Heat shock protein (65 kDa) – Stimulated proliferation of human saphenous vein smooth muscle cells is inhibited by thapsigargin

Aim: Heat shock protein (HSP) expression has been widely implicated in atherogenesis. Vein graft failure following coronary artery bypass graft surgery (CABG) involves medial thickening and neointima formation, a phenomenon mediated by vascular smooth muscle cell (VSMC) proliferation. Superimposed atherogenesis can then result in vein graft failure in as many as 50% of cases within ten years after surgery. In order to explore the possible role of HSP in mediating vein graft disease, the effect of HSP 65 kDA on the proliferation of VSMCS obtained from human saphenous vein was investigated. The role of calcium was also studied using thapsigargin which blocks the release of calcium from intracellular storage pools. Methods: VSMCs were grown from human saphenous vein using standard culture methods. Cells were grown to confluence using DMEM + 10% Foetal Calf serum (FCS). When confluent, cells were trypsinised and cultured onto 96-well plates and rendered quiescent with 0.4% FCS. HSP 65kDA, over a range of concentrations (with and without 10 nM thapsigargin) was added to the cells and proliferation stimulated with 10% FCS and further incubated for 48 hours. Proliferation was assessed by the uptake of 5-bromo-2′ deoxyurindine (Brdu) using colorimetric ELISA and cell counts from which% changes in proliferation were calculated. Results: HSP elicited a dose-dependent increase in the proliferation of VSMCs (table 1) an effect completely inhibited by the presence of 10 nM thapsigargin (table 1). This concentration of thapsigargin has previously been shown to inhibit VSMC proliferation through depletion of intracellular calcium pools. Conclusions: Since HSP 65 kDa stimulates the proliferation of VSMCs, they may play a role in graft thickening and neointima formation as well as superimposed atherogenesis. As this may result in late vein graft failure, further investigations into the role of HSP expression in vein graft disease is warranted. The inhibition of this HSP-mediated effect by thapsigargin consolidates the crucial role of calcium in mediating VSMC proliferation and that inhibition of this event represents a potential therapeutic strategy for the prevention of vein graft thickening