The ELISA data of mouse serum showed that mice treated with apoptotic cell-pulsed DCs and a boost of apoptotic cells produced anti-ssDNA and anti-dsDNA antibodies (Fig

The ELISA data of mouse serum showed that mice treated with apoptotic cell-pulsed DCs and a boost of apoptotic cells produced anti-ssDNA and anti-dsDNA antibodies (Fig. their tolerance with apoptotic cell-pulsed dendritic cells for the follow-up of autoantibody levels. The mice in the CD25+ cell-depleted group had higher titres of anti-double-strand/single-strand DNA antibodies than those of the isotype control antibody-treated group. These findings indicated that CD4+CD25+ T cells might be involved in the regulatory mechanism of autoantibody production. (compared to CD25+ Treg cell suppression 005 (D2WF1, = 4; BWF1, = 5). Gene expression profile of CD4+CD25+ T cells in D2WF1 and BWF1 mice We further analysed the expression of some specific genes which are relative to the development and the function of Brivudine Treg cells, such as Foxp3, IL-10 and TGF-. Cells were isolated from splenocytes of both mice at different ages. The purity of CD4+CD25+ T cells was greater than 90%, and that of CD4+CD25? T cells was greater than 95% (data not shown). The mRNA expression of freshly isolated cells was quantified by real-time PCR. We found that the mRNA expression of Foxp3 was predominant in CD4+CD25+ T cells. Foxp3 expression in BWF1 CD4+CD25+ T cells was comparable to that in D2WF1 CD4+CD25+ T cells. The expression level of Foxp3 in CD4+CD25? T cells was used as Brivudine a negative control. Brivudine However, we found that Foxp3 expression in BWF1 CD4+CD25? T cells was higher than that in D2WF1 CD4+CD25? T cells (Fig. 2a). Open in a separate windows Fig 2 Gene expression profiles of CD4+CD25+ cells and CD4+CD25? cells isolated from DBA-2 NZW F1 (D2WF1) mice or NZB NZW F1 (BWF1) mice at different ages. The relative mRNA expression of forkhead ACAD9 box P3 (Foxp3) (a), interleukin (IL)-10 (b) and transforming growth factor (TGF)- (c) was determined by normalizing the expression of each target gene to hypoxanthine phosphoribosyltransferase (HPRT). Bars show mean s.d. * 005, ? 001 (= 5). Moreover, TGF- expression in BWF1 CD4+CD25+ T cells was also normal (Fig. 2c). Both BWF1 CD4+CD25+ and CD4+CD25? T cells expressed higher levels of IL-10 than D2WF1 CD4+CD25+ and CD4+CD25? T cells, and this phenomenon was Brivudine more significant in aged BWF1 mice at 6C11 months of age (Fig. 2b). The suppressive function of CD4+CD25+ T cells from D2WF1 and BWF1 mice To determine the suppressive function of CD4+CD25+ T cells isolated from both mice, different numbers of D2WF1 CD4+CD25+ or CD4+CD25? T cells were added to the culture of CD4+CD25? T cells to suppress their proliferation. It was found that CD4+CD25+ T cells from both D2WF1 (Fig. 3a) and BWF1 mice (Fig. 3b) inhibited the proliferation of autologous CD4+CD25? T cells at a ratio of 025 : 1, but CD4+CD25? T cells did not. The 1 : 1 ratio of CD25+ cells to CD25? cells was then used to compare the activities of D2WF1 CD4+CD25+ T cells with BWF1 CD4+CD25+ T cells. Data showed that CD4+CD25+ T cells have similar suppressive capability and the mean inhibition of D2WF1 and BWF1 mice were 508% and 548%, respectively (Fig. 3c). Open in a separate windows Fig 3 The suppressive function of CD4+CD25+ cells isolated from DBA-2 NZW F1 (D2WF1) mice and NZB NZW F1 (BWF1) mice. Different numbers of CD4+CD25+ T cells or CD4+CD25? cells from D2WF1 mice (a) or BWF1 mice (b) were added to the culture of CD4+CD25? T cells stimulated with autologous T-depleted splenocytes and anti-CD3 monoclonal antibody. (c) CD4+CD25+ T cells from D2WF1F1 mice and BWF1 mice suppressed the proliferation of autologous CD4+CD25? cells at the ratio of 1 1 : 1. Bars show mean s.d. (= 3). CD4+CD25+ T cells play a role in controlling autoantibody production The data above indicate that CD4+CD25+ T cells in BWF1 mice had a deficiency in cell numbers, but had a normal expression level of Foxp3 and normal suppressive functions. To investigate further if the deficiency of Treg cell numbers in BWF1 mice causes the development of lupus-like disease, we depleted CD25+ cells in.