Abstract: In multiple sclerosis (MS), the immune damage to the central nervous system results from the net balance between self-reactive and immunoregulatory cells, among other factors. We identified novel perforin-expressing regulatory B-cells (BReg) in patients with clinically isolated syndrome, significantly enriched within the cerebrospinal fluid when compared to peripheral blood, of memory B cell phenotype (CD19+CD25+, CD19+CD25+FoxP3+ and CD19+FoxP3+, p=0.007, p=0.06 and p=0.03, respectively). These BReg subsets were also higher in relapsing–remitting MS during relapse symptoms than in non-clinically active MS patients. Suppressive effects by CD19+CD25+hi BReg on CD4+ T cell proliferation seem to be mediated at least in part by perforin/granzyme pathway. To our knowledge, this is the first report that shows cytolytic perforin/granzyme granule storage in B cells; the interesting point is its involvement on BReg cell immunosuppressive mechanisms, similarly to that in TReg cells. Our data may extend the understanding of pathophysiological processes in MS immunoregulation. [Copyright &y& Elsevier]