Abstract: AIM To isolate and culture human aortic valve interstitial cells and to identify their morphology, subtype and membrane molecules distribution. METHODS The endothelial cell layers on the dissected valves were removed and the remaining interstitial layers were minced and digested with collagenase I. Cells were then seeded and maintained on culture dishes. Cells were observed under light microscopy and cellular α-smooth muscle actin was detected with confocal staining microscopy. The expressed interferon(IFN)-γ and tumor necrosis factior(TNF)-α receptors were analyzed by flow cytometry with their corresponding monoclonal antibodies. RESULTS The isolated valves interstitial cells were proved to be myofibroblast. More IFN-γ receptors and TNF-α receptors were expressed on rheumatic aortic valves interstitial cells than on non-rheumatic cells and their expression was patients’ sera dependent. CONCLUSION The valves interstitial cells culture system has been established successfully and the membrane molecules differ according to their origins.