The mitogen-activated protein kinase (MAPK) pathways transmit signals from the cell membrane to the nucleus. Activation of MAPK cascades may play a role in malignant transformation. We hypothesized that enhanced expression of one or more of these pathways would occur in human lung cancers. Using Western blot analysis of tissue homogenates from resected non- small cell lung cancers and matched non-neoplastic lung tissue, we determined that only activated p38 was consistently increased in tumor compared with normal tissue. In vitro kinase assays confirmed that the levels of activated MAPK correlated with the activity of the enzymes, and immunohistochemical analysis confirmed the cellular localization of the activated MAPKs. We incubated a lung cancer cell line in a hypoxic chamber to simulate the hypoxic environment in solid lung tumors, but found no increase in p38 activation. Contrary to our expectations, ERK and JNK, the MAPK pathways traditionally associated with cell growth and perhaps malignant transformation, were not consistently activated in the human lung tumor samples. However, p38, a MAPK usually associated with stress responses, growth arrest, and apoptosis, was activated in all of the human lung cancer samples, suggesting an additional role for this pathway in malignant cell growth or transformation.
Glucocorticoids inhibit the proliferation of various cell types, but the mechanism of this inhibition remains unclear. We investigated the effect of dexamethasone on non-small cell lung cancer cell growth and cell cycle progression. We showed that dexamethasone suppresses the proliferation of A549 and Calu-1 cells, with accumulation of cells in G1/G0 stage of the cell cycle, as determined by fluorescence-activated cell sorter analysis. Western blot analysis confirmed that this is associated with hypophosphorylation of retinoblastoma protein. Using Western blot analysis and in vitro kinase assays, we found that dexamethasone results in decreased activity of CDK2 and 4, decreased levels of cyclin D, E2F, and Myc, and increased levels of the CDK inhibitor p21(Cip1). In addition, we found that dexamethasone decreases activity of extracellular signal-related kinase (ERK)/mitogen-activated protein kinase (MAPK). The kinetics of all these changes indicate that inhibition of the ERK/MAPK pathway precedes the cell cycle effects, suggesting that regulation of this MAPK-signaling pathway may be an alternative mechanism for glucocorticoid-induced cell cycle arrest and growth inhibition.
We have shown that anti-RTA RNA aptamer can protect against ricin ribotoxicity in cell-based luciferase and cell cytotoxicity assays. Hence, RNA aptamer that inhibits RTA enzymatic activity represents a novel class of nucleic acid inhibitor that has the potential to be developed as a therapeutic agent for the treatment of ricin intoxication.
Mutations in the p53 gene have been implicated to play an important role in the development of various human cancers. To evaluate the importance of p53 in lung cancer, a transgenic mouse model was established by utilizing the Clara cell secretory protein (CCSP) promoter to target the expression of a dominant-negative mutant form of p53 (dnp53) in the lung. In two transgenic CCSP-dnp53 founder lines, the dnp53 protein was expressed exclusively in the lungs. The incidence of spontaneous lung cancer in 18-month-old transgenic mice was 45%, whereas that in age-matched control mice was 20%. The relative risk of lung tumors in CCSP-dnp53 mice was 2.3 times that of wild-type mice (exact confidence limits of 0.69, 17.5). In addition to the increased incidence of spontaneous lung tumor, these mice were more susceptible to the development of lung adenocarcinoma after exposure to benzo(a)pyrene (BaP). Six months after intratracheal instillation of benzo(a)pyrene, the tumor incidence in wild-type and CCSP-dnp53 mice was 39% and 73%, respectively. The risk of lung tumors was 25.3 times greater in BaP-treated mice adjusted for transgene expression (95% confidence limits of 3.29, 678, mid-p corrected). These results suggest that p53 function is important for protecting mice from both spontaneous and BaP-induced lung cancers.
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