Adenocarcinoma Engraftment: A Method to Implant Cancer Cells into the Murine Lungs

Abstract

Source: Stevens, L.E. et al, Pre-Conditioning the Airways of Mice with Bleomycin Increases the Efficiency of Orthotopic Lung Cancer Cell Engraftment. J. Vis. Exp. (2018).

This video describes a method to engraft lung adenocarcinoma cells in murine lungs by preconditioning the airways with fibrosis-inducing agent–bleomycin. This method significantly enhances orthotopic engraftment of lung cancer cells into murine lungs

Protocol

All procedures involving animals have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Bleomycin Treatment Plan to inject mice intratracheally with bleomycin 14 days prior to tumor cell engraftment. Thaw bleomycin stock on ice 2 h prior to injection. Once thawed, dilute bleomycin to the desired working concentration (0.02 U/50 µL or 0.005 U/50 µL), and keep it on ice. <stron…

Representative Results

Table 1: List of recommended doses of bleomycin for mouse strains tested. Mouse Strain Cell Line Number of cells injected % engraftment p value Vehicle</stron…

Materials

Rodent Intubation stand	Braintree Scientific	RIS-100	Recommended stand for intratracheal injection
Bleomycin	Sigma	B5507-15UN	CAUTION Health hazard GHS08
Exel Catheter 24G	Fisher	1484121	Remove needle. For intratracheal injection
Ketamine (Ketaset inl 100 mg/mL C3N 10 mL)	Butler Schein	56344	To anesthetize mice
Xylazine	Butler Schein	33198	To anesthetize mice
Syringe 1ml	BD / Fisher	14-823-434	For intraperitoneal injections
Puralube Veterinary Ophthalmic Ointment	BUTLER ANIMAL HEALTH COMPANY LLC	8897	To prevent eye dryness while under anesthesia
MI-150 ILLUMINATOR 150W MI-150	DOLAN-JENNER INDUSTRIES	MI-150 / EEG2823M	To illuminate and visualize trachea
Graefe Forceps, 2.75 (7 cm) long serrat	Roboz	RS-5111	For intratracheal injection
Flask T/C 75cm sq canted neck, blue cap	Fisher / Corning	353135
Phosphate Buffered Saline	Life Technologies	14190-144
0.25% Trypsin-EDTA	Life Technologies	25200-056
Pipet-Lite Pipette, Unv. SL-200XLS+	METTLER-TOLEDO INTERNATIONAL	17014411
Countess Automated Cell Counter	Life Technologies	AMQAX1000
D-Luciferin powder	Perkin Elmer Health Sciences Inc	122799	For luminescent imaging. Reconstitute powder with PBS for a working concentration of 15mg/ mL. Protect from Light
IVIS Spectrum Xenogen Bioluminiscence	Perkin Elmer Health Sciences Inc	124262	For in vivo bioluminescence imaging
B6129SF1/J mice	Jackson Laboratories	101043
NIH-H2030 cells	ATCC	CRL-5914
PC9 cells	Nguyen DX et al. Cell. 2009;138:51–62
Living image software	Perkin Elmer Health Sciences Inc	128113	For in vivo bioluminescence analysis