Pro-atherogenic actions of signal transducer and activator of transcription 1 serine 727 phosphorylation in LDL receptor deficient mice via modulation of plaque inflammation

Atherosclerosis is a chronic inflammatory disorder of the vasculature regulated by cytokines. We have previously shown that extracellular signal-regulated kinase-1/2 (ERK1/2) plays an important role in serine 727 phosphorylation of signal transducer and activator of transcription-1 (STAT1) transactivation domain, which is required for maximal interferon-γ signaling, and the regulation of modified LDL uptake by macrophages in vitro. Unfortunately, the roles of ERK1/2 and STAT1 serine 727 phosphorylation in atherosclerosis are poorly understood and were investigated using ERK1 deficient mice (ERK2 knockout mice die in utero) and STAT1 knock-in mice (serine 727 replaced by alanine; STAT1 S727A). Mouse Atherosclerosis RT² Profiler PCR Array analysis showed that ERK1 deficiency and STAT1 S727A modification produced significant changes in the expression of 18 and 49 genes, respectively, in bone marrow-derived macrophages, with 17 common regulated genes that included those that play key roles in inflammation and cell migration. Indeed, ERK1 deficiency and STAT1 S727A modification attenuated chemokine-driven migration of macrophages with the former also impacting proliferation and the latter phagocytosis. In LDL receptor deficient mice fed a high fat diet, both ERK1 deficiency and STAT1 S727A modification produced significant reduction in plaque lipid content, albeit at different time points. The STAT1 S727A modification additionally caused a significant reduction in plaque content of macrophages and CD3 T cells and diet-induced cardiac hypertrophy index. In addition, there was a significant increase in plasma IL-2 levels and a trend toward increase in plasma IL-5 levels. These studies demonstrate important roles of STAT1 S727 phosphorylation in particular in the regulation of atherosclerosis-associated macrophage processes in vitro together with plaque lipid content and inflammation in vivo, and support further assessment of its therapeutical potential

Protective effects of a unique combination of nutritionally active ingredients on risk factors and gene expression associated with atherosclerosis in C57BL/6J mice fed a high fat diet

Atherosclerosis, an inflammatory disorder of the vasculature and the underlying cause of cardiovascular disease, is responsible for one in three global deaths. Consumption of active food ingredients such as omega-3 polyunsaturated fatty acids, flavanols and phytosterols have many beneficial effects on cardiovascular disease. However, their combined actions on risk factors for atherosclerosis remains poorly understood. We have previously shown that a formulation containing each of these active components at physiologically relevant doses modulated several monocyte/macrophage processes associated with atherosclerosis in vitro, including inhibition of cytokine-induced pro-inflammatory gene expression, chemokine-driven monocyte migration, expression of M1 phenotype markers, and promotion of cholesterol efflux. The objective of the present study was to investigate whether the protective actions of the formulation extended in vivo and to delineate the potential underlying mechanisms. The formulation produced several favourable changes, including higher plasma levels of HDL and reduced levels of macrophages and myeloid-derived suppressor cells in the bone marrow. The mRNA expression of liver-X-receptor-α, peroxisome proliferator-activated receptor-γ and superoxide dismutase-1 was induced in the liver and that of interferon-γ and the chemokine (C-X-C motif) ligand 1 decreased, thereby suggesting potential mechanisms for many beneficial effects. Other changes were also observed such as increased plasma levels of triglycerides and lipid peroxidation that may reflect potential activation of brown fat. This study provides new insights into the protective actions and the potential underlying mechanisms of the formulation in vivo, particularly in relation to risk factors together with changes in systemic inflammation and hepatic lipid alterations associated with atherosclerosis and metabolic syndrome, and supports further assessments in human trials

Alternative polyadenylation variants of the RNA binding protein, HuR: abundance, role of AU-rich elements and auto-Regulation

The RNA-binding protein, HuR, is involved in the stabilization of AU-rich element-containing mRNAs with products that are involved in cell-cycle progression, cell differentiation and inflammation. We show that there are multiple polyadenylation variants of HuR mRNA that differ in their abundance, using both bioinformatics and experimental approaches. A polyadenylation variant with distal poly(A) signal is a rare transcript that harbors functional AU-rich elements (ARE) in the 3′UTR. A minimal 60-nt region, but not a mutant form, fused to reporter-3′UTR constructs was able to downregulate the reporter activity. The most predominant and alternatively polyadenylated mature transcript does not contain the ARE. HuR itself binds HuR mRNA, and upregulated the activity of reporter from constructs fused with ARE-isoform and the HuR ARE. Wild-type tristetraprolin (TTP), but not the zinc finger mutant TTP, competes for HuR binding and upregulation of HuR mRNA. The study shows that the HuR gene codes for several polyadenylation variants differentially regulated by AU-rich elements, and demonstrates an auto-regulatory role of HuR