Preclinical studies frequently lack predictive value for human conditions. Human cell-based disease models that reflect patient heterogeneity may reduce the high failure rates of preclinical research. Herein, we investigated the impact of primary cell age and body region on skin homeostasis, epidermal differentiation, and drug uptake. Fibroblasts derived from the breast skin of female 20- to 30-year-olds or 60- to 70-year-olds and fibroblasts from juvenile foreskin (<10 years old) were compared in cell monolayers and in reconstructed human skin (RHS). RHS containing aged fibroblasts differed from its juvenile and adult counterparts, especially in terms of the dermal extracellular matrix composition and interleukin-6 levels. The site from which the fibroblasts were derived appeared to alter fibroblast-keratinocyte crosstalk by affecting, among other things, the levels of granulocyte-macrophage colony-stimulating factor. Consequently, the epidermal expression of filaggrin and e-cadherin was increased in RHS containing breast skin fibroblasts, as were lipid levels in the stratum corneum. In conclusion, the region of the body from which fibroblasts are derived appears to affect the epidermal differentiation of RHS, while the age of the fibroblast donors determines the expression of proteins involved in wound healing. Emulating patient heterogeneity in preclinical studies might improve the treatment of age-related skin conditions.
- MeSH
- buněčná diferenciace MeSH
- dospělí MeSH
- epidermální buňky cytologie metabolismus MeSH
- fibroblasty metabolismus patologie MeSH
- hojení ran MeSH
- homeostáza MeSH
- kultivované buňky MeSH
- kůže anatomie a histologie cytologie MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladý dospělý MeSH
- předkožka cytologie MeSH
- primární buněčná kultura MeSH
- prsy anatomie a histologie cytologie MeSH
- senioři MeSH
- stárnutí buněk fyziologie MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
Aging depicts one of the major challenges in pharmacology owing to its complexity and heterogeneity. Thereby, advanced glycated end-products modify extracellular matrix proteins, but the consequences on the skin barrier function remain heavily understudied. Herein, we utilized transmission electron microscopy for the ultrastructural analysis of ribose-induced glycated reconstructed human skin (RHS). Molecular and functional insights substantiated the ultrastructural characterization and proved the relevance of glycated RHS beyond skin aging. In particular, electron microscopy mapped the accumulation and altered spatial orientation of fibrils and filaments in the dermal compartment of glycated RHS. Moreover, the epidermal basement membrane appeared thicker in glycated than in non-glycated RHS, but electron microscopy identified longitudinal clusters of the finest collagen fibrils instead of real thickening. The stratum granulosum contained more cell layers, the morphology of keratohyalin granules decidedly differed, and the stratum corneum lipid order increased in ribose-induced glycated RHS, while the skin barrier function was almost not affected. In conclusion, dermal advanced glycated end-products markedly changed the epidermal morphology, underlining the importance of matrix⁻cell interactions. The phenotype of ribose-induced glycated RHS emulated aged skin in the dermis, while the two to three times increased thickness of the stratum granulosum resembled poorer cornification.
- MeSH
- bazální membrána účinky léků ultrastruktura MeSH
- buněčná diferenciace účinky léků MeSH
- epidermis účinky léků ultrastruktura MeSH
- fibroblasty účinky léků ultrastruktura MeSH
- keratinocyty účinky léků ultrastruktura MeSH
- kůže účinky léků ultrastruktura MeSH
- lidé MeSH
- produkty pokročilé glykace metabolismus MeSH
- ribosa farmakologie MeSH
- škára účinky léků ultrastruktura MeSH
- stárnutí kůže účinky léků MeSH
- transmisní elektronová mikroskopie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Non-melanoma skin cancer (NMSC) is the most frequent human cancer with continuously rising incidences worldwide. Herein, we investigated the molecular basis for the impaired skin barrier function of organotypic NMSC models. We unraveled disturbed epidermal differentiation by reflectance confocal microscopy and histopathological evaluation. While the presence of claudin-4 and occludin were distinctly reduced, zonula occludens protein-1 was more wide-spread, and claudin-1 was heterogeneously distributed within the NMSC models compared with normal reconstructed human skin. Moreover, the cancer altered stratum corneum lipid packing and profile with decreased cholesterol content, increased phospholipid amount, and altered ceramide subclasses. These alterations contributed to increased surface pH and to 1.5 to 2.6-fold enhanced caffeine permeability of the NMSC models. Three topical applications of ingenol mebutate gel (0.015%) caused abundant epidermal cell necrosis, decreased Ki-67 indices, and increased lactate dehydrogenase activity. Taken together, our study provides new biological insights into the microenvironment of organotypic NMSC models, improves the understanding of the disease model by revealing causes for impaired skin barrier function in NMSC models at the molecular level, and fosters human cell-based approaches in preclinical drug evaluation.
- MeSH
- diterpeny farmakologie MeSH
- esterasy metabolismus MeSH
- fibroblasty metabolismus MeSH
- keratinocyty metabolismus MeSH
- kofein farmakologie MeSH
- kokultivační techniky MeSH
- kůže účinky léků metabolismus MeSH
- lidé MeSH
- metabolismus lipidů MeSH
- nádorové buněčné linie MeSH
- nádory kůže metabolismus MeSH
- proteiny těsného spoje metabolismus MeSH
- viabilita buněk účinky léků MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
