OBJECTIVETo investigate the effects of TcpC on human umbilical vascular endothelial cells (HUVECs) and its mechanisms.

METHODSHUVECs were co-cultured with TcpC secreting wild-type E. coli strain CFT073 (TcpC(wt)) or tcpc gene-deleted CFT073 mutant strain (TcpC(mut)) in transwell system,respectively. Apoptosis of HUVECs was analyzed by Annexin-V/PI double staining. Mitochondrial membrane depolarization was detected by JC-1 staining. Expression of apoptosis-related proteins in HUVECs was determined by Western blot.

RESULTSHUVECs showed morphological changes after co-cultured with TcpC(wt) for 24 h: the cells became detached and cell debris increased,and cell number was also decreased when compared to HUVECs co-cultured with TcpC(mut). The apoptosis of HUVEC cells co-cultured with TcpC(wt) for 24 h significantly increased,compared to that of control group and TcpC(mut) group (60.1% 9.7% compared with 9.0% 1.3% and 16.9% 0.4%,respectively, P<0.05); meanwhile the mitochondrial depolarization of HUVECs co-cultured with TcpC(wt) was significantly increased,compared to that in control and TcpC(mut) groups (64.5% 0.9% compared with 14.5% 2.1% and 15.6% 3.3%, respectively,P<0.05). Cleavage of PARP and inhibition of Mcl-1 and XIAP expression were seen in HUVECs co-cultured with TcpC(wt),but not in groups of control and TcpC(mut).

CONCLUSIONTcpC secreted from CFT073 can induce apoptosis of HUVECs through mitochondrial pathway, in which PARP is cleaved and Mcl-1 and XIAP expressions are inhibited.