Interaction between castanospermine an immunosuppressant and cyclosporin A in rat cardiac transplantation

Abstract

Aim: To investigate the interaction between castanospermine and cyclosporin A (CsA) and to provide an explanation for it.

Methods: The alkaloid castanospermine was prepared from the seeds of Castanospermum austral consistently achieving purity. Rat heterotopic cardiac transplantation and mixed lymphocyte reactivity were done using genetically inbred strains of PVG (donor) and DA (recipient). For the mixed lymphocyte reaction stimulator cells were irradiated with 3000 rads using a linear accelerator. Cyclosporin A was administered by gavage and venous blood collected 2 h later (C2). The blood levels of CsA (Neoral) were measured by immunoassay which consisted of a homogeneous enzyme assay (EMIT) on Cobas Mira. Statistical analyses of interactions were done by an accelerated failure time model with Weibull distribution for allograft survival and logistic regression for the mixed lymphocyte reactivity.

Results: Castanospermine prolonged transplant survival times as a function of dose even at relatively low doses. Cyclosporin A also prolonged transplant survival times as a function of dose particularly at doses above 2 mg/kg. There were synergistic interactions between castanospermine and CsA in the prolongation of cardiac allograft survival for dose ranges of CsA by castanospermine of (0 to 2) mg/kg by (0 to 200) mg/kg (HR = 0.986; 95%CI: 0.981-0.992; P < 0.001) and (0 to 3) mg/kg by (0 to 100) mg/kg (HR = 0.986; 95%CI: 0.981-0.992; P < 0.001) respectively. The addition of castanospermine did not significantly increase the levels of cyclosporin A on day 3 or day 6 for all doses of CsA. On the contrary, cessation of castanospermine in the presence of CsA at 2 mg/kg significantly increased the CsA level (P = 0.002). Castanospermine inhibited mixed lymphocyte reactivity in a dose dependent manner but without synergistic interaction.

Conclusion: There is synergistic interaction between castanospermine and CsA in rat cardiac transplantation. Neither the mixed lymphocyte reaction nor the metabolism of CsA provides an explanation.

Keywords: Cardiac transplantation; Castanospermine; Cyclosporin A; Mixed lymphocyte reaction; Positive interaction.

Figure 1

Cardiac graft survivals in rats treated with a range of doses of castanospermine only, a range of doses of cyclosporin A only or a combination of both. The doses of CAST and CsA are given in mg/kg per day. When the two drugs are used together the survival is greater than the sum of the two drugs alone (P < 0.001 when dose of CsA and dose of CAST are treated as continuous variables): Compare CsA 2 mg/kg alone plus CAST 100 mg/kg alone with the combination of CsA 2 mg/kg and CAST 100 mg/kg. CAST: Castanospermine; CsA: Cyclosporin A.

Figure 2

Mean number of lymphocytes for increasing doses of cyclosporin A by dose of castanospermine. The doses of CAST and CsA are given in umol/L. There is a reduction in lymphocyte count for increasing doses of CsA or increasing doses of CAST. The absolute reduction in lymphocytes for a given dose of CsA decreases with decreasing doses of CAST. CAST: Castanospermine; CsA: Cyclosporin A.

Figure 3

Natural logarithm of the mean number of lymphocytes for increasing doses of cyclosporin A by dose of castanospermine. The doses of CAST and CsA are given in umol/l. There is a dose dependent reduction in the logarithm of the lymphocyte count for CsA alone (P < 0.001) or for CAST alone (P < 0.001). But when the reduction is analysed there is not a synergistic interaction (P = 0.89). CAST: Castanospermine; CsA: Cyclosporin A.