1. Epidermal SR-A Complexes Are Lipid Raft Based and Promote Nucleic Acid Nanoparticle Uptake
- Author
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Eric W. Roth, Xiao Qi Wang, Amy S. Paller, Adam Ponedal, Michael Bonkowski, Chad A. Mirkin, Thomas R. Holmes, and Qian Song
- Subjects
0301 basic medicine ,Keratinocytes ,Primary Cell Culture ,Human skin ,Dermatology ,Endocytosis ,Biochemistry ,Article ,03 medical and health sciences ,0302 clinical medicine ,Membrane Microdomains ,Nucleic Acids ,medicine ,Humans ,Scavenger receptor ,Receptors, Immunologic ,Molecular Biology ,Lipid raft ,Cells, Cultured ,Heat-Shock Proteins ,Gene knockdown ,integumentary system ,Chemistry ,Membrane Proteins ,Scavenger Receptors, Class A ,Cell Biology ,Cell biology ,030104 developmental biology ,medicine.anatomical_structure ,030220 oncology & carcinogenesis ,Gene Knockdown Techniques ,Spherical nucleic acid ,Nucleic acid ,Nanoparticles ,Epidermis ,Keratinocyte - Abstract
Scavenger receptors clear pathogens, transport lipid, and mediate polyanionic ligand uptake in macrophages, but their expression and role in skin is poorly understood. Although the epidermal barrier typically excludes nucleic acid entry, topically applied, spherically-arranged oligonucleotide nanoconjugates (spherical nucleic acids or SNAs) penetrate mouse skin, 3D skin equivalents, and human skin. We explored the mechanism of SNA uptake in normal human epidermal keratinocytes (NHEKs) and 3D skin equivalents. NHEKs and 3D raft treatment with class A scavenger receptor (SR-A) inhibitors reduced SNA uptake by >80%. Human epidermis expresses SR-As scavenger receptor class A type 3 (SCARA3) and, to a lesser extent, macrophage receptor with collagenous structure (MARCO). Simultaneous lentiviral knockdown of SCARA3 and MARCO reduced SNA uptake in NHEKs and 3D rafts after topical application, affirming a role for SR-As in SNA uptake and 3D raft penetration. Incubation of NHEKs at 4°C or with sodium azide prevented SNA uptake, suggesting active endocytosis. Endocytosis inhibitors, immunofluorescence, immunoprecipitation, and knockdown studies localized functional SR-As to flotillin-1 (FLOT-1)-containing lipid rafts throughout the epidermis and caveolin-1 (CAV-1)-containing rafts only in the upper epidermis. These studies suggest a central role for SR-A complexes in epidermal lipid rafts in mediating uptake of nucleic acid-laden nanoparticles.
- Published
- 2020