ÖZ
Amaç
Hücre içi miR-379-5p taşınması için optimize edilmiş polietilenimin (PEI) temelli bir nanotaşıyıcı geliştirilmesi ve bu sistemin, KRAS-wild tip kolorektal kanser hücrelerinde onkojenik hedefler olan metadherin (MTDH) ve Forkhead box P2 (FOXP2) üzerindeki baskılayıcı etkinliğinin değerlendirilmesi amaçlanmıştır.
Yöntemler
PEI-miRNA nanokompleksleri, farklı azot/fosfat (N:P) oranlarında sentezlenmiş ve dinamik ışık saçılımı, zeta potansiyel ölçümleri ile taramalı elektron mikroskobu (SEM) kullanılarak karakterize edilmiştir. Sitotoksisite, optimal terapötik konsantrasyonu belirlemek amacıyla Caco-2 hücrelerinde MTT testi ile değerlendirilmiştir. Gen susturma etkinliği ve hücre içi alım, gerçek zamanlı ters transkripsiyon kantitatif polimeraz zincir reaksiyonu ile nicel olarak analiz edilmiştir.
Bulgular
N:P oranı 20:1 olan formülasyon, yaklaşık 254 nm ortalama hidrodinamik çap, kompakt küresel morfoloji ve yüksek pozitif zeta potansiyeli (+56,9 mV) ile en uygun fizikokimyasal özellikleri göstermiştir. Optimize edilen 50 nM konsantrasyonda, nanokompleksler uygun hücre canlılığını korurken miR-379-5p’nin hücre içinde anlamlı düzeyde birikimini sağlamıştır. Buna bağlı olarak, bu taşıma stratejisi çıplak miRNA uygulamasına kıyasla MTDH ve FOXP2 ekspresyonunda güçlü bir azalma sağlamıştır.
Sonuç
Optimize edilmiş PEI-miRNA nanokompleksleri, taşınma engellerini etkili biçimde aşarak Caco-2 hücrelerinde başarılı gen susturulmasını mümkün kılmıştır. miR-379-5p düzenleyici ekseninin yeniden kurulması ve FOXP2’nin baskılanması yoluyla bu sistem, kolorektal malignitelerde hedefe yönelik RNA girişimi (RNAi) temelli uygulamalar için umut verici bir moleküler platform oluşturmaktadır.
Anahtar Kelimeler:
Kolorektal kanser, polietilenimin, miR-379-5p, gen taşınımı, nanopartiküller, RNA girişimi
Kaynaklar
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