Characterization of Innate Immune Responses of Human Endothelial Cells Induced by Porphyromonas gingivalis and Their Derived Outer Membrane Vesicles.

Atherosclerosis, a chronic inflammatory disease of the blood vessels, is one of the most common causes of morbidity and mortality world-wide. Involvement of Porphyromonas gingivalis in atherosclerosis is supported by observations from epidemiological, clinical, immunological, and molecular studies. Previously we reported that P. gingivalis vesicles have a much higher invasive efficiency than their originating cells. Here, we further compare the role of P. gingivalis cells and their vesicles in expression of chemoattractant proteins including CXCL1, CXCL2, and CXCL8, and adhesive molecules such as E-selectin in human umbilical vein endothelial cells (HUVECs). Both P. gingivalis 33277 cells and vesicles were able to up-regulate expression of these molecules, while the vesicles acted as more potent inducers of the inflammatory response associated with the development of atherosclerosis, consequently resulting in significant monocyte adhesion to a monolayer of HUVECs. Interestingly, we found that elevated expression of CXCL8 and E-selectin in endothelial cells induced by P. gingivalis correlated with the invasive ability of P. gingivalis cells and vesicles. Non-invasive bacterial cells and vesicles had no effect on expression of these genes. This study highlights the potential risk of P. gingivalis cells and vesicles in initiation of atherosclerosis and provides a potential target for the development of novel therapeutics against bacteria-associated atherosclerosis.

Inhibition of endoplasmic reticulum stress and atherosclerosis by 2-aminopurine in apolipoprotein e-deficient mice.

We previously reported that the apolipoprotein (apo) B48-carrying lipoproteins obtained from apoE knockout (apoE (-/-) ) mice, so called E(-)/B48 lipoproteins, transformed mouse macrophages into foam cells and enhanced the phosphorylation of eukaryotic translation initiation factor 2 α (eIF-2 α ). Furthermore, the eIF-2 α phosphorylation inhibitor, 2-aminopurine (2-AP), attenuated E(-)/B48 lipoprotein-induced foam cell formation. The present report studied the effect of 2-AP on atherosclerosis in apoE (-/-) mice. Our results showed that the level of food intake, bodyweight, plasma cholesterol, and triglycerides was comparable in apoE (-/-) mice treated with or without 2-AP. However, the mean size of atherosclerotic lesions in the aorta sinus as well as the surface area of the entire aorta of 2-AP-treated apoE (-/-) mice were reduced by about 55% and 39%, respectively, compared to samples from untreated control apoE (-/-) mice. In addition, the 2-AP-treated apoE (-/-) mice showed a significant decrease in glucose-regulated protein 78 (GRP78) and phosphorylated eIF-2 α in their aortic samples as compared to levels in untreated control apoE (-/-) mice. These observations suggest that endoplasmic reticulum stress is a causal mechanism for the development of atherosclerosis in apoE (-/-) mice and that therapeutic strategies can be developed for using eIF-2 α phosphorylation inhibitors, such as 2-AP, to prevent or treat atherosclerosis.

Regulation of the activity and expression of aryl hydrocarbon receptor by ethanol in mouse hepatic stellate cells.

BACKGROUND: During the course of alcohol-induced liver damage, hepatic stellate cells are transformed into proliferative, fibrogenic, and contractile myofibroblasts. Aryl hydrocarbon receptor (AhR) is a transcription factor that controls the expression of genes involved in the metabolism of xenobiotics, inflammation, cell proliferation, and death. METHODS: Immortal mouse hepatic stellate cells (MHSCs) were isolated from transgenic mice that expressed a thermolabile SV40 tumor antigen. Quantitative real-time reverse transcription polymerase chain reaction assays, Western blot analysis, promoter activity assays, and chromatin immunoprecipitation analyses were performed for studying the effect of ethanol (EtOH) on AhR expression and transcriptional activity. RESULTS: Treatment of MHSCs with 50 to 200 mM EtOH for 6 hours induced AhR nuclear translocation, enhanced the promoter activity of cytochrome P450 (CYP) 1A1, increased the amount of AhR bound to the promoter of CYP1A1 and 1B1, and up-regulated the mRNA expression of these AhR target genes in a dose-dependent manner. In contrast, EtOH exposure down-regulated AhR mRNA and protein expression. Similarly, benzo(a)pyrene (BaP) at 10 nM reduced AhR and increased CYP1A1 and 1B1 mRNAs. Pretreatment of MHSCs with 50 mM EtOH for 7 days diminished the capacity of MHSCs to express CYP1A1 and 1B1 induced by a 200 mM EtOH challenge, or by 10 nM BaP. However, the up-regulatory effect of EtOH on solute carrier family 16, member 6 (SLC16a6) was unaffected by EtOH pretreatment. Similar to EtOH, dimethyl sulfoxide (DMSO) at concentrations of 50 to 100 mM down-regulated AhR and up-regulated CYP1A1 mRNA expression in a dose-dependent manner. CONCLUSIONS: These data, for the first time, demonstrate that EtOH activates MHSC AhR and down-regulates its expression. Chronic EtOH pretreatment lowers the availability of AhR, and specifically diminishes the inducibility of CYP genes. The effect on AhR appears to not be an EtOH-specific response, as DMSO alone (and possibly other organic solvents) was also able to activate AhR.

Retardation of atherosclerosis by overexpression of catalase or both Cu/Zn-superoxide dismutase and catalase in mice lacking apolipoprotein E.

Oxidative stress has been suggested to potentiate atherogenesis. However, studies that have investigated the effect of antioxidants on atherosclerosis showed inconsistent results, ie, atherosclerosis was either retarded or not changed by dietary antioxidants. This report directly examined the effect of overexpressing Cu/Zn-superoxide dismutase (Cu/Zn-SOD) and/or catalase on atherosclerosis and lipid peroxidation in mice lacking apolipoprotein E (ApoE-/-). Based on lipid staining of the en face of the aorta tree and the serial sections of the proximal aorta, ApoE-/- mice overexpressing catalase or both Cu/Zn-SOD and catalase had smaller and relatively early stages of atherosclerotic lesions (eg, foam cells and free lipids) when compared with ApoE-/- mice, who developed more advanced lesions (eg, fibrous caps and acellular areas). In addition, the retarded development of atherosclerosis was correlated with a reduced F2-isoprostanes in the plasma and aortas in ApoE-/- mice overexpressing catalase or both Cu/Zn-SOD and catalase. In contrast, the levels of F2-isoprostanes and atherosclerosis in the ApoE-/- mice overexpressing Cu/Zn-SOD alone were comparable to ApoE-/- control mice. These observations implied that endogenously produced hydrogen peroxide, but not superoxide anions, contributed to the formation of oxidized lipids and the development of atherosclerosis in ApoE-/- mice.