The skin is our largest organ. Ageing of the skin leads to its thinning, loss of barrier function, loss of wound healing efficiency and increased irritability. These are not just 'cosmetic' problems but are extremely important for health and wellbeing in adult and old age. Recent progress in the field including work from the applicants has indicated a major role for cellular ageing (cell senescence) as a cause of age-associated dysfunction and disease in many organs. In fact, this research has shown that suppressing cellular senescence or outright killing the senescent cells that accumulate during ageing can prolong healthy life expectancy in mice. However, it is neither clear whether such a senolytic approach can actually 'rejuvenate' skin nor whether it might work in man. The present project aims to answer these questions. It is well known that senescent cells accumulate in ageing skin. Previously, in work together with Procter&Gamble, largely funded by BBSRC, we have generated a suite of human-based in-vitro and ex-vivo skin models, in which we can modify the numbers of senescent cells. We will now use these tools to examine the consequences of senescent skin cell accumulation for skin function. We know already that skin containing more senescent cells becomes thinner and less able to act as an efficient barrier. We hypothesize that this occurs because senescent cells compromise the differentiation capabilities of the two major cell types in skin: fibroblasts in the dermis and keratinocytes in the epidermis. We will test whether reduction of senescent cell numbers results in more 'youthful' (papillary) dermal fibroblasts and in better proliferation and/or differentiation of epidermal keratinocytes. We will develop and validate a very novel technology to identify senescent cells and to characterize their interactions with neighbouring cells in tissues by combination of an unprecedented number of functional markers in a single image at subcellular resolution. We will examine whether prior reduction of senescent cell numbers in samples of aged skin improves their ability to heal after wounding. Finally, we will test novel candidate substances for senolytic drugs in our human skin models. Presently, not more than a handful of such candidate drugs are known. Our collaboration partner used an in-silico systems pharmacology approach to predict novel senolytics, which we validate in a cell culture screen in a separate industry collaboration. Promising novel candidates will be transferred into the present project and their effect onto skin cell differentiation, skin thickness, barrier function and wound healing efficiency will be established. Our project will show for the first time whether it is possible to ameliorate ageing in a human organ (model) by reducing numbers of senescent cells. It will develop a novel technique to visualise senescent cells in their tissue context with unprecedented specificity and thus improve the understanding of their tissue-specific impact. The project will further provide improved screening tools for the development of scientifically evidence-based skin anti-ageing interventions, and it will validate novel senolytic drugs for use in human skin.