*Purpose: Antibody-mediated rejection (AMR) is a refractory rejection after donor-specific antibody (DSA)-positive or blood-type incompatible organ transplantation. Since AMR significantly deteriorates transplant outcomes, there is an urgent need to develop new therapeutic interventions for this intractable rejection.

*Methods: I. Development of a rat AMR model: Orthotopic liver transplantation (OLT) was performed from male Dark Agouti rats (DA, 240-260g) to male Lewis rats (LEW, 280-300g). In the pre-sensitized group (Group-PS), LEW were pre-sensitized by preceding skin transplantation from DA (2 x 2 cm²) 4-6 weeks prior to LT, while sham procedure was performed in non-sensitized controls (Group-NS). Tacrolimus was daily administered (0.1 mg/kg/day) up to a week or until sacrifice after LT to suppress cellular rejections.II. Evaluation of the efficacy of anti-C5 antibody (Anti-C5): Either rat Anti-C5 (20 mg/kg) or vehicle was administered intravenously on post-transplant days (PTD)-0 and -3. Blood and liver tissues were sampled on PTD-1, -3, -7, and -100 in the following 4 groups: Group-NS, -PS, -NS+Anti-C5, and -PS+Anti-C5 (n =5 each).

*Results: I. Group-PS showed increased anti-donor (DA) antibody titers (IgG: 87,100 vs 40,900, P <0.001 and IgM: 30,500 vs22,800, P<0.05) and more C4d deposition in transplanted livers than that in Group-NS (C4d-score: 1.45 vs 0.25, P<0.001) on PTD-7. Serum transaminase levels were significantly higher in Group-PS than in -NS (ALT: 447 vs 223 IU/L, P=0.01 on PTD-1). In particular, biliary damages (ALP: 237 vs 57 IU/L, P <0.001; total bile acid: 334 vs 133 nmol/mL, P<0.001; and total bilirubin: 4.0 vs 0.3 mg/dL, P<0.001) were all characteristically and significantly deteriorated on PTD-7 in Group-PS. Coagulation disorders (PT-INR, P=0.04) and thrombocytopenia (P<0.01) were also noted in Group-PS. Moreover, histopathological grading was significantly deteriorated by skin pre-sensitization (C4d+h-score: 1.9 vs 0.4, P<0.001 on PTD-3 and 4.4 vs 1.2, P<0.001 on PTD-7). Collectively, Group-PS satisfied all the Banff criteria for AMR diagnosis.II. Intrahepatic biliary damages were significantly ameliorated by anti-C5 administration (Group-PS vs Group-PS+Anti-C5 on PTD-7, bilirubin: 4.0 vs 0.7, P<0.001 [Fig.1]; TBA: 334 vs 200, P=0.01; ALP: 237 vs 88, P<0.001), followed by improved PT-INR (P=0.04 on PTD-7). Histopathological evaluation demonstrated significant protection by anti-C5 administration (h-score on PTD-1, -3, and -7: 0.7 vs 0.0, P<0.001; 1.9 vs 0.15, P<0.001; and 3.0 vs 0.5, P<0.001, respectively).

*Conclusions: We developed a rat model of LT-AMR that meets all the diagnostic criteria for AMR, and demonstrated the efficacy of C5-targeted inhibition for this refractory rejection.

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