ABSTRACT
<p><b>OBJECTIVE</b>To investigate the effects of Toll/interleukin 1 receptor domain-containing protein(TcpC)on macrophages and its mechanisms.</p><p><b>METHODS</b>Murine macrophage J774A cells were co-cultured with TcpC producing wild type E. coli strain CFT073 (TcpC(wt)) or tcpc gene-deleted CFT073 mutant (TcpC(mut)) in Transwell system, respectively. Apoptosis of J774A cells co-cultured with TcpC(wt) or TcpC(mut) was analyzed by Annexin/PI double staining. The levels of reactive oxygen species (ROS) in J774A cells were determined by DCFH-DA staining after treatment with TcpC(wt) or TcpC(mut) at 6 h, 12 h,24 h or 36 h. After the ROS was scavenged by N-acetylcysteine (NAC), the changes of J774A cell apoptosis were also examined. The expression of caspase-3 in J774A cells co-cultured with TcpC(wt) or TcpC(mut) in the presence or absence of 0.1 mmol NAC was detected by Western blot.</p><p><b>RESULTS</b>J774A cells co-cultured with TcpC(wt) for 24 h or 36 h showed significantly increased apoptosis (27.39% ± 4.05% and 28.45% ± 4.55%,respectively) when compared to control group (7.96% ± 1.63% and 10.55% ± 1.44%,P<0.01) or TcpC(mut) group (11.45% ± 2.77% and 19.26%± 2.89%,P<0.01). Levels of ROS in J774A cells treated with TcpC(wt) for 24 h (108.8 ± 9.73) or 36 h (100.3 ± 10.11) were significantly higher than those in control group (56.8 ± 4.11 and 52.8 ± 4.42,P<0.01) or TcpC(mut) (69.7 ± 5.66 and 62.6 ± 4.56, P < 0.01). The pro-apoptotic effects of TcpC(wt) on J774A cells were reversed by 0.1 or 1 mMol NAC treatment. Expression of caspase-3 in J774A cells co-cultured with TcpC(wt) (0.43 ± 0.04) decreased significantly when compared to control group (0.75 ± 0.08,P<0.05) or TcpC(mut) group (0.80 ± 0.12,P<0.05). However,total caspase-3 expression was restored in J774A cells co-cultured with TcpC(wt) in the presence of 0.1 mmol NAC (0.80 ± 0.09).</p><p><b>CONCLUSION</b>TcpC can promote ROS production in macrophages,hereby inducing macrophage apoptosis.</p>
Subject(s)
Animals , Mice , Acetylcysteine , Pharmacology , Apoptosis , Caspase 3 , Metabolism , Escherichia coli , Metabolism , Escherichia coli Proteins , Pharmacology , Macrophages , Metabolism , Reactive Oxygen Species , Metabolism , Virulence Factors , PharmacologyABSTRACT
<p><b>OBJECTIVE</b>To investigate the effects of TcpC on human umbilical vascular endothelial cells (HUVECs) and its mechanisms.</p><p><b>METHODS</b>HUVECs 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.</p><p><b>RESULTS</b>HUVECs 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).</p><p><b>CONCLUSION</b>TcpC secreted from CFT073 can induce apoptosis of HUVECs through mitochondrial pathway, in which PARP is cleaved and Mcl-1 and XIAP expressions are inhibited.</p>
Subject(s)
Humans , Apoptosis , Cells, Cultured , Escherichia coli , Metabolism , Escherichia coli Proteins , Pharmacology , Human Umbilical Vein Endothelial Cells , Pathology , Membrane Potential, Mitochondrial , Myeloid Cell Leukemia Sequence 1 Protein , Metabolism , Poly(ADP-ribose) Polymerases , Metabolism , Virulence Factors , Pharmacology , X-Linked Inhibitor of Apoptosis Protein , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To study the effects of HKL-4 on physiological changes during growth of leaves.</p><p><b>METHOD</b>Using licorice (Glycyrrhiza uralensis) as material, the effects of HKL-4 on active oxygen metabolism and photochemical efficiency in licorice leaf were determined under field condition.</p><p><b>RESULT AND CONCLUSION</b>The contents of chlorophyll, activity of SOD and CAT increased, while the MDA contents in leaves decreased. The senescence was delayed, so that the photochemical efficiency (Fv/Fm) was increasing comparing to the control.</p>