Ultrasound-Guided Intramural Inoculation of Orthotopic Bladder Cancer Xenografts: A Novel High-Precision Approach

Abstract

Orthotopic bladder cancer xenografts are essential for testing novel therapies and molecular manipulations of cell lines in vivo. Current xenografts rely on tumor cell inoculation by intravesical instillation or direct injection into the bladder wall. Instillation is limited by the lack of cell lines that are tumorigenic when delivered in this manner. The invasive model inflicts morbidity on the mice by the need for laparotomy and mobilization of the bladder. Furthermore this procedure is complex and time-consuming. Three bladder cancer cell lines (UM-UC1, UM-UC3, UM-UC13) were inoculated into 50 athymic nude mice by percutaneous injection under ultrasound guidance. PBS was first injected between the muscle wall and the mucosa to separate these layers, and tumor cells were subsequently injected into this space. Bioluminescence and ultrasound were used to monitor tumor growth. Contrast-enhanced ultrasound was used to study changes in tumor perfusion after systemic gemcitabine/cisplatin treatment. To demonstrate proof of principle that therapeutic agents can be injected into established xenografts under ultrasound guidance, oncolytic virus (VSV) was injected into UM-UC3 tumors. Xenograft tissue was harvested for immunohistochemistry after 23–37 days. Percutaneous injection of tumor cells into the bladder wall was performed efficiently (mean time: 5.7 min) and without complications in all 50 animals. Ultrasound and bioluminescence confirmed presence of tumor in the anterior bladder wall in all animals 3 days later. The average tumor volumes increased steadily over the study period. UM-UC13 tumors showed a marked decrease in volume and perfusion after chemotherapy. Immunohistochemical staining for VSV-G demonstrated virus uptake in all UM-UC3 tumors after intratumoral injection. We have developed a novel method for creating orthotopic bladder cancer xenograft in a minimally invasive fashion. In our hands this has replaced the traditional model requiring laparotomy, because this model is more time efficient, more precise and associated with less morbidity for the mice.

Suppression of progranulin expression inhibits bladder cancer growth and sensitizes cancer cells to cisplatin

“…We then assessed growth as previously described [20, 21]. After 20 days, tumors derived from UMUC-3/shScr control cells reached an average of 200 mm 3 of tumor volume (Figure 7A); in contrast, the progranulin depleted UMUC-3/shPGRN showed a significantly reduced tumor growth (*** P < 0.001, Figure 7A).…”

“…Briefly, 22 nine weeks old female athymic nude mice (NU-Foxn1 nu , Harlan Laboratories, Indianapolis, IN) with the average weight of 24.3 g were assigned into 2 groups by weight. Tumor inoculation was performed by ultrasound-guided injection according to the technique developed in our laboratories [20, 21]. Mice were anesthetized with 2.5% isoflurane plus meloxicam 1 mg/ Kg (Boehringer Ingelheim Vetmedica, St. Joseph, MO) subcutaneously.…”

High-Resolution Ultrasound Allows Percutaneous Initiation and Surveillance of Prostate Cancer in an Orthotopic Murine Model

“…Recently, a urologic research group demonstrated successful percutaneous injection and ultrasound guidance of intramural orthotopic bladder cancer xenografts with consistent tumor engraftment in different cell lines. Furthermore, and of great interest, they were able to perform intratumoral injections in all mice of a control group, demonstrating the potential of these techniques for ultrasound-guided injections [23]. …”

Effect of contrast media on urinary cytopathology specimens

“…Orthotopic murine xenograft muscle invasive bladder cancer model 13 All animal procedures were performed according to the guidelines of the Canadian Council on Animal Care (CCAC). The protocol was approved by the University of British Columbia animal care committee (protocol number A10-0192).…”

“…The protocol was approved by the University of British Columbia animal care committee (protocol number A10-0192). Briefly, five 10-week-old female athymic nude mice (Harlan Labs, Indianapolis, IN, U.S.) were anesthetized and UM-UC3 luciferase firefly-expressing cells were suspended in Matrigel ® (BD Biosciences, Mississauga, ON, Canada) and injected into the submucosa of the bladder wall via ultrasound guidance 13 ( Fig. 1A).…”