ORIGANUM MINUTIFLORUM'UN NB2A NÖROBLASTOMA HÜCRELERİNE APOPTOTİK VE SİTOTOKSİK ETKİSİ

Öz

Amaç: Bu çalışmanın amacı, Türkiye'de endemik bir tıbbi bitki olan Origanum minutiflorum'dan elde edien Oregano yağının NB2a nöroblastoma hücrelerine sitotoksik ve apoptotik etkilerini incelemektir.

Gereç ve Yöntem: Hücre proliferasyonu, apoptoz, İndüklenebilir ve endotelyal nitrik oksit sentazın ekspresyonu sırasıyla MTT, TUNEL ve immünohistokimya ile belirlendi.

Sonuç ve Tartışma: Hem Oregano yağı hem de doksorubisin derişimleri artırıldığında NB2a hücrelerinin canlılığı kademeli olarak azaldı. IC50 dozu Oregano yağı için 10.75 ul / ml doksorubisin için 5µM olarak belirlendi. Ayrıca, Oregano yağı ve doksorubisin ile muamele edildikten sonra NB2a hücrelerinde indüklenebilir ve endotelyal nitrik oksit sentazın ekspresyonu ve apoptotik hücre sayısı anlamlı olarak yükseldi. Bu ajanların ikili uygulaması ile indüklenebilir ve endotelyal nitrik oksit sentaz ekspresyon seviyelerinde ve apoptotik hücre sayısında dikkate değer bir artış olmuştur. Ek olarak, Oregano yağı, doksorubisinin apoptotik etkisini arttırmıştır. Sonuç olarak, bulgularımız O. minutiflorum’a ait Oregano yağının NB2a hücreleri üzerinde sitotoksik ve apoptotik bir etkiye sahip olduğuna ve bu etkilerin Oregano yağının indüklenebilir ve endotelyal nitrik oksit sentaz düzeylerini arttırıcı etkileri ile ilişkili olabileceğine işaret etmektedir.

Anahtar Kelimeler

• apoptoz,sitotoksisite,nöroblastom,nitrik oksit sentaz,Origanum minutiflorum

APOPTOTIC AND CYTOTOXIC EFFECT OF ORIGANUM MINUTIFLORUM ON NB2A NEUROBLASTOMA CELLS

Öz

Objective: The aim of the present study is to investigate the cytotoxic and apoptotic effects of oregano oil from Origanum minutiflorum, which is an endemic medicinal plant in Turkey, on NB2a neuroblastoma cells.

Material and Method: Cell proliferation, apoptosis, and expression of inducible and endothelial nitric oxide synthase have been determined by MTT, TUNEL, and immunohistochemistry, respectively.

Result and Discussion: The cell viability was significantly decreased gradually in NB2a cells when the concentrations of both oregano oil and doxorubicin were increased. IC50 doses were found to be10.75 µl/ml for oregano oil and 5µM for doxorubicin. Furthermore, expression of inducible and endothelial nitric oxide synthase, also the number of apoptotic cells was significantly enhanced in NB2a cells after treatment with oregano oil and doxorubicin. There was a remarkable increase in inducible and endothelial nitric oxide synthase expression levels and in the number of apoptotic cells by the dual application of these agents. In addition, oregano oil enhanced the apoptotic effect of doxorubicin. In conclusion, our results indicated that oregano oil of O. minutiflorum has a cytotoxic and apoptotic effect on NB2a cells and these effects may be related to its enhancing effects on inducible and endothelial nitric oxide synthase levels.

Anahtar Kelimeler

• apoptosis,cytotoxicity,neuroblastoma,nitric oxide synthase,Origanum minutiflorum

Kaynakça

1. Oke, F., & Aslim, B. (2010). Biological potentials and cytotoxicity of various extracts from endemic Origanum minutiflorum O. Schwarz & PH Davis. Food and Chemical Toxicology, 48(6), 1728-1733.
2. Kiliçgün, H., Korkmaz, M., & Kılıçgün, H. (2014). Hepatoprotective and antidiabetic activity of origanum minutiflorum grown wild in Turkey. Bothalia Journal, 44, 3.
3. Ozen, F., Ekinci, F. Y., & Korachi, M. (2014). The inhibition of Helicobacter pylori infected cells by Origanum minutiflorum. Industrial Crops and Products, 58, 329-334.
4. Ozkan, G., Baydar, H., & Erbas, S. (2010). The influence of harvest time on essential oil composition, phenolic constituents and antioxidant properties of Turkish oregano (Origanum onites L.). Journal of the Science of Food and Agriculture, 90(2), 205-209.
5. Sari AB, Ustuner-Aydal O. (2018). Antioxidant and immunostimulant effects of origanum minutiflorum o. schwarz et. ph davis in rainbow trout. Fresenius Environmental Bulletin, 27, 1013-21.
6. Baydar, H. (2005). The effects of different harvest dates on essential oil content and essential oil composition in Origanum minutiflorum O. Akdeniz Üniversitesi Ziraat Fakültesi Dergisi, 18(2), 175-178.
7. Dadalioǧlu, I., & Evrendilek, G. A. (2004). Chemical compositions and antibacterial effects of essential oils of Turkish oregano (Origanum minutiflorum), bay laurel (Laurus nobilis), Spanish lavender (Lavandula stoechas L.), and fennel (Foeniculum vulgare) on common foodborne pathogens. Journal of agricultural and food chemistry, 52(26), 8255-8260.
8. Vardar‐Ünlü, G., Ünlü, M., Dönmez, E., & Vural, N. (2007). Chemical composition and in vitro antimicrobial activity of the essential oil of Origanum minutiflorum O Schwarz & PH Davis. Journal of the Science of Food and Agriculture, 87(2), 255-259.

9. Fakir, H., Korkmaz, M., & Icel, B. (2016). Medicinal plants traditionally used for pain alleviation in Antalya province, Turkey. Studies on Ethno-Medicine, 10(3), 314-324.
10. Marchese, A., Orhan, I. E., Daglia, M., Barbieri, R., Di Lorenzo, A., Nabavi, S. F., ... & Nabavi, S. M. (2016). Antibacterial and antifungal activities of thymol: A brief review of the literature. Food chemistry, 210, 402-414.
11. Elmastas, M., Celik, S. M., Genc, N., Aksit, H., Erenler, R., & Gulcin, İ. (2018). Antioxidant activity of an Anatolian herbal tea—Origanum minutiflorum: isolation and characterization of its secondary metabolites. International journal of food properties, 21(1), 374-384.
12. Soliman, A. M., Desouky, S., Marzouk, M., & Sayed, A. A. (2016). Origanum majorana attenuates nephrotoxicity of cisplatin anticancer drug through ameliorating oxidative stress. Nutrients, 8(5), 264.
13. Bower, A. M., Real Hernandez, L. M., Berhow, M. A., & De Mejia, E. G. (2014). Bioactive compounds from culinary herbs inhibit a molecular target for type 2 diabetes management, dipeptidyl peptidase IV. Journal of agricultural and food chemistry, 62(26), 6147-6158.
14. Akyil, D., Oktay, S., Liman, R., Eren, Y., & Konuk, M. (2012). Genotoxic and mutagenic effects of aqueous extract from aerial parts of Achillea teretifolia. Turkish Journal of Biology, 36(4), 441-448.
15. Franz, C., Baser, K. H. C., & Windisch, W. (2010). Essential oils and aromatic plants in animal feeding–a European perspective. A review. Flavour and Fragrance Journal, 25(5), 327-340.
16. Smith, S. L., Fishwick, J., McLean, W. G., Edwards, G., & Ward, S. A. (1997). Enhanced in vitro neurotoxicity of artemisinin derivatives in the presence of haemin. Biochemical pharmacology, 53(1), 5-10.
17. Deliloglu-Gurhan, I., Tuglu, I., Vatansever, H. S., Ozdal-Kurt, F., Ekren, H., Taylan, M., & Sen, B. H. (2006). The effect of osteogenic medium on the adhesion of rat bone marrow stromal cell to the hydroxyapatite. Saudi Med J, 27(3), 305-11.
18. Özdal-Kurt, F., Tuğlu, I., Vatansever, H. S., Tong, S., Şen, B. H., & Deliloğlu-Gürhan, S. I. (2016). The effect of different implant biomaterials on the behavior of canine bone marrow stromal cells during their differentiation into osteoblasts. Biotechnic & Histochemistry, 91(6), 412-422.
19. Tuğlu, İ., Özdal-Kurt, F., Koca, H., Sarac, A., Barut, T., Kazanç, A. (2010). The contribution of differentiated bone marrow stromal stem cell-loaded biomaterial to treatment in critical size defect model in rats. Kafkas Üniversitesi Veteriner Fakültesi Dergisi, 16(5).
20. Torre, L. A., Bray, F., Siegel, R. L., Ferlay, J., Lortet‐Tieulent, J., & Jemal, A. (2015). Global cancer statistics, 2012. CA: a cancer journal for clinicians, 65(2), 87-108.
21. Demir, E., Kaya, B., Marcos, R., Cenkci, S. K., & Çetin, H. (2013). Investigation of the genotoxic and antigenotoxic properties of essential oils obtained from two Origanum species by Drosophila wing SMART assay. Turkish Journal of Biology, 37(2), 129-138.
22. Sarkar, D., Sharma, A., & Talukder, G. (1994). Chlorophyll and chlorophyllin as modifiers of genotoxic effects. Mutation Research/Reviews in Genetic Toxicology, 318(3), 239-247.
23. Özkan, A., & Erdoğan, A. (2011). A comparative evaluation of antioxidant and anticancer activity of essential oil from Origanum onites (Lamiaceae) and its two major phenolic components. Turkish Journal of Biology, 35(6), 735-742.
24. Erenler, R., Sen, O., Aksit, H., Demirtas, I., Yaglioglu, A. S., Elmastas, M., & Telci, İ. (2016). Isolation and identification of chemical constituents from Origanum majorana and investigation of antiproliferative and antioxidant activities. Journal of the Science of Food and Agriculture, 96(3), 822-836.
25. Dadalioǧlu, I., & Evrendilek, G. A. (2004). Chemical compositions and antibacterial effects of essential oils of Turkish oregano (Origanum minutiflorum), bay laurel (Laurus nobilis), Spanish lavender (Lavandula stoechas L.), and fennel (Foeniculum vulgare) on common foodborne pathogens. Journal of agricultural and food chemistry, 52(26), 8255-8260.
26. García-Beltrán, J. M., & Esteban, M. A. (2016). Properties and applications of plants of Origanum Sp. Genus. SM. J. Biol, 2, 1006-1015.
27. Can Baser, K. H. (2008). Biological and pharmacological activities of carvacrol and carvacrol bearing essential oils. Current pharmaceutical design, 14(29), 3106-3119.
28. Marrelli, M., Statti, G. A., & Conforti, F. (2018). Origanum spp.: an update of their chemical and biological profiles. Phytochemistry reviews, 17(4), 873-888.
29. Kubatka, P., Kello, M., Kajo, K., Kruzliak, P., Výbohová, D., Mojžiš, J., ... & Péč, M. (2017). Oregano demonstrates distinct tumour-suppressive effects in the breast carcinoma model. European journal of nutrition, 56(3), 1303-1316.
30. Suntres, Z. E., Coccimiglio, J., & Alipour, M. (2015). The bioactivity and toxicological actions of carvacrol. Critical reviews in food science and nutrition, 55(3), 304-318.
31. Forstermann, U., & Sessa, W. C. (2012). Nitric oxide synthases: regulation and function. Eur Heart J, 33(7), 829-37.
32. Zhu, Y., Jiang, H., Chen, Z., Lu, B., Li, J., Peng, Y., & Shen, X. (2018). The genetic association between iNOS and eNOS polymorphisms and gastric cancer risk: a meta-analysis. OncoTargets and therapy, 11, 2497.
33. Calabrese, V., Mancuso, C., Calvani, M., Rizzarelli, E., Butterfield, D. A., & Stella, A. M. G. (2007). Nitric oxide in the central nervous system: neuroprotection versus neurotoxicity. Nature Reviews Neuroscience, 8(10), 766-775.
34. Hibbs, J. B., Taintor, R. R., & Vavrin, Z. (1987). Macrophage cytotoxicity: role for L-arginine deiminase and imino nitrogen oxidation to nitrite. Science, 235(4787), 473-476.
35. Brookes, P. S., Salinas, E. P., Darley-Usmar, K., Eiserich, J. P., Freeman, B. A., Darley-Usmar, V. M., & Anderson, P. G. (2000). Concentration-dependent effects of nitric oxide on mitochondrial permeability transition and cytochrome crelease. Journal of Biological Chemistry, 275(27), 20474-20479.
36. Moriya, R., Uehara, T., & Nomura, Y. (2000). Mechanism of nitric oxide‐induced apoptosis in human neuroblastoma SH‐SY5Y cells. FEBS letters, 484(3), 253-260.