Inhibicija proliferacije, invazije i migracije ćelija kolorektalnog karcinoma i indukcija epitelno-mezenhimalne tranzicije utišavanjem TFF1 gena
Sažetak
Uvod/Cilj. Identifikacija aberantnih gena povezanih sa kolorektalnim karcinomom (colorectal cancer – CRC) i razumevanje njihovih patogenih mehanizama je od značaja za ranu dijagnozu bolesti i određivanje odgovarajuće terapije. Cilj rada bio je da se ispita uticaj trefoil faktora 1 (TFF1) na proliferaciju, invaziju, i migraciju ćelija CRC. Metode. Ekspresija TFF1 u 40 uzoraka tkiva CRC procenjena je pomoću imunohistohemije. Pored toga, ekspresija TFF1 u CRC ćelijskim linijama HCT116 i Caco-2 određena je pomoću analize kvantitativne lančane reakcije polimerazom u realnom vremenu (quantitative polymerase chain reaction – qPCR) i Western blot analize. Zatim je Caco-2 ćelijska linija transfektovana malom interferirajućom ribonukleinskom kiselinom upakovanom u lentivirus u cilju utišavanja ekspresije TFF1. Za procenu proliferacije ćelija i sposobnosti formiranja kolonija korišćeni su esej cell counting kit-8 i test formiranja kolonija ćelija, dok su Transwell esej i test zaceljivanja rana korišćeni za procenu invazivne i migratorne sposobnosti ćelija. Nakon toga, nivoi ekspresije E-kaderina i vimentina procenjeni su pomoću qPCR i Western blot analize kako bi se ispitala veza između ekspresije TFF1 i epitelno-mezenhimalne tranzicije. Rezultati. Pozitivna stopa ekspresije TFF1 u tkivima CRC bila je 35% (14/40). U HCT116 ćelijskoj liniji detektovani su umereni nivoi ekspresije TFF1, dok su u Caco-2 ćelijskoj liniji zapaženi značajno viši nivoi ekspresije. U Caco-2 ćelijama zabeležen je smanjen nivo TFF1 informacione ribonukleinske kiseline i proteina nakon utišavanja TFF1. Zatim je uočeno da se smanjuje sposobnost proliferacije i formiranja kolonija kod Caco-2 ćelija, kao i njihove invazivne i migratorne sposobnosti. Štaviše, utišavanje ekspresije TFF1 dovelo je do povećanja ekspresije E-kaderina i smanjenja ekspresije vimentina. Zaključak. Utišavanje ekspresije TFF1 može inhibirati proliferaciju, migraciju i invaziju CRC ćelija Caco-2. Podsticanjem epitelno-mezenhimalne tranzicije TFF1 može igrati značajnu ulogu u nastanku, razvoju i metastaziranju CRC.
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