Involvement of the lipoprotein receptor LRP1 in AMP-IBP5-mediated migration and proliferation of human keratinocytes and fibroblasts

Abstract

© 2020 Japanese Society for Investigative Dermatology Background: Antimicrobial peptide derived from insulin-like growth factor binding protein-5 (AMP-IBP5) is a potent antimicrobial agent that possesses various immunomodulatory activities. The parent protein of AMP-IBP5, IGFBP-5, has been shown to exert its effects via an insulin-like growth factor-1 receptor-independent mechanism, including binding to multifunctional low-density lipoprotein receptor-related protein 1 (LRP1), which contributes to several biological processes involved in skin wound healing. Objectives: To investigate whether LRP1 is involved in AMP-IBP5-induced migration and proliferation of human epidermal keratinocytes and dermal fibroblasts. Methods: The mRNA expression of LRP1 and IGFBP-5 was assessed by quantitative real-time PCR, whereas Western blotting was used to evaluate the protein expression. Production of cytokines was determined by ELISA. Cell migration was measured by the scratch wound assay, whereas cell proliferation was analyzed using the BrdU labeling assay. MAPK activation was determined by Western blotting. Results: We found that AMP-IBP5 markedly induced the migration and proliferation of keratinocytes and fibroblasts, and this effect was reversed by specific siRNA and neutralizing antibody targeting the LRP1 receptor. In addition, LRP1 was upregulated by lipopolysaccharide, flagellin and AMP-IBP5 in keratinocytes and fibroblasts. LRP1 knockdown also inhibited MAPK pathway activation, which was required for AMP-IBP5-mediated cell migration and proliferation, as evidenced by the specific inhibitors for extracellular signal-regulated kinase, c-Jun N-terminal kinase and p38. Conclusions: Our results suggest that LRP1 expressed in human epidermal keratinocytes and dermal fibroblasts contributes to AMP-IBP5-mediated cell migration and proliferation, supporting its crucial role in cutaneous wound healing process.