Regulatory T-cells play an important role in the regulation of the immune response and the mediation of dominant immunologic tolerance. We have previously shown that these cells are elevated in tumors and blood of patients with glioblastoma multiforme. Heme oxygenase-1, a rate-limiting enzyme in heme catabolism, has also been shown to accumulate during glioma progression and to play a critical role in FoxP3 mediated immune suppression. In this study, we investigated the correlation between FoxP3 and HO-1 expression in patients with various grades of astrocytoma (WHO grade II-IV). Using qualitative and quantitative reverse transcriptase-polymerase chain reaction and quantitative flow cytometry analyses, we analyzed FoxP3 and HO-1 expression in 19 patients with different grades of astrocytoma. We observed the highest level of FoxP3 expression in patients with grade IV tumors (11.54 +/- 1.95%) vs. grade III (6.74 +/- 0.19%) or grade II (2.53 +/- 0.11%) (P < 0.05). Moreover, in grade IV tumors, the frequency of HO-1 mRNA expression in CD4+ CD25+ cells was 11.8 +/- 2.45% vs. 7.42 +/- 0.31% in grade III and 2.33 +/- 0.12% in grade II. Tumor infiltrating Treg stained positively with anti-HO-1 antibody. The expression of HO-1 correlated with CD4+ CD25+ FoxP3+ infiltration (r = 0.966). Our results confirm that HO-1 expressing Treg accumulate during glioma progression. This study also suggests that HO-1 mRNA expression is linked to the induction of Foxp3 in CD4+ CD25+ glioma infiltrating Treg. These findings support the suppressive role played by regulatory T-cells in the growth of malignant brain tumors.