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1Department of Comparative Medicine, Tulane University, 2Department of Chemical and Biomolecular Engineering, Tulane University
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Bladder tumors are established in female mice in a minimally invasive fashion through catheterization, local cauterization, and subsequent adhesion of carcinoma cells to the burn sites.
Dobek, G. L., Godbey, W. T. An Orthotopic Model of Murine Bladder Cancer. J. Vis. Exp. (48), e2535, doi:10.3791/2535 (2011).
In this straightforward procedure, bladder tumors are established in female C57 mice through the use of catheterization, local cauterization, and subsequent cell adhesion. After their bladders are transurethrally catheterized and drained, animals are again catheterized to permit insertion of a platinum wire into bladders without damaging the urethra or bladder. The catheters are made of Teflon to serve as an insulator for the wire, which will conduct electrical current into the bladder to create a burn injury. An electrocautery unit is used to deliver 2.5W to the exposed end of the wire, burning away extracellular layers and providing attachment sites for carcinoma cells that are delivered in suspension to the bladder through a subsequent catheterization. Cells remain in the bladder for 90 minutes, after which the catheters are removed and the bladders allowed to drain naturally. The development of tumor is monitored via ultrasound. Specific attention is paid to the catheterization technique in the accompanying video.
3. Animal Orthotopic Tumor Model
The instillation of the bladder tumor model described here is a modification of the procedure described by Gunther et al.2
5. Representative Results:
Tumor tissue will begin to appear 3-5 days after the innoculation. In our experience, in a group of 12 mice, 50% or more routinely have a detectable tissue mass within the bladder (0.5 - 1.0 μl, using V=π/6·L·W·H) by day five. When using MB 49 cells, tumors will continue to grow and euthanasia will be required by day 24 if the mice go untreated. Euthanasia is recommended when mice become lethargic or cachectic, display ruffled fur, hunching, and non-responsive behavior, or when they lose 20% of their initial body weight.
The tumors grow quickly and will have necrotic centers after they reach a mass of 200 mg or more,3 ostensibly because the cells in the developing tumor proliferate faster than what can be serviced by the angiogenic blood supply. One can also indirectly monitor tumor progression via the level of hematurea, which is presumably related to rapid angiogenesis within developing tumors. It was consistently noted that the urine of untreated tumor-bearing mice contained blood by day 14. The concentration of red blood cells appeared to increase over time in these mice, and the volumes of urine collected diminished. Tumors in excess of 50 mg can also be palpated with relative ease, typically by 14 days post innoculation.
This minimally invasive method of establishing orthotopic bladder tumors in female mice can also be applied for cell types other than MB49, although differing results should be anticipated. When less-aggressive tumor cells are used, increasing the wattage slightly (using 0.5 W increments to establish the new optimum) will yield an area more amenable to tumor cell attachment, and longer bladder incubation times for the instilled cells will yield a greater amount of cell attachment at the burn sites (unpublished data). Although some protocols recommend delivering 10 W via electrocautery,2 this amount of power should be considered an upper limit, and perhaps be avoided altogether. Increasing the concentration of cells delivered can also increase the chances of success when other cell types are used.
Experiments on animals were performed in accordance with the guidelines and regulations set forth by the Institutional Care and Use Committee of Tulane University.
This work was funded by the National Science Foundation CAREER award (CBET-0846395) and the Department of Comparative Medicine, Tulane University.
|C57Bl/6J mice||Jackson Laboratory||000664||3-5-wk-old|
|Polycarbonate cages and ventilated rack||Lab Products||Super Mouse 750||327mm x
|Hardwood Bedding||Harlan Laboratories||Laboratory –grade SaniChips|
|Rodent diet||Harlan Laboratories||Rodent Diet 2019|
|IV Catheter||MWI Veterinary Supply||009231||24 G x ¾ in.|
|Electrocautery unit||Bovie Medical Corporation||Aaron 900|
|Ultrasound unit||Sonosite||180 Plus (Vet)|
|MB49 cell line||Wake Forest University Baptist Medical Center||N.A.|
|Fetal Bovine Serum||Gemini Bio Products||100106 500A57B|
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