Spherical Skin Model: Stratified Co-Culture of Fibroblasts and Keratinocytes on Spherical Beads Toward Compound...

Advanced skin models are critical for pursuing non-animal approaches in drug and cosmetic testing. However, existing 3D models remain complex and time-consuming, which limits their adoption. Spherical skin model (SSM) is presented, a platform that balances biological fidelity with experimental robustness. The SSM is based on a core-shell structure where the dermal core is modeled by embedding human fibroblasts into collagen microcarriers (150  $\mu m$ ), while the epidermal shell is formed by outer layers of immortalized keratinocytes. The collagen beads are generated using droplet microfluidics to enable rapid and reproducible production. The biological relevance of SSM is revealed through elevated expression of epidermal differentiation markers (loricrin, involucrin, keratin 1, keratin 10) and the dermal-epidermal junction marker collagen VII. The barrier function is validated by permeability assays that show strong exclusion of fluorescent dextran above 4 kDa. Moreover, their usefulness for screening is shown by identifying a dose-dependent effect of vitamins in reducing oxidative stress and apoptosis against tert-butyl hydroperoxide. As such, this 3D microphysiological model recapitulates key structural, molecular, and functional features of human skin while offering rapid generation, scalability, and compatibility with high-throughput applications in dermatological and cosmetic research.