MATERIALS AND METHODS

DCs were isolated from the spleen of Wistar-Furth rats by an enzymatic method, according to Chaux et al. 3 By labelling DCs with two specific MoAb (OX62 and ED5, Realef, Paris, France), ffow cytometry analysis showed an 85% to 90% purity of the isolated cells. The yield ranged from 5 105 to 2 106 cells per spleen. Anti CD4 MoAb (ER2, Diaclone, Besançon, France), an IgG2a Ab, is directed against Lewis CD4 molecule. It is nondepleting, modulates about 20% of CD4 molecules of rat spleen T cells, and does not fix the rat complement. It strongly inhibits cellular proliferation in unidirectional mixed splenocyte culture in the Lewis responder-Wistar Furth stimulating (MSCu) combination. A single dose of 3 mg/rat results in a 100% opsonization of circulating CD4 T cells for 12 hours, but only for 6 hours for the CD4 splenocytes. This single dose induces a 48-hour plasma concentration which blocks T cells functions according to in vitro studies. Four groups of 5 Lewis rats were used: a control group, a group receiving only anti-CD4 MoAb (3 mg/rat), a group receiving simultaneously DCs (2 106/rat) and anti-CD4 MoAb, and a group receiving DCs only. The DCs and ER2 were infused by IV (penial vein). Before injection, the DCs were labeled with PKH26 (Sigma, St Quentin Fallavier, France), a chromophore which marked the DCs for more than 100 days without modifying their functional activity. MSCu were performed between Lewis responders and Wistar-Furth stimulating at day 30. Microchimerism was sought in spleen, skin, and thymus using the ffuorescence of previous PKH26 labelling. In addition, the origin of DCs was controlled in splenocyte cell suspensions by ffow cytometry using a MoAb specific of class II molecules of the Wistar-Furth, the RT1Bu/Du (Pharmingen, Montrouge, France). The nonparametric Mann–Whitney test was used for statistical analysis.