Long-Time Survival of Bone Allografts in Short-Time Immunosuppressed Rats

For reconstruction of massive bone defects, non-vascularized bone grafts are problematic because of bone resorption, stress fracture, and nonunion. Available vascularized autografts seldom match the shape and mechanical properties of the missing bones. An ideal replacement would be a living bone allograft. At present, the survival of non-autogenous transplants relies on modulation of the immune system, using either long-term immunosuppression (IS) or induction of tolerance. The authors described a method of surgical neoangiogenesis to achieve a long-term survival of a bone allograft with a short-term IS.

Eighty-seven female Dark Agouti (donors) and male Piebald Virol Glaxo rats (recipients) were used. The femur was harvested with its vascular pedicle. The proximal and distal articular surfaces were resected and the medullary canal reamed. The vessels were anastomosed to the right femoral vessels of the recipient. The graft was wrapped in silicone sheeting and placed in a subcutaneous abdominal pocket. The left saphenous arteriovenous (AV) bundle was harvested and implanted into the medullary canal. The rats were randomly allocated to one of four groups (1: non IS/AV bundle ligated; 2: non IS/AV patent; 3: IS/AV ligated; 4:IS/AV patent). IS in Groups 3 and 4 was maintained for 2 weeks with FK506. All of the rats were sacrificed at 18 weeks.

Blood flow in the graft was measured using the hydrogen washout method. Microfil was infused for microangiography. The bones were decalcified and cleared with a modified Spalteholz technique. Capillary density was measured using digital photographs and image analysis software (SCION Image). Histologic analysis quantified bone viability and degree of rejection.

Blood flow in Groups 2 (0.06 ± 0.07 ml/min/ml) and 4 (0.14 ± 0.09) was significantly higher, compared to the control Groups 1 (0.006 ± 0.02) and 3 (0.009 ± 0.03). Blood flow in Group 4 was significant higher, compared to Group 2. The same result was seen for the capillary density (1: 1.98% ± 3.6%; 2: 10.81 ± 6.43; 3: 8.00 ± 8.23%; 4: 28.63 ± 16.75). The histologic grading of bone necrosis was severe in Group 1, moderate to severe in Group 2, moderate in Group 3, and mild to moderate in Group 4.

While short-term allograft survival can be maintained with immunosuppression, life-long use carries significant risks. This experimental study demonstrated that neoangiogenesis from implanted host-derived AV bundles, combined with short-term immunosuppression, maintained blood flow in vascularized bone allografts. This offers a future potential for clinical application.