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shRNA inhibits anchorage independent growth and invasion of human non-small cell lung cancer cell line H460SM
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Abstract:
Objective: To investigate the effect of lentivirus-mediated integrin α6 (ITGA6) shRNA on growth and invasion of human non-small cell lung cancer (NSCLC) H460SM cells. Methods: The expression levels of ITGA6 in 9 human NSCLC cell lines were evaluated by quantitative-PCR (Q-PCR) and Western blotting. Lentiviral ITGA6 shRNA were transfected into H460SM cells and ITGA6expression was detected by Q-PCR and Western blotting; the cellular morphology change was observed under a microscope; cell proliferation was detected by MTS assay; anchorage-independent growth was determined by colony formation assay in soft agar; apoptosis and cell cycle were analyzed by flow cytometry; and cell invasion and migration were determined by invasion and migration assay. Results: ITGA6 was expressed in 7 NSCLC cell lines. A significantly higher level of ITGA6 expression was seen in H460SM cells compared with the parental H460 cells (\[8.75±0.09\] vs \[5.78±0.26\], P<0.01). After H460SM cells were stably transfected with lentiviral ITGA6 shRNA (H460SM-75, H460SM-76 cells), a significant down-regulation of ITGA6 expression was observed in H460SM-75 and H460SM-76 cells compared with negative control H460SM-NS cells (\[1.00±0.01\] vs \[0.11±0.04\], \[0.22±004\], P<0.01). The cell viability of H460SM-75 and H460SM-76 cells had no difference with H460SM-NS cells (P>0.05), as well as the apoptotic rate, cell proliferative index and proportion of G0/G1 phase cells as compared with the negative control (P>0.05). The colony formation rate of H460SM-75 and H460SM-76 cells was significantly decreased as compared with H460SM-NS cells (\[20.81±1.38\]% vs \[18.87±1.47\]%, \[18.85±1.11\]%, P<0.05), and the migration rate (\[100.00±0.50\]% vs \[43.92±0.41\]%, \[24.10±0.33\]%, P<0.01) and invasion rate (\[100.00±6.74\]% vs \[7.04±2.96\]%, \[4.68±0.27\]%, P<0.01) were also significantly decreased. Conclusion: Knockdown of ITGA6 expression can inhibit anchorage-independent cell growth, migration and invasion of NSCLC cells, but have no effect on cell proliferation and apoptosis.
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Project Supported:
Project supported by the National Natural Science Foundation of China (No. 81060177), the Applied and Basic Research Foundation of Yunnan Province (No. 2009CD181), the Scientific Research Foundation for Returned Overseas Chinese Scholars of State Personnel Ministry (No. 412-2010), and the Science and Technology Medical Research Foundation o f Yunnan Province (No. 2009NS079)
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