PGE2 and other lipids in rheumatic diseases

Despite numerous options for treatment of rheumatic diseases, there is an unfulfilled clinical need for therapeutic strategies that can reduce inflammation and prevent tissue destruction. Lipid mediators (eicosanoids and fatty acids (FA)) are involved in the regulation of inflammatory processes and contribute to the pathogenesis of rheumatic diseases. Thus, selective targeting of the lipid mediators might enable improved antiinflammatory treatment. Microsomal prostaglandin synthase (mPGES) -1 produces prostaglandin E2 (PGE2) at sites of inflammation in rheumatic diseases. Inhibitors of mPGES-1 have been proposed as a more selective anti-inflammatory treatment retaining the therapeutic potential of non-steroidal anti-inflammatory drugs (NSAIDs) but with less severe side effects associated with NSAIDs. However, the impact of mPGES-1 inhibition on different pathological and physiological processes is not completely elucidated. Moreover, chronic inflammation might cause dysregulation of lipid and FA metabolism that may contribute to skeletal muscle weakness in patients with polymyositis (PM) and dermatomyositis (DM). The major aim of this thesis was to gain better understanding of the regulation of PGE2 and other lipid mediators in RA, PM and in DM to improve treatment of patients. First, we have determined the catalytic mechanism of mPGES-1 activity by site-directed mutagenesis (Paper I). The amino acid residues arginine (Arg) 126 and aspartate (Asp) 49 were identified as essential for the catalytic activity of mPGES-1, as when exchanged, the enzyme variants lost their enzymatic activity. Previous high-resolution structural studies predicted a role for serine (Ser) 127 in the enzymatic activity of mPGES-1. In contrast, we have demonstrated that Ser127, as well as Arg73, do not seem to be significant to the catalytic mechanism because when exchanged, their variants retained considerable activity. These results are of relevance for the development of the new generation of mPGES-1 inhibitors. Further, we studied whether mPGES-1 deletion might be beneficial for reducing inflammation via the suppression of platelet functions (Paper II). Platelet activation, the formation of platelet-leukocyte aggregates, and release of platelet-derived microparticles (PMP) were significantly reduced in mPGES-1 KO mice compared to WT after lipopolysaccharide (LPS) treatment. In addition, KO mice displayed a significant decrease in platelet aggregation ex vivo. The reduced activation of platelets may contribute to antiinflammatory effect and cardiovascular safety of mPGES-1 inhibitors. In Paper III, we investigated effects of mPGES-1, PGIS, and cyclooxygenase (COX) -2 on vascular and renal pathways associated with asymmetric dimethylarginine (ADMA) and endothelial nitric oxide synthase (eNOS). WT mice treated with COX-2 inhibitor displayed no change in the plasma levels of cardioprotective prostacyclin (PGI2), while mPGES-1 KO mice showed significantly higher PGI2 levels in the plasma. In contrast to COX-2 inhibition, mPGES-1 deletion had no effect on genes responsible for the production or breakdown of ADMA in the kidney. Plasma creatinine and ADMA were elevated in mice treated with COX-2 inhibitor or PGIS KO mice but unaltered in mPGES-1 KO mice. Furthermore, the deletion of mPGES-1 significantly improved the eNOS-driven dilator response to acetylcholine in the aorta. These data further confirmed the cardioprotective effects of mPGES-1 deletion suggesting selective inhibitors of mPGES-1 as a safer alternative to NSAIDs. To clarify mechanisms involved in muscle weakness, we examined effects of the conventional immunosuppressive treatment on global gene expression profiles in skeletal muscle from PM and DM patients (Paper IV). The genes related to immune response and inflammation including the interferon and the inflammasome pathways were downregulated by treatment. The genes involved in muscle tissue remodeling and growth were negatively affected by treatment. The immunosuppressive treatment caused an induction of gene markers of fast type II fibers. Furthermore, the fiber composition of the muscle tissue from patients was switched towards type II fibers after treatment. Importantly, the expression of genes involved in lipid metabolism was altered, signifying a probable lipotoxic effect on muscles, that at least partly might explain the persistent muscle weakness and fatigue observed in PM and DM patients despite treatment. To confirm dysregulated lipid metabolism in myositis patients, we analyzed lipid and FA profiles in serum from patients with PM and DM in comparison to healthy individuals and response to immunosuppressive treatment (Paper V). FA composition of total serum lipids was changed in myositis patients compared to healthy individuals. In myositis patients, the levels of palmitic 16:0 acid was significantly higher while the levels of arachidonic 20:4(n- 6) acid was significantly lower. The levels of serum lipid species within phosphatidylcholine (PC), lysophosphatidylcholine (LPC) and triglycerides (TG) were also significantly changed in myositis patients compared to healthy individuals. Immunosuppressive treatment resulted in increased serum levels of C20:2(n-6) acid and C20:5(n-3) acids as well as in the changed serum PC, phosphatidylethanolamine (PE) and LPC profiles in myositis patients. In conclusion, in this thesis, we have provided new knowledge on the catalytic mechanism and the impact of mPGES-1 on inflammation and cardiovascular safety. Furthermore, we have demonstrated that lipid metabolism is altered in PM and DM patients and might contribute to disease pathogenesis

Effects on muscle tissue remodeling and lipid metabolism in muscle tissue from adult patients with polymyositis or dermatomyositis treated with immunosuppressive agents.

BACKGROUND: Polymyositis (PM) and dermatomyositis (DM) are autoimmune muscle diseases, conventionally treated with high doses of glucocorticoids in combination with immunosuppressive drugs. Treatment is often dissatisfying, with persisting muscle impairment. We aimed to investigate molecular mechanisms that might contribute to the persisting muscle impairment despite immunosuppressive treatment in adult patients with PM or DM using gene expression profiling of repeated muscle biopsies. METHODS: Paired skeletal muscle biopsies from six newly diagnosed adult patients with DM or PM taken before and after conventional immunosuppressive treatment were examined by gene expression microarray analysis. Selected genes that displayed changes in expression were analyzed by Western blot. Muscle biopsy sections were evaluated for inflammation, T lymphocytes (CD3), macrophages (CD68), major histocompatibility complex (MHC) class I expression and fiber type composition. RESULTS: After treatment, genes related to immune response and inflammation, including inflammasome pathways and interferon, were downregulated. This was confirmed at the protein level for AIM-2 and caspase-1 in the inflammasome pathway. Changes in genes involved in muscle tissue remodeling suggested a negative effect on muscle regeneration and growth. Gene markers for fast type II fibers were upregulated and fiber composition was switched towards type II fibers in response to treatment. The expression of genes involved in lipid metabolism was altered, suggesting a potential lipotoxic effect on muscles of the immunosuppressive treatment. CONCLUSION: The anti-inflammatory effect of immunosuppressive treatment was combined with negative effects on genes involved in muscle tissue remodeling and lipid metabolism, suggesting a negative effect on recovery of muscle performance which may contribute to persisting muscle impairment in adult patients with DM and PM

Pulsed-field Gel Electrophoresis for Salmonella Infection Surveillance, Texas, USA, 2007

To identify sources of transmission for area clusters, in 2007 the Houston Department of Health and Human Services conducted an 8-month study of enhanced surveillance of Salmonella infection. Protocol included patient interviews and linking the results of interviews to clusters of pulsed-field gel electrophoresis patterns detected by the local PulseNet laboratory

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance

INTRODUCTION Investment in Africa over the past year with regard to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing has led to a massive increase in the number of sequences, which, to date, exceeds 100,000 sequences generated to track the pandemic on the continent. These sequences have profoundly affected how public health officials in Africa have navigated the COVID-19 pandemic. RATIONALE We demonstrate how the first 100,000 SARS-CoV-2 sequences from Africa have helped monitor the epidemic on the continent, how genomic surveillance expanded over the course of the pandemic, and how we adapted our sequencing methods to deal with an evolving virus. Finally, we also examine how viral lineages have spread across the continent in a phylogeographic framework to gain insights into the underlying temporal and spatial transmission dynamics for several variants of concern (VOCs). RESULTS Our results indicate that the number of countries in Africa that can sequence the virus within their own borders is growing and that this is coupled with a shorter turnaround time from the time of sampling to sequence submission. Ongoing evolution necessitated the continual updating of primer sets, and, as a result, eight primer sets were designed in tandem with viral evolution and used to ensure effective sequencing of the virus. The pandemic unfolded through multiple waves of infection that were each driven by distinct genetic lineages, with B.1-like ancestral strains associated with the first pandemic wave of infections in 2020. Successive waves on the continent were fueled by different VOCs, with Alpha and Beta cocirculating in distinct spatial patterns during the second wave and Delta and Omicron affecting the whole continent during the third and fourth waves, respectively. Phylogeographic reconstruction points toward distinct differences in viral importation and exportation patterns associated with the Alpha, Beta, Delta, and Omicron variants and subvariants, when considering both Africa versus the rest of the world and viral dissemination within the continent. Our epidemiological and phylogenetic inferences therefore underscore the heterogeneous nature of the pandemic on the continent and highlight key insights and challenges, for instance, recognizing the limitations of low testing proportions. We also highlight the early warning capacity that genomic surveillance in Africa has had for the rest of the world with the detection of new lineages and variants, the most recent being the characterization of various Omicron subvariants. CONCLUSION Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve. This is important not only to help combat SARS-CoV-2 on the continent but also because it can be used as a platform to help address the many emerging and reemerging infectious disease threats in Africa. In particular, capacity building for local sequencing within countries or within the continent should be prioritized because this is generally associated with shorter turnaround times, providing the most benefit to local public health authorities tasked with pandemic response and mitigation and allowing for the fastest reaction to localized outbreaks. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century

Deletion of mPGES-1 affects platelet functions in mice

Abstract Microsomal prostaglandin E 2 synthase-1 (mPGES-1) constitutes an essential player in inflammation and is involved in the pathogenesis of rheumatoid arthritis. Platelets participate in the regulation of inflammatory processes by the release of proinflammatory mediators and platelet-derived microparticles (PMPs). However, the role of the inducible mPGES-1/PGE 2 pathway in platelet functions has not been investigated. In the present study we report a significant impact of mPGES-1 on platelet functions during inflammation. Wild-type (WT) and mPGES-1 −/− knockout (KO) mice were stimulated with lipopolysaccharide (LPS) for 24 h. Platelet counts and activation were assessed by flow cytometry analysing CD62P-CD154 expression, PMP numbers, platelet-leukocyte aggregates and platelet aggregation. The accumulation of platelets and fibrinogen in the liver was analysed by immunofluorescent staining. In native platelets from WT and mPGES-1 KO mice, there were no differences among the investigated functions. After LPS treatment, the number of platelets was significantly decreased in WT, but not in KO mice. Platelet activation, platelet-leukocyte aggregates and PMP numbers were all significantly lower in KO mice compared with WT mice after LPS treatment. In addition, KO mice displayed a significant reduction in platelet aggregation ex vivo. In the liver of LPS-stimulated WT and KO mice, there were no differences in platelet accumulation, although the percentage of total vessel area in the KO liver was significantly lower compared with the WT one. Our results demonstrate that systemic inhibition of mPGES-1 prevents platelet activation, which should have important implications with regard to the cardiovascular safety of mPGES-1 inhibitors

Business-driven Management of Differentiated Services

Abstract — Several intra- and inter-domain quality of service (QoS) provisioning mechanisms for IP networks have been researched and developed using Differentiated Services (DiffServ) technology. However, the incremental efforts needed to manage DiffServ networks from a business-oriented viewpoint have received relatively little attention. This paper addresses this gap and presents a framework for achieving business-driven QoS provisioning in DiffServ over MPLS networks. We provide a rich model of SLA and business indicators as well as reasonable mapping functions that define, with key degrees of importance, the impact of business indicators over service management policies. Business and service indicators are used to control static and dynamic admission of services. The paper advances the state of the art by considering the influence of business-level objectives on the policy refinement process. We evaluate and discuss the effectiveness of our approach through a simulation environment that we developed over OPNET

Targeted lipidomics analysis identified altered serum lipid profiles in patients with polymyositis and dermatomyositis

Abstract Background Polymyositis (PM) and dermatomyositis (DM) are severe chronic autoimmune diseases, characterized by muscle fatigue and low muscle endurance. Conventional treatment includes high doses of glucocorticoids and immunosuppressive drugs; however, few patients recover full muscle function. One explanation of the persistent muscle weakness could be altered lipid metabolism in PM/DM muscle tissue as we previously reported. Using a targeted lipidomic approach we aimed to characterize serum lipid profiles in patients with PM/DM compared to healthy individuals (HI) in a cross-sectional study. Also, in the longitudinal study we compared serum lipid profiles in patients newly diagnosed with PM/DM before and after immunosuppressive treatment. Methods Lipidomic profiles were analyzed in serum samples from 13 patients with PM/DM, 12 HI and 8 patients newly diagnosed with PM/DM before and after conventional immunosuppressive treatment using liquid chromatography tandem mass spectrometry (LC-MS/MS) and a gas-chromatography flame ionization detector (GC-FID). Functional Index (FI), as a test of muscle performance and serum levels of creatine kinase (s-CK) as a proxy for disease activity were analyzed. Results The fatty acid (FA) composition of total serum lipids was altered in patients with PM/DM compared to HI; the levels of palmitic (16:0) acid were significantly higher while the levels of arachidonic (20:4, n-6) acid were significantly lower in patients with PM/DM. The profiles of serum phosphatidylcholine and triacylglycerol species were changed in patients with PM/DM compared to HI, suggesting disproportionate levels of saturated and polyunsaturated FAs that might have negative effects on muscle performance. After immunosuppressive treatment the total serum lipid levels of eicosadienoic (20:2, n-6) and eicosapentaenoic (20:5, n-3) acids were increased and serum phospholipid profiles were altered in patients with PM/DM. The correlation between FI or s-CK and levels of several lipid species indicate the important role of lipid changes in muscle performance and inflammation. Conclusions Serum lipids profiles are significantly altered in patients with PM/DM compared to HI. Moreover, immunosuppressive treatment in patients newly diagnosed with PM/DM significantly affected serum lipid profiles. These findings provide new evidence of the dysregulated lipid metabolism in patients with PM/DM that could possibly contribute to low muscle performance