The accumulation of senescent cells with age has been linked to numerous age-associated pathologies, and may be a result of phenotypic changes such the senescence-associated secretory phenotype (SASP), which disrupts tissue homeostasis and can act in a paracrine manner on neighbouring cells. 
Previously, primary human epithelial cells and fibroblasts were successfully reverted from a deeply senescent (DS) to an early proliferating (EP) phenotype. Comparing EP, DS and senescence reversal cells allowed us to more deeply understand the molecular mechanisms of senescence. To investigate these mechanisms within the context of ageing, a 2D model of senescence has been established by serially passaging human dermal fibroblasts (HDFs) to DS, which display a panel of well-established senescence markers including SA-β-Gal, p16 and p21. HDFs were also induced into premature senescence using UVB and similarly exhibited senescence markers. 
To examine how senescence contributes to the ageing in a tissue-like environment, a 3D living skin equivalent (LSE) of ageing is in development. EP or mixed EP/DS HDFs are suspended in a dermal matrix while proliferating primary human epidermal keratinocytes are seeded on the surface to create an epidermis. Using this model, we can determine paracrine influences of DS HDFs on the skin phenotype, and, in the future, will investigate if by reversing senescence the skin ageing phenotype can be alleviated. LSEs using EP HDFs show epidermal stratification and spatial expression of proteins corresponding to human skin. High-throughput data extraction platforms have allowed us to quantitatively analyse phenotypic changes between LSEs using immunofluorescence. The latest findings with regards to the contribution of senescent fibroblasts to skin ageing will be presented.

Development of a 3D living skin equivalent to explore the influence of senescence on the skin ageing phenotype