Abstract
BackgroundLung cancer is the leading cause of cancer-related morbidity and mortality all over the world. Surgery resection, radiotherapy, chemotherapy, immunotherapy and combined treatments have been discovered and well established for treatments. However, low survival rate of five years after clinical treatments mainly due to recurrence of stress-resistant cancer cells calls for better understanding and new ideas. Our project aimed to understand the forming process of stress resistant lung cancer cells after radiotherapy.MethodsTwo classic non-small cell lung cancer (NSCLC) cell lines A549 and H1299 initially were radiated with a 137Cs gamma-ray source with doses ranging from 0 to 12 Gy to generate radiation-resistant cancer cells. 8 Gy of radiation was regard as a standard dosage since it provides effective killing as well as good amount of survivals. The expression levels of autophagy-related proteins including Beclin-1, LC3-II and p62 were studied and measured by both western blot and quantitative real-time polymerase chain reaction (real-time RT-PCR).ResultsIncreased Beclin-1, LC3-II and decreased p62 have been observed in radiation-resistant cells indicating elevated autophagy level. Decreased miR-191 in radiation-resistant cells performed by Taqman qRT-PCR also has been seen. Two binding sites between Beclin-1 and miR-191 suggest potential association between.ConclusionsIt is reasonable to speculate that inhibition of miR-191 expression in lung cancer cells would contribute to the establishment of radiation-resistant cells via mediating cellular autophagy. Therefore, miR-191 is a potential target for therapy in treating radiation-resistant lung cancer.
Highlights
Lung cancer is the leading cause of cancer-related mortality and morbidity and it is one of the predominant lifethreatening conditions among cancers
It is reasonable to speculate that inhibition of miR-191 expression in lung cancer cells would contribute to the establishment of radiation-resistant cells via mediating cellular autophagy
MiR-191 is a potential target for therapy in treating radiation-resistant lung cancer
Summary
Cell culturing and proliferation Cell culturing and established models of radiation-resistant cell lines A549 and H1299 cells were harvested by exposing to trypsin. 100 μl of cell suspension (5000 cells/well) obtained from 2, 4, 7, 10, 14, 18 and 21 days after radiation treatment were dispensed into a 96-well plate This plate was pre-incubated for 24 hours in a humidified incubator (e.g., at 37°C, 5% CO2). The membrane was incubated in primary antibody (Beclin-1, GAPDH, Abcam) with a 1/1000 dilution in blocking buffer (50 mM Tris base; 100 mM NaCl; 0.02% Tween 20; and 3% BSA) overnight. The membrane was rinsed by TTBS (0.1% Tween 20, 10 mM Tris base, 100 mM NaCl, pH 7.5) for three times before adding secondary antibody (Abcam) with 1/5000 dilution in blocking buffer for 2 hours. Taqman real-time qRT-PCR was performed to detect the relative levels of miR-191 in radiation resistant and non-treatment cells.
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
