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Cryptococcus neoformans is included in the World Health Organization fungal priority pathogen list, complied to expedite improved research and public-health interventions. The limited number of available antifungal drugs, their associated toxicity, and the emergence of drug-resistant strains make the development of new therapeutic strategies mandatory. Pattern-recognition receptors (PRRs) from the host's innate immune system constitute a potential source of new antimicrobial agents. CD5 and CD6 are lymphoid members of the ancient scavenger receptor cysteine-rich superfamily (SRCR-SF) which bind pathogen-associated molecular patterns (PAMPs) of fungal and bacterial origin. Evidence supports the concept that such binding maps to 11-mer sequences present in each of their three SRCR extracellular domains. Herein, we have designed synthetic peptides containing tandems of such 11-mer sequences (namely CD5-T and CD6-T) and analyzed their C. neoformans-binding properties in vitro. Our results show both inhibitory effects on fungal growth and an ability to impact capsule formation and titanization, two critical virulence factors of C. neoformans involved in immune evasion. These effects hold promise for CD5-T and CD6-T peptides as single or adjuvant therapeutic agents against cryptococcosis. [ABSTRACT FROM AUTHOR]

Polyunsaturated fatty acids (PUFAs) have been extensively studied for their health benefits because they can be oxidized by lipoxygenases to form bioactive oxylipins. In this study, we investigated the impact of double bond placement on the kinetic properties and product profiles of human platelet 12-lipoxygenase (h12-LOX), human reticulocyte 15-lipoxygenase-1 (h15-LOX-1), and human endothelial 15-lipoxygenase-2 (h15-LOX-2) by using 22-carbon (C22) fatty acid substrates with differing double bond content. With respect to kcat/KM values, the loss of Δ4 and Δ19 led to an 18-fold loss of kinetic activity for h12-LOX, no change in kinetic capability for h15-LOX-1, but a 24-fold loss for h15-LOX-2 for both C22-FAs. With respect to the product profiles, h12-LOX produced mainly 14-oxylipins. For h15-LOX-1, the 14-oxylipin production increased with the loss of either Δ4 and Δ19, however, the 17-oxylipin became the major species upon loss of both Δ4 and Δ19. h15-LOX-2 produced mostly the 17-oxylipin products throughout the fatty acid series. This study also investigated the effects of various 17-oxylipins on platelet activation. The results revealed that both 17(S)-hydroxy-4Z,7Z,10Z,13Z,15E,19Z-DHA (17-HDHA) and 17-hydroxy-4Z,7Z,10Z,13Z,15E-DPAn6 (17-HDPAn6) demonstrated anti-aggregation properties with thrombin or collagen stimulation. 17-hydroxy-7Z,10Z,13Z,15E,19Z-DPAn3 (17-HDPAn3) exhibited agonistic properties, and 17-hydroxy-7Z,10Z,13Z,15E-DTA (17-HDTA) showed biphasic effects, inhibiting collagen-induced aggregation at lower concentrationsbut promoting aggregation at higher concentrations. Both 17-hydroxy-13Z,15E,19Z-DTrA (17-HDTrA), and 17-hydroxy-13Z,15E-DDiA (17-HDDiA) induced platelet aggregation. In summary, the number and placement of the double bonds affect platelet activation, with the general trend being that more double bonds generally inhibit aggregation, while less double bonds promote aggregation. These findings provide insights into the potential role of specific fatty acids and their metabolizing LOX isozymes with respect to cardiovascular health.

Purpose of the Review: Atherosclerotic plaques result from imbalanced lipid metabolism and maladaptive chronic immune responses. Class B scavenger receptors are lipid transporters and regulators of their metabolism. The purpose of this review is to explore recent structural findings of these membrane-associated receptors, with particular focus on their higher-order oligomeric organization and impact on lipid transport. Recent Findings: Class B scavenger receptors have evidence for oligomerization, with recent efforts placed on identifying residues and motifs responsible for mediating this process. The first studies correlating scavenger receptor oligomerization to function are described. Summary: This review highlights two emerging hypotheses regarding the function of scavenger receptor oligomerization. The first is a hydrophobic channel created by self-association of receptors to promote transport. The second hypothesis suggests that homo-oligomerization stabilizes receptors, prevents internalization and thereby promotes transport indirectly. Novel computational and in vitro experimental techniques with purified receptors are also described. [ABSTRACT FROM AUTHOR]

Lipopolysaccharide (LPS) in blood circulation causes endotoxemia and is linked to various disease conditions. Current treatments focus on preventing LPS from interacting with its receptor Toll-like receptor 4 (TLR4) and reducing inflammation. However, our body has a natural defense mechanism: reticuloendothelial cells in the liver rapidly degrade and inactivate much of the circulating LPS within minutes. But this LPS clearance mechanism is not perfect. Excessive LPS that escape this clearance mechanism cause systemic inflammatory damage through TLR4. Despite its importance, the role of reticuloendothelial cells in LPS elimination is not well-studied, especially regarding the specific cells, receptors, and mechanisms involved. This gap hampers the development of effective therapies for endotoxemia and related diseases. This review consolidates the current understanding of LPS clearance, narrates known and explores potential mechanisms, and discusses the relationship between LPS clearance and LPS signaling. It also aims to highlight key insights that can guide the development of strategies to reduce circulating LPS by way of bolstering host defense mechanisms. Ultimately, we seek to provide a foundation for future research that could lead to innovative approaches for enhancing the body's natural ability to clear LPS and thereby lower the risk of endotoxin-related inflammatory diseases, including sepsis. [ABSTRACT FROM AUTHOR]

The influence of chronic diseases on various facets of macrophage cellular senescence is poorly understood. This study evaluated the impact of chronic hyperglycemia on the induction of cellular senescence and subsequent immunosurveillance functions in RAW264.7 macrophages. Macrophages were cultured under normal glucose (NG; 5 mM), high glucose (HG; 20 mM), and very high glucose (VHG; 40 mM) conditions and assessed for markers of cellular senescence. Hyperglycemia induced strong upregulation of SA-β-gal activity, and loss of PCNA and Lamin B1 gene expression while markers of cell cycle arrest generally decreased. Non-significant changes in SASP-related proteins were observed while ROS levels slightly decreased and mitochondrial membrane potential increased. Protein concentration on the exosome membrane surface and their stability appeared to increase under hyperglycemic conditions. However, when macrophages were exposed to the secretory media (SM) of senescent preadipocytes, a dramatic increase in the levels of all inflammatory proteins was recorded especially in the VHG group that was also accompanied by upregulation of NF-κB and NLRP3 gene expression. SM treatment to hyperglycemic macrophages activated the TLR-2/Myd88 pathway but decreased the expression of scavenger receptors RAGE, CD36, and Olr-1 while CD44 and CXCL16 expression increased. On exposure to LPS, a strong upregulation in NO, ROS, and inflammatory cytokines was observed. Together, these results suggest that primary markers of cellular senescence are aberrantly expressed under chronic hyperglycemic conditions in macrophages with no significant SASP activation. Nonetheless, hyperglycemia strongly deregulates macrophage functions leading to impaired immunosurveillance of senescent cells and aggravation of inflamm-aging. This work provides novel insights into how hyperglycemia-induced dysfunctions can impact the potency of macrophages to manage senescent cell burden in aging tissues. [ABSTRACT FROM AUTHOR]

Lipid metabolism is a complex process crucial for energy production resulting in high levels of acyl-coenzyme A (acyl-CoA) molecules in the cell. Acyl-CoAs have also been implicated in inflammation, which could be possibly linked to lipoxygenase (LOX) biochemistry by the observation that an acyl-CoA was bound to human platelet 12-lipoxygenase via cryo-EM. Given that LOX isozymes play a pivotal role in inflammation, a more thorough investigation of the inhibitory effects of acyl-CoAs on lipoxygenase isozymes was judged to be warranted. Subsequently, it was determined that C18 acyl-CoA derivatives were the most potent against h12-LOX, human reticulocyte 15-LOX-1 (h15-LOX-1), and human endothelial 15-LOX-2 (h15-LOX-2), while C16 acyl-CoAs were more potent against human 5-LOX. Specifically, oleoyl-CoA (18:1) was most potent against h12-LOX (IC50 = 32 μM) and h15-LOX-2 (IC50 = 0.62 μM), stearoyl-CoA against h15-LOX-1 (IC50 = 4.2 μM), and palmitoleoyl-CoA against h5-LOX (IC50 = 2.0 μM). The inhibition of h15-LOX-2 by oleoyl-CoA was further determined to be allosteric inhibition with a Ki of 82 +/- 70 nM, an α of 3.2 +/- 1, a β of 0.30 +/- 0.07, and a β/α = 0.09. Interestingly, linoleoyl-CoA (18:2) was a weak inhibitor against h5-LOX, h12-LOX, and h15-LOX-1 but a rapid substrate for h15-LOX-1, with comparable kinetic rates to free linoleic acid (kcat = 7.5 +/- 0.4 s-1, kcat/KM = 0.62 +/- 0.1 µM-1s-1). Additionally, it was determined that methylated fatty acids were not substrates but rather weak inhibitors. These findings imply a greater role for acyl-CoAs in the regulation of LOX activity in the cell, either through inhibition of novel oxylipin species or as a novel source of oxylipin-CoAs.

The antiplatelet effect of polyunsaturated fatty acids is primarily attributed to its metabolism to bioactive metabolites by oxygenases, such as lipoxygenases (LOX). Platelets have demonstrated the ability to generate 15-LOX-derived metabolites (15-oxylipins); however, whether 15-LOX is in the platelet or is required for the formation of 15-oxylipins remains unclear. This study seeks to elucidate whether 15-LOX is required for the formation of 15-oxylipins in the platelet and determine their mechanistic effects on platelet reactivity. In this study, 15-HETrE, 15-HETE, and 15-HEPE attenuated collagen-induced platelet aggregation, and 15-HETrE inhibited platelet aggregation induced by different agonists. The observed anti-aggregatory effect was due to the inhibition of intracellular signaling including αIIbβ3 and protein kinase C activities, calcium mobilization, and granule secretion. While 15-HETrE inhibited platelets partially through activation of peroxisome proliferator-activated receptor β (PPARβ), 15-HETE also inhibited platelets partially through activation of PPARα. 15-HETrE, 15-HETE, or 15-HEPE inhibited 12-LOX in vitro, with arachidonic acid as the substrate. Additionally, a 15-oxylipin-dependent attenuation of 12-HETE level was observed in platelets following ex vivo treatment with 15-HETrE, 15-HETE, or 15-HEPE. Platelets treated with DGLA formed 15-HETrE and collagen-induced platelet aggregation was attenuated only in the presence of ML355 or aspirin, but not in the presence of 15-LOX-1 or 15-LOX-2 inhibitors. Expression of 15-LOX-1, but not 15-LOX-2, was decreased in leukocyte-depleted platelets compared to non-depleted platelets. Taken together, these findings suggest that 15-oxylipins regulate platelet reactivity; however, platelet expression of 15-LOX-1 is low, suggesting that 15-oxylipins may be formed in the platelet through a 15-LOX-independent pathway.

Human 15-lipoxygenases (LOX) are critical enzymes in the inflammatory process, producing various pro-resolution molecules, such as lipoxins and resolvins, but the exact role each of the two 15-LOXs in these biosynthetic pathways remains elusive. Previously, it was observed that h15-LOX-1 reacted with 5S-HETE in a non-canonical manner, producing primarily the 5S,12S-diHETE product. To determine the active site constraints of h15-LOX-1 in achieving this reactivity, amino acids involved in the fatty acid binding were investigated. It was observed that R402L did not have a large effect on 5S-HETE catalysis, but F414 appeared to π-π stack with 5S-HETE, as seen with AA binding, indicating an aromatic interaction between a double bond of 5S-HETE and F414. Decreasing the size of F352 and I417 shifted oxygenation of 5S-HETE to C12, while increasing the size of these residues reversed the positional specificity of 5S-HETE to C15. Mutants at these locations demonstrated a similar effect with 7S-HDHA as the substrate, indicating that the depth of the active site regulates product specificity for both substrates. Together, these data indicate that of the three regions proposed to control positional specificity, π-π stacking and active site cavity depth are the primary determinants of positional specificity with 5S-HETE and h15-LOX-1. Finally, the altered reactivity of h15-LOX-1 was also observed with 5S-HEPE, producing 5S,12S-diHEPE instead of 5S,15S-diHEPE (aka resolvin E4 (RvE4). However, h15-LOX-2 efficiently produces 5S,15S-diHEPE from 5S-HEPE. This result is important with respect to the biosynthesis of the RvE4 since it obscures which LOX isozyme is involved in its biosynthesis. Future work detailing the expression levels of the lipoxygenase isoforms in immune cells and selective inhibition during the inflammatory response will be required for a comprehensive understanding of RvE4 biosynthesis.

Background and Purpose: Atherosclerosis induced by cyclosporine A (CsA), an inhibitor of the calcineurin/nuclear factor of activated T cells (NFAT) pathway, is a major concern after organ transplantation. However, the atherosclerotic mechanisms of CsA remain obscure. We previously demonstrated that calcineurin/NFAT signalling inhibition contributes to atherogenesis via suppressing microRNA‐204 (miR‐204) transcription. We therefore hypothesised that miR‐204 is involved in the development of CsA‐induced atherosclerosis. Experimental Approach: ApoE−/− mice with macrophage‐miR‐204 overexpression were generated to determine the effects of miR‐204 on CsA‐induced atherosclerosis. Luciferase reporter assays and chromatin immunoprecipitation sequencing were performed to explore the targets mediating miR‐204 effects. Key Results: CsA alone did not significantly affect atherosclerotic lesions or serum lipid levels. However, it exacerbated high‐fat diet‐induced atherosclerosis and hyperlipidemia in C57BL/6J and ApoE−/− mice, respectively. miR‐204 levels decreased in circulating monocytes and plaque lesions during CsA‐induced atherosclerosis. The upregulation of miR‐204 in macrophages inhibited CsA‐induced atherosclerotic plaque formation but did not affect serum lipid levels. miR‐204 limited the CsA‐induced foam cell formation by reducing the expression of the scavenger receptors SR‐BII and CD36. SR‐BII was post‐transcriptionally regulated by mature miR‐204‐5p via 3′‐UTR targeting. Additionally, nuclear‐localised miR‐204‐3p prevented the CsA‐induced binding of Ago2 to the CD36 promoter, suppressing CD36 transcription. SR‐BII or CD36 expression restoration dampened the beneficial effects of miR‐204 on CsA‐induced atherosclerosis. Conclusion and Implications: Macrophage miR‐204 ameliorates CsA‐induced atherosclerosis, suggesting that miR‐204 may be a potential target for the prevention and treatment of CsA‐related atherosclerotic side effects. [ABSTRACT FROM AUTHOR]

Cryptococcus neoformans is included in the World Health Organization fungal priority pathogen list, complied to expedite improved research and public-health interventions. The limited number of available antifungal drugs, their associated toxicity, and the emergence of drug-resistant strains make the development of new therapeutic strategies mandatory. Pattern-recognition receptors (PRRs) from the host’s innate immune system constitute a potential source of new antimicrobial agents. CD5 and CD6 are lymphoid members of the ancient scavenger receptor cysteine-rich superfamily (SRCR-SF) which bind pathogen-associated molecular patterns (PAMPs) of fungal and bacterial origin. Evidence supports the concept that such binding maps to 11-mer sequences present in each of their three SRCR extracellular domains. Herein, we have designed synthetic peptides containing tandems of such 11-mer sequences (namely CD5-T and CD6-T) and analyzed their C. neoformans-binding properties in vitro. Our results show both inhibitory effects on fungal growth and an ability to impact capsule formation and titanization, two critical virulence factors of C. neoformans involved in immune evasion. These effects hold promise for CD5-T and CD6-T peptides as single or adjuvant therapeutic agents against cryptococcosis.

Lipopolysaccharide (LPS) in blood circulation causes endotoxemia and is linked to various disease conditions. Current treatments focus on preventing LPS from interacting with its receptor Toll-like receptor 4 (TLR4) and reducing inflammation. However, our body has a natural defense mechanism: reticuloendothelial cells in the liver rapidly degrade and inactivate much of the circulating LPS within minutes. But this LPS clearance mechanism is not perfect. Excessive LPS that escape this clearance mechanism cause systemic inflammatory damage through TLR4. Despite its importance, the role of reticuloendothelial cells in LPS elimination is not well-studied, especially regarding the specific cells, receptors, and mechanisms involved. This gap hampers the development of effective therapies for endotoxemia and related diseases. This review consolidates the current understanding of LPS clearance, narrates known and explores potential mechanisms, and discusses the relationship between LPS clearance and LPS signaling. It also aims to highlight key insights that can guide the development of strategies to reduce circulating LPS by way of bolstering host defense mechanisms. Ultimately, we seek to provide a foundation for future research that could lead to innovative approaches for enhancing the body’s natural ability to clear LPS and thereby lower the risk of endotoxin-related inflammatory diseases, including sepsis.

BackgroundDeletions and duplications of the multigenic 16p11.2 and 22q11.2 copy number variant (CNV) regions are associated with brain-related disorders including schizophrenia, intellectual disability, obesity, bipolar disorder, and autism spectrum disorder (ASD). The contribution of individual CNV genes to each of these identified phenotypes is unknown, as well as the contribution of these CNV genes to other potentially subtler health implications for carriers. Hypothesizing that DNA copy number exerts most effects via impacts on RNA expression, we attempted a novel in silico fine-mapping approach in non-CNV carriers using both GWAS and biobank data.MethodsWe first asked whether gene expression level in any individual gene in the CNV region alters risk for a known CNV-associated behavioral phenotype(s). Using transcriptomic imputation, we performed association testing for CNV genes within large genotyped cohorts for schizophrenia, IQ, BMI, bipolar disorder, and ASD. Second, we used a biobank containing electronic health data to compare the medical phenome of CNV carriers to controls within 700,000 individuals in order to investigate the full spectrum of health effects of the CNVs. Third, we used genotypes for over 48,000 individuals within the biobank to perform phenome-wide association studies between imputed expressions of individual 16p11.2 and 22q11.2 genes and over 1500 health traits.ResultsUsing large genotyped cohorts, we found individual genes within 16p11.2 associated with schizophrenia (TMEM219, INO80E, YPEL3), BMI (TMEM219, SPN, TAOK2, INO80E), and IQ (SPN), using conditional analysis to identify upregulation of INO80E as the driver of schizophrenia, and downregulation of SPN and INO80E as increasing BMI. We identified both novel and previously observed over-represented traits within the electronic health records of 16p11.2 and 22q11.2 CNV carriers. In the phenome-wide association study, we found seventeen significant gene-trait pairs, including psychosis (NPIPB11, SLX1B) and mood disorders (SCARF2), and overall enrichment of mental traits.ConclusionsOur results demonstrate how integration of genetic and clinical data aids in understanding CNV gene function and implicates pleiotropy and multigenicity in CNV biology.

OBJECTIVE: To investigate the role of adipocyte Pcpe2 (procollagen C-endopeptidase enhancer 2) in SR-BI (scavenger receptor class BI)-mediated HDL-C (high-density lipoprotein cholesterol) uptake and contributions to adipose lipid storage. APPROACH AND RESULTS: Pcpe2, a glycoprotein devoid of intrinsic proteolytic activity, is believed to participate in extracellular protein-protein interactions, supporting SR-BI- mediated HDL-C uptake. In published studies, Pcpe2 deficiency increased the development of atherosclerosis by reducing SR-BI-mediated HDL-C catabolism, but the biological impact of this deficiency on adipocyte SR-BI-mediated HDL-C uptake is unknown. Differentiated cells from Ldlr-/-/Pcpe2-/- (Pcpe2-/-) mouse adipose tissue showed elevated SR-BI protein levels, but significantly reduced HDL-C uptake compared to Ldlr-/- (control) adipose tissue. SR-BI-mediated HDL-C uptake was restored by preincubation of cells with exogenous Pcpe2. In diet-fed mice lacking Pcpe2, significant reductions in visceral, subcutaneous, and brown adipose tissue mass were observed, despite elevations in plasma triglyceride and cholesterol concentrations. Significant positive correlations exist between adipose mass and Pcpe2 expression in both mice and humans. CONCLUSIONS: Overall, these findings reveal a novel and unexpected function for Pcpe2 in modulating SR-BI expression and function as it relates to adipose tissue expansion and cholesterol balance in both mice and humans.

Cancer management still requires more potent and safer treatments, of which immunomodulatory receptors on the lymphocyte surface have started to show promise in new cancer immunotherapies (e.g., CTLA-4 and PD-1). CD6 is a signal-transducing transmembrane receptor, mainly expressed by all T cells and some B and NK cell subsets, whose endogenous ligands (CD166/ALCAM, CD318/CDCP-1, Galectins 1 and 3) are overexpressed by malignant cells of different lineages. This places CD6 as a potential target for novel therapies against haematological and non-haematological malignancies. Recent experimental evidence for the role of CD6 in cancer immunotherapies is summarised in this review, dealing with diverse and innovative strategies from the classical use of monoclonal antibodies to soluble recombinant decoys or the adoptive transfer of immune cells engineered with chimeric antigen receptors. [ABSTRACT FROM AUTHOR]

Stabilin‐1 (Stab1) and Stabilin‐2 (Stab2) are two major scavenger receptors of liver sinusoidal endothelial cells that mediate removal of diverse molecules from the plasma. Double‐knockout mice (Stab‐DKO) develop impaired kidney function and a decreased lifespan, while single Stabilin deficiency or therapeutic inhibition ameliorates atherosclerosis and Stab1‐inhibition is subject of clinical trials in immuno‐oncology. Although POSTN and TFGBI have recently been described as novel Stabilin ligands, the dynamics and functional implications of these ligands have not been comprehensively studied. Immunofluorescence, Western Blotting and Simple Western™ as well as in situ hybridization (RNAScope™) and qRT‐PCR were used to analyze transcription levels and tissue distribution of POSTN and TGFBI in Stab‐KO mice. Stab‐POSTN‐Triple deficient mice were generated to assess kidney and liver fibrosis and function in young and aged mice. TGFBI and POSTN protein accumulated in liver tissue in Stab‐DKO mice and age‐dependent in glomeruli of Stabilin‐deficient mice despite unchanged transcriptional levels. Stab‐POSTN‐Triple KO mice showed glomerulofibrosis and a reduced lifespan comparable to Stab‐DKO mice. However, alterations of the glomerular diameter and vascular density were partially normalized in Stab‐POSTN‐Triple KO. TGFBI and POSTN are Stabilin‐ligands that are deposited in an age‐dependent manner in the kidneys and liver due to insufficient scavenging in the liver. Functionally, POSTN might partially contribute to the observed renal phenotype in Stab‐DKO mice. This study provides details on downstream effects how Stabilin dysfunction affects organ function on a molecular and functional level. [ABSTRACT FROM AUTHOR]

Atherosclerotic plaques develop from the accumulation of macrophage-derived foam cells via the uptake of modified low-density lipoprotein (LDL). CD36 and CD204 are the principal scavenger receptors responsible for the uptake of modified LDL. Although glucocorticoids are suspected to exacerbate atherosclerosis, the precise mechanisms have not been fully elucidated. We investigated the effects of long-term treatment (2 weeks) with both a natural glucocorticoid (hydrocortisone, HC, 1 μM) and a synthetic glucocorticoid (dexamethasone, Dex, 100 nM) on murine bone marrow-derived macrophages using flow cytometry and western blotting. Treatment with HC and Dex enhanced CD204 expression but not CD36 expression and acetylated LDL (Ac-LDL) uptake. Treatment with HC and Dex also induced the phosphorylation of extracellular signal-regulated kinase (ERK). The Dex-induced enhancement in CD204 expression and Ac-LDL uptake were suppressed by an inhibitor of the mitogen-activated protein kinase (MAPK)/ERK kinase. These results suggest that glucocorticoids activate the MAPK/ERK pathway, which enhances CD204 expression and results in increased uptake of Ac-LDL in macrophages. The MAPK/ERK pathway in macrophages might be a key target to prevent atherosclerosis that is worsened by glucocorticoids.

Production of nitric oxide (NO) by LPS-activated macrophages is due to a complex cellular signaling initiated by TLR4 that leads to the transcription of IFN-β, which activates IRF-1 and STAT-1, as well as to the activation of NF-κB, required for iNOS transcription. High concentrations of LPS can also be uptaken by scavenger receptors (SRs), which, in concert with TLR4, leads to inflammatory responses. The mechanisms by which TLR4 and SRs interact, and the pathways activated by this interaction in macrophages are not elucidated. Therefore, our main goal was to evaluate the role of SRs, particularly SR-A, in LPS-stimulated macrophages for NO production. We first showed that, surprisingly, LPS can induce the expression of iNOS and the production of NO in TLR4−/− mice, provided exogenous IFN-β is supplied. These results indicate that LPS stimulate receptors other than TLR4. The inhibition of SR-A using DSS or neutralizing antibody to SR-AI showed that SR-A is essential for the expression of iNOS and NO production in stimulation of TLR4 by LPS. The restoration of the ability to express iNOS and produce NO by addition of rIFN-β to inhibited SR-A cells indicated that the role of SR-AI in LPS-induced NO production is to provide IFN-β, probably by mediating the internalization of LPS/TLR4, and the differential inhibition by DSS and neutralizing antibody to SR-AI suggested that other SRs are also involved. Our results reinforce that TLR4 and SR-A act in concert in LPS activation and demonstrated that, for the production of NO, it does mainly by synthesizing IRF-3 and also by activating the TRIF/IRF-3 pathway for IFN-β production, essential for LPS-mediated transcription of iNOS. Consequently STAT-1 is activated, and IRF-1 is expressed, which together with NF-κB from TLR4/MyD88/TIRAP, induce iNOS synthesis and NO production. TLR4 and SRs act in concert activating IRF-3 to transcribe IFN-β and activate STAT-1 to produce NO by LPS-activated macrophages. • LPS-activated TLR4-deficient cells are able to produce NO provided IFN-β is supplied. • SRs are indispensable for TLR4-mediated NO production in macrophages. • SR-AI acts in concert with TLR4 to internalize LPS and activate TRIF/IRF-3 pathway. • SR-AI is needed for TLR4-mediated expression of IFN-β, crucial for iNOS expression. • SR-AI is needed for activation of STAT-1 and expression of IRF-1, crucial for iNOS expression. [ABSTRACT FROM AUTHOR]

ObjectivesThis study sought to develop an integrative positron emission tomography (PET) with magnetic resonance imaging (MRI) procedure for accurate atherosclerotic plaque phenotyping, facilitated by clinically approved and nanobody radiotracers.BackgroundNoninvasive characterization of atherosclerosis remains a challenge in clinical practice. The limitations of current diagnostic methods demonstrate that, in addition to atherosclerotic plaque morphology and composition, disease activity needs to be evaluated.MethodsWe screened 3 nanobody radiotracers targeted to different biomarkers of atherosclerosis progression, namely vascular cell adhesion molecule (VCAM)-1, lectin-like oxidized low-density lipoprotein receptor (LOX)-1, and macrophage mannose receptor (MMR). The nanobodies, initially radiolabeled with copper-64 (64Cu), were extensively evaluated in Apoe-/- mice and atherosclerotic rabbits using a combination of in vivo PET/MRI readouts and ex vivo radioactivity counting, autoradiography, and histological analyses.ResultsThe 3 nanobody radiotracers accumulated in atherosclerotic plaques and displayed short circulation times due to fast renal clearance. The MMR nanobody was selected for labeling with gallium-68 (68Ga), a short-lived radioisotope with high clinical relevance, and used in an ensuing atherosclerosis progression PET/MRI study. Macrophage burden was longitudinally studied by 68Ga-MMR-PET, plaque burden by T2-weighted MRI, and neovascularization by dynamic contrast-enhanced (DCE) MRI. Additionally, inflammation and microcalcifications were evaluated by fluorine-18 (18F)-labeled fluorodeoxyglucose (18F-FDG) and 18F-sodium fluoride (18F-NaF) PET, respectively. We observed an increase in all the aforementioned measures as disease progressed, and the imaging signatures correlated with histopathological features.ConclusionsWe have evaluated nanobody-based radiotracers in rabbits and developed an integrative PET/MRI protocol that allows noninvasive assessment of different processes relevant to atherosclerosis progression. This approach allows the multiparametric study of atherosclerosis and can aid in early stage anti-atherosclerosis drug trials.

Our lack of understanding of the immune microenvironment in canine osteosarcoma (cOSA) has limited the identification of potential immunotherapeutic targets. In particular, our ability to utilize readily available tissue from a dog's primary tumour to predict the type and extent of immune response in their pulmonary metastatic lesions is unknown. We, therefore, collected 21 matched pairs of primary tumours and pulmonary metastatic lesions from dogs with OSA and performed immunohistochemistry to quantify T-lymphocyte (CD3), FOXP3+ cell, B-lymphocyte (Pax-5), and CD204+ macrophage infiltration. We found that T-lymphocytes and FOXP3+ infiltrates in primary tumours positively correlated with that of metastatic lesions (ρ = 0.512, P = 0.038 and ρ = 0.698, P = 0.007, respectively), while a strong trend existed for CD204+ infiltrates (ρ = 0.404, P = 0.087). We also observed T- and B-lymphocytes, and CD204+ macrophages to be significantly higher in a dog's pulmonary metastasis compared to their primary tumour (P = 0.018, P = 0.018, P = 0.016, respectively), while FOXP3+ cells were only significantly higher in metastases when all primary tumour and metastasis lesions were compared without pairing (P = 0.036). Together, these findings suggest that the metastatic immune microenvironment may be influenced by that of the primary cOSA, and that primary tumour immune biomarkers could potentially be applied to predict immunotherapeutic responses in gross metastatic disease. We, therefore, provide a rationale for the treatment of cOSA pulmonary metastases with immunotherapeutics that enhance the anti-tumour activity of these immune cells, particularly in dogs with moderate to high immune cell infiltration in their primary tumours.

An active form of cathelicidin antimicrobial peptide, LL-37, has immunomodulatory and stimulatory effects, though the specific pathways are not clear. The purpose of this study was to identify the cellular pathways by which LL-37 amplifies the inflammation induced by damage-associated molecular patterns (DAMPs). We performed DNA microarray, reverse transcription polymerase chain reaction, immunoblotting, and proximity ligation assays using cultured keratinocytes treated with LL-37 and/or the DAMP poly(I:C), a synthetic double-stranded RNA. In contrast to the combination of LL-37 and poly(I:C), LL-37 alone induced genes related to biological metabolic processes such as VEGFA and PTGS2 (COX-2). Inhibition of FPR2, a known receptor for cathelicidin, partially suppressed the induction of VEGFA and PTGS2. Importantly, VEGFA and PTGS2 induced by LL-37 alone were diminished by the knockdown of scavenger receptors including SCARB1 (SR-B1), OLR1 (SR-E1), and AGER (SR-J1). Moreover, LL-37 alone, as well as the combination of LL-37 and poly(I:C), showed proximity to the scavenger receptors, indicating that LL-37 acts via scavenger receptors and intermediates between them and poly(I:C). These results showed that the broad function of cathelicidin is generally dependent on scavenger receptors. Therefore, inhibitors of scavenger receptors or non-functional mock cathelicidin peptides may serve as new anti-inflammatory and immunosuppressive agents. [ABSTRACT FROM AUTHOR]

Atherosclerosis is a chronic inflammation characterized by macrophage infiltration, lipid deposition, and arterial wall thickening. Prevention of atherosclerosis by nutraceuticals is gaining attention. Myricetin, a dietary flavonol, is claimed to possess anti-atherosclerosis properties. We studied myricetin's effect on the atherosclerosis-associated molecular mechanism. Cytotoxicity and proliferation testing to check the viability of myricetin-treated THP-1 macrophages and monocyte migration study in the presence and absence of myricetin was performed. The whole transcriptome analysis was conducted using the Affymetrix microarray platform. The Partek genomics suite for detecting differentially expressed genes (DEGs) and ingenuity pathway analysis was used to identify canonical pathways. Cytotoxicity assays exhibited no significant toxicity in THP-1 macrophages treated with different myricetin concentrations (10–200 μM). Genome-wide expression profiling revealed 58 DEGs (53 upregulated and 5 downregulated) in myricetin-treated THP-1 macrophages. Pathway analysis revealed inhibition of LXR/RXR activation and angiogenesis inhibition by thrombospondin-1 and activated phagocytosis in myricetin-treated THP-1 macrophages. The cytotoxicity assay shows myricetin as a safe phytochemical. In vitro and in silico pathway studies on THP-1 macrophages showed that they can inhibit THP-1 monocyte migration and alter the cholesterol efflux mediated via LXR/RXR signaling. Therefore, myricetin could help in the prevention of cell infiltration in atherosclerotic plaque with reduced risk of stroke or brain damage. [ABSTRACT FROM AUTHOR]

Abstract Scavenger receptors belong to a superfamily of proteins that are structurally heterogeneous and encompass the miscellaneous group of transmembrane proteins and soluble secretory extracellular domain. They are functionally diverse as they are involved in various disorders and biological pathways and their major function in innate immunity and homeostasis. Numerous scavenger receptors have been discovered so far and are apportioned in various classes (A-L). Scavenger receptors are documented as pattern recognition receptors and known to act in coordination with other co-receptors such as Toll-like receptors in generating the immune responses against a repertoire of ligands such as microbial pathogens, non-self, intracellular and modified self-molecules through various diverse mechanisms like adhesion, endocytosis and phagocytosis etc. Unlike, most of the scavenger receptors discussed below have both membrane and soluble forms that participate in scavenging; the role of a potential scavenging receptor Angiotensin-Converting Enzyme-2 has also been discussed whereby only its soluble form might participate in preventing the pathogen entry and replication, unlike its membrane-bound form. This review majorly gives an insight on the functional aspect of scavenger receptors in host defence and describes their mode of action extensively in various immune pathways involved with each receptor type. Video abstract

Microglia are intrinsic components of the brain immune system and are activated in many central nervous system disorders. The ability to noninvasively image these cells would provide valuable information for both research and clinical applications. Today, most imaging probes for activated microglia are mainly designed for positron emission tomography (PET) and target translocator proteins that also reside on other cerebral cells. The PET images obtained are not specific for microglia-driven inflammation. Here, we describe a potential PET/MRI multimodal imaging probe that selectively targets the scavenger receptor class A (SR-A) expressed on activated microglia. These sulfated dextran-coated iron oxide (SDIO) nanoparticles are avidly taken up by microglia and appear to be nontoxic when administered intravenously in a mouse model. Intravenous administration of this SDIO demonstrated visualization by T2∗ -weighted MRI of microglia activated by intracerebral administration of tumor necrosis factor alpha (TNF-α). The contrast was significantly enhanced by SDIO, whereas there was little to no contrast change in animals treated with nontargeted nanoparticles or untreated controls. Thus, SR-A targeting represents a promising strategy to image activated microglia in the brain.

Increased consumption of Western-type diets and environmental insults lead to wide-spread increases in the plasma levels of saturated fatty acids and lipoprotein oxidation. The aim of this study is to examine whether palmitate and minimally modified low-density lipoprotein (mmLDL) exert an additive effect on macrophage activation. We found that CXCL2 and TNF-α secretion as well as ERK and p38 phosphorylation were additively increased by co-treatment of J774 macrophages with palmitate and mmLDL in the presence of lipopolysaccharide (LPS). Furthermore, the analysis of differentially expressed genes using the KEGG database revealed that several pathways, including cytokine-cytokine receptor interaction, and genes were significantly altered. These results were validated with real-time PCR, showing upregulation of Il-6, Csf3, Il-1β, and Clec4d. The present study demonstrated that palmitate and mmLDL additively potentiate the LPS-induced activation of macrophages. These results suggest the existence of synergistic mechanisms by which saturated fatty acids and oxidized lipoproteins activate immune cells.

An ability of poorly differentiated cells of different genesis, including tumor stem-like cells (TSCs), to internalize extracellular double-stranded DNA (dsDNA) fragments was revealed in our studies. Using the models of Krebs-2 murine ascites carcinoma and EBV-induced human B-cell lymphoma culture, we demonstrated that dsDNA internalization into the cell consists of several mechanistically distinct phases. The primary contact with cell membrane factors is determined by electrostatic interactions. Firm contacts with cell envelope proteins are then formed, followed by internalization into the cell of the complex formed between the factor and the dsDNA probe bound to it. The key binding sites were found to be the heparin-binding domains, which are constituents of various cell surface proteins of TSCs—either the C1q domain, the collagen-binding domain, or domains of positively charged amino acids. These results imply that the interaction between extracellular dsDNA fragments and the cell, as well as their internalization, took place with the involvement of glycocalyx components (proteoglycans/glycoproteins (PGs/GPs) and glycosylphosphatidylinositol-anchored proteins (GPI-APs)) and the system of scavenger receptors (SRs), which are characteristic of TSCs and form functional clusters of cell surface proteins in TSCs. The key provisions of the concept characterizing the principle of organization of the "group-specific" cell surface factors of TSCs of various geneses were formulated. These factors belong to three protein clusters: GPs/PGs, GIP-APs, and SRs. For TSCs of different tumors, these clusters were found to be represented by different members with homotypic functions corresponding to the general function of the cluster to which they belong. [ABSTRACT FROM AUTHOR]

Scavenger receptors are part of a complex surveillance system expressed by host cells to efficiently orchestrate innate immune response against bacterial infections. Stabilin-1 (STAB-1) is a scavenger receptor involved in cell trafficking, inflammation, and cancer; however, its role in infection remains to be elucidated. Listeria monocytogenes (Lm) is a major intracellular human food-borne pathogen causing severe infections in susceptible hosts. Using a mouse model of infection, we demonstrate here that STAB-1 controls Lm-induced cytokine and chemokine production and immune cell accumulation in Lm-infected organs. We show that STAB-1 also regulates the recruitment of myeloid cells in response to Lm infection and contributes to clear circulating bacteria. In addition, whereas STAB-1 appears to promote bacterial uptake by macrophages, infection by pathogenic Listeria induces the down regulation of STAB-1 expression and its delocalization from the host cell membrane. We propose STAB-1 as a new SR involved in the control of Lm infection through the regulation of host defense mechanisms, a process that would be targeted by bacterial virulence factors to promote infection.

Background and Objectives: Cardiovascular (CV) risk is elevated in rheumatoid arthritis (RA). RA patient plasma causes pro-atherogenic derangements in cholesterol transport leading to macrophage foam cell formation (FCF). The TARGET randomized clinical trial compares CV benefits of 2 RA drug regimens. Hydoxychloroquine (HCQ) is a key medication used in TARGET. This study examines effects of HCQ on lipid transport to elucidate mechanisms underlying TARGET outcomes and as an indicator of likely HCQ effects on atherosclerosis in RA. Materials and Methods: THP1 human macrophages were exposed to media alone, IFNγ (atherogenic cytokine), HCQ, or HCQ + IFNγ. Cholesterol efflux protein and scavenger receptor mRNA levels were quantified by qRT-PCR and corresponding protein levels were assessed by Western blot. FCF was evaluated via Oil-Red-O and fluorescent-oxidized LDL. Intracellular cholesterol and efflux were quantified with Amplex Red assay. Results: With the exception of a decrease in the efflux protein cholesterol 27-hydroxylase in the presence IFNγ at all HCQ concentrations, no significant effect on gene or protein expression was observed upon macrophage exposure to HCQ and this was reflected in the lack of change in FCF and oxidized LDL uptake. Conclusions: HCQ did not significantly affect THP1 macrophage cholesterol transport. This is consistent with TARGET, which postulates superior effects of anti-TNF agents over sulfasalazine + HCQ. [ABSTRACT FROM AUTHOR]

Enhanced bioactive anti-oxidant formulations are critical for treatment of inflammatory diseases, such as atherosclerosis. A hallmark of early atherosclerosis is the uptake of oxidized low density lipoprotein (oxLDL) by macrophages, which results in foam cell and plaque formation in the arterial wall. The hypolipidemic, anti-inflammatory, and antioxidative properties of polyphenol compounds make them attractive targets for treatment of atherosclerosis. However, high concentrations of antioxidants can reverse their anti-atheroprotective properties and cause oxidative stress within the artery. Here, we designed a new class of nanoparticles with anti-oxidant polymer cores and shells comprised of scavenger receptor targeting amphiphilic macromolecules (AMs). Specifically, we designed ferulic acid-based poly(anhydride-ester) nanoparticles to counteract the uptake of high levels of oxLDL and regulate reactive oxygen species generation (ROS) in human monocyte derived macrophages (HMDMs). Compared to all compositions examined, nanoparticles with core ferulic acid-based polymers linked by diglycolic acid (PFAG) showed the greatest inhibition of oxLDL uptake. At high oxLDL concentrations, the ferulic acid diacids and polymer nanoparticles displayed similar oxLDL uptake. Treatment with the PFAG nanoparticles downregulated the expression of macrophage scavenger receptors, CD-36, MSR-1, and LOX-1 by about 20-50%, one of the causal factors for the decrease in oxLDL uptake. The PFAG nanoparticle lowered ROS production by HMDMs, which is important for maintaining macrophage growth and prevention of apoptosis. Based on these results, we propose that ferulic acid-based poly(anhydride ester) nanoparticles may offer an integrative strategy for the localized passivation of the early stages of the atheroinflammatory cascade in cardiovascular disease.Statement of significanceFuture development of anti-oxidant formulations for atherosclerosis applications is essential to deliver an efficacious dose while limiting localized concentrations of pro-oxidants. In this study, we illustrate the potential of degradable ferulic acid-based polymer nanoparticles to control macrophage foam cell formation by significantly reducing oxLDL uptake through downregulation of scavenger receptors, CD-36, MSR-1, and LOX-1. Another critical finding is the ability of the degradable ferulate-based polymer nanoparticles to lower macrophage reactive oxygen species (ROS) levels, a precursor to apoptosis and plaque escalation. The degradable ferulic acid-based polymer nanoparticles hold significant promise as a means to alter the treatment and progression of atherosclerosis.

The study aimed to achieve enhanced targeted cytotoxicity and cell-internalization of cisplatin-loaded deoxyribonucleic acid-nanothread (CPT-DNA-NT), mediated by scavenger receptors into HeLa cells. DNA-NT was developed with stiff-topology utilizing circular-scaffold to encapsulate CPT. Atomic force microscopy (AFM) characterization of the DNA-NT showed uniformity in the structure with a diameter of 50–150 nm and length of 300–600 nm. The successful fabrication of the DNA-NT was confirmed through native-polyacrylamide gel electrophoresis analysis, as large the molecular-weight (polymeric) DNA-NT did not split into constituting strands under applied current and voltage. The results of cell viability confirmed that blank DNA-NT had the least cytotoxicity at the highest concentration (512 nM) with a viability of 92% as evidence of its biocompatibility for drug delivery. MTT assay showed superior cytotoxicity of CPT-DNA-NT than that of the free CPT due to the depot release of CPT after DNA-NT internalization. The DNA-NT exhibited targeted cell internalizations with the controlled intracellular release of CPT (from DNA-NT), as illustrated in confocal images. Therefore, in vitro cytotoxicity assessment through flow cytometry showed enhanced apoptosis (72.7%) with CPT-DNA-NT (compared to free CPT; 64.4%). CPT-DNA-NT, being poly-anionic, showed enhanced endocytosis via scavenger receptors.

DNA-nanotechnology-based nano-architecture scaffolds based on circular strands were designed in the form of DNA-nanowires (DNA-NWs) as a polymer of DNA-triangles. Circularizing a scaffold strand (84-NT) was the critical step followed by annealing with various staple strands to make stiff DNA-triangles. Atomic force microcopy (AFM), native polyacrylamide gel electrophoresis (PAGE), UV-analysis, MTT-assay, flow cytometry, and confocal imaging were performed to assess the formulated DNA-NWs and cisplatin (CPT) loading. The AFM and confocal microscopy images revealed a uniform shape and size distribution of the DNA-NWs, with lengths ranging from 2 to 4 μm and diameters ranging from 150 to 300 nm. One sharp band at the top of the lane (500 bp level) with the loss of electrophoretic mobility during the PAGE (native) gel analysis revealed the successful fabrication of DNA-NWs. The loading efficiency of CPT ranged from 66.85% to 97.35%. MTT and flow cytometry results showed biocompatibility of the blank DNA-NWs even at 95% concentration compared with the CPT-loaded DNA-NWs. The CPT-loaded DNA-NWs exhibited enhanced apoptosis (22%) compared to the apoptosis (7%) induced by the blank DNA-NWs. The release of CPT from the DNA-NWs was sustained at < 75% for 6 h in the presence of serum, demonstrating suitability for systemic applications. The IC50 of CPT@DNA-NWs was reduced to 12.8 nM CPT, as compared with the free CPT solution exhibiting an IC50 of 51.2 nM. Confocal imaging revealed the targetability, surface binding, and slow internalization of the DNA-NWs in the scavenger-receptor-rich cancer cell line (HepG2) compared with the control cell line.

The scavenger receptors (SRs) gene family is considered as the membrane-associated pattern recognition receptors that plays important roles in the immune responses of organisms. However, there is currently limited research on the systematic identification of the SRs gene family in teleost and their role in the innate immunity of S. schegelii. In this study, we identified and annotated 15 SRs genes in S. schegelii. Through phylogenetic analysis, analysis of conserved domains, gene structure, and motif composition, we found that SRs gene family within different classes were relatively conserved. Additionally, we used qRT-PCR to analyze the expression patterns of SRs genes in immune-related tissues from healthy and Acinetobacter johnsonii -infected S. schegelii. The results showed that SRs genes exhibited different tissue expression patterns and the expression of SRs genes significantly changed after A. johnsonii infection. These results provided a valuable basis for further understanding of the functions of SRs in the innate immune response of S. schegelii. • 15 SRs genes from eight classes were identified for the first time in black rockfish (Sebastes schlegelii). • Revealed the relative conservatism of the SRs gene family in Sebastes schlegelii. • SRs genes exhibited different tissue expression patterns. • The expression of some SRs genes significantly changed after Acinetobacter johnsonii infection. [ABSTRACT FROM AUTHOR]

Deposition of fibrillar forms of amyloid β-protein (Aβ) is commonly found in patients with Alzheimer's disease (AD) associated with cognitive decline. Impaired clearance of Aβ species is thought to be a major cause of late-onset sporadic AD. Aβ secreted into the extracellular milieu can be cleared from the brain through multiple pathways, including cellular uptake in neuronal and non-neuronal cells. Recent studies have showed that the naturally-occurring polyphenol amentoflavone (AMF) exerts anti-amyloidogenic effects. However, its effects on metabolism and cellular clearance of Aβ remain to be tested. In the present study, we demonstrated that AMF significantly increased the cellular uptake of both Aβ1-40 and Aβ1-42, but not inverted Aβ42-1 in mouse neuronal N2a cells. Though AMF promoted internalization of cytotoxic Aβ1-42, it significantly reduced cell death in our assay condition. Our data further revealed that the internalized Aβ is translocated to lysosomes and undergoes enzymatic degradation. The saturable kinetic of Aβ uptake and our pharmacologic experiments showed the involvement of receptor-mediated endocytosis, in part, through the class A scavenger receptors as a possible mechanism of action of AMF. Taken together, our findings indicate that AMF can lower the levels of extracellular Aβ by increasing their cellular uptake and clearance, suggesting the therapeutic potential of AMF for the treatment of AD. [ABSTRACT FROM AUTHOR]

Oxidative stress and inflammation cannot be considered as diseases themselves; however, they are major risk factors for the development and progression of the pathogenesis underlying many illnesses, such as cancer, neurological disorders (including Alzheimer's disease and Parkinson's disease), autoimmune and metabolic disorders, etc. According to the results obtained from extensive studies, oxidative stress–induced biomolecules, such as advanced oxidation protein products, advanced glycation end products, and advanced lipoxidation end products, are critical for an accelerated level of inflammation and oxidative stress–induced cellular damage, as reflected in their strong affinity to a wide range of scavenger receptors. Based on the limitations of antioxidative and anti-inflammatory molecules in practical applications, targeting such interactions between harmful molecules and their cellular receptors/signaling with advances in gene engineering technology, such as CRISPR or TALEN, may prove to be a safe and effective alternative. In this review, we summarize the findings of recent studies focused on the deletion of scavenger receptors under oxidative stress as a development in the therapeutic approaches against the diseases linked to inflammation and the contribution of advanced glycation end products (AGEs), advanced lipid peroxidation products (ALEs), and advanced oxidation protein products (AOPPs). [ABSTRACT FROM AUTHOR]

Scavenger receptors (SR) are not only pattern recognition receptors involved in the immune response against pathogens but are also important receptors exploited by different virus to enter host cells, and thus represent targets for antiviral therapy. The high mutation rates of viruses, as well as their small genomes are partly responsible for the high rates of virus resistance and effective treatments remain a challenge. Most currently approved formulations target viral-encoded factors. Nevertheless, host proteins may function as additional targets. Thus, there is a need to explore and develop new strategies aiming at cellular factors involved in virus replication and host cell entry. SR-virus interactions have implications in the pathogenesis of several viral diseases and in adenovirus-based vaccination and gene transfer technologies, and may function as markers of severe progression. Inhibition of SR could reduce adenoviral uptake and improve gene therapy and vaccination, as well as reduce pathogenesis. In this review, we will examine the crucial role of SR play in cell entry of different types of human virus, which will allow us to further understand their role in protection and pathogenesis and its potential as antiviral molecules. The recent discovery of SR-B1 as co-factor of SARS-Cov-2 (severe acute respiratory syndrome coronavirus 2) entry is also discussed. Further fundamental research is essential to understand molecular interactions in the dynamic virus-host cell interplay through SR for rational design of therapeutic strategies. [ABSTRACT FROM AUTHOR]

The retinal phagocytic machinery resembles the one used by macrophages to clear apoptotic cells. However, in the retina, the permanent contact between photoreceptor outer segments (POS) and retinal pigment epithelial (RPE) cells requires a tight control of this circadian machinery. In addition to the known receptors synchronizing POS internalization, several others are expressed by RPE cells. Notably, scavenger receptor CD36 has been shown to intervene in the internalization speed. We thus investigated members of the scavenger receptor family class A SR-AI and MARCO and class B CD36, SR-BI and SR-B2/LIMP-2 using immunoblotting, immunohisto- and immunocytochemistry, lipid raft flotation gradients, phagocytosis assays after siRNA/antibody inhibition, RT-qPCR and western blot analysis along the light:dark cycle. All receptors were expressed by RPE cell lines and tissues and colocalized with POS, except SR-BI. All receptors were associated with lipid rafts, and even more upon POS challenge. SR-B2/LIMP-2 inhibition suggested a role in the control of the internalization speed similar to CD36. In vivo, MARCO and CD36 displayed rhythmic gene and protein expression patterns concomitant with the phagocytic peak. Taken together, our results indicate that CD36 and SR-B2/LIMP-2 play a direct regulatory role in POS phagocytosis dynamics, while the others such as MARCO might participate in POS clearance by RPE cells either as co-receptors or via an indirect process. [ABSTRACT FROM AUTHOR]

Scavenger receptors form a superfamily of membrane-bound receptors that bind and internalize different types of ligands, including pro-atherogenic oxidized low-density lipoproteins (oxLDLs). In vitro studies have indicated a role for the liver sinusoidal endothelial cell receptors stabilin 1 (stab1) and 2 (stab2) in oxLDL clearance. In this study, we evaluated the potential role of stab1 and stab2 in lipoprotein uptake in zebrafish, an upcoming model for studying cholesterol metabolism and atherosclerosis. Lipoproteins were injected in the duct of Cuvier of wild-type (ABTL) or stab1 and stab2 mutant (stab1 −/− stab2 −/−) zebrafish larvae at 3 days post-fertilization. To examine the effect of stabilin deficiency on lipoprotein and cholesterol metabolism, zebrafish larvae were challenged with a high cholesterol diet (HCD; 4% w/w) for 10 days. Lipoprotein injections showed impaired uptake of both LDL and oxLDL into the vessel wall of caudal veins of stab1 −/− stab2 −/− zebrafish, which was paralleled by redistribution to tissue macrophages. Total body cholesterol levels did not differ between HCD-fed stab1 −/− stab2 −/− and ABTL zebrafish. However, stab1 −/− stab2 −/− larvae exhibited 1.4-fold higher mRNA expression levels of ldlra involved in (modified) LDL uptake, whereas the expression levels of scavenger receptors scarb1 and cd36 were significantly decreased. We have shown that stabilins 1 and 2 have an important scavenging function for apolipoprotein B-containing lipoproteins in zebrafish and that combined deficiency of these two proteins strongly upregulates the clearance of lipoproteins by macrophages within the caudal vein. Our current study highlights the use of zebrafish as model to study lipoprotein metabolism and liver sinusoidal endothelial cell function. [Display omitted] • Stabilin 1 and 2 are key players in zebrafish lipoprotein metabolism. • Stabilin 1 and 2 double deficiency induces macrophage oxidized low-density lipoproteins (oxLDLs) uptake in zebrafish. • Stabilin 1/2 double knockout zebrafish are resistant to vascular lipid deposition. • Zebrafish are a good model to study liver sinusoidal endothelial cell function. [ABSTRACT FROM AUTHOR]

Lectin-like oxidized low-density lipoprotein (LDL) receptor-1 (LOX-1) is an endothelial receptor for oxidized LDL. Increased expression of LOX-1 has been demonstrated in atherosclerotic lesions and diabetic vasculopathy. In this study, we investigate the expression of LOX-1 receptor in sickle cell disease (SCD) vasculopathy. Expression of LOX-1 in brain vascular endothelium is markedly increased and LOX-1 gene expression is upregulated in cultured human brain microvascular endothelial cells by incubation with SCD erythrocytes. Also, the level of circulating soluble LOX-1 concentration is elevated in the plasma of SCD patients. Increased LOX-1 expression in endothelial cells is potentially involved in the pathogenesis of SCD vasculopathy. Soluble LOX-1 concentration in SCD may provide a novel biomarker for risk stratification of sickle cell vascular complications.

OBJECTIVE: Approximately 30% of autogenous vein grafts develop luminal narrowing and fail because of intimal hyperplasia or negative remodeling. We previously found that vein graft cells from patients who later develop stenosis proliferate more in vitro in response to growth factors than cells from patients who maintain patent grafts. To discover novel determinants of vein graft outcome, we have analyzed gene expression profiles of these cells using a systems biology approach to cluster the genes into modules by their coexpression patterns and to correlate the results with growth data from our prior study and with new studies of migration and matrix remodeling. METHODS: RNA from 4-hour serum- or platelet-derived growth factor (PDGF)-BB-stimulated human saphenous vein cells obtained from the outer vein wall (20 cell lines) was used for microarray analysis of gene expression, followed by weighted gene coexpression network analysis. Cell migration in microchemotaxis chambers in response to PDGF-BB and cell-mediated collagen gel contraction in response to serum were also determined. Gene function was determined using short-interfering RNA to inhibit gene expression before subjecting cells to growth or collagen gel contraction assays. These cells were derived from samples of the vein grafts obtained at surgery, and the long-term fate of these bypass grafts was known. RESULTS: Neither migration nor cell-mediated collagen gel contraction showed a correlation with graft outcome. Although 1188 and 1340 genes were differentially expressed in response to treatment with serum and PDGF, respectively, no single gene was differentially expressed in cells isolated from patients whose grafts stenosed compared with those that remained patent. Network analysis revealed four unique groups of genes, which we term modules, associated with PDGF responses, and 20 unique modules associated with serum responses. The yellow and skyblue modules, from PDGF and serum analyses, respectively, correlated with later graft stenosis (P = .005 and P = .02, respectively). In response to PDGF, yellow was also associated with increased cell growth. For serum, skyblue was also associated with inhibition of collagen gel contraction. The hub genes for yellow and skyblue (ie, the gene most connected to other genes in the module), scavenger receptor class A member 5 (SCARA5) and suprabasin (SBSN), respectively, were tested for effects on proliferation and collagen contraction. Knockdown of SCARA5 increased proliferation by 29.9% ± 7.8% (P < .01), whereas knockdown of SBSN had no effect. Knockdown of SBSN increased collagen gel contraction by 24.2% ± 8.6% (P < .05), whereas knockdown of SCARA5 had no effect. CONCLUSIONS: Using weighted gene coexpression network analysis of cultured vein graft cell gene expression, we have discovered two small gene modules, which comprise 42 genes, that are associated with vein graft failure. Further experiments are needed to delineate the venous cells that express these genes in vivo and the roles these genes play in vein graft healing, starting with the module hub genes SCARA5 and SBSN, which have been shown to have modest effects on cell proliferation or collagen gel contraction.

AIMS:We recently reported that immunosuppression with FTY720 improves cardiac function and extends longevity in Hypomorphic ApoE mice deficient in scavenger receptor Type-BI expression, also known as the HypoE/SR-BI(–/–) mouse model of diet-induced coronary atherosclerosis and myocardial infarction (MI). In this study, we tested the impact of FTY720 on cardiac dysfunction in HypoE/SR-BI(–/–) mice that survive MI and subsequently develop chronic heart failure. METHODS/RESULTS:HypoE/SR-BI(–/–) mice were bred to Mx1-Cre transgenic mice, and offspring were fed a high-fat diet (HFD) for 3.5 weeks to provoke hyperlipidemia, coronary atherosclerosis, and recurrent MIs. In contrast to our previous study, hyperlipidemia was rapidly reversed by inducible Cre-mediated gene repair of the HypoE allele and switching mice to a normal chow diet. Mice that survived the period of HFD were subsequently given oral FTY720 in drinking water or not, and left ventricular (LV) function was monitored using serial echocardiography for up to 15 weeks. In untreated mice, LV performance progressively deteriorated. Although FTY720 treatment did not initially prevent a decline of heart function among mice 6 weeks after Cre-mediated gene repair, it almost completely restored normal LV function in these mice by 15 weeks. Reversal of heart failure did not result from reduced atherosclerosis as the burden of aortic and coronary atherosclerosis actually increased to similar levels in both groups of mice. Rather, FTY720 caused systemic immunosuppression as assessed by reduced numbers of circulating T and B lymphocytes. In contrast, FTY720 did not enhance the loss of T cells or macrophages that accumulated in the heart during the HFD feeding period, but it did enhance the loss of B cells soon after plasma lipid lowering. Moreover, FTY720 potently reduced the expression of matrix metalloproteinase-2 and genes involved in innate immunity-associated inflammation in the heart. CONCLUSIONS:Our data demonstrate that immunosuppression with FTY720 prevents postinfarction myocardial remodeling and chronic heart failure.

The scavenger receptor superfamily represents a highly diverse collection of evolutionarily-conserved receptors which are known to play key roles in host homeostasis, the most prominent of which is the clearance of unwanted endogenous macromolecules, such as oxidized low-density lipoproteins, from the systemic circulation. Members of this family have also been well characterized in their binding and internalization of a vast range of exogenous antigens and, consequently, are generally considered to be pattern recognition receptors, thus contributing to innate immunity. Several studies have implicated scavenger receptors in the pathophysiology of several inflammatory diseases, such as Alzheimer's and atherosclerosis. Hepatic resident cellular populations express a diverse complement of scavenger receptors in keeping with the liver's homeostatic functions, but there is gathering interest in the contribution of these receptors to hepatic inflammation and its complications. Here, we review the expression of scavenger receptors in the liver, their functionality in liver homeostasis, and their role in inflammatory liver disease and cancer. [ABSTRACT FROM AUTHOR]