The WP’s progress towards the objectives in the reporting period are summarized below.
WP2 “The regenerative matrix”, different components were prepared to be included in bioscaffolds, e.g. type III collagen and fractions of elastin hydrolysates. First prototypes of 3D collagen-based bioscaffolds were constructed and characterized. For extracellular matrix production by fibroblasts of different origins, protocols were optimized and initial characterizations performed. Three Early-stage researchers (ESRs) participated in a laboratory secondment at ADBC in Beverwijk to study the behaviour of various fibroblasts on biomaterials. Another ESR established a protocol to decellularize dermis of mouse and spiny mouse. During two distinct ESR secondments at HMGU, the basis was laid for collaborative proteomics studies. For this, the host ESR is setting up a histology-directed workflow for the molecular analysis of ECM proteins in an animal model for skin regeneration and in vitro cultured human foetal/adult fibroblasts while performing pilot experiments.
WP3 “Regenerative non-scarring cells”, one ESR has been trained spiny mouse handling and was able to adapt existing histological techniques protocols of tissue to the particularities of Acomys skin. For human foetal, adult, or eschar fibroblasts, protocols regarding e.g. RNA isolation, RT-qPCR and transcriptome sequencing were optimized for mesenchymal cell populations of different origins. To study the human fibroblast lineages accountable for scarless skin regeneration, another ESR successfully established a human induced pluripotent stem cells (iPSC)-derived skin organoid model. In addition, differentiation protocols for blood-derived monocytes to differentiate into pro-inflammatory or anti-inflammatory macrophage subsets were optimized.
WP4 “Introduction of skin appendages and sensory nerves”, focus lies on 1) human sweat gland stem cells, 2) human hair proto-follicles, and 3) human nerve cells. Human iPSC-derived skin organoids were used to isolate and propagate sweat gland stem cells, where sweat gland-specific markers proved that after long-time culture (day 120) the fate choice towards sweat gland formation was achieved. Human hair proto-follicle studies applied hair follicle dermal cell cultures in 2D and bilayered 3D spheroids. To investigate the neurovascular link, human endothelial cells and nerve cells are being included into collagen hydrogels to create a vascular link with nerve cells. For this purpose, human dermal microvascular endothelial cells were isolated and sorted from human skin biopsies and commercial human iPSCs cultures were established.
WP5 “Towards translation into medical devices and ATMPs”, prototypes of an intact three-dimensional collagen biomatrix were produced by different approaches and compounds and the most promising prototype was defined according to key opinion leader meetings, internal evaluation, and market needs. In addition, the first protocol for a melanocyte-containing tissue-engineered skin construct was designed and executed.
In WP6 “Education and training”, ESRs are trained in complementary and specific research skills. Nine training courses on personal skills and four workshops on interrelated fields such as biomaterials, skin tissue engineering, hair biology and regeneration were provided.
