UVR modulates macrophage recruitment and phenotype in the resolution phase of human skin inflammation in vivo

Macrophages (Ms) have an essential role in mediating inflammatory responses. While Ms immediately infiltrate the skin after UVR, it is unknown if and how Ms mediate the resolution phase. To investigate this in vivo, photoprotected human skin was exposed to 3xMED broadband UVR (n=13; 20-58y, phototype I-III) and M distribution was analysed 14 days post UVR. We analysed the pan-M marker CD68 by IHC. Consistent with reports CD68+ cells increased over baseline after 24h (193%; p<0.01). However, M levels remained elevated on D7 (84%; p<0.01) and D10 (46%;p<0.01) post UVR, suggesting they participate in late phases of the sunburn response. To determine if the elevation of CD68+ cells in the resolution phase is attributed to an expansion of the M2 anti-inflammatory population of Ms, immunofluorescence analysis of the M2 marker, CD206, was performed with CD68 (n=3). This revealed the number of CD206+CD68+ cells rose at D4 post UVR and peaked on D10 (219% over D0). The hydroxyl fatty acid, 12-HETE, acts as a chemoattractant for Ms in human skin. We hypothesised 12-HETE could recruit Ms in the skin after UVR. We screened for lipoxygenase enzymes responsible for 12-HETE production and identified ALOX12, -12B and -15 mRNA is expressed in the human epidermis. We further analysed 12-HETE in suction blister fluid post UVR by LC-MS/MS and found that 12-HETE levels significantly correlated with number of CD68+ cells over 14 days (r2=0.98; p<0.001). Collectively we provide evidence that Ms are potentially recruited by UVR stimulated epidermal 12-HETE production and maintained in the late phases of the sunburn response, with the M2 Ms specifically elevated in the resolution phase. Modulation of M recruitment and phenotype could provide a novel strategy to abrogate human UVR skin damage.