Autoantigens ADAMTSL5 and LL37 are significantly upregulated in active Psoriasis and localized with keratinocytes, dendritic cells and other leukocytes

Abstract

Psoriasis is a common immune-mediated disease that affects 2%-4% of individuals in North America and Europe. In the past decade, advances in research have led to an improved understanding of immune pathways involved in the pathogenesis of psoriasis and has spurred the development of targeted therapeutics. Recently, three psoriasis autoantigens have been described: cathelicidin (LL37), a disintegrin and metalloprotease domain containing thrombospondin type 1 motif-like 5 (ADAMTSL5), and lipid antigens generated by phospholipase A2 (PLA2) group IVD (PLA2G4D). It is important to establish the expression, regulation and therapeutic modulation of these psoriasis autoantigens. In this study, we performed immunohistochemistry and two-colour immunofluorescence on non-lesional and lesional psoriasis skin to characterize ADAMTSL5 and LL37, and their co-expression with T cells, dendritic cells, neutrophils and macrophages, which are the main immune cells that drive this disease. Our results showed that ADAMTSL5 and LL37 are significantly (P<.05) increased in lesional skin and are co-expressed by many dendritic cells, macrophages and some T cells in the dermis. Gene expression analysis showed significant (P<.05) upregulation of LL37 in lesional skin and significant downregulation following treatment with etanercept. ADAMTSL5 and LL37 are also significantly decreased by IL-17 or TNF-α blockade, suggesting feed-forward induction of psoriasis autoantigens by disease-related cytokines.

Keywords: ADAMTSL5; LL37/Cathelicidin; dendritic cells; macrophages; psoriasis.

Figure 1

(a) Representative immunohistochemistry images for ADAMTSL5 and LL-37 in lesional skin showed increased expression in keratinocytes and dermal cells compared to NL skin and (b) significant (p<0.05) increase in number of ADAMTSL5⁺ and LL-37⁺ cells in lesional compared to non-lesional skin. Scale bar = 100 μm. Error bars represent standard errors of the mean and p values are as indicated.

Figure 2

(a) Representative two-color immunofluorescence on non-lesional and lesional skin for ADAMTSL5 (red) with CD11c (green) showing co-localization of many ADAMTSL5⁺ cells with CD11c⁺ dendritic cells in the superficial dermis and DEJ. (b) ADAMTSL5 (red) with CD163 (green) showing co-localization of many ADAMTSL5⁺ cells with CD163⁺ macrophages in the superficial and deep dermis. (c) There were also some ADAMTSL5⁺ cells that co-localized with CD3⁺ T-cells in the dermal aggregates. Dermal cells staining positive for both ADAMTSL5 and CD11c, CD163 or CD3 appear as yellow. Scale bar = 100 μm.

Figure 3

(a) Representative two-color immunofluorescence on non-lesional and lesional skin for LL37 (clone OSX12)(green) with CD11c (red) showing co-localization of many LL37⁺ cells with CD11c⁺ dendritic cells near the DEJ. (b) LL37 (clone H7)(red) with CD163 (green) showing co-localization of many LL37⁺ cells with CD163⁺ macrophages in the superficial and deep dermis. (c) Some LL37⁺ cells (red) co-localized with CD3⁺ T-cells (green) in the dermal aggregates. Dermal cells staining positive for both LL-37 and CD11c, CD163 or CD3 appear as yellow. Scale bar = 100 μm.

Figure 4

(a) Representative immunohistochemistry images for ADAMTSL5 and LL-37 on baseline non-lesional, lesional and post-treatment lesional skin of patients treated with Etanercept for 3 months showed increased expression of both antigens in Lesional skin compared to NL skin. (b) Cell counts showed significant (p<0.05) increase of ADAMTSL5⁺ and LL-37⁺ cells in lesional compared to NL skin, and significant decrease in post-treatment compared to baseline lesional skin. (c) qRT-PCR showed significant downregulation of ADAMTSL5 in LS compared to NL skin and upregulation in post-treatment compared to baseline lesional skin. (d) Representative immunohistochemistry images for ADAMTSL5 and LL-37 (clone OSX12) on baseline and post-treatment lesional skin of patients treated with Ixekizumab for 6 weeks showed increased expression of both antigens in Lesional skin compared to NL skin. Scale bar = 100 μm. (e) In patients treated with Brodalumab for 6 weeks, there was significant upregulation in gene expression of LL-37 in lesional compared to NL skin and significant downregulation in post-treatment compared to baseline lesional skin. Shown are average normalized log expression values for non-lesional, lesional and post-treatment (n=6). Error bars represent standard errors of the mean and p values are as indicated.