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Effect of gene transfer of tumor necrosis factor receptors into human lung carcinoma cell line
H Ohara1, Y Hasegawa, T Kawabe
1First Department of Internal Medicine, Nagoya University School of Medicine.
Abstract:
The human lung adenocarcinoma cell line A549 is known to be resistant to tumor necrosis factor alpha (TNF-alpha)-mediated tumor cell lysis in spite of the expression of 55 kDa TNF receptor (TNF-R55) mRNA and its cell surface protein. In this study, we investigated the mechanism of TNF-alpha resistance and the role of two types of TNF receptors (TNF-R55 and TNF-R75 (75 kDa TNF receptor)). TNF-R55 or TNF-R75 cDNA was transfected into A549 cells. In addition, a C-terminal deletion mutant of TNF-R75 which lacks the intracellular domain of TNF-R75 was also transfected into A549 cells. We assessed the TNF-alpha-mediated tumor cell lysis of these transfected clones, and found that the cytotoxic effect increased in transfected clones highly expressing TNF- R55, but not in low-expression clones. As for TNF-R75, the cytotoxic effect of TNF-alpha was observed in TNF-R75-transfected clones even when expression was low. Furthermore, the cytotoxic effect was also observed in clones transfected with the deletion mutant of TNF-R75, as well as the complete TNF-R75. These results indicate that a certain level of expression of TNF-R75 is necessary for obtaining TNF-alpha-mediated tumor cell lysis in the absence of TNF-R75. On the other hand, the expression of TNF-R75 strongly induces TNF-alpha-mediated cytotoxicity through TNF-R55 in the absence of an intracellular signal via TNF-R75.
Insights
Lung adenocarcinoma cells resistant to tumor necrosis factor alpha (TNF-alpha) lysis were studied. Introducing TNF receptor 75 (TNF-R75) enhanced TNF-alpha-mediated cell death, suggesting its crucial role in overcoming resistance.
Area of Science:
- Immunology
- Cell Biology
- Oncology
Background:
- A549 lung adenocarcinoma cells exhibit resistance to tumor necrosis factor alpha (TNF-alpha)-induced cell death.
- These cells express TNF receptor 55 (TNF-R55) mRNA and protein, yet remain resistant to TNF-alpha.
- The roles of TNF-R55 and TNF receptor 75 (TNF-R75) in this resistance mechanism require further investigation.
Purpose of the Study:
- To investigate the mechanism underlying TNF-alpha resistance in A549 cells.
- To elucidate the specific roles of TNF-R55 and TNF-R75 in TNF-alpha-mediated cytotoxicity.
- To determine if transfection with TNF-R55 or TNF-R75 can restore sensitivity to TNF-alpha.
Main Methods:
- A549 cells were transfected with cDNA encoding TNF-R55, TNF-R75, or a C-terminal deletion mutant of TNF-R75.
- Transfected clones were assessed for TNF-alpha-mediated tumor cell lysis.
- Expression levels of transfected TNF receptors were evaluated.
Main Results:
- High expression of transfected TNF-R55 increased TNF-alpha-mediated cytotoxicity, while low expression did not.
- TNF-alpha-mediated cytotoxicity was observed in TNF-R75-transfected clones even at low expression levels.
- Cytotoxicity was also observed with a TNF-R75 mutant lacking its intracellular domain, indicating its signaling is not solely dependent on intracellular interaction.
Conclusions:
- A minimum level of TNF-R75 expression is necessary for TNF-alpha-mediated lysis in A549 cells lacking endogenous TNF-R75 signaling.
- TNF-R75 expression can induce TNF-alpha-mediated cytotoxicity through TNF-R55, even without intracellular signaling via TNF-R75.
- These findings highlight the complex interplay between TNF receptors in overcoming TNF-alpha resistance in lung adenocarcinoma.