Protein-bound P-cresol inhibits human umbilical vein endothelial cell proliferation by inducing cell cycle arrest at G0/G1.

P-cresol is a typical protein-bound uremic toxin, which is retained in patients with renal failure. It is not known whether protein-bound P-cresol exhibits the toxicity in humans. This study aims to investigate the endothelial toxicity of protein-bound P-cresol. Cultured human umbilical vein endothelial cells (HUVEC) were treated with unbound or human serum albumin-bound (HSA, 4 g/dL), P-cresol (0, 20, 40, 80 μg/mL) for 24, 48, 72 h, respectively. Cell viability was determined by using cell counting kit-8 (CCK-8) assay. Cell apoptosis and cell cycle were assessed by using flow cytometry. The expression of cell cycle proteins in HUVEC were analyzed by using western blot and double immunofluorescent labeling assay. The results indicated that the viability of HUVEC was dose- and time-dependently inhibited by the protein-bound P-cresol (77.56% inhibition at 72 h, P<0.05) and unbound P-cresol (80.65% inhibition at 72 h, P<0.05). Most HUVECs were arrested at G0/G1 phase by both protein-bound P-cresol (79.63% inhibition at 72 h, P<0.05) and unbound P-cresol (81.27% at 72 h, P<0.05). Both protein-bound and unbound P-cresol enhanced the expression of p21Cip1 (0.62 and 0.60, both P<0.05) and suppressed the expression of cyclin D1 (0.49 and 0.53, both P<0.05) in a dose-dependent manner. In conclusion, unbound and protein-bound P-cresol inhibit the HUVEC proliferation by inducing cell cycle arrest at G0/G1 phase in a dose- and time-dependent manner, which associates with the up-regulation of p21Cip1 and down-regulation of cyclin D1.