Wound healing revised: A novel reepithelialization mechanism revealed by in vitro and in silico models

Kai Safferling,1,2 Thomas Sütterlin,1,2 Kathi Westphal,1,2 Claudia Ernst,1,2 Kai Breuhahn,3 Merlin James,1,2 Dirk Jäger,1,2Niels Halama,1,2 and Niels Grabe1,2, 1Hamamatsu Tissue Imaging and Analysis Center, BIOQUANT, and 2Department of Medical Oncology, National Center for Tumor Diseases, University of Heidelberg,69117 Heidelberg, Germany, 3Institute of Pathology, University Hospital of Heidelberg, 69120 Heidelberg, Germany

Wound healing is a complex process in which a tissue’s individual cells have to be orchestrated in an efficient and robust way. We integrated multiplex protein analysis, immunohistochemical analysis, and whole-slide imaging into a novel medium-throughput platform for quantitatively capturing proliferation, differentiation, and migration in large numbers of organotypic skin cultures comprising epidermis and dermis. Using fluorescent time-lag staining, we were able to infer source and final destination of keratinocytes in the healing epidermis. This resulted in a novel extending shield reepithelialization mechanism, which we confirmed by computational multicellular modeling and perturbation of tongue extension. This work provides a consistent experimental and theoretical model for epidermal wound closure in 3D, negating the previously proposed concepts of epidermal tongue extension and highlighting the so far underestimated role of the surrounding tissue. Based on our findings, epidermal wound closure is a process in which cell behavior is orchestrated by a higher level of tissue control that 2D monolayer assays are not able to capture.


EFT-400, wound healing, reepithelialization, wound closure, IL-1alpha, IL-3, IL-6, IL-8, MCP-1, and granulocyte colony-stimulating factor (G-CSF), paracrine signaling, epidermal proliferation, KI-67, desmoglein 1, occludin, E-cadherin, connexin 43, actin, and P-cadherin, biopsy punch, phospho-histone 3, laminin-5, TUNEL assay, cytokeratin 10, involucrin, filaggrin,  cytokeratin 14, CMFDA, cell tracker green, cell tracker red, CMTPX

Materials Tested

Punch biopsy wound

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