Regenerative medicine approaches based on mesenchymal stem cells (MSCs) are being investigated to treat several aging-associated diseases, including age-related macular degeneration (AMD). Loss of retinal pigment epithelium (RPE) cells occurs early in AMD, and their transplant has the potential to slow disease progression.The human RPE contains a subpopulation of cells - adult RPE stem cells (RPESCs) - that are capable of self-renewal and of differentiating into RPE cells in vitro. However, age-related MSC changes involve loss of function and acquisition of a senescence-associated secretory phenotype (SASP), which can contribute to the maintenance of a chronic state of low-grade inflammation in tissues and organs.In a previous study we isolated, characterized, and differentiated RPESCs. Here, we induced replicative senescence in RPESCs and tested their acquisition of the senescence phenotype and the SASP as well as the differentiation ability of young and senescent RPESCs.Senescent RPESCs showed a significantly reduced proliferation ability, high senescence-associated β-galactosidase activity, and SASP acquisition. RPE-specific genes were downregulated and p21 and p53 protein expression was upregulated.These findings document the effects of senescence and SASP acquisition on RPESC differentiation ability and highlight the need for a greater understanding of their role in AMD pathogenesis.