Investigation of the anticancer, antiproliferative and apoptotic effects of pine bark extract in ovarian (MDAH-2774) cancer cell culture model

Abstract

Objective: Gynecological cancers represent the fourth most common type of cancer worldwide. The high-performance liquid chromatography (HPLC) analysis revealed the presence of phenolic compounds, including catechin, epicatechin, taxifolin and phenolic acids in pine bark. The anticarcinogenic, antiproliferative and apoptotic effects of pine bark extracts on the human ovarian cancer cell line (MDAH-2774) were investigated. Materials and Methods: Gene expression was determined by means of the real-time reverse transcription-polymerase chain reaction (RT-PCR) method. A TdT-mediated dUTP Nick-End labelling (TUNEL) test was conducted to ascertain the extent of apoptosis. The IC₅₀ dose in MDAH-2774 cells was determined to be 117.6 μg/ml by 48 hours. Results: Notable alterations were discerned in the expression of caspase 3, 9, and 10 genes, which are implicated in programmed cell death, as well as in genes such as Bax, Bcl-2, and Bcl-Xl, which play a pivotal role in regulating the mitochondrial apoptosis pathway. A notable increase in the proportion of apoptotic cells was observed in the dose group. A significant reduction in invasion capacity was also identified. It has been demonstrated that pine bark extract can inhibit the migration capacity of ovarian cancer cells and colony formation. Conclusion: The findings suggest that pine bark extract exerts anticarcinogenic, antiproliferative and apoptotic effects on the ovarian cancer cell line.

Keywords

• Ovarian cancer cell (MDAH-2774)
• Cell culture techniques
• Apoptosis
• Neoplasms

References

1. WHO. https://www.who.int/news/item/01-02-2024-globalcancer- burden-growing—amidst-mounting-need-forservices [Internet]. 2022. Available from: https://www.who. int/news/item/01-02-2024-global-cancer-burden-growing— amidst-mounting-need-for-services Accessed on 13.12.2024.
2. Bray F, Ferlay J, Soerjomataram I, Siegel RL, Torre LA, Jemal A. Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin 2018;68:394-424. doi: 10.3322/ caac.21492.
3. Coburn SB, Bray F, Sherman ME, Trabert B. International patterns and trends in ovarian cancer incidence, overall and by histologic subtype. Int J Cancer 2017;140:2451-60. doi: 10.1002/ijc.30676.
4. Yoneda A, Lendorf ME, Couchman JR, Multhaupt HAB. Breast and ovarian cancers: a survey and possible roles for the cell surface heparan sulfate proteoglycans. J Histochem Cytochem 2012;60:9-21. doi: 10.1369/0022155411428469.
5. Badgwell D, Bast RC. Early detection of ovarian cancer. Dis Markers 2007;23:397-410. doi: 10.1155/2007/309382.
6. Jacobs IJ, Menon U. Progress and challenges in screening for early detection of ovarian cancer. Mol Cell Proteomics 2004;3:355-66. doi: 10.1074/mcp.R400006-MCP200
7. Momenimovahed Z, Tiznobaik A, Taheri S, Salehiniya H. Ovarian cancer in the world: epidemiology and risk factors. Int J Womens Health 2019;11:287-99. doi: 10.2147/IJWH. S197604.
8. Sun Z-L, Tang Y-J, Wu W-G, et al. AZD1480 can inhibit the biological behavior of ovarian cancer SKOV3 cells in vitro. Asian Pac J Cancer Prev 2013;14:4823-7. doi: 10.7314/ apjcp.2013.14.8.4823.

9. Elbe H, Yigitturk G, Cavusoglu T, Baygar T, Ozgul Onal M, Ozturk F. Comparison of ultrastructural changes and the anticarcinogenic effects of thymol and carvacrol on ovarian cancer cells: which is more effective? Ultrastruct Pathol 2020;44:193-202. doi: 10.1080/01913123.2020.1740366.
10. Achmad AB, Proboningrat A, Ansori ANM, et al. Stem bark ethanolic extract of Pinus merkusii induces caspase 9-mediated apoptosis in HeLa cells. Open Vet J 2024;14:2628- 33. doi: 10.5455/OVJ.2024.v14.i10.12.
11. Proboningrat A, Ansori ANM, Fadholly A, Putri N, Kusala MKJ, Achmad AB. First report on the cytotoxicity of pinus merkusii bark extract in widr, a human colon carcinoma cell line. Res J Pharm Technol.2021;14:1685-8. doi: 10.5958/0974- 360X.2021.00299.7
12. Sobral MV, Xavier AL, Lima TC, de Sousa DP. Antitumor activity of monoterpenes found in essential oils. ScientificWorldJournal 2014;2014:953451. doi: 10.1155/2014/953451.
13. Yesil-Celiktas O, Ganzera M, Akgun I, Sevimli Gür C, Korkmaz K, Bedir E. Determination of polyphenolic constituents and biological activities of bark extracts from different Pinus species. J Sci Food Agric 2009;89:1339-45. doi.org/10.1002/ JSFA.3591
14. Packer L, Rimbach G, Virgili F. Antioxidant activity and biologic properties of a procyanidin-rich extract from pine (Pinus maritima) bark, pycnogenol. Free Radic Biol Med 1999;27:704-24. doi: 10.1016/s0891-5849(99)00090-8.
15. Romani A, Ieri F, Turchetti B, Mulinacci N, Vincieri FF, Buzzini P. Analysis of condensed and hydrolysable tannins from commercial plant extracts. J Pharm Biomed Anal 2006;41:415-20. doi: 10.1016/j.jpba.2005.11.031.
16. D’Andrea G. Pycnogenol: a blend of procyanidins with multifaceted therapeutic applications? Fitoterapia 2010;81:724-36. doi: 10.1016/j.fitote.2010.06.011.
17. Maimoona A, Naeem I, Saddiqe Z, Jameel K. A review on biological, nutraceutical and clinical aspects of French maritime pine bark extract. J Ethnopharmacol 2011 ;133:261- 77. doi: 10.1016/j.jep.2010.10.041.
18. Apetrei CL, Tuchilus C, Aprotosoaie AC, Oprea A, Malterud KE, Miron A. Chemical, antioxidant and antimicrobial investigations of Pinus cembra L. bark and needles. Molecules 2011;16:7773-88. doi: 10.3390/molecules16097773.
19. Ku S, Mun SP. Antioxidant properties of monomeric, oligomeric, and polymeric fractions in hot water extract from Pinus radiata bark. Wood Sci Technol 2007 Jan;42:47–60. doi: 10.1007/s00226-007-0150-9.
20. Yu L, Zhao M, Wang J shui, et al. Antioxidant, immunomodulatory and anti-breast cancer activities of phenolic extract from pine (Pinus massoniana Lamb) bark. Innov Food Sci Emerg Technol 2008;9:122-8. doi. org/10.1016/j.ifset.2007.06.006
21. Sebaugh JL. Guidelines for accurate EC50/IC50 estimation. Pharm Stat 2011 ;10:128-34. doi: 10.1002/pst.426.
22. Dodurga Y, Seçme M, Eroğlu C, et al. Investigation of the effects of a sulfite molecule on human neuroblastoma cells via a novel oncogene URG4/URGCP. Life Sci 2015 ;143:27-34. doi: 10.1016/j.lfs.2015.10.005.
23. Fahrioğlu U, Dodurga Y, Elmas L, Seçme M. Ferulic acid decreases cell viability and colony formation while inhibiting migration of MIA PaCa-2 human pancreatic cancer cells in vitro. Gene 2016;576(1 Pt 3):476-82. doi: 10.1016/j. gene.2015.10.061.
24. Eroğlu C, Seçme M, Bağcı G, Dodurga Y. Assessment of the anticancer mechanism of ferulic acid via cell cycle and apoptotic pathways in human prostate cancer cell lines. Tumour Biol 2015;36:9437-46. doi: 10.1007/s13277-015- 3689-3
25. Seçme M, Eroğlu C, Dodurga Y, Bağcı G. Investigation of anticancer mechanism of oleuropein via cell cycle and apoptotic pathways in SH-SY5Y neuroblastoma cells. Gene 2016;585:93-9. doi: 10.1007/s13277-015-3689-3
26. Esmaeili-Mahani S, Falahi F, Yaghoobi MM. Proapoptotic and antiproliferative effects of thymus caramanicus on human breast cancer cell line (MCF-7) and ıts ınteraction with anticancer drug vincristine. Evid Based Complement Alternat Med 2014;2014:893247. doi: 10.1155/2014/893247.
27. Piccart MJ, Bertelsen K, Stuart G, et al. Long-term follow-up confirms a survival advantage of the paclitaxel-cisplatin regimen over the cyclophosphamide-cisplatin combination in advanced ovarian cancer. Int J Gynecol Cancer 2003;13 Suppl 2:144-8. doi: 10.1111/j.1525-1438.2003.13357.x.
28. Aggarwal BB, Shishodia S. Molecular targets of dietary agents for prevention and therapy of cancer. Biochem Pharmacol 2006;71:1397-421. doi: 10.1016/j.bcp.2006.02.009.
29. Arunasree KM. Anti-proliferative effects of carvacrol on a human metastatic breast cancer cell line, MDA-MB 231. Phytomedicine 2010;17:581-8. doi: 10.1016/j. phymed.2009.12.008.
30. Şahin F, Avcı ÇB, Avcu F, et al. Red grape seed extract and its compound resveratrol exert cytotoxic effect to various lines. human cancer lines. Turk J Hematol 2007;24:102-9.
31. Alexa E, Sumalan RM, Danciu C, et al. Synergistic antifungal, allelopatic and anti-proliferative potential of salvia officinalis L., and thymus vulgaris l. essential oils. Molecules 2018;23:185. doi: 10.3390/molecules23010185.
32. Palabiyik SS, Karakus E, Halici Z, et al. The protective effects of carvacrol and thymol against paracetamol-induced toxicity on human hepatocellular carcinoma cell lines (HepG2). Hum Exp Toxicol 2016;35:1252-63. doi: 10.1177/0960327115627688.
33. Frontela-Saseta C, López-Nicolás R, González-Bermúdez CA, Martínez-Graciá C, Ros-Berruezo G. Anti-inflammatory properties of fruit juices enriched with pine bark extract in an in vitro model of inflamed human intestinal epithelium: The effect of gastrointestinal digestion. Food Chem Toxicol 2013;53:94-9. doi: 10.1016/j.fct.2012.11.024.
34. Cretu E, Karonen M, Salminen J-P, et al. In vitro study on the antioxidant activity of a polyphenol-rich extract from Pinus brutia bark and its fractions. J Med Food 2013;16:984-91. doi: 10.1089/jmf.2013.0050
35. Rao YK, Geethangili M, Fang SH, Tzeng YM. Antioxidant and cytotoxic activities of naturally occurring phenolic and related compounds: A comparative study. Food Chem Toxicol 2007;45:1770-6. doi: 10.1016/j.fct.2007.03.012.
36. Mao P, Zhang E, Chen Y, et al. Pinus massoniana bark extract inhibits migration of the lung cancer A549 cell line. Oncol Lett 2017;13:1019-23. doi: 10.3892/ol.2016.5509.
37. Ma H, Liu B, Feng D, et al. Pinus massoniana bark extract selectively induces apoptosis in human hepatoma cells, possibly through caspase-dependent pathways. Int J Mol Med 2010;25:751-9. doi: 10.3892/ijmm_00000401.
38. Ma H, Lai F, Xie H, Wang J, Wang H. Involvement of the Bcl-2 family members in Pinus massoniana bark extract induced apoptosis in HeLa cells. Phytother Res 2008 ;22:1472-6. doi: 10.1002/ptr.2496.
39. Meeran SM, Katiyar SK. Grape seed proanthocyanidins promote apoptosis in human epidermoid carcinoma A431 cells through alterations in Cdki-Cdk-cyclin cascade, and caspase-3 activation via loss of mitochondrial membrane potential. Exp Dermatol 2007 ;16:405-15. doi: 10.1111/j.1600- 0625.2007.00542.x.
40. Cui Y-Y, Xie H, Qi K-B, He Y-M, Wang J-F. Effects of Pinus massoniana bark extract on cell proliferation and apoptosis of human hepatoma BEL-7402 cells. World J Gastroenterol 2005;11:5277-82. doi: 10.3748/wjg.v11.i34.5277.
41. Chen Q, Peng W, Xu A. Apoptosis of a human non-small cell lung cancer (NSCLC) cell line, PLA-801, induced by acutiaporberine, a novel bisalkaloid derived from Thalictrum acutifolium (Hand.-Mazz.) Boivin. Biochem Pharmacol 2002;63:1389-96. doi: 10.1016/s0006-2952(02)00871-7.
42. Deb DD, Parimala G, Saravana Devi S, Chakraborty T. Effect of thymol on peripheral blood mononuclear cell PBMC and acute promyelotic cancer cell line HL-60. Chem Biol Interact 2011;193:97-106. doi: 10.1016/j.cbi.2011.05.009.
43. Huynh HT, Teel RW. Selective induction of apoptosis in human mammary cancer cells (MCF-7) by pycnogenol. Anticancer Res 2000;20:2417-20.
44. Huang WW, Yang JS, Lin CF, Ho WJ, Lee MR. Pycnogenol induces differentiation and apoptosis in human promyeloid leukemia HL-60 cells. Leuk Res 2005;29:685-92. doi: 10.1016/j. leukres.2004.10.006.
45. Zalewski M, Kulbacka J, Saczko J, Drag-Zalesinska M, Choromanska A. Valspodar-modulated chemotherapy in human ovarian cancer cells SK-OV-3 and MDAH- 2774. Bosn J Basic Med Sci 2019;19:234-41. doi: 10.17305/ bjbms.2019.4073.
46. Qi Y, Li X, Zhao S, Han Y. Value of porous titanium alloy plates for chest wall reconstruction after resection of chest wall tumors. Asian Pac J Cancer Prev 2014;15:4535-8. doi: 10.7314/ apjcp.2014.15.11.4535.
47. Sajid A, Manzoor Q, Iqbal M, Tyagi AK, Sarfraz RA, Sajid A. Pinus Roxburghii essential oil anticancer activity and chemical composition evaluation. EXCLI J 2018;17:233-45. doi: 10.17179/excli2016-670.
48. Ren P, Ren X, Cheng L, Xu L. Frankincense, pine needle and geranium essential oils suppress tumor progression through the regulation of the AMPK/mTOR pathway in breast cancer. Oncol Rep 2018;39:129-37. doi: 10.3892/or.2017.6067.