EpiDerm Full Thickness Skin model and related experiments

The EpiDerm Full Thickness Skin tissue EFT-412 (1.0 cm² surface area) consists of normal, human epidermal keratinocytes and dermal fibroblasts (Fig. 3A, Lower Panel). Both types of cells are cultured to form a multilayered model of the human dermis and epidermis that are mitotically and metabolically active. EFT-412 skin tissue is an ideal in vitro system to study the anti-aging efficacy of cosmetic ingredients and formulations, and is applied in the following experiments.

Tissue viability assay: After overnight pre-incubation, the tissues were transferred to 6-well plates containing 2.5 ml/well fresh medium supplemented with vehicle (PBS) or MB at various concentrations from 0.1 μM to 10 μM. The culture medium was changed daily. After two weeks incubation, tissue viability by MTT assay was performed as described above.

Tissue histological analysis: After two weeks incubation with vehicle or MB, the EFT-412 issues were fixed in 10% formalin solution overnight and transferred in PBS solution on next day. The fixed tissues were sent back to MatTek for histological processing and H&E staining. The IHC staining with the anti-elastin antibody was performed on the cross sections following Abcam IHC-paraffin protocol.

Hydration test: DPM 9003 device (NOVA, USA) attached to a 4-mm sensor probe was used to measure the impedance of EFT-412 skin tissues. Briefly, after two weeks’ incubation in vehicle- or MB-containing medium, the EFT-412 tissue insert was removed from the 6-well plate and placed on a sterile pad in the culture hood. The tissue surface was pat dried with sterile cotton-tipped swabs and then air-dried for additional two minutes. The impedance of four-quarter circles was measured clockwise with the average value representing the hydration of each tissue.