3D-Human dermis equivalent as in vitro biological platform for in vitro assessment of photodamage

In this study we validated our 3D-human dermis equivalent (3D-HDE), as an in vitro biological platform to study ECM modification under simulated solar exposure and as screening platform for testing cosmetic agents. 3D-HDE has been realized by means of a tissue engineering bottom-up approach. Due to endogenous and three-dimensional features of this tissue model is possible to analyze structure, composition and ECM assembly, tissue remodeling due to damage and photo-protective action of active compound. Samples have been investigated by Multiphoton Microscopy (MPM), immunofluorescence analyses (IF) and histological analyses. Second harmonic generation (SHG) imaging has been exploited to assess modification in endogenous collagen assembly with and without UV irradiation. Texture analysis has been performed by Grey Level Co-occurrence Matrix on the SHG images in order to assess the correlation length (LC) a parameters indicating the coarsening degree of the collagen matrix. The results revealed that 3D-HDE UVA irradiated suffered a strong disappearance of fibroblasts; nuclei with irregular morphology and ECM alterations. Furthermore, IF test showed an up-regulation of both MMP-1 and MMP-9 and histological analyses by Picro Sirius Red staining showed a decrease of both mature and immature collagen. According to literature data on photoaged skin, 3D-HDE UVA irradiated showed an increase in Lc respect to control, it means that collagen degeneration causing change in textural features of collagen matrix. Moreover, active compound tested as photoprotectans show reduce the damage due to UV irradiation both at cellular and ECM level. Taken together these results highlight that our model is unique in the capability to recapitulate in vitro relevant phenomena occurring in the native dermis during human daily life such as chronic exposure to sunlight.