Interleukin 18 Regulate Calcification Of Valvular Interstitial Cells In Vitro

Background and Aim: Calcific aortic valve disease is the main heart valve disease in the elderly.Valvular interstitial cells(VICs)play an important role in the process of valve calcification.Interleukin 18(IL-18)is expressed in stenosis aortic valves and is positively related with the clinical severity of aortic stenosis.However,the role of IL-18 in aortic valve calcification remains unclear.This study isolated and culture VICs in vitro.We examined whether IL-18 promotes myofibroblast and/or osteoblast transdifferention of VICs.Materials and Methods: Porcine valvular interstitial cells were isolated from fresh harvested porcine aortic valves by collagenase digestion.Immunocytochemical staining was used to identify the purities of cell populations.PVICs were cultured in osteogenic media or in normal media and Alkaline phosphatase(ALP)staining was performed.Then PVICs were treated with IL-18.Gene and protein expression of myofibroblastic and osteoblastic markers were tested and nuclear factor κB(NF-κB)phosphorylation was also analyzed.Alkaline phosphatase(ALP)staining and activity assay was also performed.Results: Enzymatic digestion methods isolated the PVICs in vitro successfully.PVICs were positive stained for α-SMA and vimentin and negative stained for CD31.Activity of ALP was higher in experimental group than that of control group after 21 days of osteogenic induction.Our experiments demonstrated that IL-18 significantly enhanced alpha-smooth muscle actin(α-SMA)gene and protein expression.IL-18 treatment also promoted the expression of osteopontin(OPN)and runtrelated transcription factor 2(Runx2)m RNA,although OPN and Runx2 protein expressionswere not changed.IL-18 could induce ALP activity in the presence of conditioning medium.We also demonstrated that IL-18 markedly enhanced NF-κB p65 phosphorylation in VICs.Conclusions: Together these results suggest that IL-18 promotes the myofibroblast differentiation of VICs and accelerates calcification in the presence of conditioning medium via the NF-κB pathway.