Cellular senescence can be activated in response to noxious environmental stimuli. A senescent-like phenotype has been detected in the peritoneal mesothelium of mice exposed to high intraperitoneal glucose. We have sought to examine whether high glucose (HG) can induce the senescence program in human peritoneal mesothelial cells (HPMC) in vitro. Senescence of omentum-derived HPMC was induced by serial passages. Cells were cultured in media containing either 5 mM glucose, 30 mM glucose, or 5 mM glucose and 25 mM mannitol (M) for osmotic control. Compared with HPMC cultured in low glucose, the growth rate of cells exposed to HG was significantly decreased so that the cells reached fewer population doublings before entering senescence. Exposure to HG led to increased expression of senescence-associated beta-galactosidase (SA-beta-Gal) and of the cell cycle inhibitors p21(Waf1) and p27(Kip1). Late-passage HPMC exposed to HG displayed marked hypertrophy and released increased amounts of fibronectin and TGF-beta1. These effects were absent from HPMC treated with equimolar M. Exposure of early-passage HPMC to exogenous recombinant TGF-beta1 induced a senescence marker SA-beta-Gal in a dose-dependent manner and mimicked other senescence-associated alterations induced by HG. The addition of anti-TGF-beta1 neutralizing antibody partially reduced the activation of HG-induced SA-beta-Gal. These results indicate that chronic exposure to elevated glucose may result in TGF-beta1-mediated accelerated senescence of HPMC in vitro, which may hypothetically contribute to the peritoneal membrane dysfunction during peritoneal dialysis in vivo.