The development of intimal hyperplasia (IH) after coronary artery bypass graft (CABG) surgery significantly reduces the success rate of this procedure. Transforming growth factor-beta (TGFβ) promotes IH; however the underlying signalling mechanisms remain to be fully elucidated. We aimed to perform mechanistic characterisation of the ALK1 and ALK5 TGFβ signalling pathways in the context of vein graft failure. Wound healing and proliferation assays were performed in primary human saphenous vein SMCs (HSVSMC) treated with rTGFβ1 ± small molecule inhibitors of ALK1 (K02288) or ALK5 (SB525334). Although no effect on proliferation was observed, HSVSMC migration was significantly increased in the presence of SB525334, whereas K02288 treatment significantly reduced migration (p < 0.05 and p < 0.001, respectively). Gene expression analysis revealed that SB525334 treatment enhanced the expression of matrix metalloproteinase-9 (MMP-9), whereas treatment with K02288 reduced MMP-9 expression. The expression of pSmad2/3 and pSmad1/5/8 (the ALK5/ALK1 signalling mediators, respectively) was assessed in sections from mouse and pig vein grafts immediately prior to and 28 days after implantation. pSmad2/3 expression within SMCs was observed in both pre-implantation and grafted vein sections. However, pSmad1/5/8 expression was only detected within SMCs following grafting. Importantly, we also detected pSmad2/3 and pSmad1/5/8 expression within neointimal SMCs in failed vein grafts from Scottish CABG patients, indicating activation of both the ALK5 and ALK1 pathways. Overall, our data suggests that the ALK1/Smad1/5/8 TGFβ signalling pathway is activated following vein grafting and induces specific transcriptional changes to promote VSMC migration. Further mechanistic characterisation is required in order to effectively target TGFβ-mediated IH therapeutically.
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