Figure 3: Reintroduction of PLCδ1 in PLCδ1^−/− keratinocytes restores normal IL-17 levels and granulocyte counts.

(a) Structure of the Foxn1::PLCδ1 gene. Poly A: the bovine growth hormone polyadenylation sequence. (b) PCR analysis of genomic DNA from the tails of wild-type and Foxn1::PLCδ1 Tg mice. Products derived from Foxn1::PLCδ1 and endogenous PLCδ1. (c) Immunoblotting of PLCδ1 and β-actin in tissues from control, PLCδ1^−/− (KO), and Tg/KO mice. (d) Skin stained with antibody against PLCδ1 (red) and Hoechst stain (blue). Dotted lines denote the dermal-epidermal border. Scale bar, 50 μm. (e) Haematoxylin-eosin (HE) stained dorsal skin sections. Scale bar, 50 μm. (f) The skin was stained with antibodies against CD3 (green) or F4/80 (green) and Hoechst (blue). Scale bar, 100 μm. (g) IL-17 mRNA expression in the skin of control, PLCδ1^−/− (KO), and Tg/KO mice determined by real-time RT–PCR. All values are normalized to GAPDH. Results are displayed as arbitrary units (expression in the skin of KO mice=1). Mean±s.e.m. (n=5). (h) Macroscopic appearance of ILNs in control and Tg/KO mice. Scale bar, 1 mm. (i) Representative FACS profiles of intracellular IL-17 in ILNs. Cells were stimulated and intracellular IL-17 was detected (n=4). Three independent experiments were performed. (j) Absolute numbers of IL-17⁺ cells. Mean±s.e.m. (n=4). The combined results from three independent experiments are displayed. (k) IL-17 concentration in serum measured by ELISA. Mean±s.e.m. (n=3). (l) Representative FACS profiles of CD11b⁺ Gr-1⁺ granulocytes in the PBL, spleen, and BM of Tg/KO mice (n=4). (d–l) 8–12-week-old mice were used. (d–j, l) Heterozygotes with Foxn1::PLCδ1 were used as controls. Statistical significance was assessed using a Student′s t-test. **P<0.01. ND, not detected.