Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin

İNCESU, Zerrin; KILIÇASLAN, Seda Mehtap SARI; AYRIM, Aysun; APAYDIN, ELİF

doi:10.4274/tjps.35220

Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin

Seda Mehtap SARI KILIÇASLAN, (Anadolu Üniversitesi, Eğitim Fakültesi, İlköğretim Bölümü, Eskişehir, Türkiye)

Aysun AYRIM, (Eskişehir Osmangazi Üniversitesi, Yaşam Bilimleri Enstitüsü, Eskişehir, Türkiye)

Elif APAYDIN, (Anadolu Üniversitesi, Yaşam Bilimleri Enstitüsü, Eskişehir, Türkiye)

Zerrin İNCESU (Anadolu Üniversitesi, Eczacılık Fakültesi, Biyokimya Anabilim Dalı, Eskişehir, Türkiye)

Turkish Journal of Pharmaceutical Sciences

0 0

Yıl: 2018 Cilt: 15 Sayı: 1 Sayfa Aralığı: 50 - 56 Metin Dili: İngilizce DOI: 10.4274/tjps.35220 İndeks Tarihi: 19-08-2020

Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin

Öz:

Objectives: To investigate the effects of intracellular calcium (Ca2+) mobilization, β-catenin and Akt signal pathways after the binding of metastaticovarian cells to fibronectin.Materials and Methods: The expression levels of α4β1 and αvβ6 integrin were determined using α4, β1, αv, and β6 antibodies using flow cytometryon PEO-1 cells. The effect of [Ca2+]i on cell adhesion capacity was investigated using RTCA after stimulating PEO-1 cells using thapsigargin andtunicamycin. The binding rate of PEO-1 cells to fibronectin was also investigated in the presence of either different concentrations of cardamonin,which inhibits the accumulation of β-catenin, or different concentrations of FPA 124, which is a specific inhibitor for the PKB/Akt signal pathway,using RTCA.Results: RTCA analysis results showed that increasing [Ca2+]i through leakage of the calcium pool was strongly effective on PEO-1 cell binding tofibronectin. Extracellular calcium influx also reduced the binding of PEO-1 cells. Cell binding to fibronectin was also inhibited with a ratio of 64%in the presence of 100 µM cardamonin compared with untreated control cells. Finally, it was found that PKB/Akt inhibition with 15 µM FPA 124decreased the binding of PEO-1 cells to fibronectin with a ratio of 88% compared with untreated control cells.Conclusion: PEO-1 cell binding to fibronectin via integrins could be related to intracellular Ca2+ mobilization and Akt signaling.

Anahtar Kelime:

Ca2+ Mobilizasyonu ve Akt İnhibisyonu ile PEO-1 Hücrelerinin Fibronektine Bağlanmasında Azalma

Öz:

Amaç: Metastatik ovaryum kanser hücrelerinin fibronektine bağlanmasından sonra, hücre içi Ca2+ mobilizasyonu, β-katenin ve Akt sinyal yollarının etkilerinin araştırılmasıdır. Gereç ve Yöntemler: α4β1 ve αvβ6 integrinlerin ekspresyon düzeyleri α4, β1, αv vçe β6 antikorları kullanılarak akım sitometrisi ile PEO-1 hücrelerinde tespit edilmiştir. [Ca2+]’un hücre adezyon kapasitesine etkisi, PEO-1 hücrelerinin tapsigargin ve tunikamisin ile uyarılmasından sonra RTCA kullanılarak araştırıldı. PEO-1 hücrelerinin fibronektine bağlanma oranı β-kateninin toplanmasını inhibe eden farklı kardamonin konsantrasyonları varlığında veya PKB/Akt sinyal yolu için spesifik inhibitör olan FPA 124’ün farklı konsantrasyonları için RTCA kullanılarak araştırıldı. Bulgular: RTCA analiz sonuçları kalsiyum depolarından sızmasıyla artan [Ca2+]’un PEO-1 hücrelerinin fibronektine bağlanmasını güçlü bir şekilde etkilediğini göstermiştir. Hücre dışı kalsiyum akımı PEO-1 hücrelerinin bağlanmasını azaltmıştır. 100 µM kardamonin varlığında muamele edilmemiş kontrol hücrelerine göre hücrenin fibronektine bağlanması %64 oranında inhibe olmuştur. Son olarak, 15 µM FPA 124 ile PKB/Akt inhibisyonu da PEO-1 hücrelerinin fibronektine bağlanmasını %88 oranında düşürdüğü bulunmuştur. Sonuç: PEO-1 hücrelerinin integrinler aracılığı ile fibronektine bağlanmasının hücre içi Ca2+ mobilizasyonu ve Akt sinyali ile ilgili olabilir.

Anahtar Kelime:

Belge Türü: Makale Makale Türü: Araştırma Makalesi Erişim Türü: Erişime Açık

1. Manso AM, Kang SM, Ross RS. Integrins, Focal Adhesions and Cardiac Fibroblasts. J Investig Med. 2009;57:856-860.
2. Aoudjit F, Vuori K. Integrin Signalling in Cancer Cell Survival and Chemoresistance. Chemotherapy Res and Pract. 2012;2012:1-16.
3. Hirohashi S, Kanai Y. Cell adhesion system and human cancer morphogenesis. Cancer Sci. 2003;94:575-581.
4. van den Hoogen C, van der Horst G, Cheung H, Buijs JT, Pelger RC, van der Pluijm G. Integrin αv expression is required for the acquisition of a metastatic stem/progenitor cell phenotype in human prostate cancer. Am J Pathol. 2011;179:2559-2568.
5. Paoli P, Giannoni E, Chiarugi P. Anoikis molecular pathways and its role in cancer progression. Biochimica et Biophysica Acta-Molecular Cell Research. 2013;1833:3481-3498.
6. Lu JG, Li Y, Li L, Kan X. Overexpression of osteopontin and integrin αv in laryngeal and hypopharyngeal carcinomas associated with differentiation and metastasis. J Cancer Res Clin Oncol. 2011;137:1613-1618.
7. Puiffe ML, Le Page C, Filali-Mouhim A, Zietarska M, Ouellet V, Tonin PN, Chevrette M, Provencher DM, Mes-Masson AM. Characterization of ovarian cancer ascites on cell invasion, proliferation, spheroid formation, and gene expression in an in vitro model of epithelial ovarian cancer. Neoplasia. 2007;9:820-829.
8. Kenny HA, Kaur S, Coussens LM, Lengyel E. The initial steps of ovarian cancer cell metastasis are mediated by MMP-2 cleavage of vitronectin and fibronectin. J Clin Invest. 2008;118:1367-1379.
9. Bodnar L, Stanczak A, Cierniak S, Smoter M, Cichowicz M, Kozlowski W, Szczylik C, Wieczorek M, Lamparska-Przybysz M. Wnt/β-catenin pathway as a potential prognostic and predictive marker in patients with advanced ovarian cancer. J of Ovarian Research. 2014;7:16-26.
10. Hur J, Jeong S. Multitasking β-catenin: From adhesion and transcription to RNA regulation. Animal Cell Systems. 2013;17:299-305.
11. Uchino M, Kojima H, Wada K. Nuclear beta-catenin and CD44 upregulation characterize invasive cell populations in non-aggressive MCF-7 breast cancer cell. BMC Cancer. 2010;10:414-417.
12. Chau WK, Ip CK, Mak AS. C-Kit mediates chemoresistance and tumorinitiating capacity of ovarian cancer cells through activation of Wnt/betacatenin-ATP-binding cassette G2 signaling. Oncogene. 2012;10:103-107.
13. Tran NL, Adams DG, Vaillancourt RR, Heimark RL. Signal transduction from N-cadherin increases Bcl-2. Regulation of the phosphatidylinositol 3-kinase/Akt pathway by homophilic adhesion and actin cytoskeletal organization. J Biol Chem. 2002;277:32905-32914.
14. Sharma M, Chuang WW, Sun Z. Phosphatidylinositol 3-kinase/Akt stimulates androgen pathway through GSK3beta inhibition and nuclear beta-catenin accumulation. J Biol Chem. 2002;277:30935-30941.
15. Rahim S, Üren A. A real time electrical impedance based technique to measure invasion of endothelial cell monolayer by cancer cells. J Vis Exp. 2011;50:2792-2797.
16. Zhang JD. 2016. Available from: https://www.bioconductor.org/ packages/devel/bioc/vignettes/RTCA/inst/doc/aboutRTCA.pdf.
17. Sawada K, Mitra AK, Radjabi AR, Bhaskar V, Kistner EO. Tretiakova M, Jagadeeswaran S, Montag A, Becker A, Kenny HA. Loss of E-cadherin promotes ovarian cancer metastasis via α 5-integrin, which is a therapeutic target. Cancer Res. 2008;68:2329-2339.
18. Strobel T, Cannistra S. B1-Integrins partly mediated binding of ovarian cancer cells to peritoneal mesothelium in vitro. Gynecologic Oncology. 1999;73:362-367.
19. Carduner L, Leroy-Dudal J, Picot CR, Gallet O, Carreiras F, Kellouche S. Ascites-induced shift along epithelial-mesenchymal spectrum in ovarian cancer cells: enhancement of their invasive behaviour partly dependant on av integrins. Clinical and Experimental Metastasis. 2014;31:675-688.
20. Lessan K, Aguiar DJ, Oegema T, Siebenson L, Skubitz AP. CD44 and β1 integrin mediate ovarian carcinoma cell adhesion to peritoneal mesothelial cells. Am J Pathol. 1999;154:1525-1537.
21. Bignotti E, Tassi RA, Calza S, Ravaggi A, Bandiera E, Rossi E, Donzelli C, Pasinetti B, Pecorelli S, Santin AD. Gene expression profile of ovarian serous papillary carcinomas: identification of metastasis-associated genes. Am J Obstet Gynecol. 2007;196:245.
22. Wilhelm O, Hafter R, Coppenrath E, Pflanz M, Schmitt M, Graeff H. Fibrin-fibronectin compounds in human ovarian tumor ascites and their possible relation to the tumor stroma. Cancer Res. 1988;48:3507-3514.
23. Mitra AK, Sawada K, Tiwari P, Mui K, Gwin K, Lengyel E. Ligandindependent activation of c-Met by fibronectin and α(5)β(1)-integrin regulates ovarian cancer invasion and metastasis. Oncogene. 2011;30:1566-1576.
24. Tsaktanis T, Kremling H, Pavsic M, von Stackelberg R, Mack B, Fukumori A, Steiner H, Vielmuth F, Spindler V, Huang Z, Jakubowski J, Stoecklein NH, Luxenburger E, Lauber K, Lenarcic B, Gires O. Cleavage and cell adhesion properties of human epithelial cell adhesion molecule (HEPCAM). The Journal of Biological Chemistry. 2015;290:24574-24591.
25. Giorgi C, Bonova M, Pinton P. Inside the tumor: p53 modulates calcium homeostasis. Cell Cycle. 2015;147:933-934.
26. Nami B, Donmez H, Kocak N. Tunicamycin-induced endoplasmic reticulum stress reduces in vitro subpopulation and invasion of CD44+/CD24- phenotype breast cancer stem cells. Exp Toxicol Pathol. 2016;68:418-426.
27. Hwang HJ, Chung HS, Jung TW, Ryu JY, Hong HC, Seo JA, Kim SG, Kim NH, Choi KM, Choi DS, Baik SH, Yoo HJ. The dipeptidyl peptidaseIV inhibitor inhibits the expression of vascular adhesion molecules and inflammatory cytokines in HUVECs via Akt- and AMPK-dependent mechanisms. Mol Cell Endocrinol. 2015;405:25-34.
28. Leung HW, Wang Z, Yue GG, Zhao SM, Lee JK, Fung KP, Leung PC, Lau CB, Tan NH. Cyclopeptide RA-V inhibits cell adhesion and invasion in both estrogen receptor positive and negative breast cancer cells via PI3K/AKT and NF-κB signaling pathways. Biochim Biophys Acta. 2015;1853:1827-1840.
29. Piegeler T, Schlapfer M, Dulli RO, Schwartz DE, Borgeat A, Minshall RD, Beck-Schimmer B. Clinically relevant concentrations of lidocaine and ropivacaine inhibit TNFa-induced invasion of lung adenocarcinoma cells in vitro by blocking the activation of Akt and focal adhesion kinase. Br J Anaesth. 2015;115:784-791.
30. Gwak H, Kim S, Dhanasekaran DN, Song YS. Resveratrol triggers ER stress-mediated apoptosis by disrupting N-linked glycosylation of proteins in ovarian cancer cells. Cancer Lett. 2016;371:347-353.
31. Yousif NG. Fibronectin promotes migration and invasion of ovarian cancer cells through up-regulation of FAK–PI3K/Akt pathway. Cell Biol Int. 2013;38:85-91.
32. Xing H, Weng D, Chen G, Tao W, Zhu T, Yang X, Meng L, Wang S, Lu Y, Ma D. Activation of fibronectin/PI-3K/Akt2 leads to chemoresistance to docetaxel by regulating survivin protein expression in ovarian and breast cancer cells. Cancer Lett. 2008;261:108-119.
33. Fang D, Hawke D, Zheng Y, Xia Y, Meisenhelder J, Nika H, Mills GB, Kobayashi R, Hunter T, Lu Z. Phosphorylation of beta-catenin by AKT promotes beta-catenin transcriptional activity. J Biol Chem. 2007;282:11221-11229.
34. Luu HH, Zhang R, Haydon RC, Rayburn E, Kang Q, Si W, Park JK, Wang H, Peng Y, Jiang W, He TC. Wnt/beta-catenin signalling pathway as a novel cancer drug target. Curr Cancer Drug Targets. 2004;4:653-671.
35. Pramanik KC, Fofaria NM, Gupta P, Ranjan A, Kim SH, Srivastava SK. Inhibition of β-catenin signaling suppresses pancreatic tumor growth by disrupting nuclear β-catenin/TCF-1 complex: critical role of STAT-3. Oncotarget. 2015;6:11561-11574.
36. Verma UN, Surabhi RM, Schmaltieg A, Becerra C, Gaynor RB. Small Interfering RNAs Directed against β-Catenin Inhibit the in vitro and in vivo Growth of Colon Cancer Cells. Clin Cancer Res. 2003;9:1291-1300.

APA	KILIÇASLAN S, AYRIM A, APAYDIN E, İNCESU Z (2018). Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin. , 50 - 56. 10.4274/tjps.35220
Chicago	KILIÇASLAN Seda Mehtap SARI,AYRIM Aysun,APAYDIN ELİF,İNCESU Zerrin Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin. (2018): 50 - 56. 10.4274/tjps.35220
MLA	KILIÇASLAN Seda Mehtap SARI,AYRIM Aysun,APAYDIN ELİF,İNCESU Zerrin Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin. , 2018, ss.50 - 56. 10.4274/tjps.35220
AMA	KILIÇASLAN S,AYRIM A,APAYDIN E,İNCESU Z Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin. . 2018; 50 - 56. 10.4274/tjps.35220
Vancouver	KILIÇASLAN S,AYRIM A,APAYDIN E,İNCESU Z Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin. . 2018; 50 - 56. 10.4274/tjps.35220
IEEE	KILIÇASLAN S,AYRIM A,APAYDIN E,İNCESU Z "Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin." , ss.50 - 56, 2018. 10.4274/tjps.35220
ISNAD	KILIÇASLAN, Seda Mehtap SARI vd. "Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin". (2018), 50-56. https://doi.org/10.4274/tjps.35220

APA	KILIÇASLAN S, AYRIM A, APAYDIN E, İNCESU Z (2018). Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin. Turkish Journal of Pharmaceutical Sciences, 15(1), 50 - 56. 10.4274/tjps.35220
Chicago	KILIÇASLAN Seda Mehtap SARI,AYRIM Aysun,APAYDIN ELİF,İNCESU Zerrin Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin. Turkish Journal of Pharmaceutical Sciences 15, no.1 (2018): 50 - 56. 10.4274/tjps.35220
MLA	KILIÇASLAN Seda Mehtap SARI,AYRIM Aysun,APAYDIN ELİF,İNCESU Zerrin Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin. Turkish Journal of Pharmaceutical Sciences, vol.15, no.1, 2018, ss.50 - 56. 10.4274/tjps.35220
AMA	KILIÇASLAN S,AYRIM A,APAYDIN E,İNCESU Z Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin. Turkish Journal of Pharmaceutical Sciences. 2018; 15(1): 50 - 56. 10.4274/tjps.35220
Vancouver	KILIÇASLAN S,AYRIM A,APAYDIN E,İNCESU Z Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin. Turkish Journal of Pharmaceutical Sciences. 2018; 15(1): 50 - 56. 10.4274/tjps.35220
IEEE	KILIÇASLAN S,AYRIM A,APAYDIN E,İNCESU Z "Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin." Turkish Journal of Pharmaceutical Sciences, 15, ss.50 - 56, 2018. 10.4274/tjps.35220
ISNAD	KILIÇASLAN, Seda Mehtap SARI vd. "Calcium Mobilization and Inhibition of Akt Reduced the Binding of PEO-1 Cells to Fibronectin". Turkish Journal of Pharmaceutical Sciences 15/1 (2018), 50-56. https://doi.org/10.4274/tjps.35220