Cyclosporine and sirolimus inhibit calcium channels in rat aorta vascular smooth muscle cells

Diogo Maio1, Elisa Cairrao1, Flavio Reis2, Ignacio Verde1. 1University of Beira Interior (UBI), Head of Health Sciences Research Centre (CICS-UBI), Av. Infante D. Henrique, 6200-506 Covilha, Portugal, Portugal, 2University of Coimbra, Laboratory of Pharmacology & Experimental Therapeutics, IBILI, Medicine Faculty, 3000-548 Coimbra, Portugal

Post transplantation hypertension produces serious cardiovascular risks that jeopardize a long-lasting, successful organ transplant. Cyclosporine (CsA) is among the most widely used immunosuppressant for preventing graft rejection and autoimmune diseases. However, its clinical use is hampered by its significant nephrotoxicity and has also been implicated in the development of endothelial dysfunction and transplant vasculopathy. Sirolimus (rapamycin, SRL), is a macrocyclin lactone with a novel mechanism of immunosuppressive action. Its high degree of synergy with CsA would not only more efficiently prevent rejection but also allow minimization of CsA-induced toxicity. However, little is yet known concerning the mechanisms underlying the protection and whether the vasomotor effects are distinct from those reported for CsA. Preliminary studies using animal models have provided conflicting results. The aim of this work was to determine the vascular effects of SRL and/or CsA in A7r5 cells from rat aorta. We used the whole cell patch clamp and the planar cell surface area (PCSA) techniques to analyse L-type calcium channels (LTCC) activity and cell contractility, respectively.

The relaxation responses induced by CsA and SRL on noradrenaline (NA, 1 μ M), serotonin (5-HT, 1 μ M) and Bay-K (0.1nM) contractions were assessed using PCSA. CsA relaxed to a maximum of 140% over the contractile effect of 5-HT, 120% over NA and 110% over Bay-K effects. SRL relaxed to a maximum of 100% over the contractile effects of 5-HT and NA and to a maximum of 125% over Bay-K effect. Both compounds relaxed the effect of the contractile agents (45-140%). The electrophysiological analysis showed that CsA (8nM-12 μM) and SRL (10nM – 10 μM) inhibit LTCC. The addition of SRL (10 μM) to CsA (4 μM) potentiated the LTCC inhibition.

Our data suggest that SRL and CsA cause relaxation of A7r5 cells through the inhibition of LTCC.

Understanding the molecular mechanisms of CsA and SRL is important for designing specific immunosuppressant that could efficiently control blood pressure and improve long term transplant conditions.