MCF 7 Hücre Hattında 4- Aminopiridin ve Paklitakselin Sinerjistik Etkileri

Year 2019, Volume: 9 Issue: 4, 321 - 327, 31.12.2019

Esra Münire Cüce Aydoğmuş , Gunseli Ayse Inhan Garıp

https://doi.org/10.33808/clinexphealthsci.557797

Abstract

Amaç: Çalışmamızda, meme
kanseri tedavisinde yaygın olarak kullanılan Paklitaksel’in (PTX) etkinliğinin
4-aminopiridin (4-AP) kullanılarak arttırılması hedeflenmiştir.

Yöntemler: Çalışmamızda, L929
(ATCC CRL-6364) ve MCF7 (ATCC- HTB 22) hücre hatları kullanıldı. IC₅₀
ve canlılık tripan mavisi metoduyla, hücre döngüsü analizleri Cdk2 ve Histon (H3)
seviyeleri ile ve membran potansiyeli ölçümleri Dibac4(3)
[Bis-(1,3-dibutylbarbituric acid)trimethine oxonol] kullanılarak floresan
mikroplaka okuyucuda belirlendi.

Bulgular: Yapılan deneylerde,
her iki ajan için IC₅₀ değerleri belirlendi ve bu değerler kombine
edildi. Kombinasyonlarda iki farklı PTX konsantrasyonu kullanıldı. Kombinasyon
sonucunda, L929 hücrelerinde 4-AP (4 mM) ve PTX (5 nM), 4-AP (4 mM) ve PTX (7.5
nM) kombinasyonunun canlılığı sırasıyla % 17 ± 8.08,  % 45 ±
3.18, MCF 7 hücrelerinde ise sırasıyla %60 ± 3.7, %74 ± 2.6 azalttığı
belirlenmiştir. Membran potansiyeli (Vm) ölçümlerinde, L929 hücreleri için 4-AP
ve 5 nM- 7.5 nM PTX değerlerinde depolarizasyon, kombinasyonlar için
hiperpolarizasyon gözlemlenirken, MCF7 hücrelerinde 4-AP,7.5nM PTX ve 4-AP (4
mM) + PTX (5 nM) için depolarizasyon, 5 nM PTX ve 4-AP (4 mM) + PTX (7.5 nM)
için hyperpolarizasyon gözlemlenmiştir. Cdk2 ve H3 seviyeleri ölçümlerinde, MCF
7 hücreleri için ağırlıklı olarak G1 fazında, L929 hücreleri ağırlıklı olarak
G2/ M fazında duraklama görülmüştür.

Sonuç: MCF-7 meme kanseri
hücrelerinde 4-AP ve PTX in beraber kullanılmasıyla canlılıkta azalma
beklendiği şekilde artarken, sağlıklı hücre hattı olarak kullanılan L929
hücrelerinde antagonist etki görülmüştür. Bu veriler, 4- AP ile PTX
kombinasyonunun kanser hücrelerini seçici olarak etkilediğini ve dolayısıyla
umut vadeden bir yaklaşım olduğunu göstermektedir. 

Keywords

4- aminopiridine, Paklitaksel, MCF7 meme kanseri hücre hattı

Synergistic Effects of 4-Aminopyridine and Paclitaxel on MCF 7 Cell Line

Abstract

Objectives: Aim of this study is to increase the effectiveness of paclitaxel (PTX) with the use of 4 – aminopyridine (4-AP) on breast cancer cell line MCF-7.
Methods: In this study, L-929 (ATCC CRL-6364) and MCF-7 (ATCC – HTB 22) cell lines were used. IC50 and survival values were determined by trypan blue exclusion; cell cycle analysis was determined by measuring levels of Cdk2 and Histone (H3) and plasma membrane potential (Vm) measurements were performed using fluorescent Bis-(1,3-dibutylbarbituric acid) trimethine oxonol (DiBaC4(3)).
Results: IC50 values were determined for two agents and these values were combined. Combination treatments ie. “4-AP (4 mM) + PTX (5 nM)” and “4-AP (4 mM) + PTX (7.5 nM)” decreased viability 17% ± 8.08 and 45% ± 3.18, respectively for L-929 cells and decreased viability 60%± 3.7 and 74%± 2.6, respectively for MCF-7 cells. For L-929 cells, plasma membrane potential measurements resulted in depolarization for 4-AP, PTX (5 nM) and PTX (7.5 nM), and resulted in hyperpolarization for the combinations. For MCF-7 cells, plasma membrane potential measurements resulted in depolarization for 4-AP, PTX (7.5 nM) and 4-AP + PTX (5 nM), and resulted in hyperpolarization for PTX (5 nM) and 4-AP + PTX (7.5 nM). Changes of Cdk2 and H3 levels showed mostly G1 arrest for MCF 7 cells and G2/M arrest for L-929 cells.
Conclusions: Combination treatments increased the cell death for MCF-7 cells. But, combination treatments didn’t show synergistic effect on L-929 which is accepted as a non-cancerous cell. These data showed that use of 4-AP in combination with the anticancer agent paclitaxel is a promising approach for cancer treatment.

Keywords

4- aminopyridine, Paclitaxel, MCF 7 breast cancer cell line

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