Construction and Influence of Human Nuclear Factor-κB p65 shRNA Lentiviral Vector on Malignant Biological Behavior of Lung Cancer Cells
Background and objective Nuclear factor-κB is an important transcription factor and is closely associated with a variety of malignant tumors. The biological behavior of lung tumor cells can be reversed by inhibiting the expression of NF-κBp65 directly or indirectly. Nuclear factor-κBp65 gene shRNA r...
| Main Authors: | , , |
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| Format: | Article |
| Language: | zho |
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Chinese Anti-Cancer Association; Chinese Antituberculosis Association
2014-05-01
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| Series: | Chinese Journal of Lung Cancer |
| Subjects: | |
| Online Access: | http://dx.doi.org/10.3779/j.issn.1009-3419.2014.05.02 |
| _version_ | 1818207201837187072 |
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| author | Hongjuan GUO Guangfa ZHU Chunting WU |
| author_facet | Hongjuan GUO Guangfa ZHU Chunting WU |
| author_sort | Hongjuan GUO |
| collection | DOAJ |
| description | Background and objective Nuclear factor-κB is an important transcription factor and is closely associated with a variety of malignant tumors. The biological behavior of lung tumor cells can be reversed by inhibiting the expression of NF-κBp65 directly or indirectly. Nuclear factor-κBp65 gene shRNA recombinant plasmids were constructed and then infected with A549 cells. New stable cell lines were selected, and the ability of migration and adhesion was identified. Methods Both scramble control sequence and interference sequence (shRNA) of human nuclear factor-κBp65 were designed and synthesized to build recombinant plasmids, with BamH I site at the 5′ end and Xho I and EcoR I sites at the 3′ end. A549 cells were infected, and stable transfection strains were selected by puromycin. Western blot and qRT-PCR methods were applied to assess the interference efficient of NF-κBp65 and the protein expression level of IκBα. Transwell and MTT assays were carried out to analyze the ability of migration and adhesion of A549 cells separately. Results Recombinant plasmids were successfully built, and A549/NF-κB p65 scramble and A549/NF-κB p65 shRNA stable transfection strains were also successfully screened. Both mRNA and protein expression levels of NF-κBp65 showed that A549/NF-κBp65 shRNA cells decreased compared with A549/NF-κB p65 scramble cells and A549 cells, whereas the protein level of IκBα significantly increased. Both migration and adhesion abilities were also reduced. Conclusion In this study, both mRNA and protein expression levels of NF-κBp65 were effectively suppressed by RNA interference technique. NF-κBp65 inhibition can significantly reduce the migration and adhesion ability of A549 cells. |
| first_indexed | 2024-12-12T04:25:10Z |
| format | Article |
| id | doaj.art-ddebb3f2908d4924b66fea7575f200c6 |
| institution | Directory Open Access Journal |
| issn | 1009-3419 |
| language | zho |
| last_indexed | 2024-12-12T04:25:10Z |
| publishDate | 2014-05-01 |
| publisher | Chinese Anti-Cancer Association; Chinese Antituberculosis Association |
| record_format | Article |
| series | Chinese Journal of Lung Cancer |
| spelling | doaj.art-ddebb3f2908d4924b66fea7575f200c62022-12-22T00:38:13ZzhoChinese Anti-Cancer Association; Chinese Antituberculosis AssociationChinese Journal of Lung Cancer1009-34192014-05-0117536937710.3779/j.issn.1009-3419.2014.05.02Construction and Influence of Human Nuclear Factor-κB p65 shRNA Lentiviral
Vector on Malignant Biological Behavior of Lung Cancer CellsHongjuan GUO0Guangfa ZHU1Chunting WU2Department of Respiratory and Critical Care Medicine, Beijing Anzhen Hospital, Capital Medical University; Beijing Institute of Heart, Lung and Blood Vessel Diseases, Beijing 100029, ChinaDepartment of Respiratory and Critical Care Medicine, Beijing Anzhen Hospital, Capital Medical University; Beijing Institute of Heart, Lung and Blood Vessel Diseases, Beijing 100029, ChinaDepartment of Respiratory and Critical Care Medicine, Beijing Anzhen Hospital, Capital Medical University; Beijing Institute of Heart, Lung and Blood Vessel Diseases, Beijing 100029, ChinaBackground and objective Nuclear factor-κB is an important transcription factor and is closely associated with a variety of malignant tumors. The biological behavior of lung tumor cells can be reversed by inhibiting the expression of NF-κBp65 directly or indirectly. Nuclear factor-κBp65 gene shRNA recombinant plasmids were constructed and then infected with A549 cells. New stable cell lines were selected, and the ability of migration and adhesion was identified. Methods Both scramble control sequence and interference sequence (shRNA) of human nuclear factor-κBp65 were designed and synthesized to build recombinant plasmids, with BamH I site at the 5′ end and Xho I and EcoR I sites at the 3′ end. A549 cells were infected, and stable transfection strains were selected by puromycin. Western blot and qRT-PCR methods were applied to assess the interference efficient of NF-κBp65 and the protein expression level of IκBα. Transwell and MTT assays were carried out to analyze the ability of migration and adhesion of A549 cells separately. Results Recombinant plasmids were successfully built, and A549/NF-κB p65 scramble and A549/NF-κB p65 shRNA stable transfection strains were also successfully screened. Both mRNA and protein expression levels of NF-κBp65 showed that A549/NF-κBp65 shRNA cells decreased compared with A549/NF-κB p65 scramble cells and A549 cells, whereas the protein level of IκBα significantly increased. Both migration and adhesion abilities were also reduced. Conclusion In this study, both mRNA and protein expression levels of NF-κBp65 were effectively suppressed by RNA interference technique. NF-κBp65 inhibition can significantly reduce the migration and adhesion ability of A549 cells.http://dx.doi.org/10.3779/j.issn.1009-3419.2014.05.02NF-κBp65IκBαLentiviral vectorStable transfection cell lineMigrationAdhesion |
| spellingShingle | Hongjuan GUO Guangfa ZHU Chunting WU Construction and Influence of Human Nuclear Factor-κB p65 shRNA Lentiviral Vector on Malignant Biological Behavior of Lung Cancer Cells Chinese Journal of Lung Cancer NF-κBp65 IκBα Lentiviral vector Stable transfection cell line Migration Adhesion |
| title | Construction and Influence of Human Nuclear Factor-κB p65 shRNA Lentiviral
Vector on Malignant Biological Behavior of Lung Cancer Cells |
| title_full | Construction and Influence of Human Nuclear Factor-κB p65 shRNA Lentiviral
Vector on Malignant Biological Behavior of Lung Cancer Cells |
| title_fullStr | Construction and Influence of Human Nuclear Factor-κB p65 shRNA Lentiviral
Vector on Malignant Biological Behavior of Lung Cancer Cells |
| title_full_unstemmed | Construction and Influence of Human Nuclear Factor-κB p65 shRNA Lentiviral
Vector on Malignant Biological Behavior of Lung Cancer Cells |
| title_short | Construction and Influence of Human Nuclear Factor-κB p65 shRNA Lentiviral
Vector on Malignant Biological Behavior of Lung Cancer Cells |
| title_sort | construction and influence of human nuclear factor κb p65 shrna lentiviral
vector on malignant biological behavior of lung cancer cells |
| topic | NF-κBp65 IκBα Lentiviral vector Stable transfection cell line Migration Adhesion |
| url | http://dx.doi.org/10.3779/j.issn.1009-3419.2014.05.02 |
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