DataSheet_1_Circulating T Cell Activation and Exhaustion Markers Are Associated With Radiation Pneumonitis and Poor Survival in Non-Small-Cell Lung Cancer.docx

IntroductionPersistent inflammation and immune activation in the lungs are associated with adverse outcomes such as radiation pneumonitis (RP) and poor survival in non-small-cell lung cancer (NSCLC) patients. However, it is unknown how this is reflected by leukocyte activation markers in serum.ObjectiveThe aim was to evaluate the serum levels of activation of different leukocyte subsets and to examine those in relation to the pathogenesis of RP and survival in NSCLC.MethodsWe analyzed the serum levels of MPO, sCD25, sTIM-3, sPD-L1, sCD14, sCD163, CCL19 and CCL21 in 66 inoperable NSCLC patients with stage IA-IIIA disease. The patients were treated with stereotactic body radiation therapy (SBRT) or concurrent chemoradiation therapy (CCRT), followed by regular blood sampling for 12 months after treatment and for 5 years for survival.ResultsNineteen (29%) patients developed RP, which occurred more frequently and earlier in patients receiving CCRT than in those receiving SBRT. Increases in sCD25, sTIM-3 and CCL21 levels were observed at the last 6 months of follow-up in patients who had RP after SBRT. Patients who had RP after CCRT had higher sTIM-3 levels during the first 3 months of follow-up. Baseline sCD25 was independently associated with both 2- and 5-year mortality outcomes, while baseline sTIM-3 was independently associated with 2-year mortality.ConclusionWe showed that T cell activation and exhaustion markers such as sCD25 and sTIM-3 are enhanced in patients developing RP and are associated with poor survival in NSCLC.</p

Table_1_Relationship between fibroblast growth factor in plasma and carotid plaque neovascularization: a pilot study.docx

BackgroundUnstable atherosclerotic carotid plaques with intraplaque neovascularization (IPN) carry a substantial risk for ischemic stroke. Conventional ultrasound methods fall short in detecting IPN, where superb microvascular imaging (SMI) has emerged as a promising tool for both visualizing and quantification. High levels of fibroblast growth factor 23 (FGF-23) have, in observational studies, been suggested as related to cardiovascular morbidity and mortality. The association of FGF-23 to atherosclerotic carotid plaque instability remains relatively unexplored.MethodsA cohort of twenty-nine patients with ≥50% atherosclerotic carotid stenosis underwent conventional carotid ultrasound, SMI, and blood tests, including measurement of FGF-23 in plasma. Nineteen patients were characterized as symptomatic and ten as asymptomatic.ResultsOur major findings were: i) Higher FGF-23 levels were strongly correlated with increased SMI-assessed IPN. ii) Neo-vessel count recorded by quantitative SMI was positively correlated to increased FGF-23 levels, but not with basic FGF levels. (iii) In contrast, traditional risk factors for plaque instability exhibited no noteworthy associations with SMI-assessed IPN or with FGF-23 levels.ConclusionThis pilot study suggest the potential of FGF-23 as a valuable marker for neovascularization and atherosclerotic carotid plaque instability as a risk factor for ischemic stroke. Further research involving larger cohorts and prospective data is necessary to understand FGF-23’s role in this context comprehensively.</p

Additional file 1: Table S1. of Severe hypertriglyceridemia in Norway: prevalence, clinical and genetic characteristics

Inclusion and exclusion criteria for patients with sHTG for detailed analyses. Table S2. List of lipid-related genes analyzed in peripheral blood mononuclear cells from patients with severe hypertriglyceridemia. Table S3. Baseline biochemical characteristic of 65 patients with severe hypertriglyceridemia. Table S4. Combinations of medications used at end of study among the 65 patients with hypertriglyceridemia. Table S5. Characteristics of the six patients with hypertriglyceridemia subjected to in-depth molecular analyses. (PDF 258 kb

Additional file 1: Table S1. of Severe hypertriglyceridemia in Norway: prevalence, clinical and genetic characteristics

Inclusion and exclusion criteria for patients with sHTG for detailed analyses. Table S2. List of lipid-related genes analyzed in peripheral blood mononuclear cells from patients with severe hypertriglyceridemia. Table S3. Baseline biochemical characteristic of 65 patients with severe hypertriglyceridemia. Table S4. Combinations of medications used at end of study among the 65 patients with hypertriglyceridemia. Table S5. Characteristics of the six patients with hypertriglyceridemia subjected to in-depth molecular analyses. (PDF 258 kb

The accumulation and subcellular redistribution of IL-1β-induced chemokines in HUVECs depends on inhibition of isoprenylation.

<p>HUVECs were pretreated with atorvastatin together with medium (black bars), mevalonate (white bars), squalene (grey bars) or GGPP (hatched bars) before IL-1β-stimulation. In the histogram the four bars of the same color represent cells pretreated with increasing doses of atorvastatin (from the left: 0, 1, 5 and 30 µM). Chemokine levels were measured by ELISA for GRO-α, IL-8, MCP-1 (A) and immunostaining for GRO-α (B). * p<0.05, ** p<0.01 and *** p<0.001 from IL-1β-treated control cells without atorvastatin.</p

GRO-α localizes to CD63 positive multivesicular bodies in atorvastatin treated cells.

<p>Thawed cryo sections of fixed HUVECs pretreated without (A) or with 10 µM atorvastatin (B-D) were either single labeled (A-B) with anti-GRO-α antibody followed by 15 nm protein A gold (large arrows in insets), or double labeled (C-D) with anti-GRO-α antibody followed by 15 nm protein A gold (large arrows) and anti-CD63 antibody followed by 10 nm protein A gold (small arrows). Both in control cells (A) and atorvastatin-treated cells (B) labeling for GRO-α was found in the Golgi region (G) and small electron dense vesicles (insets in A and B). In atorvastatin-treated cells labeling for GRO-α was also found in large electron dense, CD63-positive multivesicular bodies (C-D). Note that the electron lucent CD63 positive multivesicular body in the lower left corner of D is GRO-α negative. Scale bars are 100 nm.</p

Atorvastatin induces colocalization of IL-1β-induced chemokines and E-selectin in endocytic compartments.

<p>HUVECs were pretreated with 5 µM atorvastatin (A-B, D, F, G) or medium (C, E, G) before IL-1β-stimulation, fixation and immunostaining for GRO-α (A-H), IL-8 (A), MCP-1 (B), E-selectin (C-D), EEA-1 (E-F) and CD63 (G-H). Yellow arrows in panel H outlines colocalization of GRO-α and CD63. Scale bars are 10 µm.</p

Statins Affect the Presentation of Endothelial Chemokines by Targeting to Multivesicular Bodies

<div><h3>Background</h3><p>In addition to lowering cholesterol, statins are thought to beneficially modulate inflammation. Several chemokines including CXCL1/growth-related oncogene (GRO)-α, CXCL8/interleukin (IL)-8 and CCL2/monocyte chemoattractant protein (MCP)-1 are important in the pathogenesis of atherosclerosis and can be influenced by statin-treatment. Recently, we observed that atorvastatin­treatment alters the intracellular content and subcellular distribution of GRO-α in cultured human umbilical vein endothelial cells (HUVECs). The objective of this study was to investigate the mechanisms involved in this phenomenon.</p> <h3>Methodology/ Principal Findings</h3><p>The effect of atorvastatin on secretion levels and subcellular distribution of GRO-α, IL-8 and MCP-1 in HUVECs activated by interleukin (IL)-1β were evaluated by ELISA, confocal microscopy and immunoelectron microscopy. Atorvastatin increased the intracellular contents of GRO-α, IL-8, and MCP-1 and induced colocalization with E-selectin in multivesicular bodies. This effect was prevented by adding the isoprenylation substrate GGPP, but not the cholesterol precursor squalene, indicating that atorvastatin exerts these effects by inhibiting isoprenylation rather than depleting the cells of cholesterol.</p> <h3>Conclusions/ Significance</h3><p>Atorvastatin targets inflammatory chemokines to the endocytic pathway and multivesicular bodies and may contribute to explain the anti-inflammatory effect of statins at the level of endothelial cell function.</p> </div

Antibodies used for immunostainings.

<p>Antibodies used for immunostainings.</p

Baseline characteristics.

<p>Data are given as median (min-max) except when percentage is indicated.</p><p>HbA1c: n = 18 (high HDL); FFA: n = 18 (high HDL); LDL: n = 14 (low HDL).</p><p>Low and High HDL groups.</p