Abstract
OBJECTIVE: Global DNA hypomethylation in rheumatoid arthritis synovial fibroblasts (RASF) contributes to their intrinsic activation. The aim is to explore that an increased polyamine metabolism is associated with a decreased level of S-adenosylmethionine (SAM), causing the global DNA hypomethylation. METHODS: RASF (n = 12) and osteoarthritis synovial fibroblasts (OASF, n = 6) were isolated from synovial tissues. The cells were stained for adenosylmethionine decarboxylase (AMD), spermine/spermidine N1-acetyltransferase (SSAT1), polyamine modulated factor binding protein 1 (PMFBP1), solute carrier3A2 (SC3A2), DNA methyltransferase1 (Dnmt1), integrin α9, and β1, were analyzed by flow cytometry. Nuclear 5-methylcytosine (5-MeC) was measured by flow cytometry, diacetylspermine (DASp) in cell culture supernatants and cell extracts were determined by ELISA and SAM was measured in cell extracts by fluorometry. RESULTS: SSAT1, AMD and PMFBP1 were significantly (p < 0.05) increased in RASF, compared to OASF. DASp in cell culture supernatants and SC3A2 were significantly elevated (p < 0.01) in RASF. The levels of SAM in cell culture extracts, as well as of Dnmt1 protein and 5-MeC were significantly reduced (p < 0.001) in RASF. Parameters of the polyamines metabolism negatively correlated with SAM, Dnmt1 and 5-MeC. Supplementation of RASF with SAM increased Dnmt1 and 5-MeC, whereas integrin β1 was decreased. CONCLUSION: These data clearly show that intrinsic elevations of PMFBP1 and SSAT1 enhance the catabolism and recycling of polyamines in RASF and suggest that a high consumption of SAM by this pathway is an important factor contributing to the global DNA hypomethylation in these cells.
Karouzakis, E