FK506 can activate transforming growth factor-beta signalling in vascular smooth muscle cells and promote proliferation.

Aims. FK506 binding protein (FKBP) 12 is an inhibitor of transforming growth factor (TGF)-b type I receptors. Several lines of evidence support the view that TGF-b stimulates vascular smooth muscle cell (VSMC) proliferation and matrix accumulation. We investigated the effect of FK506 on cellular proliferation and on matrix protein production in primary human VSMCs. Methods. We measured cell proliferation with flow cytometry using BrdU incorporation and fluorimetrically by measuring DNA concentration with Hoechst 33258. Western blot assay of whole cell lysates was used to measure the levels of signaling proteins involved in proliferative pathways, namely: b-catenin, pErk, pAkt, pmTOR and cyclin D1. Collagen synthesis was also investigated by Western blot. The TGF-b signal was studied by both Western blot and confocal microscopy. We used the SiRNA technique for FKBP12 gene silencing. Results. Our results show that FK506 stimulates VSMC proliferation and collagen type I production. FK506 enhanced b-catenin levels and activated the Erk, Akt and mTOR kinases, which are important effectors of proliferation. Accordingly, cyclin D1 expression was increased. We also demonstrate that FK506 activates the TGF-b signal in VSMCs and that, through this mechanism, it stimulates cell proliferation. Conclusion. FK506 can act as a growth factor for VSMCs.