Autoimmune hepatitis (AIH), a severe hepatopathy characterized by hypergammaglobulinaemia, autoantibodies and interface hepatitis, is occasionally associated with systemic autoimmune manifestations [systemic lupus erythematosus (SLE); mixed connective tissue disease (MCTD)]. In both AIH and SLE/MCTD numerical and/or functional impairment of regulatory T-cells (T-regs) is believed to favour autoimmunity. To investigate whether immune-tolerance breakdown profiles differ in patients with AIH and SLE/MCTD, isolated AIH or systemic autoimmunity, we studied phenotypic and functional features of T-regs in 10 patients with AIH-SLE/MCTD, 22 with AIH, 12 with SLE and 20 healthy subjects. Compared to health, CD4 pos CD25 pos cells were decreased in number and expressed high levels of the CD127 activation marker in all three disease groups; in AIH-SLE/MCTD and in SLE they displayed low levels of FOXP3. In AIH-SLE/MCTD, they also contained a high proportion of IFNγ positive cells, indicating a Th1 profile. Similarly, in AIH-SLE/MCTD, CD4 pos CD25 pos CD25 high T-regs were reduced in number and contained an increased proportion of activated CD127 pos and IFNγ pos cells. Such skewing towards a Th1 profile was also present at effector level, as a high frequency of IFNγ-producing cells was observed within AIH-SLE/MCTD CD4 pos CD25 neg responder cells. Impairment in suppressor function both of CD4 pos CD25 pos cells and CD4 pos CD25 pos CD127 neg T-regs was observed in all three autoimmune conditions, but while addition of CD4 pos CD25 pos CD127 neg T-regs decreased CD4 pos CD25 neg responder cell proliferation in healthy subjects and partially in AIH patients, it had no effect in AIH-SLE/MCTD and SLE patients.In conclusion, in AIH-SLE/MCTD T-regs display a distinctive phenotypic and functional signature, characterized by marked activation, elevated IFNγ production and by a profound impairment of suppressive function, suggesting that multiple autoimmune manifestations may derive from a complex defect of immune-regulation.