The establishment of a lung colonization assay for circulating tumor cell visualization in lung tissues

Tsung Cheng Lin, Ying Chih Liao, Wen-Tsan Chang, Cheng Han Yang, Li Hsin Cheng, Megan Cheng, Hung-Chi Cheng

Research output: Contribution to journalArticle

Abstract

Metastasis is the major cause of cancer death. The role of circulating tumor cells (CTCs) in promoting cancer metastasis, in which lung colonization by CTCs critically contributes to early lung metastatic processes, has been vigorously investigated. As such, animal models are the only approach that captures the full systemic process of metastasis. Given that problems occur in previous experimental designs for examining the contributions of CTCs to blood vessel extravasation, we established an in vivo lung colonization assay in which a long-term-fluorescence cell-tracer, carboxyfluorescein succinimidyl ester (CFSE), was used to label suspended tumor cells and lung perfusion was performed to clear non-specifically trapped CTCs prior to lung removal, confocal imaging, and quantification. Polymeric fibronectin (polyFN) assembled on CTC surfaces has been found to mediate lung colonization in the final establishment of metastatic tumor tissues. Here, to specifically test the requirement of polyFN assembly on CTCs for lung colonization and extravasation, we performed short term lung colonization assays in which suspended Lewis lung carcinoma cells (LLCs) stably expressing FN-shRNA (shFN) or scramble-shRNA (shScr) and pre-labeled with 20 μM of CFSE were intravenously inoculated into C57BL/6 mice. We successfully demonstrated that the abilities of shFN LLC cells to colonize the mouse lungs were significantly diminished in comparison to shScr LLC cells. Therefore, this short-term methodology may be widely applied to specifically demonstrate the ability of CTCs within the circulation to colonize the lungs.

Original languageEnglish
Article numbere56761
JournalJournal of Visualized Experiments
Volume2018
Issue number136
DOIs
Publication statusPublished - 2018 Jun 18

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Circulating Neoplastic Cells
Tumors
Assays
Visualization
Cells
Tissue
Lung
Lewis Lung Carcinoma
Small Interfering RNA
Fibronectins
Neoplasm Metastasis
Esters
Neoplasms
Blood vessels
Design of experiments
Inbred C57BL Mouse
Labels
Animals
Blood Vessels
Cause of Death

All Science Journal Classification (ASJC) codes

  • Neuroscience(all)
  • Chemical Engineering(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

Cite this

Lin, Tsung Cheng ; Liao, Ying Chih ; Chang, Wen-Tsan ; Yang, Cheng Han ; Cheng, Li Hsin ; Cheng, Megan ; Cheng, Hung-Chi. / The establishment of a lung colonization assay for circulating tumor cell visualization in lung tissues. In: Journal of Visualized Experiments. 2018 ; Vol. 2018, No. 136.
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The establishment of a lung colonization assay for circulating tumor cell visualization in lung tissues. / Lin, Tsung Cheng; Liao, Ying Chih; Chang, Wen-Tsan; Yang, Cheng Han; Cheng, Li Hsin; Cheng, Megan; Cheng, Hung-Chi.

In: Journal of Visualized Experiments, Vol. 2018, No. 136, e56761, 18.06.2018.

Research output: Contribution to journalArticle

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