekspresyon seviyeleri üzerindeki etkilerini değerlendirmektir. Bu amaçla mantar örnekleri güneş ışığı görmeden kurutulduktan sonra Soxhlet cihazı ile metanol kullanılarak ekstraktlar hazırlandı. Ekstrelerin sitotoksik etkileri MTT testi ile değerlendirildi. Gerçek zamanlı PCR ile, dört apoptotik gen bölgesinin (Hrk, Bax, Apaf1 ve casp3) ekspresyon seviyelerini değerlendirildi. MTT testinin sonuçları, elde edilen ekstrelerin doza ve zamana bağlı bir şekilde sitotoksik bir etki gösterdiğini göstermiştir. Bununla birlikte P.ostreatus’tan elde edilen metanolik özütlerin apoptotik hücre ölümüyle ilişkili gen bölgelerinin ekspresyon seviyelerinde upregülasyona neden olduğu bulundu. Sonuç olarak, bu çalışma ile, P. ostreatus'un kolorektal kanser üzerinde potansiyel bir terapötik etkiye sahip olduğunu ve bu durumun farklı kanser türleri ve hücre dizileri ile ilgili diğer çalışmalarla uyumlu olduğunu göstermektedir. Bu çalışma, ekstraktaki aktif maddeleri tanımlamaya ve hücre ölümünün moleküler yollarını bulmaya devam edecek gelecekteki çalışmalar için öncü bir çalışmadır.

Keywords

• cDNA
• MTT
• RT-PCR
• Kavak Mantarı
• Türkiye

Pleurotus ostreatus (Jacq.) P. Kumm. Extract Alters the Expression of Some Apoptosis Related Genes

Abstract

Mushrooms have been used for food and medicinal purposes since ancient times. Especially mushrooms with therapeutic effects attract the attention of many research groups. Besides, it is thought that the active compounds derived from fungi could potentially be a valuable source of new anticancer agents. This study aims to evaluate the effects of methanolic extract of Pleurotus ostreatus (Jacq.) P. Kumm. on the expression levels of some genes important in the intrinsic pathway in apoptosis. For this purpose, after the mushroom samples were dried without sunlight, extracts were prepared via Soxhlet apparatus by using methanol. Cytotoxic effects of the extracts were evaluated with the MTT test. Real-time PCR was performed to evaluate the expression levels of the four apoptotic genes (Hrk, Bax, Apaf1 and casp3). The results of the MTT assay showed that the extracts obtained show a cytotoxic effect in a dose and time-dependent manner. Also, methanolic extracts from P. ostreatus were found to cause upregulation in expression levels of genes which related apoptotic cell death. In conclusion, this study shows that P. ostreatus has a potential therapeutic effect on colorectal cancer and is compatible with other studies of different types of cancer and cell lines. This study is a pioneering study for future studies that will continue to identify the active substances in the extract and find the molecular pathways of cell death.

Keywords

• cDNA
• MTT
• RT-PCR
• Oyster mushroom
• Turkey

References

1. Altunkaynak, B. Z., & Özbek, E. (2008). Programlanmış hücre ölümü: Apoptoz nedir. Tıp Araştırmaları Dergisi, 6(2), 93-104.
2. Arora, S., & Tandon, S. (2015). Mushroom extracts induce human colon cancer cell (COLO-205) death by triggering the mitochondrial apoptosis pathway and G0/G1-phase cell cycle arrest. Arch Iran Med, 18(5):284-95.
3. Barros, L., Baptista, P., Estevinho, L. M., & Ferreira, I. C. (2007). Bioactive properties of the medicinal mushroom Leucopaxillus giganteus mycelium obtained in the presence of different nitrogen sources. Food Chemistry, 105(1), 179-186.
4. Blagodatski, A., Yatsunskaya, M., Mikhailova, V., Tiasto, V., Kagansky, A., & Katanaev, V. L. (2018). Medicinal mushrooms as an attractive new source of natural compounds for future cancer therapy. Oncotarget, 9(49), 29259.
5. Burlacu, A. (2003). Regulation of apoptosis by Bcl‐2 family proteins. Journal of Cellular and Molecular Medicine, 7(3), 249-257.
6. Czabotar, P. E., Lessene, G., Strasser, A., & Adams, J. M. (2014). Control of apoptosis by the BCL-2 protein family: implications for physiology and therapy. Nature reviews Molecular Cell Biology, 15(1), 49-63.
7. Ebrahimi, A., Atashi, A., Soleimani, M., Mashhadikhan, M., Barahimi, A., & Maghari, A. (2018). Anti-invasive and antiproliferative effects of Pleurotus ostreatus extract on acute leukemia cell lines. Journal of Basic and Clinical Physiology and Pharmacology, 29(1), 95-102.
8. Ekowati, N., Mumpuni, A. & Muljowati, J. S. (2017). Effectiveness of Pleurotus ostreatus Extract Through Cytotoxic Test and Apoptosis Mechanism of Cervical Cancer Cells. Biosaintifika: Journal of Biology & Biology Education, 9(1), 148-155.

9. Fan, L., Pan, H., Soccol, A. T., Pandey, A., & Soccol, C. R. (2006). Advances in mushroom research in the last decade. Food Technology and Biotechnology, 44(3), 303-311.
10. Fulda, S. and Debatin, K. (2006) Extrinsic versus intrinsic apoptosis pathways in anticancer chemotherapy. Oncogene, 25, 4798-4811.
11. Gortat, A., Sancho, M., Mondragón, L., Messeguer, À., Pérez-Payá, E., & Orzáez, M. (2015). Apaf1 inhibition promotes cell recovery from apoptosis. Protein & Cell, 6(11), 833-843.
12. Gu, Y. H., & Sivam, G. (2006). Cytotoxic effect of oyster mushroom Pleurotus ostreatus on human androgen-independent prostate cancer PC-3 cells. Journal of Medicinal Food, 9(2), 196-204.
13. Inohara, N., Ding, L., Chen, S., & Núñez, G. (1997). Harakiri, a novel regulator of cell death, encodes a protein that activates apoptosis and interacts selectively with survival‐promoting proteins Bcl‐2 and Bcl‐XL. The EMBO journal, 16(7), 1686-1694.
14. Jedinak, A., & Sliva, D. (2008). Pleurotus ostreatus inhibits proliferation of human breast and colon cancer cells through p53-dependent as well as p53-independent pathway. International Journal of Oncology, 33(6), 1307-1313.
15. Kim, H. G., Yoon, D. H., Lee, W. H., Han, S. K., Shrestha, B., Kim, C. H., ... & Song, W. O. (2007). Phellinus linteus inhibits inflammatory mediators by suppressing redox-based NF-κB and MAPKs activation in lipopolysaccharide-induced RAW 264.7 macrophage. Journal of Ethnopharmacology, 114(3), 307-315.
16. Lavi, I., Friesem, D., Geresh, S., Hadar, Y., & Schwartz, B. (2006). An aqueous polysaccharide extract from the edible mushroom Pleurotus ostreatus induces anti-proliferative and pro-apoptotic effects on HT-29 colon cancer cells. Cancer letters, 244(1), 61-70.
17. Lindequist, U., Niedermeyer, T. H., & Jülich, W. D. (2005). The pharmacological potential of mushrooms. Evidence-Based Complementary and Alternative Medicine, 2(3), 285-299.
18. Mizuno, T., Saito, H., Nishitoba, T., & Kawagishi, H. (1995). Antitumor‐active substances from mushrooms. Food Reviews International, 11(1), 23-61.
19. Montero, J., Sarosiek, K. A., DeAngelo, J. D., Maertens, O., Ryan, J., Ercan, D., ... & Jänne, P. A. (2015). Drug-induced death signaling strategy rapidly predicts cancer response to chemotherapy. Cell, 160(5), 977-989.
20. Nagata, S. (2000). Apoptotic DNA fragmentation. Experimental Cell Research, 256(1), 12-18.
21. Oltvai, Z. N., Milliman, C. L., & Korsmeyer, S. J. (1993). Bcl-2 heterodimerizes in vivo with a conserved homolog, Bax, that accelerates programmed cell death. Cell, 74(4), 609-619.
22. Polyakov, V. Y., Kir’yanov, G. I., Gerasimenya, V. P., Orlov, A. E., Lazareva, E. M., Murasheva, M. I., & Chentsov, Y. S. (2007). Synergistic effects of mycelial fungus (Pleurotus ostreatus) extracts and some cytostatic drugs on proliferation and apoptosis in transformed human cells. Biochemistry (Moscow) Supplement Series A: Membrane and Cell Biology, 1(4), 301-309.
23. Sarikurkcu, C., Tepe, B., & Yamac, M. (2008). Evaluation of the antioxidant activity of four edible mushrooms from the Central Anatolia, Eskisehir–Turkey: Lactarius deterrimus, Suillus collitinus, Boletus edulis, Xerocomus chrysenteron. Bioresource Technology, 99(14), 6651-6655.
24. Spanos, S., Rice, S., Karagiannis, P., Taylor, D., Becker, D. L., Winston, R. M. L., & Hardy, K. (2002). Caspase activity and expression of cell death genes during development of human preimplantation embryos. Reproduction-Cambridge-, 124(3), 353-363.
25. Synytsya, A., Míčková, K., Synytsya, A., Jablonský, I., Spěváček, J., Erban, V., ... & Čopíková, J. (2009). Glucans from fruit bodies of cultivated mushrooms Pleurotus ostreatus and Pleurotus eryngii: Structure and potential prebiotic activity. Carbohydrate polymers, 76(4), 548-556.
26. Wasser, S. P. (2002). Medicinal mushrooms as a source of antitumor and immunomodulating polysaccharides. Applied Microbiology and Biotechnology, 60(3), 258-274.
27. Xu, T., B Beelman, R., & D Lambert, J. (2012). The cancer preventive effects of edible mushrooms. Anti-Cancer Agents in Medicinal Chemistry (Formerly Current Medicinal Chemistry-Anti-Cancer Agents), 12(10), 1255-1263.