Dolph Houben

80 CHAPTER 4 Post-Operative Care All animals received two weeks of prophylactic antibiotic therapy with Enrofloxacin 7.5mg/kg (Baytril, Bayer Healthcare LLC, Shawnee Mission, KS) and Ceftiofur 100mg/kg (Excede, Pfizer Inc, New York, NY). Direct weight-bearing was allowed after recovering from anesthesia. Daily observation permitted administration of appropriate analgesics, used until no evidence of pain was observed. The animals were individually housed and received standard feed and water ad libitum. Pigs were anesthetized at 2, 4, 6, 10 and 20 weeks for wound treatment, ultrasound assessment of vascular pedicle patency/thrombosis and radiography. At two weeks the immunosuppression was stopped, wound dressing changed, and the central venous catheter was surgically removed. Calcein (20mg/kg) and Oxytetracycline (20mg/kg) were administered by intramuscular injection 10 days apart prior to sacrifice. Sacrifice Procedure After the 20 week survival period all animals were anesthetized (Telazol + Xylazine, IM), and euthanized with intravenous administration of phenobarbital as recommended by the Panel on Euthanasia of the American Veterinary Medical Association and performed according to NIH guidelines under the direction of the Institutional Animal Care and Use Committee. The procedure was performed in a sterile manner. The femoral vessels were dissected and injected with Microfil (Microfil, MV-122, Flow Tech, Carver, MA). A Micro-CT of the whole tibia was made with all soft tissue attached, after removing the plates and screws. Next, the tibia was further dissected, and the allotransplant segment removed and divided into segments with an oscillating saw allowing multiple analyses of the allotransplant. For control purposes, the contralateral tibia was also removed, scanned, embedded and analyzed. Matched segments of contralateral normal and allotransplant bone were used for biomechanical analysis. Imaging Studies Ultrasound Doppler ultrasound imaging (Vivid 7 dimension, GE Medical Systems, Horten, Norway) was used to evaluate the allogenic vascular pedicle patency over time. Since the allogenic vascular pedicle to the transplant was tunneled subcutaneously for anastomosis to the femoral vessels, biweekly assessment of the proximal pedicle patency was possible. We identified the proximal femoral artery and end to side anastomosis first with color Doppler imaging. By following the end of the anastomosis and lining up the skin mark, we were able to identify the subcutaneous vascular pedicle. The pedicle was considered patent if a triphasic Doppler signal could be generated and color Doppler imaging showed a pulsating subcutaneous artery. Studies were performed until pedicle rejection, determined at 2,4, 6 and 10 weeks postoperatively.