[Experimental study on characterization of normal canine bladder transitional epithelial cells cultured in vitro].
OBJECTIVE
To explore an effective method to culture and purify canine bladder transitional epithelial cells.


METHODS
Bladder tissue was obtained from healthy puppy under sterile conditions. Bladder mucosa was removed from the remaining tissue with fine scissor and minced into small pieces, and then were dissociated into single cell suspensions with 0. 125% trypsin. The bladder epithelial cells were cultured in defined keratinocyte serum free medium. The cells were passaged and purified by 0.05% trypsin and 0.02% EDTA. Morphological characterization were studied under inverted phase contrast microscope and transmission electron microscope. Expression of cell specific marker protein was assessed by immunohistochemistry.


RESULTS
Canine bladder transitional epithelial cells could be efficiently cultivated and expanded in serum-free medium without fibroblast contamination. The cells could be passaged 4-6 times without a distinguished decrease in cell proliferation. The cells were characterized by well-developed microfilament and desmosome junction under transmission electron microscope. Immunohistochemical staining with broadly reacting anti-cytokeratin antibodies (AE1/AE3) confirmed the epithelial phenotype of the cells. Different generations of cells showed diploid cells.


CONCLUSIONS
A large number of bladder transitional epithelial cells can be obtained from small bladder tissue with our digestion method. The cultured bladder epithelial cells can be proliferated to sufficient quantities for further reconstructive purposes.
Han
Ping
P
Division of Stem Cell and Tissue Engineering, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu Sichuan, 610041, PR China. Hanp2001a@sohu.com
Yang
Zhiming
Z
Li
Xiuqun
X
D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002453:Cell Cycle; D049109:Cell Proliferation; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D004285:Dogs; D004847:Epithelial Cells / Q000166:cytology* / Q000648:ultrastructure; D005260:Female; D005434:Flow Cytometry; D007150:Immunohistochemistry; D008297:Male; D023822:Tissue Engineering / Q000379:methods*; D001743:Urinary Bladder / Q000166:cytology*