Monosodium Urate Crystals Bind with Idiotype Protein and Promote Dendritic Cells to Induce Cytotoxic T Cells in Vitro

Abstract

Monosodium urate (MSU) crystals have been studied to act as a key substance in local immunoreactions. MSU released from damaged cells works as an endogenous danger signal to antigen-presenting cells. MSU crystals evoke specific cell immunity and work as an adjuvant in a mouse model. The crystals also have another unique characteristic to bind with positively charged proteins, which could help to deliver some antigens into human dendritic cells (DCs). We focused on the application of MSU crystals as a not only an adjuvant but also as a carrier of positively charged antigenic protein to induce human cytotoxic T cells (CTLs) efficiently in vitro. We confirmed that MSU crystals facilitated human DCs to express the maturation marker, CD83, deliver (Fab′)2 attaching to the crystals. In order to determine whether MSU crystals facilitate the T-cell proliferation activity of DCs, the proliferative effects of DCs on allogeneic CD4+ cells were investigated. DCs pulsed with MSU crystals significantly facilitated the proliferation of allogeneic CD4+ cells when compared to DCs alone. The stimulation index (SI) was 2.5 ± 0.1 and 1.7 ± 0.1, respectively. When using DCs pulsed with the Fab attached to MSU crystals, the proliferation of CD4+ cells was significantly greater than when using DCs pulsed with Fab alone. The SI was 2.6 ± 0.2 and 1.9 ± 0.1, respectively. No significant differences were seen in the proliferation of allogeneic CD4+ cells between DCs pulsed with the Fab attached to MSU and DCs pulsed with MSU alone. We selected the multiple myeloma IM-9 cell line and its product idiotype (Id) protein as an ideal pair of target cells and positively charged tumor-specific antigen, respectively. After sensitizing DCs derived from HLA-A matched volunteers pulsed with tumor-specific monoclonal IgG-Fab fragments (IM-9 Fab) attached to MSU crystals, the CD8+ T cells stimulated by the DCs killed significantly more target cells (38.5 ± 3.5%, n=4) than those stimulated by DCs pulsed with IM-9 Fab alone (3.5 ± 7.5%). These cytotoxic effects of CD8+ cells stimulated by the DCs pulsed with IM-9 Fab attached to MSU crystals were reduced (3.6 ± 1.7%) when MSU crystals were pre-coated with fetal bovine serum to block to bind with IM-9 Fab. For efficient induction of CTLs, it is necessary for Id proteins to attach to MSU crystals. MSU crystals have some advantages of a protein carrier binding with positively charged proteins and delivering antigenic protein into DCs, as well as an adjuvant promoting DC maturation and inducing CTLs.