Non‐depleting anti‐CD4, but not anti‐CD8, antibody induces long‐term survival of xenogeneic and allogeneic hearts in α1,3‐galactosyltransferase knockout (GT‐Ko) mice

The anti‐galactose‐α1,3‐galactose (Gal) antibody (Ab) response following pig‐to‐human transplantation is vigorous and largely resistant to currently available immunosuppression. The recent generation of GT‐Ko mice provides a unique opportunity to study the immunological basis of xenograft‐elicited anti‐Gal Ab response in vivo, and to test the efficacy of various strategies at controlling this Ab response [1]. In this study, we compared the ability of non‐depleting anti‐CD4 and anti‐CD8 to control rejection and antibody production in GT‐Ko mice following xenograft and allograft transplantation. Hearts from baby Lewis rat or C3H mice were transplanted heterotopically into GT‐Ko. Non‐depleting anti‐CD4 (YTS177) and anti‐CD8 (YTS105) Abs were used at 1 mg/mouse, and given as four doses daily from day −2 to 1 then q.o.d. till day 21. Xenograft rejection occurred at 3 to 5 days post‐transplantation in untreated GT‐Ko recipients, and was histologically characterized as vascular rejection. Anti‐CD4, but not anti‐CD8, Ab treatment prolonged xenograft survival to 68 to 74 days and inhibited anti‐Gal Ab as well as xeno‐Ab production. In four of the five hearts from anti‐CD4 mAbs‐treated GT‐Ko mice, we observed classic signs of chronic rejection, namely, thickened intima in the lumen of vessels, significant IgM deposition, fibrosis and modest mononuclear cell infiltrate of Mac‐1 + macrophages and scattered T cells (CD8 > CD4). Xenograft rejection in untreated, as well as anti‐CD4‐ and anti‐CD8‐treated, recipients was associated with increased intragraft IL‐6, IFN‐γ and IL‐10 mRNA. C3H allografts were rejected in 7 to 9 days by untreated GT‐Ko mice and were histologically characterized as cellular rejection. Treatment with anti‐CD4 and anti‐CD8 mAb resulted in graft survivals of > 94.8 and 11.8 days, respectively. Anti‐CD4 mAb treatment resulted in a transient inhibition of alloreactive and anti‐Gal Ab production. The presence of circulating alloreactive and anti‐Gal Abs at > 50 days post‐transplant was associated with significant IgM and IgG deposition in the graft. Yet, in the anti‐CD4 mAb‐treated group, the allografts showed no signs of rejection at the time of sacrifice (> 100 days post‐transplantation). All rejected allografts had elevated levels of intragraft IL‐6, IFN‐γ and IL‐10 mRNA, while the long‐surviving anti‐CD4‐treated allografts had reduced mRNA levels of these cytokines. Collectively, our studies suggest that the elicited xeno‐antibody production and anti‐Gal Ab production in GT‐Ko mice are CD4 + T‐cell dependent. The majority of xenografts succumbed to chronic rejection, while allografts survived with minimal histological change, despite elevated levels of circulating alloAbs. Thus, immunosuppression with anti‐CD4 mAb therapy induces long‐term survival of allografts more effectively than to xenografts.