Evaluation of direct intramural injection to the bladder wall as a method for developing orthotopic tumor models

Abstract Background Bladder cancer poses a great burden on society and its high rate of recurrence and treatment failure necessitates use of appropriate animal models to study its pathogenesis and test novel treatments. Orthotopic models are superior to other types since they provide a normal microenvironment. Four methods are described for developing bladder cancer models inside the animal's bladder. Direct intramural injection is one of these methods and is widely used. However, its efficacy in model development has not yet been studied. We aimed to evaluate the efficacy and success rate of the direct intramural injection method of developing an orthotopic model for the study of bladder cancer. Method Tumor cell lines were prepared in four microtubes. Aliquots of 200 × 10 3 cells were injected through a 27 gauge needle into the ventral wall of the bladders of 4 male and 4 female BALB/c mice following a midline 1 cm laparotomy incision. In addition, 1 million cells from each microtube were injected into the flanks of control mice. To prevent infection and alleviate pain, 5 mg/kg enrofloxacin and 2.5 mg/kg flunixin meglumine, respectively, were injected subcutaneously. Results Tumors formed in all mice, resulting in 100% take rate and zero post‐operation mortality. Surgery time was ≤15 min per mouse. In two mice, tumors were found in the peritoneal space as well. Conclusion Direct intramural injection is a rapid, reliable, and reproducible method for developing orthotopic models of bladder cancer. It can be done on both male and female mice and only requires readily available surgical tools. However, needle track can result in cell spillage and peritoneal tumors.