摘要

Objectives: 1: Develop a 3D in vitro tri-layered model simulating epidermis, papillary and reticular dermis with a porous structure mimicking key human skin functions without using living cells. 2: Quantify effects of sodium hyaluronate, hydrolyzed collagen, elastin, and ceramides on elasticity, firmness, hydration, barrier function, and pore size. 3: Assess impact of ingredient localization in specific layers versus the whole model. 4: Validate predictive value by comparison with available clinical data to support anti-aging claims. Introduction: Anti-aging ingredients play an essential role in aesthetic medicine due to their ability to improve firmness, elasticity, hydration, and barrier function. Conventional efficacy testing often relies on animal models, which pose ethical and regulatory concerns. Materials / method: Each ingredient was incorporated independently into the membrane layer corresponding to its natural site of action in human skin and compared to its inclusion in all three layers. Elasticity, firmness, hydration, barrier function and pore area variation were measured using biophysical assays adapted to the model. Results: The membrane successfully differentiated ingredient-specific effects. Inclusion of the ingredient in the target layer versus all layers yielded distinct outcomes, highlighting the importance of both the layer and the concentration used. Results obtained with the membrane model were consistent with previously published clinical efficacy data, when available, supporting the model’s predictive value. Conclusion: The tri-layered chitosan membrane enables reproducible, ethical, and layer-specific testing of anti-aging ingredients. This innovative approach serves as a reliable alternative to animal models and holds potential for the preclinical evaluation of topical, injectable, and oral applications, aligning with the global transition toward animal-free testing strategies.