Abstract

Canlı hücrelerin uzun süre boyunca çok düşük sıcaklıklarda saklanması işlemi kriyo-korunma olarak adlandırılır. Kriyo-koruma işlemi biyoteknoloji, klinik çalışmalar ve hayvan veya bitki hücreleriyle ilgili birçok çalışmada çok önemli bir rol oynadığından dolayı, kriyo-korumada kullanılan ajanların araştırılması son derece mühimdir. Kriyo- koruma ajanları, hücresel metabolizma ve biyofizyoloji üzerindeki etkileri nedeniyle uzun süreli kriyo-korumanın ardından hücresel canlılığın korunmasını sağlarlar. Mevcut çalışmada; hücre canlılık testi, koloni oluşturma testi ve gerçek zamanlı polimeraz zincir reaksiyonu tekniklerinden yararlanılarak, borun kriyo-koruma üzerindeki etkisi, insan akciğer kanser hücre hattı, A549 ve fare fibroblast hücre hattı, L929 kullanılarak araştırılmıştır. Hücre dondurma ortamını 15 µg/ml bor ile desteklemenin hücre canlılığı üzerine olumlu etki ettiği gözlemlenmiştir. Ayrıca, tekrar eden dondurma - çözme döngüleri ve uzun süreli kriyo-koruma sonucunda, gen anlatım profilleri değişen A549 ve L929 hücre hatlarının, bor takviyesi sonrasında, programlı hücre ölümüyle alakalı genlerinin anlatımında azalma, hücre çoğalması ile ilgili genlerinde de artış gözlemlenmiştir. Sonuçlarımız göstermiştir ki bor kaynağı olarak sodium pentaborat pentahidrat, kanser veya sağlıklı hücrelerin canlılıklarını kaybetmeksizin dondurulmalarını ve hücrelerin uzun süreli saklanmaları için son derece önemli bir kriyo-koruyucu ajan olarak kullanılabilir.

BORON INCREASES THE VIABILITY OF HUMAN CANCER AND MURINE FIBROBLAST CELLS AFTER LONG TIME OF CRYOPRESERVATION

Abstract

Through the process of cryopreservation, cells are stored at very low temperature for a long time to decrease the biological and chemical reactions in viable cells. In this process, the administration of cryoprotective agents is crucial since cryopreservation is regarded as a leading process in various research fields such as biotechnology, clinical medicine and maintenance of both animal and plant cells. Even after a long time of storage in very low temperatures, a recovery is achieved by cryo-preservative agents that act on cellular metabolism and biophysiology of cells. In the current study, the effect of boron on cryopreservation of human lung cancer cell line, A549, and murine fibroblast cell line, L929, was investigated with the help of cell viability assay, colony forming unit assay and RT-PCR analysis. 15 µg/ml boron supplemented freezing medium was found to indicate a positive effect on cell viability. Moreover, gene expression profiles of A549 and L929 cell lines have been altered. The levels of apoptosis related genes decreased while proliferation related gene levels increased significantly after repeated freeze-thaw cycles or long period of freezing. As indicated through our results, sodium pentaborate pentahydrate, as a boron source, might be a crucial cryoprotective agent for cryo-protection and bio-banking of cancer and healthy cells while keeping their viability and functionality.

Keywords

• Boron
• cryopreservation
• cryoprotective agents
• cancer cells
• fibroblast cells

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