Figure 5.

Effect of donor APC and DC depletion on Th17 mediated alloimmune response. Splenocytes were isolated from the recipients (DBA/2 mice) of skin and hind limb grafts-derived from untreated (WT), APC depleted (CL; clodronate treated) and cDC depleted (zDC; DT treated zDC-DTR) donor mice at POD 6 and analyzed for the expression of IL-17 and Hellios in CD4⁺ T cells using flow cytometry. The bar graphs indicate frequencies of Tregs (CD4⁺Hellios⁺) (A) and Th17 (CD4⁺IL17⁺) (B) cells, respectively. The CD3⁺ T cells were gated on CD3⁺CD45⁺ cells. The gating strategies are described in Supplementary Figure 4 . All values are given in % of the described population on the ordinate/”y-axis”. Serum was isolated from the blood samples of the skin and hind limb recipients at POD 6 and subjected to multiplex analysis for the levels of Th17-related cytokines IL17E/IL-25 and lymphotoxin-α. The representative bar graphs are shown (C). (D) Quantitative RT-PCR analysis of IL17/IL-25 and lymphotoxin-α expression in the skin and hind limb grafts at POD 6. (n=8 skin Tx, n=5 hind limb Tx). *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001.