Collective movement of epithelial cells drives essential multicellular organization during various fundamental physiological processes encompassing embryonic morphogenesis, cancer and wound healing. Yet the molecular mechanism that ensures the coordinated movement of many cells remains elusive. Here we show that a tumour suppressor protein, merlin, coordinates collective migration of tens of cells, by acting as a mechanochemical transducer. In a stationary epithelial monolayer and also in three-dimensional human skin, merlin localizes to cortical cell-cell junctions. During migration initiation, a fraction of cortical merlin relocalizes to the cytoplasm. This relocalization is triggered by the intercellular pulling force of the leading cell and depends on the actomyosin-based cell contractility. Then in migrating cells, taking its cue from the intercellular pulling forces, which show long-distance ordering, merlin coordinates polarized Rac1 activation and lamellipodium formation on the multicellular length scale. Together, these results provide a distinct molecular mechanism linking intercellular forces to collective cell movements in migrating epithelia.

Most molecular hallmarks of cellular senescence have been identified in studies of cells aged in vitro by driving them into replicative or stress-induced senescence. Comparatively, less is known about the characteristic features of cells that have aged in vivo. Here we provide a systematic molecular analysis of normal human dermal fibroblasts (NHDFs) that were isolated from intrinsically aged human skin of young versus middle aged versus old donors. Intrinsically aged NHDFs in culture exhibited more frequently nuclear foci positive for p53 binding protein 1 and promyelocytic leukemia protein reminiscent of 'DNA segments with chromatin alterations reinforcing senescence (DNA-SCARS)'. Formation of such foci was neither accompanied by increased DNA double strand breaks, nor decreased cell viability, nor telomere shortening. However, it was associated with the development of a secretory phenotype, indicating incipient cell senescence. By quantitative analysis of the entire secretome present in conditioned cell culture supernatant, combined with a multiplex cytokine determination, we identified 998 proteins secreted by intrinsically aged NHDFs in culture. Seventy of these proteins exhibited an age-dependent secretion pattern and were accordingly denoted 'skin aging-associated secreted proteins (SAASP)'. Systematic comparison of SAASP with the classical senescence-associated secretory phenotype (SASP) revealed that matrix degradation as well as proinflammatory processes are common aspects of both conditions. However, secretion of 27 proteins involved in the biological processes of 'metabolism' and 'adherens junction interactions' was unique for NHDFs isolated from intrinsically aged skin. In conclusion, fibroblasts isolated from intrinsically aged skin exhibit some, but not all, molecular hallmarks of cellular senescence. Most importantly, they secrete a unique pattern of proteins that is distinct from the canonical SASP and might reflect specific processes of skin aging.

Experimental analysis and computational modeling of epidermal wound closure in 3D suggests an important role for surrounding tissue in determining epithelial cell movement and fate.

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Immune cell infiltrates within SCC and at defined ROIs of tumour-surrounding skin in OTRs differ markedly in their composition and microanatomical distribution compared with tumours arising in immunocompetent patients. Our findings substantially broaden the understanding of how long-term systemic immunosuppression modulates the local inflammatory microenvironment in the skin and at the site of invasive SCC.

Cutaneous regeneration utilizes paracrine feedback mechanisms to fine-tune the regulation of epidermal keratinocyte proliferation and migration. However, it is unknown how fibroblast-derived hepatocyte growth factor (HGF) affects these mutually exclusive processes in distinct cell populations. We here show that HGF stimulates the expression and phosphorylation of the microtubule-destabilizing factor stathmin in primary human keratinocytes. Quantitative single cell- and cell population-based analyses revealed that basal stathmin levels are important for the migratory ability of keratinocytes in vitro; however, its expression is moderately induced in the migration tongue of mouse skin or organotypic multi-layered keratinocyte 3D cultures after full-thickness wounding. In contrast, clearly elevated stathmin expression is detectable in hyperproliferative epidermal areas. In vitro, stathmin silencing significantly reduced keratinocyte proliferation. Automated quantitative and time-resolved analyses in organotypic cocultures demonstrated a high correlation between Stathmin/phospho-Stathmin and Ki67 positivity in epidermal regions with proliferative activity. Thus, activation of stathmin may stimulate keratinocyte proliferation, while basal stathmin levels are sufficient for keratinocyte migration during cutaneous regeneration.

Integrating organotypic skin cultures, whole-slide scanning and subsequent image processing provides bioinformatics with a novel source of spatial expression data. We here used this approach to quantitatively describe the effect of treating organotypic skin cultures with sodium dodecyl sulphate in a non-corrosive concentration. We first measured the differentiation-related spatial expression gradient of Heat-Shock-Protein 27 in a time series of up to 24 h. Secondly, a multi-dimensional tissue classifier for predicting skin irritation was developed based on abstract features of these profiles. We obtained a high specificity of 0.94 and a sensitivity of 0.92 compared with manual classification. Our results demonstrate that the integration of tissue cultures, whole-slide scanning and image processing is well suited for both the standardized data acquisition for systems biological tissue models and a highly robust classification of tissue responses.

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