Fig. 6: CGRP inhibits langerin-mediated HSV-2 infection of human LCs and induces atypical langerin double trimers.

a, b Untreated or CGRP-treated MDLCs were pre-incubated for 1 h at 37 °C with nAbs to nectin-1, nectin-2, HVEM or langerin vs. matched isotype non-neutralizing control Abs, or pre-incubated with heparinase II/III vs. medium. Cells were then pulsed for 2 h with HSV-2 (isolates 6 at MOI = 0.5 and isolate 8 at MOI = 2.5) and infection was examined 24 h pi by flow cytometry. Results represent mean ± SEM percentages of HSV-2 infection inhibition (a, n = 8) or normalized against untreated cells (b, n = 6). c Untreated or CGRP-treated inner foreskin epidermal cell suspensions were pre-incubated for 1 h at 37 °C with langerin nAb, pulsed for 24 h with HSV-2 (isolate 7 at MOI = 10.0), and infection was examined 48 h pi by flow cytometry. Results represent mean ± SEM (using n = 3 tissues from different individuals) normalized percentages of HSV-2 infection. d–f Untreated or CGRP-treated MDLCs were pulsed with HSV-2 (isolate 4 at MOI = 0.1 and isolate 7 at MOI = 0.5), alone or in the presence of dynasore, NH₄Cl or bafilomycin A1 as indicated, and infection was measured 24 h pi by flow cytometry. Results represent mean ± SEM (n = 5) normalized percentages of HSV-2 infection. In all graphs, *p < 0.0500, **p < 0.0050 and ***p < 0.0005; two-tailed Student’s t-test. g Representative Western blot showing langerin expression in untreated (lane 1), untreated / DSG cross-linked (lane 2), CGRP-treated (lane 3), and CGRP-treated / DSG cross-linked (lane 4) MDLCs. Langerin appears as ∼47 kDa monomers, ∼120 kDa trimers and ∼240 double trimers. Pie charts (representative of n = 4 experiments) show quantitative analysis of the proportions of each langerin form (p = 0.0038, ANOVA, untreated vs. CGRP-treated).