This study was undertaken to investigate whether plasmacytoid dendritic cells (PDC) are involved in the generation of a higher proportion of CD4(+) and CD25(+) regulatory T (Treg) cells in chronic hepatitis B virus (HBV) infection compared with healthy patients. The amount, phenotype, and function of Treg cells in CD4(+) T cells primed by PDC from 46 chronic HBV patients, 25 healthy controls, and 10 individuals with resolved HBV infection were studied by flow cytometry, reverse transcription polymerase chain reaction, enzyme-linked immunosorbent assay, and proliferation assay. CD4(+) T cells primed by PDC from chronic HBV patients were more effective than CD4(+) T cells primed by PDC from the healthy controls and the resolved HBV patients in suppressing the hepatitis B core antigen-specific proliferation and the interferon production. The interleukin-10 and transforming growth factor-beta1 could also be detected in the supernatants of the PDC-primed CD4(+) T cells. A higher percentage of Treg cells, defined as CD4,CD25, CD45RO, and cytotoxic T-lymphocyte-associated antigen 4-positive cells, was detected within the population of CD4(+) T cells primed by PDC from chronic HBV patients compared with the healthy controls and individuals with resolved HBV infection. Accordingly, CD25(+) Treg cells from PDC-primed CD4(+) T cells displayed a high Fox P3 messenger RNA level. Depleting CD4(+) and CD25(+) Treg cells from CD4(+) T cells primed by PDC from the chronic HBV patients, the healthy volunteers, and the resolved HBV patients made PDC-primed CD4(+) T lose the capability in suppressing HBV-specific T cells. PDC from the patients with chronic HBV infection induced the generation of a higher proportion of CD4(+) and CD25(+) Treg cells compared with the healthy patients.