613 Everolimus induces HLA-G expression in human coronary artery smooth muscle cells: Impact on graft rejection and cardiac allograft vasculopathy

Abstract

Human leukocyte antigen-G (HLA-G) is a non-classical MHC I protein that plays an important role in immune tolerance. Its expression in serum and endomyocardial biopsies of heart transplant patients is associated with a lower incidence of graft rejection and cardiac allograft vasculopathy (CAV). However, the mechanisms of HLA-G expression remain vague. Previous clinical studies found that patients treated with everolimus expressed significantly higher levels of HLA-G in the blood plasma. Our objective was to determine if everolimus upregulates HLA-G in human coronary artery smooth muscle cells (HCASMC), explaining a potential mechanism of HLA-G expression after heart transplant. We also tested if the soluble HLA-G inhibits HCASMC proliferation, a main cause of intimal hyperplasia associated with CAV. Commercially available HCASMC (n = 4) were cultured in media containing 5% fetal bovine serum. After reaching full confluence, the cells were exposed to various doses of everolimus (0.1-100 ng/ml) for 24 hours. HLA-G expression was assessed by Western blot. We also cultured HCASMC (n = 8) in conditioned media from Jeg-3 cells, choriocarcinoma cells known to release soluble HLA-G. Cell proliferation was measured at various time points (24-120 hours) post treatment. There was a significant dose-dependent increase in HLA-G expression in the cells treated with 100ng/ml of everolimus when compared to the control group (P = 0.012) (Figure) . Interestingly, there was no significant difference in proliferation between the cells treated with HLA-G conditioned media and the control group. We have shown for the first time that everolimus induces HLA-G expression in HCASMC in a dose-dependent manner. We have also shown that soluble HLA-G does not inhibit HCASMC proliferation in an in-vitro setting. Induction of HLA-G expression in HCASMC may represent a promising and novel therapeutic strategy to protect against graft rejection after heart transplantation. Although higher HLA-G levels protect against graft rejection, our results suggest that HLA-G does not inhibit SMC proliferation, implying that other cellular pathways may play a role in reducing CAV formation post transplant.