Silymarin flavonoids and cyclosporin A: Effective co-therapy?

Milk thistle (Silybum marianum) is a native plant of Mediterranean Sea areas. Silymarin, an extract from its seeds, is used in various liver diseases for its hepatoprotective effects. It contains silymarin flavonoids (SF) such as silybin, silydianin and silychristin. Cyclosporin A (CsA) is widely used as an immunosuppressant in organ transplantations. Its other applications in many autoimmune diseases were demonstrated. Unfortunately CsA has obligatory negative effects: hepatotoxicity and nephrotoxicity. P-glycoprotein (Pgp) is the main target of CsA in plasma membrane and high affinity of CsA to this membrane transporter is connected with CsA toxicity. The aim of this study was to confirm the hypothesis, that SF compete with CsA for binding sites of membrane proteins in vitro and could protect liver cells against the entry of CsA toxic concentration. Rat hepatocytes were isolated by collagenase perfusion. They were incubated with silybin, silydianin, silychristin and CsA and 3H-labeled CsA was always added. The amounts of ligands bound in hepatocytes were estimated after vacuum filtration of cell suspension with help of radioligand depletion by using the scintillation method. Software GraphPad Prism4 was used for interpretation and statistic evaluation of data. All tested SF depleted 3H-CsA from its bound in hepatocytes. The mechanism of interaction between SF and CsA was evidently competitive. The highest affinity for the target protein has silychristin, IC50 being 0.14 ?M (95% confidence interval for log (IC50) [M] -7.03 to -6.69). IC50 for silybin and silydianin were estimated: 0.22 and 0.23 ?M, resp. (95% confidence intervals for log (IC50) [M] -6.91 to -6.42 and -6.93 to -6.33 resp.). The existence of competitive interaction between SF and CsA in hepatocyte membrane was confirmed and with high probability it is connected with one receptor, Pgp. It is possible to say, that the SF-protection of liver cells by patients with CsA-therapy could be effective.