TLR2 stimuli enhance HIV susceptibility in mLCs and resident epidermal LCs. mLCs were stimulated via TLR2 using heat-killed Gram⁺ bacteria (HKLM; 0.2, 1, 5 × 10⁸/mL), synthetic agonists (Pam₃CSK₄, 0.2, 1, 5 μg/mL; FSL1, 2, 10, 50 μg/mL), or Gram⁺ bacterial components (LTA, 0.4, 2, 10 μg/mL; PGN, 0.2, 1, 5 μg/mL) for 24 hours at various concentrations, as has been described in “Methods” (A-C). Cells were stimulated via TLR2 before (B) and after (C) HIV exposure. The expression of CD86 (A) and percentage of positive cells for HIV p24 (B,C) were assessed in langerin⁺ CD11c⁺ mLCs or CD11c⁺ mDCs (MFI, mean fluorescence intensity). Epithelial sheets obtained from suction blister roofs were preincubated with 5 × 10⁸/mL HKLM or 20 μg/mL poly(I:C) for 2 hours, and then exposed to R5 HIV (D). The emigrating cells from the epidermal sheets were collected 3 days after HIV exposure, and HIV-infected epidermal LCs were assessed by HIV p24 intracellular staining (D). Representative FACS analyses of CD11c and p24 mAb double-stained cells are shown. For the assessment of HIV transmission from LCs to CD4⁺ T cells, emigrated LCs were collected 3 days after HIV exposure and washed, and then 10⁴ LCs were cocultured with 2 × 10⁶ allogeneic CD4⁺ T cells for 12 days. p24 protein levels in culture supernatants were assessed by ELISA on the indicated days (E). Results are shown as means plus or minus SD (n = 3). *P < .05; **P < .01. All data shown represent at least 2 separate experiments.