In vivo (Athens, Greece) · 2025
Development and Characterization of a Murine Lung Adenocarcinoma Cell Line With High Thoracic Pleural Metastatic Potential.
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Liao Liwei, Xu Weidong, Li Jia, Li Jiaye, Li Rui, Li Ruixia, Li Chang, Zheng Ziwen, Deng Mingming, Miao Jinrui, Wang Zilin, Zhang Qin, Bian Yiding, Wang Kai, Wang Han, Hou Gang
Аннотация
BACKGROUND/AIM: Pleural metastasis and malignant pleural effusion (MPE) are common complications of lung adenocarcinoma. Patients with MPE have poor outcomes, with overall survival ranging from 5 to 11.4 months. The lack of established cell lines and stable animal models of pleural metastasis has limited studies on the underlying mechanisms of MPE development. In this study, we aimed to develop a murine lung adenocarcinoma cell line with high thoracic pleural metastatic potential.
MATERIALS AND METHODS: Luciferase-tagged Lewis lung carcinoma (LLC) cells were implanted into the pleural cavity of C57bl/6 mice, with five rounds of subsequent extraction from pleural foci and reinjection into the pleural cavity. The metastatic properties of the established cell line were verified in vivo by evaluating the metastatic burden and MPE volume (n=5). In vitro, the metastatic ability of cell lines was assessed by scratch assay, transwell migration assay, cell-matrix adhesion assay and cell-cell adhesion assay (3-5 replicates). The transcription profile was characterized by mRNA sequencing. Differential analysis and KEGG enrichment were performed to show their distinctions. Differential genes were verified by quantitative real-time PCR (qPCR).
RESULTS: An LLC subpopulation with high thoracic pleural metastatic potential was generated and named LLC-PLM. In vivo, compared with parental LLC (LLC-P), LLC-PLM demonstrated a greater incidence of MPE and greater MPE volumes. In vitro, LLC-PLM demonstrated increased metastatic capacity and augmented adhesion capacities, compared to LLC-P. Transcriptomic analysis revealed that pathways related to adhesion, migration, and membrane signaling were notably enriched and activated in LLC-PLM cells. Relative genes were obviously activated, including Lamc2, Col4a3, Col6a3, Col1a1, Itga2 and Itga1.
CONCLUSION: We successfully established a murine cell line LLC-PLM that can serve as a valuable tool for studying pleural metastasis and MPE.
MATERIALS AND METHODS: Luciferase-tagged Lewis lung carcinoma (LLC) cells were implanted into the pleural cavity of C57bl/6 mice, with five rounds of subsequent extraction from pleural foci and reinjection into the pleural cavity. The metastatic properties of the established cell line were verified in vivo by evaluating the metastatic burden and MPE volume (n=5). In vitro, the metastatic ability of cell lines was assessed by scratch assay, transwell migration assay, cell-matrix adhesion assay and cell-cell adhesion assay (3-5 replicates). The transcription profile was characterized by mRNA sequencing. Differential analysis and KEGG enrichment were performed to show their distinctions. Differential genes were verified by quantitative real-time PCR (qPCR).
RESULTS: An LLC subpopulation with high thoracic pleural metastatic potential was generated and named LLC-PLM. In vivo, compared with parental LLC (LLC-P), LLC-PLM demonstrated a greater incidence of MPE and greater MPE volumes. In vitro, LLC-PLM demonstrated increased metastatic capacity and augmented adhesion capacities, compared to LLC-P. Transcriptomic analysis revealed that pathways related to adhesion, migration, and membrane signaling were notably enriched and activated in LLC-PLM cells. Relative genes were obviously activated, including Lamc2, Col4a3, Col6a3, Col1a1, Itga2 and Itga1.
CONCLUSION: We successfully established a murine cell line LLC-PLM that can serve as a valuable tool for studying pleural metastasis and MPE.