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The aim of the present study was to determine whether it was possible to repair urethral defects with a material of adipose mesenchymal stem cells (ADMSCs)-porous silk fibroin (SF). A total of 39 male New Zealand white rabbits were randomly divided into a control group, an SF group and a bromodeoxyuridine (BrdU)-labeled ADMSCs-SF group (SSF group; n=13/group). Defects were made by resecting the posterior urethral wall. The defects in the SF and SSF groups were repaired using SF and BrdU-labeled ADMSCs-SF materials respectively. Then the anterior wall was sutured, and the urethral catheter was retained for 3 weeks following surgery. The catheter was rinsed with nitrofurazone once a day. The cells with positive expressions of factor VIII related antigen (FVIII-RAg), α-smooth muscle actin (α-SMA) and pan-cytokeratin (AE1/AE3) were detected by immunohistochemical assay, and the distributions of BrdU positive cells and macrophages were observed. Urethrography was performed prior to and following surgery. All rabbits had normal urethral morphologies prior to surgery. The incidence rates of postoperative complications in the control, SF and SSF groups were 76.92 (7/13), 23.07 (3/13) and 15.38% (2/13), respectively (P<0.05). The number of positive macrophages in the SSF group was significantly lower than that of the SF group 4 weeks following surgery (P<0.05). In the SSF group, BrdU positive cells were scattered within the SF material following surgery, primarily at the intersection between the SF material and the urethra. The number of FVIII-RAg positive cells in the SSF and SF groups were significantly different (P<0.05), which were also significantly higher than that of control group (P<0.01). The number of α-SMA positive cells in the SSF and SF groups were significantly different (P<0.05), and these values also significantly exceeded those exhibited by the control group (P<0.01). In addition, the SSF and SF groups had positive staining of AE1/AE3. Similar to normal urethral mucosa, the cytoplasm was stained brownish yellow (P<0.05). It is thus feasible to repair urethral defects using ADMSCs-SF material. [ABSTRACT FROM AUTHOR]