Anti-phosphorylcholine antibodies in cardiovascular disease

Cardiovascular disease (CVD) is the leading cause of morbidity and mortality in the industrialized countries and a growing concern for the developing world. The underlying cause of almost all CVD is atherosclerosis, a process that is characterized by a low-grade inflammation in the artery walls. This inflammation is believed to be initiated by components of low density lipoproteins which, upon oxidation, display pro-inflammatory self-neo epitopes such as malonyldialdehyde (MDA) and phosphorylcholine (PC). Antibodies directed against the PC-epitope (anti-PC) have been researched for several decades but the relatively recent realization that these antibodies also recognize oxidized phospholipids has revolutionized the field and opened up a whole new avenue of investigation. Anti-PC has been shown to aid the clearance of apoptotic cells and prevent the formation of foam cells by clearing oxidized low density lipoprotein. Murine studies with PC-vaccination have shown strong beneficial effects on experimental atherosclerosis in vivo and human epidemiology has consistently linked anti-PC insufficiency to CVD. All humans have detectable anti-PC IgM in serum, though the concentration varies greatly between individuals. Our group has previously shown that people with low serum/plasma levels of anti-PC IgM have increased risk of CVD and the subgroup analysis had indicated that this association was particular strong with regard to the incidence of stroke. In paper I, we tested this hypothesis in a stroke material from northern Sweden. A significant association between low plasma level of anti-PC IgM at baseline and incident stroke was seen for the whole group at anti-PC levels below the 30th percentile (OR 1.62; CI 1.11 to 2.35). Analyses of gender-specific associations indicated fairly strong associations for females, especially at the lowest 30th percentile (OR 2.65; CI 1.41 to 4.95). However, no association was noted for men. Paper II focused on the properties of different anti-PC antibody classes/subclasses. We report that anti-PC IgM, IgA and IgG1 (but not IgG2) were negatively associated with IMT-progression, which is a surrogate marker for atherosclerosis development. Examination of binding profiles revealed that the protective isotypes (IgM, IgA and IgG1) have a different fine-specificity than the non-protective IgG2. Analysis of serum samples taken four years apart in study participants showed that anti-PC IgM titers, essentially, do not change over time. In this paper, we also demonstrate that anti-PC IgM inhibits LPC-induced cell death in vitro and propose that this is yet another protective mechanism. For paper III, I had successfully designed a PC-specific probe to identify, isolate and characterize PC-reactive B cells from ten healthy human donors. We found that all ten had mounted somatically mutated antibodies towards PC utilizing a broad variety of immunoglobulin-genes. PC-reactive B cells were primarily found in the IgM+ memory subset though significant numbers were also detected among naïve, IgG+ and CD27+CD43+ B cells. From the isolated B cells, we derived several human monoclonal antibodies (mAbs) with proven PC specificity. In conclusion, the future of CVD treatment lies with immunomodulation and the premise of this thesis represents one avenue of research in the quest for novel diagnostic tools and improved treatment options. My work has focused on understanding the properties and molecular ontogeny of human anti-PC antibodies. Though this thesis represents one important step in the direction of clinical application, much more research is needed before we will see use of anti-PC mAbs or PC-vaccination in the treatment of patients

Study on administration of 1,5-anhydro-D-fructose in C57BL/6J mice challenged with high-fat diet

1,5-Anhydro-D-fructose (AF) is a mono-saccharide directly formed from starch and glycogen by the action of α-1,4-glucan lyase (EC 4.2.2.13). Our previous study has indicated that AF increases glucose tolerance and insulin secretion in NMRI mice after administration through a gastric gavage in a single dose at 150 mg per mouse. In this study, we used high-fat feeding of C57BL/6J mice to examine the influence of long-term administration of AF on glucose-stimulated insulin secretion in vivo and in vitro. We found that 8-weeks of high-fat feeding increased body weight, fasting blood glucose and insulin levels in C57BL/6J mice when compared to mice fed normal diet. Impaired glucose tolerance was also observed in mice receiving 8-weeks of high-fat diet. In contrast, AF (1.5 g/kg/day), administered through drinking water for 8-weeks, did not affect body weight or food and water intake in mice fed either the high-fat or normal diet. There was no difference in basal blood glucose or insulin levels between AF-treated and control group. Oral glucose tolerance test (OGTT) showed that AF did not affect glucose-stimulated insulin secretion in mice. In in vitro studies with isolated islets, AF did not influence glucose-stimulated insulin secretion in mice receiving either high-fat or normal diet. We therefore conclude that when given through drinking water for 8 weeks at 1.5 g/kg/day, AF has no effect on glucose-stimulated insulin secretion in C57BL/6J mice challenged with a high-fat diet

Avdrag för underskott i utländska fasta driftställen inom Europeiska unionen: En analys av EU-domstolens restriktionsprövning

Det direkta beskattningsområdet har en särskild ställning inom EU-rätten då medlemsstaterna själva i stor utsträckning utformar sin skattelagstiftning. Trots medlemsstaternas relativt starka självbestämmanderätt måste lagstiftningen som utformas fortfarande iaktta och respektera de grundläggande friheterna. Intressemotsättningar som uppstår mellan å ena sidan medlemsstaternas behörighet till att beskatta och å andra sidan etableringsfriheten är ett återkommande grundproblem, och detta tydliggörs bland annat när medlemsstater begränsar möjligheten för inhemska huvudbolag att göra avdrag för underskott i utländska fasta driftställen i syfte att tillvarata trängande allmänintressen. Samtidigt kan en sådan begränsning utgöra en inskränkning av etableringsfriheten. Eftersom en positiv harmonisering på det direkta beskattningsområdet i princip kräver enhällighet får intressemotsättningarna i praktiken lösas av EU-domstolens negativa harmonisering. EU-domstolen tillämpar en restriktionsprövning där bland annat frågan om jämförbarhet, den väl avvägda fördelningen av beskattningsrätten mellan medlemsstaterna och principen om slutlig förlust fått en allt större betydelse.In the field of direct taxation Member States have retained broad sovereignty which makes it a special area within European law. The European Union and its organs lack taxing powers with regard to direct taxes, but the powers retained by the Member States must nevertheless be exercised consistently with the fundamental freedoms of the European Union. This gives rise to tension between two opposing systems, and raises a fundamental difficulty, namely the conflict between the power conferred on the Member States to tax income arising in their territory and the freedom conferred on Community nationals to establish themselves within the Community. This is particularly evident when national provisions only allow losses incurred by a local permanent establishment to be accounted for when calculating the profits and taxable income of the principal company, with regards to overriding reasons in the public interest. Solving this fundamental difficulty through positive harmonization requires unanimity which is hard to reach. In the absence of positive harmonization, the European Court of Justice plays an important role with its negative harmonization. The Court of Justice uses a discrimination-based analysis in which the question about comparability, the balanced allocation of the power to impose taxes between the different Member States and the principle of final losses have gained significance

Induction of Dendritic Cell–Mediated T-Cell Activation by Modified but Not Native Low-Density Lipoprotein in Humans and Inhibition by Annexin A5

International audienceObjective— Atherosclerosis is an inflammatory disease, where activated immunocompetent cells, including dendritic cells (DCs) and T cells are abundant in plaques. Low-density lipoprotein modified either by oxidation (oxLDL) or by human group X-secreted phospholipase A2 (LDLx) and heat shock proteins (HSP), especially HSP60 and 90, have been implicated in atherosclerosis. We previously reported that Annexin A5 inhibits inflammatory effects of phospholipids, decreases vascular inflammation and improves vascular function in apolipoprotein E −/− mice. Here, we focus on the LDLx effects on human DCs and T cells. Approach and Results— Human DCs were differentiated from peripheral blood monocytes, stimulated by oxLDL or LDLx. Naive autologous T cells were cocultured with pretreated DCs. oxLDL and LDLx, in contrast to LDL, induced DC-activation and T-cell proliferation. T cells exposed to LDLx-treated DCs produced interferon-γ, interleukin (IL)-17 but not IL-4 and IL-10. Annexin A5 abrogated LDLx effects on DCs and T cells and increased production of transforming growth factor-β and IL-10. Furthermore, IL-10 producing T cells suppressed primary T-cell activation via soluble IL-10, transforming growth factor-β, and cell–cell contact. Lentiviral-mediated shRNA knock-down HSP60 and 90 in DCs attenuated the effect of LDLx on DCs and subsequent T-cell proliferation. Experiments on DC and T cells derived from carotid atherosclerotic plaques gave similar results. Conclusions— Our data show that modified forms of LDL such as LDLx but not native LDL activate human T cells through DCs. HSP60 and 90 contribute to such T-cell activation. Annexin A5 promotes induction of regulatory T cells and is potentially interesting as a therapeutic agent

Induction of dendritic cell-mediated T-cell activation by modified but not native low-density lipoprotein in humans and inhibition by annexin a5: involvement of heat shock proteins

International audienceObjective— Atherosclerosis is an inflammatory disease, where activated immunocompetent cells, including dendritic cells (DCs) and T cells are abundant in plaques. Low-density lipoprotein modified either by oxidation (oxLDL) or by human group X-secreted phospholipase A2 (LDLx) and heat shock proteins (HSP), especially HSP60 and 90, have been implicated in atherosclerosis. We previously reported that Annexin A5 inhibits inflammatory effects of phospholipids, decreases vascular inflammation and improves vascular function in apolipoprotein E −/− mice. Here, we focus on the LDLx effects on human DCs and T cells. Approach and Results— Human DCs were differentiated from peripheral blood monocytes, stimulated by oxLDL or LDLx. Naive autologous T cells were cocultured with pretreated DCs. oxLDL and LDLx, in contrast to LDL, induced DC-activation and T-cell proliferation. T cells exposed to LDLx-treated DCs produced interferon-γ, interleukin (IL)-17 but not IL-4 and IL-10. Annexin A5 abrogated LDLx effects on DCs and T cells and increased production of transforming growth factor-β and IL-10. Furthermore, IL-10 producing T cells suppressed primary T-cell activation via soluble IL-10, transforming growth factor-β, and cell–cell contact. Lentiviral-mediated shRNA knock-down HSP60 and 90 in DCs attenuated the effect of LDLx on DCs and subsequent T-cell proliferation. Experiments on DC and T cells derived from carotid atherosclerotic plaques gave similar results. Conclusions— Our data show that modified forms of LDL such as LDLx but not native LDL activate human T cells through DCs. HSP60 and 90 contribute to such T-cell activation. Annexin A5 promotes induction of regulatory T cells and is potentially interesting as a therapeutic agent