RADIOSENSITIVITY OF HUMAN BRONCHIAL EPITHELIAL CELLS WITH GENETIC ALTERATIONS

Purpose: To analyze the proliferation rates and the radiosensitivity of hu- man bronchial epithelial cells (HBECs) with genetic alterations including human telomerase reverse transcriptase (hTERT), Cdk4 (cyclin depen- dent kinase 4), knockdown p53 and mutant K-ras overexpression. Materials and Methods: HBECs obtained from human bronchus spe- cimens were placed into short-term culture and were serially transfected with retroviral constructs containing Cdk4 and hTERT, resulting in conti- nuously growing immortalized normal human bronchial epithelial cell li- nes. These cell lines were used to produce knock-down p53, mutant K-ras and knockdown p53 plus K-ras expressing cell lines; the proliferation ra- tes and the radiosensitivity of these cell lines were evaluated. The control cells and the Cdk4, hTERT, knockdown p53 and K-ras expressing cells were irradiated (1 Gy-10 Gy) and the fraction survival was determined 1 week later. Results: The proliferation rates of the cells expressing both knockdown p53 and K-ras were higher than those of the other cell lines and show a loss of contact inhibition in vitro. The HBECs infected with both knock- down p53 and mutant K-ras were more radioresistant compared to cells infected with only mutant K-ras or knockdown p53. Conclusion: HBECs expressing knockdown p53 in addition to mutant K- ras have higher proliferation rates and additionally they lose the ability of growth arrest in response to DNA damage signals such as irradiation. Key Words: Human Bronchial Epithelial Cells, Cdk4, Htert, P53, K-Ras, Irradiation. GENETIK FARKLILIK GOSTEREN INSAN BRONŞ EPITEL HUCRELERININ RADYOTERAPIYE HASSASIYETLERI Amac: Bu calismada, insan bronsiyal epitel hucrelerinin hTERT (insan telomeraz “reverse” transkriptazi), Cdk4 (siklin bagimli kinaz 4), “knock- down” p53 ve mutant K-ras onkogeni ile enfekte edilmesiyle elde edilen hucre serilerinin hucre kulturu ortaminda buyume hizlarinin ve radyotera- piye karsi hassasiyetlerinin karsilastirilmasi amaclanmistir. Metodlar: Insan brons dokusu orneklerinden elde edilen bronsiyal hucre- ler, oncelikle Cdk4 ve hTERT ile enfekte edilerek olumsuzlestirildi. Daha sonra bu hucreler, “knockdown” p53, mutant K-ras ve “knockdown” p53 ‘le birlikte mutant K-ras eksprese eden hucre serilerinin elde edilmesinde kullanildi. Bulgular: Hem “knockdown” p53, hem de K-ras ekspresyonu gosteren hucrelerin buyume hizlarinin in vitro sartlarda diger hucre serilerine oran- la daha yuksek oldugu ve kontakt inhibisyon ozelliklerini kaybettikleri saptandi. Kontrol hucreleri ile Cdk4, hTERT, “knockdown” p53 ve K-ras ile enfekte edilmis olan hucrelere 1 Gy ile 10 Gy arasinda farkli dozlarda radyoterapi uygulandi, 1 hafta sonra canli kalan hucrelerin orani belir- lendi. Sonuc: Bu calismanin sonuclari, hem“knockdown” p53 hem de mutant K-ras ile enfekte edilen bronsiyal epitel hucrelerinin, sadece K-ras veya “knockdown” p53 ile enfekte edilenlere oranla in vitro sartlarda yuksek proliferasyon hizina sahip olduklarini ve ayrica DNA hasarina yol acan radyoterapiye cevap olarak buyume duraklamasina girme ozelliklerini kaybettiklerini ortaya koymaktadir. Anahtar Kelimeler: Insan Brons Epitel Hucreleri, Cdk4, Htert, P53, K- Ras, Radyoterapi.