Experimental limb transplantation, part i: identification of an effective tapered triple combination immunosuppressive regime

Abstract

Limb transplantation was performed across the Brown Norway to Fischer 344 histocompatibility barrier in rats to evaluate the effects of triple combination immunosuppressive therapeutic regimens. Sixty rats were divided into five groups: group I (F344 to F344) isograft controls group II (BN to F344) allograft controls received no immunosuppressive treatment. Groups III and V (BN to F344) received various exposures to tacrolimus (TRL), mycophenolate mofetil (MMF) and prednisolone (Pred) for two weeks: namely, group III: TRL 0.5 mg/kg/d; MMF 10 mg/kg/d; Pred 0.5 mg/kg/d; group IV: TRL 2 mg/kg/d, MMF 15 mg/kg/d, Pred 0.5 mg/kg/d; and group V: TRL 3 mg/kg/d; MMF 20 mg/kg/d; Pred 0.5 mg/kg/d. After 2 weeks, group III and V animals underwent a simultaneous 20% taper of Pred and MMF each further week such that by week 7 the animals were only on TRL. At this time TRL was tapered at the same rate (20% every week) to a maintenance dose of 0.6 mg/kg/d. Evidence of rejection was sought by daily visual observation for swelling, redness, erythema, edema, or skin necrosis. Salvage treatment was used only if rejection occurred after the first 7 weeks, namely, reversing to 100% of the initial TRL dose in that group for 2 weeks with a subsequent taper. Skin and muscle biopsies were obtained from grafted limbs on day 3, 13, 24, 35, and at the endpoint (9 months or uncontrollable rejection). There was no rejection in group I, while all animals showed acute rejection as expected in group II. All group III rats displayed a similar though delayed acute rejection, showing that the regimen was not therapeutic. Rats in group IV displayed the best results, namely, 10 of 12 (83%) with no rejection or side effects at 9 months. Rats in group V displayed numerous, unacceptable side effects due to overtreatment with a 1-month mortality rate of 50%. This study shows that low-dose TRL in combination with MMF and Pred may achieve excellent long-term results of composite tissue transplants. TRL can be used alone as maintenance therapy following an initial loading dose and a tapering period. Rejection is easily reversed by only temporarily increasing the TRL dose.