Meme kanseri kök hücrelerinde elajik asit ile indüklenmiş miRNA’ların ifadesi ve elajik asidin apoptoz üzerine etkisi

Hasan Onur Çağlar; Sunde Yılmaz Süslüer; Şebnem Kavaklı; Cumhur Gündüz; Biray Ertürk; Ferda Özkınay; Ayfer Haydaroğlu

doi:10.19161/etd.399234

EN TR

Ellagic acid induced expression of miRNAs in breast cancer stem cells and effect of ellagic acid on apoptosis

Abstract

Aim: miRNAs which are associated with estrogen dependent tumorigenesis of breast cancer are suppressed by ellagic acid (EA) treatment. However, modulation of expression profiles of miRNAs in breast cancer stem cells (BCSCs) after EA treatment is still unclear. In this study, it was aimed to show EA-induced apoptosis in BCSCs and to determine altered expression profiles of miRNAs after EA treatment.

Materials and Methods: Cytotoxic effects of EA on BCSCs were examined by WST-1 reagent test. Apoptosis and cell cycle analysis after EA treatment were detected by flow cytometry analysis. After EA treatment, miRNA expression profiles of BCSCs were determined by RT-PCR miRNA array.

Results: Cytotoxic effect of EA on BCSCs was found. IC50 concentration of EA at 48th and 72nd hours was 24.8 μM. EA did not induce apoptosis in BCSCs. In addition, S phase arrest was observed at 24th, 48th, 72nd hours of EA treatment. Expression profiles of 76 of totally 84 miRNA genes were detected. Expression of hsa-miR-125b-1-3p was increased as 3.59 folds by EA treatment. Expression profiles of all the others miRNAs, including members of Let-7 and miR-200 families, showed a decrease due to EA treatment.

Conclusion: In this study, it was observed that EA leads to upregulated expression of hsa-miR-125b-1-3p in contrast to downregulated expression of all other miRNAs, especially hsa-miR-485-5p and hsa-miR-328-3p, and does not induce apoptosis in BCSCs. It was suggested that BCSCs may show resistance to EA treatment due to aberrant expression profiles of miRNAs.

Keywords

Ellagic acid,breast cancer stem cells,miRNAs,apoptosis

Meme kanseri kök hücrelerinde elajik asit ile indüklenmiş miRNA’ların ifadesi ve elajik asidin apoptoz üzerine etkisi

Öz

Amaç: Östrojen bağımlı meme kanseri tümör oluşumunda etkili miRNA’lar elajik asit (EA) uygulaması ile baskılanmaktadır. Ancak, EA uygulamasından sonra meme kanseri kök hücrelerindeki (MKKH) miRNA’ların ifade profillerinin düzenlenmesi hala belirsizdir. Bu çalışmada, MKKH’lerde EA-indüklenmiş apoptozun gösterilmesi ve EA uygulamasından sonra değişen miRNA ifade profilinin belirlenmesi amaçlanmıştır.

Gereç ve Yöntem: EA’nın MKKH’lerdeki sitotoksik etkisi WST-1 testi ile incelendi. EA uygulamasından sonra apoptoz ve hücre döngüsü analizleri flow sitometri ile yapıldı. EA uygulamasından sonra, MKKH’lerin miRNA ifade profilleri RT-PCR miRNA array ile tanımlandı.

Bulgular: MKKH’ler üzerinde EA’nın sitotoksik etkisi saptandı. 48. ve 72. saatlerde EA’nın IC50 konsantrasyonu 24.8 μM’dı. EA, MKKH’lerde apoptozu indüklemedi. Bununla beraber, EA’nın 24., 48. ve 72. saat uygulamalarında S fazı bloğu gözlendi. Toplamda 84 miRNA geninin 76’sının ifade profili belirlendi. hsa-miR-125b-1-3p’nin ifadesi EA uygulanmasıyla 3.59 kat arttı. Let-7 ve miR-200 aileleri dahil diğer tüm miRNA’ların ifade profili EA uygulaması nedeniyle azalış gösterdi.

Sonuç: Bu çalışmada, EA’nın MKKH’de apoptozu etkilemediği ve hsa-miR-125b-1-3p’in ifadesini arttırırken, başta hsa-miR-485-5p ve hsa-miR-328-3p olmak üzere diğer miRNA’ların ifadesinde düşmeye neden olduğu görüldü. Anormal miRNA ifade profili nedeniyle MKKH’lerin EA uygulamasına direnç gösterebileceği düşünüldü.

Anahtar Kelimeler

Elajik asit,meme kanseri kök hücreleri,miRNA,apoptoz

Kaynakça

Siegel RL, Miller KD, Jemal A. Cancer statistics, 2015. CA Cancer J Clin 2015;65(1):5-29.
Liu H, Liu Y, Zhang JT. A new mechanism of drug resistance in breast cancer cells: Fatty acid synthase overexpression-mediated palmitate overproduction. Mol Cancer Ther 2008;7(2):263-70.
Barok M, Joensuu H, Isola J. Trastuzumab emtansine: Mechanisms of action and drug resistance. Breast Cancer Res 2014;16(2):209.
Hayes EL, Lewis-Wambi JS. Mechanisms of endocrine resistance in breast cancer: An overview of the proposed roles of noncoding RNA. Breast Cancer Res 2015;17:40.
Weigelt B, Peterse JL, van 't Veer LJ. Breast cancer metastasis: Markers and models. Nat Rev Cancer 2005;5(8):591-602.
Lee J, Kim YS, Lee J, et al. Walnut phenolic extract and Its bioactive compounds suppress colon cancer cell growth by regulating colon cancer stemness. Nutrients 2016;8(7):E439.
Ambigaipalan P, de Camargo AC, Shahidi F. Phenolic compounds of pomegranate byoproducts (outer skin, mesocarp, divider membrane) and their antioxidant activities. J Agric Food Chem 2016;64(34):6584-604.
Hacke AC, Granato D, Maciel LG, et al. Jabuticaba (Myrciaria cauliflora) seeds: Chemical characterization and extraction of antioxidant and antimicrobial compounds. J Food Sci doi: 10.1111/1750-3841.13405.

Saleem A, Husheem M, Harkonen P, Pihlaja K. Inhibition of cancer cell growth by crude extract and the phenolics of Terminalia chebula retz. fruit. J Ethnopharmacol 2002;81(3):327-36.
Wang Z, Loo WT, Wang N, et al. Effect of Sanguisorba officinalis L on breast cancer growth and angiogenesis. Expert Opin Ther Targets 2012;16 (Suppl 1):79-89.
Guruvayoorappan C, Kuttan G. (+)-Catechin inhibits tumour angiogenesis and regulates the production of nitric oxide and TNF-alpha in LPS-stimulated macrophages. Innate immunity 2008;14(3):160-74.
Iniguez-Franco F, Soto-Valdez H, Peralta E, Ayala-Zavala JF, Auras R, Gamez-Meza N. Antioxidant activity and diffusion of catechin and epicatechin from antioxidant active films made of poly(L-lactic acid). J Agric Food Chem 2012;60(26):6515-23.
Siraj MA, Shilpi JA, Hossain MG, et al. Anti-inflammatory and antioxidant activity of Acalypha hispida leaf and analysis of its major bioactive polyphenols by HPLC. Adv Pharm Bull 2016;6(2):275-83.
Chen HS, Bai MH, Zhang T, Li GD, Liu M. Ellagic acid induces cell cycle arrest and apoptosis through TGF-beta/Smad3 signaling pathway in human breast cancer MCF-7 cells. Int J Oncol 2015;46(4):1730-8.
Shi L, Gao X, Li X, et al. Ellagic acid enhances the efficacy of PI3K inhibitor GDC-0941 in breast cancer cells. Curr Mol Med 2015;15(5):478-86.
Zhang T, Chen HS, Wang LF, et al. Ellagic acid exerts anti-proliferation effects via modulation of Tgf-beta/Smad3 signaling in MCF-7 breast cancer cells. Asian Pac J Cancer Prev 2014;15(1):273-6.
Wang N, Wang ZY, Mo SL, et al. Ellagic acid, a phenolic compound, exerts anti-angiogenesis effects via VEGFR-2 signaling pathway in breast cancer. Breast Cancer Res Treat 2012;134(3):943-55.
Dawood S, Austin L, Cristofanilli M. Cancer stem cells: Implications for cancer therapy. Oncology (Williston Park) 2014;28(12):1101-7.
Salvador MA, Birnbaum D, Charafe-Jauffret E, Ginestier C. Breast cancer stem cells programs: enter the (non)-code. Brief Funct Genomics 2016;15(3):186-99.
Yu F, Yao H, Zhu P, et al. Llet-7 regulates self renewal and tumorigenicity of breast cancer cells. Cell 2007;131(6):1109-23.
Shimono Y, Mukohyama J, Nakamura S, Minami H. MicroRNA regulation of human breast cancer stem cells. J Clin Med doi: 10.3390/jcm5010002.
Munagala R, Aqil F, Vadhanam MV, Gupta RC. MicroRNA ‘signature’during estrogen-mediated mammary carcinogenesis and its reversal by ellagic acid intervention. Cancer Lett 2013;339(2):175-84.
Vlachos IS, Kostoulas N, Vergoulis T, et al. DIANA miRPath v.2.0: Investigating the combinatorial effect of microRNAs in pathways. Nucleic Acids Res 2012 [cited 04 Sept 2016]. Available from: http://nar.oxfordjournals.org/content/40/W1/W498.long
Atta Ur R, Ngounou FN, Choudhary MI, et al. New antioxidant and antimicrobial ellagic acid derivatives from Pteleopsis hylodendron. Planta Med 2001;67(4):335-9.
Venkataramanamma D, Aruna P, Singh RP. Standardization of the conditions for extraction of polyphenols from pomegranate peel. J Food Sci Technol 2016;53(5):2497-503.
Adams LS, Zhang Y, Seeram NP, Heber D, Chen S. Pomegranate ellagitannin-derived compounds exhibit antiproliferative and antiaromatase activity in breast cancer cells in vitro. Cancer Prev Res 2010;3(1):108-13.
Strati A, Papoutsi Z, Lianidou E, Moutsatsou P. Effect of ellagic acid on the expression of human telomerase reverse transcriptase (hTERT) alpha+beta+transcript in estrogen receptor-positive MCF-7 breast cancer cells. Clin Biochem 2009;42(13-14):1358-62.
Dai Z, Nair V, Khan M, Ciolino HP. Pomegranate extract inhibits the proliferation and viability of MMTV-Wnt-1 mouse mammary cancer stem cells in vitro. Oncol Rep 2010;24(4):1087-91.
Kumar D, Basu S, Parija L, et al. Curcumin and ellagic acid synergistically induce ROS generation, DNA damage, p53 accumulation and apoptosis in HeLa cervical carcinoma cells. Biomed Pharmacother 2016;81(1):31-7.
Mertens-Talcott SU, Percival SS. Ellagic acid and quercetin interact synergistically with resveratrol in the induction of apoptosis and cause transient cell cycle arrest in human leukemia cells. Cancer Lett 2005;218(2):141-51.
Garofalo M, Croce CM. Role of microRNAs in maintaining cancer stem cells. Adv Drug Deliv Rev 2015;81:53-61.
Ali Hosseini Rad SM, Bavarsad MS, Arefian E, Jaseb K, Shahjahani M, Saki N. The Role of microRNAs in Stemness of Cancer Stem Cells. Oncol Rev 2013;7(1):e8.
Zhou N, Mo YY. Roles of microRNAs in cancer stem cells. Front Biosci (Schol Ed) 2012;1(4):810-8.
Gomes BC, Rueff J, Rodrigues AS. MicroRNAs and cancer drug resistance. Methods Mol Biol 2016;1395:137-62.
Chen W, Zhou S, Mao L, et al. Crosstalk between TGF-beta signaling and miRNAs in breast cancer metastasis. Tumour Biol 2016;37(8):10011-9.
Hu X, Guo J, Zheng L, et al. The heterochronic microRNA let-7 inhibits cell motility by regulating the genes in the actin cytoskeleton pathway in breast cancer. Mol Cancer Res 2013;11(3):240-50.
Iliopoulos D, Hirsch HA, Struhl K. An epigenetic switch involving NF-kappaB, Lin28, Let-7 MicroRNA, and IL6 links inflammation to cell transformation. Cell 2009;139(4):693-706.
Iliopoulos D, Hirsch HA, Wang G, Struhl K. Inducible formation of breast cancer stem cells and their dynamic equilibrium with non-stem cancer cells via IL6 secretion. Proc Natl Acad Sci USA 2011;108(4):1397-402.
Melton C, Judson RL, Blelloch R. Opposing microRNA families regulate self-renewal in mouse embryonic stem cells. Nature 2010;463(7281):621-6.
Sun H, Ding C, Zhang H, Gao J. Let7 miRNAs sensitize breast cancer stem cells to radiationinduced repression through inhibition of the cyclin D1/Akt1/Wnt1 signaling pathway. Mol Med Rep 2016 doi: 10.3892/mmr.2016.5656
Zhao B, Chen YG. Regulation of TGF-beta Signal Transduction. Scientifica 2014;2014:874065.
Humphries B, Yang C. The microRNA-200 family: small molecules with novel roles in cancer development, progression and therapy. Oncotarget 2015;6(9):6472-98.
Gill JG, Langer EM, Lindsley RC, et al. Snail and the microRNA-200 family act in opposition to regulate epithelial-to-mesenchymal transition and germ layer fate restriction in differentiating ESCs. Stem cells 2011;29(5):764-76.
Wang G, Guo X, Hong W, et al. Critical regulation of miR-200/ZEB2 pathway in Oct4/Sox2-induced mesenchymal-to-epithelial transition and induced pluripotent stem cell generation. Proc Natl Acad Sci USA 2013;110(8):2858-63.
Shimono Y, Zabala M, Cho RW, et al. Downregulation of miRNA-200c links breast cancer stem cells with normal stem cells. Cell 2009;138(3):592-603.

Ayrıntılar

Birincil Dil

Türkçe

Konular

Sağlık Kurumları Yönetimi

Bölüm

Araştırma Makalesi

Yazarlar

Hasan Onur Çağlar ^*
Ege Üniversitesi, Sağlık Bilimleri Enstitüsü, Kök Hücre Anabilim Dalı, İzmir
0000-0002-3637-4755
Türkiye

Sunde Yılmaz Süslüer
Ege Üniversitesi Tıp Fakültesi, Tıbbi Biyoloji Anabilim Dalı, İzmir
0000-0002-0535-150X
Türkiye

Şebnem Kavaklı
Ege Üniversitesi, Fen Bilimleri Enstitüsü, Biyoteknoloji Anabilim Dalı, İzmir
0000-0001-9949-2305
Türkiye

Cumhur Gündüz
Ege Üniversitesi Tıp Fakültesi, Tıbbi Biyoloji Anabilim Dalı, İzmir
0000-0002-6593-3237
Türkiye

Biray Ertürk
Ege Üniversitesi Tıp Fakültesi, Tıbbi Genetik Anabilim Dalı, İzmir
0000-0002-0348-6267
Türkiye

Ferda Özkınay
Ege Üniversitesi Tıp Fakültesi, Tıbbi Genetik Anabilim Dalı, İzmir
0000-0001-7542-7787
Türkiye

Ayfer Haydaroğlu
Ege Üniversitesi Tıp Fakültesi, Radyasyon Onkolojisi Anabilim Dalı
0000-0001-5709-0981
Türkiye

Yayımlanma Tarihi

1 Aralık 2017

Gönderilme Tarihi

8 Eylül 2016

Kabul Tarihi

14 Aralık 2016

Yayımlandığı Sayı

Yıl 2017 Cilt: 56 Sayı: 4

DOI

https://doi.org/10.19161/etd.399234

IZ

https://izlik.org/JA86LH88JE

Kaynak Göster

RIS / Bibtex

Vancouver

1.Hasan Onur Çağlar, Sunde Yılmaz Süslüer, Şebnem Kavaklı, Cumhur Gündüz, Biray Ertürk, Ferda Özkınay, Ayfer Haydaroğlu. Meme kanseri kök hücrelerinde elajik asit ile indüklenmiş miRNA’ların ifadesi ve elajik asidin apoptoz üzerine etkisi. ETD. 01 Aralık 2017;56(4):183-92. doi:10.19161/etd.399234

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