Screening of integrins localized on the surface of human epidermal melanocytes

In vivo, melanocytes occupy three-dimensional (3D) space. Nevertheless, most experiments involving melanocytes are performed in a two-dimensional microenvironment, resulting in difficulty obtaining accurate results. Therefore, it is necessary to construct an artificial in vivo-like 3D microenvironment. Here, as a step towards engineering a precisely defined acellular 3D microenvironment supporting the maintenance of human epidermal melanocytes (HEMs), we examined the types of integrin heterodimers that are expressed transcriptionally, translationally, and functionally in HEMs. Real-time PCR and fluorescent immunoassay analyses were used to elucidate the expression of integrin α and β subunit genes at the transcriptional and translational levels, respectively. The functionality of the presumed integrin heterodimers was confirmed using attachment and antibody-inhibition assays. Among the genes encoding 12 integrin subunits (α₁, α₂, α₃, α₄, α₅, α₆, α₇, α_V, β₁, β₃, β₅, and β₈) showing significantly higher transcription levels, proteins translated from the integrin α₂, α₄, α₅, β₁, β₃, and β₅ subunit genes were detected on the surface of HEMs. These HEMs showed significantly increased adhesion to collagen I, fibronectin, laminin, and vitronectin, and functional blockade of the integrin α₂ subunits significantly inhibited adhesion to collagen I, fibronectin, and laminin. In addition, there was no significant inhibition of the adhesion to fibronectin or vitronectin in HEMs with functional blockade of the integrin α₄, α₅, or α_V subunits. These results indicate that the active integrin α₂β₁ heterodimer and the inactive integrin α₄, α₅, α_V, β₃, and β₅ subunits are all localized on the surface of HEMs.