PLAG Exerts Anti-Metastatic Effects by Interfering with Neutrophil Elastase/PAR2/EGFR Signaling in A549 Lung Cancer Orthotopic Model
Abstract
:1. Introduction
2. Results
2.1. PLAG Inhibited Metastasis of A549 Human Lung Cancer in an Orthotopic Mouse Model.
2.2. Treatment with 100 mpk PLAG Reduced Metastasis to the Brain and Intestines.
2.3. PLAG Treatment Inhibited the Growth of A549 Human Lung Cancer in Mice.
2.4. The Metastatic Properties of A549 Cells were Promoted by the Tumor-Infiltrating Neutrophil Environment and Attenuated by PLAG.
2.5. EGFR Transactivation for Metastatic Activity was Mediated by the PAR2 Receptor in the Presence of Neutrophils.
2.6. Neutrophil-Induced Metastatic Activity of A549 Cells was Dependent on PAR2 and β-Arrestin-2.
2.7. The Inhibitory Effect of PLAG on Neutrophil-Induced Metastatic Activity in A549 Cells was Dependent on GPIHBP1 and ⍺-Arrestin.
3. Discussion
4. Materials and Methods
4.1. Cell Culture
4.2. Invasion Assay to Measure the Migration Activity of Cancer Cells
4.3. Co-Culture System for Indirect Contact using Separated Chambers
4.4. Immunofluorescence Staining and Wound Healing Assay
4.5. Spheroid-Formation Assay to Examine Cancer Cell Scattering
4.6. Analysis of Protein Degradation by Ubiquitination Assay
4.7. Analysis of Secreted Proteins by ELISA
4.8. Examination by Confocal Microscopy for Lysosome Co-Localization
4.9. Analysis of Protein Assemblies by Immunoprecipitation
4.10. Analysis of Expressed Transcripts using Quantitative PCR
4.11. Gene Silencing by Small Interfering RNA (siRNA)
4.12. Lung Cancer Orthotopic Implantation Model
4.13. Overall Survival Analysis
4.14. Tumor Mass Scan by Computed Tomography
4.15. Immunohistochemistry Staining
4.16. Hematoxylin and Eosin Staining
4.17. Statistics
5. Conclusions
Author Contributions
Funding
Conflicts of Interest
References
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Kim, G.T.; Hahn, K.W.; Yoon, S.Y.; Sohn, K.-Y.; Kim, J.W. PLAG Exerts Anti-Metastatic Effects by Interfering with Neutrophil Elastase/PAR2/EGFR Signaling in A549 Lung Cancer Orthotopic Model. Cancers 2020, 12, 560. https://doi.org/10.3390/cancers12030560
Kim GT, Hahn KW, Yoon SY, Sohn K-Y, Kim JW. PLAG Exerts Anti-Metastatic Effects by Interfering with Neutrophil Elastase/PAR2/EGFR Signaling in A549 Lung Cancer Orthotopic Model. Cancers. 2020; 12(3):560. https://doi.org/10.3390/cancers12030560
Chicago/Turabian StyleKim, Guen Tae, Kyu Woong Hahn, Sun Young Yoon, Ki-Young Sohn, and Jae Wha Kim. 2020. "PLAG Exerts Anti-Metastatic Effects by Interfering with Neutrophil Elastase/PAR2/EGFR Signaling in A549 Lung Cancer Orthotopic Model" Cancers 12, no. 3: 560. https://doi.org/10.3390/cancers12030560