Type I Interferon (IFN-α/β) therapy has altered the natural course of multiple sclerosis. In this paper we evaluate the possible molecular mechanisms involved in the in vitro effects of IFN-α/β on peripheral blood mononuclear cells from patients with clinically definite Relapsing-Remitting Multiple Sclerosis. The total RNA from IFN-α, IFN-β treated cells and untreated cells was extracted and amplified for CD86, CD28, CTLA-4, TNF-α, IFN-γ, CCL2, CCR5, IL-13, MMP-9, TIMP-1, CD25, TGF-β, IL-10 and the transcriptional factor Foxp3 by Reverse Transcription-Polymerase Chain Reaction and the CD4+CD25high subset was evaluated using flow cytometry. In general, there were no significant differences concerning the modulation of the genes studied in the response to IFN-α and IFN-β treatments, which suggest a similar mechanism of action for both interferons. However, we found a significant increment in IFN-γ expression after IFN-α but not after IFN-β treatments. The in vitro treatment of mononuclear cells from multiple sclerosis patients with both interferons significantly increased the CD25 mRNA. Furthermore, we observed a CD25/Foxp3 correlation and an increment of the CD4+CD25high subset, indicating that the induction of regulatory T cells could be a crucial mechanism involved in the type I interferon effects.