Atherosclerosis and Alzheimer’s disease are wide-spread diseases with rising prevalence. Therefore it is important to uncover more about the diseases themselves and thus understanding and contain the risks they bring along. As blood lipoproteins are involved in nanoplaque formation as well as in inflammatory processes in both diseases, which are linked to PML and its interaction partners, this study concentrates on the question if PML expression is modulated by lipoproteins. As apoE4-homozygous lipoproteins cause a genetic predisposition and thus a high risk for developing Alzheimer’s disease, not only lipoproteins with pooled apoE, but also apoE4-homozygous lipoproteins were used. To determine the role of PML in inflammatory processes in atherosclerosis, endothelial cells (EA.hy926 and HUVECs) were incubated with variations of lipoproteins (of LDL, HDL or in compositions containing VLDL, IDL, HDL and different concentrations of LDL). RT-qPCR, immunoblotting and immunofluorescence analysis showed that PML expression and the number of PML nuclear bodies (PML-NBs) increased after only 3 hours of incubation with the compositions or the single lipoproteins. This increase was especially high when treated with high concentrations of LDL in a composition or as a single apoE-pooled or apoE4-homozygous lipoprotein. Moreover, the activity of protein kinase C (PKC) and the secretion of the cytokines IL-6 and IL-8 were also positively regulated by lipoprotein compositions, pooled and apoE4-homozygous LDL in a dose-dependent manner. To investigate whether PML is a component of this signalling cascade in endothelial cells, EA.hy926 cells were transfected with an expression vector encoding PML, in the presence or absence of a known PKC inhibitor (sc-3088) or PKC activator (PMA). The expression of IL-6 and IL-8 was higher in PML-overexpressing cells, suggesting that IL-6 and IL-8 act downstream of PML. Furthermore, treatment of the cells with sc-3088 demonstrated lower PML expression and PML-NB assembly, while treatment with PMA resulted in opposing effects. Combining the stimulation of the cells with blood lipoprotein compositions and inhibitor treatment or transfection with a PML vector showed that the LDL-induced effect on IL-6 and IL-8 was dependent on PKC activity and PML concentrations. Incubation with LDL for 24 hours as well as activation of PKC further led to an increase in STAT3 expression and activation, while inhibition of STAT3 activity with the inhibitor STATTIC caused PML expression to slightly decrease. Therefore, STAT3 might be another link in this signalling cascade, whereas it was shown to negatively influence the activity of PKC as a feedback. The inhibition of STAT3 activation in endothelial cells resulted in lower expression of IL-6 and IL-8, thus it is assumed that the influence of STAT3 on these cytokines is independent of PML. The LDL-induced PKC activating pathway in endothelial cells also positively affected p53 protein expression. However, experiments with PML-overexpressing cells indicate that this increase is independent of PML. In contrast, eNOS-expression was not influenced by LDL, yet eNOS expression in endothelial cells was induced by HDL. PML-overexpressing cells showed a decrease in eNOS expression. Taken together, the results of this thesis indicate that a so far non-described signalling pathway is present in endothelial cells that includes activation of PKC by LDL in an early response, leading to an increased PML expression, followed by an increase in the expression of the PML targets IL-6 and IL-8. Other factors that are (partly) involved in this signalling cascade are STAT3 and p53.
