and promoting a Th1-mediated immunity (1). So, manipulation of NK cell activation has been regarded as an important approach to the immunotherapy of cancer and infectious diseases (2). The liver is a unique organ containing the highest percentage of NK cells, with 25-30% of intrahepatic lymphocytes in human and 15-20% in mice being NK cells (3). However, in addition to its anatomic characteristics, liver is also proposed as a tolerogenic organ prone to cancer metastasis and chronic infection such as chronic hepatitis B. Therefore, the mechanistic study for the regulation of NK cell function in liver will contribute to better understand the roles of NK cells in liver immunity and design of immunotherapy for the control of liver diseases such as liver cancer.Myeloid-derived suppressor cells (MDSC), a population of CD11b ϩ Gr-1 ϩ myeloid cells at earlier stages of differentiation, represent ϳ20 -30% of normal bone marrow cells and 1-4% of all nucleated cells in spleen (4, 5). MDSC expand dramatically during tumor progression, infection, and even immunization (4, 5). MDSC have been shown to inhibit T cell proliferation and activation, suppress maturation of DCs, which together contribute to the negative regulation of immune responses and the promotion of immune escape of tumors and pathogens (6, 7). In vivo depletion of MDSC with the monoclonal anti-Gr-1 Ab can improve T cellmediated immune responses and suppress tumor growth in murine models (8). Dramatic reduction of MDSC is one of the mechanisms responsible for the potent antitumor effects of all-trans-retinoic acid used in vivo (9). Therefore, depletion of MDSC in tumor-bearing host has been proposed as a new approach for cancer immunotherapy. Although the regulation of adaptive immune response by MDSC is extensively studied, the roles of MDSC in the regulation of innate immunity, especially in the regulation of hepatic NK cell function, have not been completely elucidated.Considering that the immunosuppressive MDSC expand dramatically during tumor progression and NK cells play important roles in the antitumor immunity, we investigated the regulation of NK cell function by MDSC in orthotopic tumor models including orthotopic liver cancer-bearing mice. We show that the cytotoxicity and IFN-␥ production of NK cells in liver are decreased in all kinds of tumor-bearing mice we tested, suggesting a universal phenomenon of impairment of hepatic NK cell function in tumorbearing host. We went further to investigate the underlying mechanisms for the down-regulation of hepatic NK cell function in tumor-bearing host, and demonstrated that MDSC could inhibit NK cell cytotoxicity, NKG2D expression and IFN-␥ production through their membrane-bound TGF-␤1. Also, after interaction with MDSC, NK cells are hyporesponsive to the activating stimuli, indicating MDSC induce NK cell anergy. So, we provide new mechanistic explanation for tumor immune escape by showing the negative regulation of hepatic NK cell-mediated innate immunity by MDSC.

The underlying mechanisms of tumor-induced immune suppression need to be fully understood. Regulatory T (Treg) cells have been shown to play an important role in tumor immune escape. Until now, many subsets of Treg cells have been described that can suppress T cell response via different mechanisms. CD69 is generally regarded as one of the activating markers; however, recent studies show that CD69 may exert regulatory function in the immune response. In this study, we have identified tumor-induced CD69+CD4+CD25− T cells as a new subset of CD4+ Treg cells. CD69+CD4+CD25− T cells increase dramatically along tumor progression, with up to 40% of CD4+ T cells in the advanced tumor-bearing mice. Distinct from the previously described CD4+ Treg cell subsets, CD69+CD4+CD25− T cells express high CD122, but they do not express Foxp3 and secrete IL-10, TGF-β1, IL-2, and IFN-γ. CD69+CD4+CD25− T cells are hyporesponsive and can suppress CD4+ T cell proliferation in a cell-cell contact manner. Interestingly, the fixed CD69+CD4+CD25− T cells still have suppressive activity, and neutralizing Abs against TGF-β1 can block their suppressive activity. We found that CD69+CD4+CD25− T cells express membrane-bound TGF-β1, which mediates suppression of T cell proliferation. Furthermore, engagement of CD69 maintains high expression of membrane-bound TGF-β1 on CD69+CD4+CD25− T cells via ERK activation. Our results demonstrate that CD69+CD4+CD25− T cells act as a new subset of regulatory CD4+ T cells, with distinct characteristics of negative expression of Foxp3, no secretion of IL-10, but high expression of CD122 and membrane-bound TGF-β1. Our data contribute to the better understanding of mechanisms for tumor immune escape.