Receptor CD36 Research Articles

Dysfunction in B cell tolerance and activation in obesity.

Abstract Obesity is a complex condition that affects a large part of the global population, making it a major risk factor for premature mortality and medical complications. Obesity profoundly engages the immune system, impacting cancer immunity, infections, and autoimmunity. Additionally, obesity is associated with a low-grade chronic inflammation which exacerbates metabolic dysfunction. B cells play a significant role in obesity-associated inflammation and metabolic dysfunction, but their effect is subset-dependent and can be either protective or pathogenic. Therefore, understanding how different B cell populations contribute to these phenotypes represent a crucial goal. Our lab has identified a subset of B lymphocytes which is expanded in the adipose tissue of humans and mice during obesity. These B cells co-express the transcription factor T-bet and the integrin CD11c, and they produce IgG2c antibodies. In vivo studies with mice fed a high fat diet revealed that these Tbet+ B cells promote inflammation through the production of proinflammatory antibodies. Additionally, these B cells express high levels of the scavenger receptor CD36, a lipid-transport molecule associated with inflammation and atherosclerosis. As a potential consequence of the CD36 expression, the Tbet+ B cells maintain a higher intracellular lipid content and are capable of increased uptake of oxidized LDL compared to conventional B cells, suggesting a unique lipid metabolism. Because of their unique nature, T-bet+ B cells represent a potential link between obesity, inflammation, and autoimmunity. Identifying the mechanisms regulating their function will provide new potential targets to reduce inflammation and immunological dysfunction in obese patients. Supported by R01 AI132798

Amelioration of acute lung injury by peptides targeting scavenger receptor CD36

CD36 belongs to scavenger receptor class B and is a fatty acid translocase. Our previous studies have suggested that the receptor may be involved in regulation of responses to endotoxin and inflammation of the lungs in mice. In current studies, we have designed a group of helical peptides that interact with and modulate the activities of CD36 and tested their activities against endothelial barrier dysfunction and acute lung injuries in mice. Peptides that inhibit CD36 activities including L37pA, ELR‐B and ELR‐BP reversed or mitigated the barrier disruptive activities induced by LPS, HKSA, truncated phospholipids and histone H3 as measured by TER (trans‐endothelial electric resistance). In consistence with this, the peptides also enhanced the barrier protective activities of OxPAPC. The peptides were delivered intravenously to mice treated with LPS or HKSA to induce acute lung injuries. It has been observed that among peptides that antagonize the activities of CD36, L37pA, ELR‐B and ELR‐BP have demonstrated inhibitory effects on acute lung injuries induced by LPS characterized by reduced extravagance of inflammatory cells from circulation to bronchoalveolar lavage (BAL) and a decrease in protein concentration in BAL fluid from the peptide‐treated mice. The observation indicates that modulation of CD36 activities by peptide inhibitors is a novel strategy to reduce acute lung injuries induced by pro‐inflammatory agents.

Novel Roles of FFAR4 in Macrophage Foam Cell Formation

Free fatty acid receptor 4 (FFAR4), also known as G‐protein coupled receptor 120 (GPR120), is a long‐chain unsaturated fatty acid receptor expressed in adipocytes, endothelial cells, and macrophages. Activation of FFAR4 helps maintain metabolic homeostasis by regulating adipogenesis, insulin sensitivity, and inflammation. While FFAR4 is best known for its role its role in preventing obesity and diabetes, recent studies have demonstrated that FFAR4 may also play an important role in the development of atherosclerosis and cardiovascular disease (CVD). Given FFAR4’s importance in anti‐inflammatory signaling and high expression levels in macrophages, we designed experiments to test the hypothesis that FFAR4 prevents the development of atherosclerosis by reversing macrophage foam cell formation, a hallmark of early atherogenesis. In these studies, we isolated peritoneal macrophages from wild‐type C57/BL6J mice and incubated them with oxidized low‐density lipoprotein (oxLDL) to generate foam cells. We then investigated the effects of FFAR4 activation by GW9508 (a synthetic agonist) on lipid accumulation, cytokine secretion, and cholesterol efflux. Activation of FFAR4 by GW9508 decreased macrophage secretion of pro‐inflammatory cytokines. We also found that activation of FFAR4 with GW9508 reduced lipid accumulation in macrophages as observed by decreased Oil Red O staining and reduced cellular cholesterol content. Additionally, activation of FFAR4 by GW9508 increased [3H]‐cholesterol efflux to high‐density lipoprotein (HDL). Interestingly, the increased efflux was accompanied by decreased scavenger receptor CD36 expression (that mediates oxLDL uptake) and increased expression of ATP binding cassette transporters, ABCA1 and ABCG1 (that mediate cholesterol efflux). Taken together, our results support an exciting and novel role for FFAR4 in the reversal of foam cell formation and could emerge this receptor as a new target for treating CVD by preventing accumulation of atherosclerotic plaque.

Abstract 521: Parkin Has Non-canonical Roles In Macrophage Lipid Metabolism With Effects On Atherosclerosis

Defects in the E3 ubiquitin ligase Parkin are sine qua non of Parkinson and related neurodegenerative diseases. Loss of ubiquitination of distinct targets leads to accumulation of cytotoxic protein aggregates, loss of mitophagy, and defects in lipid metabolism. Given the critical role of these processes in plaque macrophages, we set out to define the role of Parkin in atherosclerosis. Pro-atherogenic LDLR-KO mice were transplanted with bone marrow (BM) from wild type and Parkin-KO mice and lesion formation stimulated by 4 months of Western diet feeding. Mice transplanted with wild type BM developed lipid-rich atherosclerotic plaques in both the aortic root and whole aorta replete with foam cell macrophages. In contrast, mice transplanted with Parkin-deficient BM had significantly reduced plaque burden with concomitant reductions in lipid content, foam cell macrophages, and surrogates of plaque complexity including reduced lesional apoptosis and necrotic core formation. The paradoxical atherogenic role for Parkin including development of lipid-laden foam cell macrophages led us to focus on the impact of Parkin in macrophage lipid handling. We find that Parkin mediates the ubiquitination and stabilization of scavenger receptor CD36, a key lipid transporter in macrophages. Parkin-null macrophages had reduced expression of CD36 and concomitant reductions in oxLDL uptake. Further, Parkin deficiency decreased cholesterol efflux in macrophages exacerbating macrophage lipid storage. These results suggest that the non-canonical functions of Parkin in lipid metabolism outweigh its traditional cytoprotective roles in macrophages and mediate its deleterious effects on atherosclerotic progression.

Pharmacological Modulation of B16‐F10 Murine Melanoma‐Induced TAMs Functional Phenotype via Lipoprotein‐Based Nanoparticles

Tumor‐associated macrophages (TAMs) are one of the most abundant immune cells in the tumor microenvironment (TME). Their immunosuppressive phenotype (M2) promotes tumor progression. The re‐education of TAMs in the TME to a pro‐immune phenotype (M1) is considered to enhance the effectiveness of the anti‐tumor immune response and leads to tumor regression. An effective therapeutic approach utilizing macrophage re‐polarization will likely require the selective delivery of agents to the TME to execute the M2‐to‐M1 phenotype transformation. This strategy is known to avoid adverse immune‐related events through the utilization of nanocarriers that can effectively penetrate the TME and interact with TAMs. As a drug delivery platform, reconstituted high‐density lipoprotein nanoparticles (rHDL NPs) have attracted increased interest in cancer therapeutics, owing to their biocompatibility and their capacity to transport lipophilic drugs toward their accumulation in the tumor. We and others have shown that the scavenger receptor class B type 1 (SR‐B1), at the surface of target cells, mediates the intracellular (non‐endocytic) delivery of rHDL NPs’ payload. The novel lipophilic agonist of the STimulator of INterferon Genes (STING), MSA‐2, has been used successfully to mediate the M2‐to‐M1 reversal in murine models of cancer. In this study, we aim to deliver MSA‐2 to TAMs using mannose‐decorated rHDL NPs to induce a conversion to an M1 phenotype. Because TAMs highly express the mannose receptor CD206 that facilitates the interaction of mannose with TAMs, inducing an M1 phenotype, we anticipate a successful outcome based on this strategy. We prepared and validated the retention of the PEGylated mannose moiety on the rHDL NPs. Fluorescence anisotropy and absorbance studies showed that rHDL NPs successfully encapsulated MSA‐2. IL‐4‐induced M2 macrophages and B16‐F10 murine melanoma conditioned media‐induced TAMs showed a higher SR‐B1 expression compared to M1 macrophages. The lower uptake of the surrogate dye payload by M1 macrophages (compared to M2 macrophages and TAMs) from the mannose‐decorated rHDL NPs, corroborates the importance of the SR‐B1 receptor in rHDL payload delivery. The above findings have significant therapeutic implications for the strategy of utilizing mannose‐decorated rHDL‐NPs to selectively target TAMs in the tumor microenvironment. In addition, this formulation of MSA‐2 with the mannose‐decorated rHDL NPs can be applied in a synergistic combination with mainstream immunotherapies to improve treatment outcomes for cancer patients, diagnosed with solid tumors.

Sea cucumber ether-phospholipids improve hepatic steatosis and enhance hypothalamic autophagy in high-fat diet-fed mice

As a promising group of natural bioactive lipids, ether-phospholipids (ether-PLs), exhibit the ability to attenuate high-fat diet (HFD)-induced lipid accumulation and atherosclerosis. However, the underlying mechanism is unclear. Autophagy has been implicated in the regulation of obesity. Therefore, we investigated the effects of dietary ether-PLs on hepatic steatosis and the activation of hypothalamic autophagy. HFD-fed C57BL/6J mice were orally administered with ether-PLs (150 mg/kg body weight) including plasmenyl phosphatidylethanolamine (PE-P) and plasmanyl phosphatidylcholine (PC-O) for three days or eight weeks. Ether-PLs supplementation relieved diet-induced hepatic lipid accumulation and regulated the hypothalamic peroxisome proliferator-activated receptor gamma (PPARγ) and CD36. Notably, PE-P activated hypothalamic autophagy more strongly than PC-O, with an increased ratio of microtubule-associated protein light chain 3 II/I (LC3II/I) and reduced p62/sequestosome-1 (p62) accumulation by rescuing the HFD-impaired autophagy-lysosome fusion. The phosphorylation of ULK1 mediated by Akt-mTOR and AMPK, was involved in ether-PLs activated autophagy. Furthermore, the enhanced hypothalamic autophagy promoted the production of α-melanocyte-stimulating hormone (α-MSH), which has been reported to maintain energy balance. It is concluded that ether-PLs ameliorated HFD-induced hypothalamic autophagy and ameliorated hepatic steatosis. Ether-PLs could thus be an attractive autophagy-enhancers against chronic HFD-induced obesity.

Soluble CD163 is produced by monocyte-derived and alveolar macrophages, and is not associated with the severity of idiopathic pulmonary fibrosis.

The soluble form of the membrane hemoglobin scavenger receptor CD163 (sCD163), released by shedding, is a strong marker for macrophage activation. Serum sCD163 levels rise in several acute inflammatory states and some fibrosing diseases. Monocyte-derived macrophages (MoDM) differentiated by macrophage colony-stimulating factor (M-MoDM) contribute to the pathophysiology of idiopathic pulmonary fibrosis (IPF), an irreversible and rapidly fatal interstitial lung disease. Since M-MoDM express high membrane CD163 levels, we thus postulated that sCD163 could be a relevant biomarker for macrophage activation in IPF. We found that M-MoDM constitutively released higher amounts of sCD163 (49.5 ± 24.5 ng/ml) than monocytes (0.45 ± 0.32 ng/ml) or MoDM differentiated with granulocyte macrophage-stimulating factor (2.24 ± 0.98 ng/ml). The basal production of sCD163 by M-MoDM was increased following stimulation with lipopolysaccharide (123.4 ± 54.9 ng/ml) or ATP (168.9 ± 41.8 ng/ml). The sCD163 release was controlled by metalloproteases but not through ADAM17 activation. Moreover, CD163-positive macrophages and sCD163 were detected in pulmonary tissues and alveolar fluids of Caucasian patients with IPF, respectively. IPF alveolar macrophages constitutively secreted sCD163 amounts (67.6 ± 44.6 ng/µg RNA) which were significantly higher than those released by alveolar macrophages isolated from controls (19.2 ± 7.6 ng/µg RNA) or patients with other interstitial lung disease (31.5 ± 16.6 ng/µg RNA). However, the concentrations of sCD163 in blood serum collected from 155 patients with IPF did not correlate with the severity of their disease. In conclusion, our results show that M-MoDM constituted a pertinent model to study the regulation of sCD163 production. Yet, serum sCD163 values could not provide a prognostic biomarker for IPF in our cohort.

The SC cell line as an in vitro model of human monocytes.

In vitro analysis of human macrophages is generally hampered by the necessity to differentiate them from peripheral blood monocytes. We have analyzed to which extent noncancerous SC monocytes could be used as an in vitro macrophage model. Macrophages differentiated from peripheral monocytes using standard CSF1 and CSF2 protocols for M2 and M1 precursors, respectively, were compared with THP-1-derived macrophages treated with PMA and with SC-derived macrophages differentiated either by CSF1, CSF2, or PMA according to different protocols. The optimal condition for generation of SC macrophages was treatment with PMA for 3 days, followed by 5-days culture without PMA and 24-h polarization with LPS/IFN-γ or IL-4/IL-13. Similar to THP-1, SC cells do not express the monocyte marker CD14 and differentiation to macrophages results neither in CD68 nor in CD14 expression, both of which were expressed by monocyte-derived macrophages. Similar to THP-1-macrophages, a proportion of SC macrophages can be polarized to the M1-like subtype that is characterized by higher expression of CD38, CD86, CD80, TNF-α, and IL-1ra, whereas treatment with IL4/IL13 did not lead to expression of the M2-associated receptors CD163, CD206, and only slightly increased the CD200R expression. Still, SC-M1 express much lower levels of the M1-associated markers compared with monocyte-derived M1 and no IL-1β. The data demonstrate that SC-derived macrophages differ from monocyte-derived macrophages in respect of their morphology, expression of important macrophage markers, phagocytosis. Yet, polarized SC-M1-like cells may with restrictions serve as a model for M1 macrophages, though this model does not provide significant advantages over already well-described THP-1-M1-like cells.

TRPM2 deficiency in mice protects against atherosclerosis by inhibiting TRPM2-CD36 inflammatory axis in macrophages.

Atherosclerosis is the major cause of ischemic heart disease and stroke, the leading causes of mortality worldwide. The central pathological features of atherosclerosis include macrophage infiltration and foam cell formation. However, the detailed mechanisms regulating these two processes remain unclear. Here we show that oxidative stress-activated Ca2+-permeable transient receptor potential melastatin 2 (TRPM2) plays a critical role in atherogenesis. Both global and macrophage-specific Trpm2 deletion protect Apoe−/− mice against atherosclerosis. Trpm2 deficiency reduces oxidized low-density lipoprotein (oxLDL) uptake by macrophages, thereby minimizing macrophage infiltration, foam cell formation and inflammatory responses. Activation of the oxLDL receptor CD36 induces TRPM2 activity, and vice versa. In cultured macrophages, TRPM2 is activated by CD36 ligands oxLDL and thrombospondin-1 (TSP1), and deleting Trpm2 or inhibiting TRPM2 activity suppresses the activation of CD36 signaling cascade induced by oxLDL and TSP1. Our findings establish the TRPM2-CD36 axis as a molecular mechanism underlying atherogenesis, and suggest TRPM2 as a potential therapeutic target for atherosclerosis.

Pregnane X Receptor Mediates Atherosclerosis Induced by Dicyclohexyl Phthalate in LDL Receptor-Deficient Mice.

Plastic-associated endocrine disrupting chemicals (EDCs) have been implicated in the etiology of cardiovascular disease (CVD) in humans, but the underlying mechanisms remain elusive. Dicyclohexyl phthalate (DCHP) is a widely used phthalate plasticizer; whether and how exposure to DCHP elicits adverse effects in vivo is mostly unknown. We previously reported that DCHP is a potent ligand of the pregnane X receptor (PXR) which acts as a xenobiotic sensor to regulate xenobiotic metabolism. PXR also functions in macrophages to regulate atherosclerosis development in animal models. In the current study, LDL receptor-deficient mice with myeloid-specific PXR deficiency (PXRΔMyeLDLR−/−) and their control littermates (PXRF/FLDLR−/−) were used to determine the impact of DCHP exposure on macrophage function and atherosclerosis. Chronic exposure to DCHP significantly increased atherosclerotic lesion area in the aortic root and brachiocephalic artery of PXRF/FLDLR−/− mice by 65% and 77%, respectively. By contrast, DCHP did not affect atherosclerosis development in PXRΔMyeLDLR−/− mice. Exposure to DCHP led to elevated expression of the scavenger receptor CD36 in macrophages and increased macrophage form cell formation in PXRF/FLDLR−/− mice. Our findings provide potential mechanisms underlying phthalate-associated CVD risk and will ultimately stimulate further investigations and mitigation of the adverse effects of plastic-associated EDCs on CVD risk in humans.

Plaque Evaluation by Ultrasound and Transcriptomics Reveals BCLAF1 as a Regulator of Smooth Muscle Cell Lipid Transdifferentiation in Atherosclerosis.

Understanding the processes behind carotid plaque instability is necessary to develop methods for identification of patients and lesions with stroke risk. Here, we investigated molecular signatures in human plaques stratified by echogenicity as assessed by duplex ultrasound. Lesion echogenicity was correlated to microarray gene expression profiles from carotid endarterectomies (n=96). The findings were extended into studies of human and mouse atherosclerotic lesions in situ, followed by functional investigations in vitro in human carotid smooth muscle cells (SMCs). Pathway analyses highlighted muscle differentiation, iron homeostasis, calcification, matrix organization, cell survival balance, and BCLAF1 (BCL2 [B-cell lymphoma 2]-associated transcription factor 1) as the most significant signatures. BCLAF1 was downregulated in echolucent plaques, positively correlated to proliferation and negatively to apoptosis. By immunohistochemistry, BCLAF1 was found in normal medial SMCs. It was repressed early during atherogenesis but reappeared in CD68+ cells in advanced plaques and interacted with BCL2 by proximity ligation assay. In cultured SMCs, BCLAF1 was induced by differentiation factors and mitogens and suppressed by macrophage-conditioned medium. BCLAF1 silencing led to downregulation of BCL2 and SMC markers, reduced proliferation, and increased apoptosis. Transdifferentiation of SMCs by oxLDL (oxidized low-denisty lipoprotein) was accompanied by upregulation of BCLAF1, CD36, and CD68, while oxLDL exposure with BCLAF1 silencing preserved MYH (myosin heavy chain) 11 expression and prevented transdifferentiation. BCLAF1 was associated with expression of cell differentiation, contractility, viability, and inflammatory genes, as well as the scavenger receptors CD36 and CD68. BCLAF1 expression in CD68+/BCL2+ cells of SMC origin was verified in plaques from MYH11 lineage-tracing atherosclerotic mice. Moreover, BCLAF1 downregulation associated with vulnerability parameters and cardiovascular risk in patients with carotid atherosclerosis. Plaque echogenicity correlated with enrichment of distinct molecular pathways and identified BCLAF1, previously not described in atherosclerosis, as the most significant gene. Functionally, BCLAF1 seems necessary for survival and transdifferentiation of SMCs into a macrophage-like phenotype. The role of BCLAF1 in plaque vulnerability should be further evaluated.

Pleiotropic Roles of Scavenger Receptors in Circadian Retinal Phagocytosis: A New Function for Lysosomal SR-B2/LIMP-2 at the RPE Cell Surface.

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.

CD36-Binding Amphiphilic Nanoparticles for Attenuation of Alpha Synuclein-Induced Microglial Activation.

Neuroinflammation is one of the hallmarks contributing to Parkinson's Disease (PD) pathology, where microglial activation occurs as one of the earliest events, triggered by extracellular alpha synuclein (aSYN) binding to the CD36 receptor. Here, CD36-binding nanoparticles (NPs) containing synthetic tartaric acid-based amphiphilic polymers (AMs) were rationally designed to inhibit this aSYN-CD36 binding. In silico docking revealed that four AMs with varying alkyl side chain lengths presented differential levels of CD36 binding affinity and that an optimal alkyl chain length would promote the strongest inhibitory activity towards aSYN-CD36 interactions. In vitro competitive binding assays indicated that the inhibitory activity of AM-based NPs plateaued at intermediate side chain lengths of 12- and 18-carbons, supporting the in silico docking predictions. These 12- and 18-carbon length AM NPs also had significantly stronger effects on reducing aSYN internalization and inhibiting the production of the proinflammatory molecules TNF-α and nitric oxide from aSYN-challenged microglia. All four NPs modulated the gene expression of aSYN-challenged microglia, downregulating the expression of the proinflammatory genes TNF, IL-6, and IL-1β, and upregulating the expression of the anti-inflammatory genes TGF-β and Arg1. Overall, this work represents a novel polymeric nanotechnology platform that can be used to modulate aSYN-induced microglial activation in PD.

Novel Secreted Peptides From Rhizopus arrhizus var. delemar With Immunomodulatory Effects That Enhance Fungal Pathogenesis.

Secreted fungal peptides are known to influence the interactions between the pathogen and host innate immunity. The aim of this study is to screen and evaluate secreted peptides from the fungus Rhizopus arrhizus var. delemar for their immunomodulatory activity. By using mass spectrometry and immuno-informatics analysis, we identified three secreted peptides CesT (S16), Colicin (S17), and Ca2+/calmodulin-dependent protein kinase/ligand (CAMK/CAMKL; S27). Culturing peripheral blood-derived monocytic macrophages (PBMMs) in the presence of S16 or S17 caused cell clumping, while culturing them with S27 resulted in the formation of spindle-shaped cells. S27-treated PBMMs showed cell cycle arrest at G0 phase and exhibited alternatively activated macrophage phenotype with pronounced reduction in scavenger receptors CD163 and CD206. Homology prediction indicated that IL-4/IL-13 is the immunomodulatory target of S27. Confirming this prediction, S27 initiated macrophage activation through phosphorylation of STAT-6; STAT-6 inhibition reversed the activity of S27 and reduced the formation of spindle-shaped PBMMs. Lastly, S27 treatment of PBMMs was associated with altered expression of key iron regulatory genes including hepcidin, ferroportin, transferrin receptor 1, and ferritin in a pattern consistent with increased cellular iron release; a condition known to enhance Rhizopus infection. Collectively, R. arrhizus var. delemar secretes peptides with immunomodulatory activities that support fungal pathogenesis. Targeting the IL-4/IL-13R/STAT-6 axis is a potential therapeutic approach to enhance the PBMM-mediated fungal phagocytosis. This represents a potential new approach to overcome lethal mucormycosis.

Mechanisms of selective monocyte targeting by liposomes functionalized with a cationic, arginine-rich lipopeptide

Stimulation of monocytes with immunomodulating agents can harness the immune system to treat a long range of diseases, including cancers, infections and autoimmune diseases. To this end we aimed to develop a monocyte-targeting delivery platform based on cationic liposomes, which can be utilized to deliver immunomodulators and thus induce monocyte-mediated immune responses while avoiding off-target side-effects. The cationic liposome design is based on functionalizing the liposomal membrane with a cholesterol-anchored tri-arginine peptide (TriArg). We demonstrate that TriArg liposomes can target monocytes with high specificity in both human and murine blood and that this targeting is dependent on the content of TriArg in the liposomal membrane. In addition, we show that the mechanism of selective monocyte targeting involves the CD14 co-receptor, and selectivity is compromised when the TriArg content is increased, resulting in complement-mediated off-target uptake in granulocytes. The presented mechanistic findings of uptake by peripheral blood leukocytes may guide the design of future drug delivery systems utilized for immunotherapy. Statement of significanceMonocytes are attractive targets for immunotherapies of cancers, infections and autoimmune diseases. Specific delivery of immunostimulatory drugs to monocytes is typically achieved using ligand-targeted drug delivery systems, but a simpler approach is to target monocytes using cationic liposomes. To achieve this, however, a deep understanding of the mechanisms governing the interactions of cationic liposomes with monocytes and other leukocytes is required. We here investigate these interactions using liposomes incorporating a cationic arginine-rich lipopeptide. We demonstrate that monocyte targeting can be achieved by fine-tuning the lipopeptide content in the liposomes. Additionally, we reveal that the CD14 receptor is involved in the targeting process, whereas the complement system is not. These mechanistic findings are critical for future design of monocyte-targeting liposomal therapies.

Soluble Trem2 is a negative regulator of erythrophagocytosis after intracerebral hemorrhage in a CD36 receptor recycling manner

IntroductionMicroglia and macrophages participate in hematoma clearance after intracerebral hemorrhage (ICH), thereby facilitating tissue restoration and neurological recovery. Triggering receptor expressed on myeloid cells 2 (Trem2) has been indicated as a major pathology-induced immune signaling hub on the microglial/macrophage surface. Soluble Trem2 (sTrem2), the proteolytic form of Trem2, is abundant in the body fluid and is positively correlated with the pathological process. ObjectivesIn the present study, we aimed to investigate the potential role of sTrem2 in hematoma resolution after ICH and to elucidate its underlying mechanisms. MethodsWe explored the biological functions of sTrem2 in the murine ICH brain by stereotaxic injection of recombinant sTrem2 protein or by adeno-associated virus-mediated expression. Erythrocyte phagocytosis was assessed using flow cytometry and immunofluorescence. Western blotting was performed to evaluate protein expression. Changes in behavior, sTrem2-induced down-stream pathway, and microglia were examined. ResultssTrem2 impedes hematoma resolution and impairs functional motor and sensory recovery. Interestingly, sTrem2 bypasses full-length Trem2, negatively regulating microglial/macrophage erythrophagocytosis, and promotes an inflammatory phenotype, which is associated with reduced retromer levels and impaired recycling of the pro-erythrophagocytic receptor CD36. Rescue of retromer Vps35 abolishes the phagocytosis-inhibiting effects and lysosome-dependent CD36 degradation caused by sTrem2. ConclusionThese findings indicate sTrem2 as a negative factor against microglia/macrophage-mediated hematoma and related neuronal damage clearance, provide insight into the mechanisms by which erythrophagocytosis is regulated and how it may be impaired after ICH, and suggest that the anti-proteolytic activity of Trem2 can be explored for ICH therapy.

Correction for Jeyaseelan et al., "Distinct Roles of Pattern Recognition Receptors CD14 and Toll-Like Receptor 4 in Acute Lung Injury".

Correction for Jeyaseelan et al., "Distinct Roles of Pattern Recognition Receptors CD14 and Toll-Like Receptor 4 in Acute Lung Injury".

Broad antiviral peptides against PRRSV based on novel linear epitopes on porcine CD163

PRRSV causes major economic losses to swine industry world-wide, which requires innovative antiviral agents. Porcine scavenger receptor CD163 has been identified as an essential fusion receptor for Porcine reproductive and respiratory Syndrome Virus (PRRSV) infection. In this study, novel antiviral peptides from pCD163 against PRRSV were developed based on broad neutralizing monoclonal antibodies. SRCR-5-9 of pCD163 from baculovirus efficiently binds to PRRSVs of lineage 8 and lineage 1, blocking infection in PAMs. A batch of monoclonal antibodies targeting SRCR-5-9 were generated and characterized. 8H2 and 4H7 block PRRSV infection by the disruption in viral attachment to PAMs. Virus titer reduced 100–1000 folds in average and the virus copy number decreased about 104 folds with these antibodies. Linear epitopes of 8H2 and 4H7 were individually localized in SRCR6 (1-30 aa) and PSTI(1-15aa) of pCD163. Mutations of SRCR6 NI1718KT and PST SS1314AA abolished the recognition of 8H2 and 4H7 to the corresponding region individually. Peptides derived from the linear epitopes displayed a broad inhibitory effect on PRRSVs of different lineages in a dose-dependent manner and further modulated PRRSV-related NF-κB pathway. In conclusion, these findings deepen the understanding in the interaction between PRRSV and pCD163 receptor and provide alternative universal antiviral strategies against PRRSV.

Lipopolysaccharide and ARDS caused by new coronavirus infection: hypotheses and facts

This review presents data from the literature that provide insight into the role of the lipopolysaccharide (LPS) of the Gram-negative bacteria in pathogenesis of acute respiratory distress syndrome (ARDS) caused by the novel SARS-CoV-2 coronavirus infection. ARDS is a syndrome of severe respiratory failure, an acutely occurring diffuse inflammatory lesion of lung tissue that develops as a nonspecific reaction to various direct (aspiration, inhalation of toxic gases), and systemic (sepsis, polytrauma) damaging factors and leading to development of acute respiratory failure (ARF), due to impaired structure of the lung parenchyma, disturbances in vascular permeability, decreased area of ventilated lung tissue. ARDS from coronavirus infection appears to have worse outcomes than ARDS from other causes. Mortality from typical ARDS at the intensive care units and hospitals is 35.3% and 40.0%, respectively, while the lethality rates for COVID-19-associated ARDS, ranged from 26% to 61.5%. Among patients who underwent artificial ventilation of the lungs, the mortality rates can range from 65.7% to 94%. Risk factors for poor outcomes include, e.g., older age, presence of concomitant diseases such as hypertension, cardiovascular disease and diabetes mellitus; decreased number of lymphocytes, kidney injury, and increased D-dimer levels. Death with ARDS in COVID-19 occurs as a result of respiratory failure (53%), respiratory failure combined with heart failure (33%), myocardial damage and circulatory failure (7%), or death from an unknown cause. A large number of studies show that bacterial LPS is directly or indirectly involved in all pathogenetic links of ARDS caused by the SARS-CoV-2 virus, i.e., worsening the course of inflammatory lung diseases due to decreased level of angiotensin-converting enzyme 2 (ACE2); increasing generation of reactive oxygen species (ROS) via NADPH oxidase and subsequent inactivation of endothelial nitric oxide synthase (eNOS) and decreasing bioavailability of endothelial NO, thus leading to endothelial dysfunction; interacting with proteins of surfactants. SP-A and SP-D, promoting early destruction of the cellular monolayers and lowering surface tension, interact with soluble CD14 receptor, which also has a pro-inflammatory effect on epithelial and endothelial cells, leading to p38MAPK activation via TLR4 receptors, causing degradation of IêBá protein and subsequent translocation of p65 NF-êB into the nucleus, thus inducing transcription of IL-6 and adhesion molecules (ICAM-1, VCAM-1 and E-selectin), and, as shown by Petruk et al. (2020), causing direct binding to the viral S protein in combination with LPS, thus enhancing activation of nuclear factor-kappa B (NF-êB) in monocytic THP-1 cells and cytokine responses in mononuclear blood cells. These pathophysiological mechanisms require further in-depth study in order to understand the nature of changes that occur in the patients with new SARS-CoV-2 infection.

Up-regulated serum levels of soluble CD25 and soluble CD163 in pediatric patients with SARS-CoV-2

Similar to hemophagocytic lymphohistiocytosis (HLH), some patients with SARS-CoV-2 have cytokine storm. Serum soluble interleukin-2 receptor (sCD25) and soluble CD163 (sCD163) are potential diagnostic biomarkers for HLH that help in guiding its treatment. This study was the first to investigate serum sCD25 and sCD163 levels in SARS-CoV-2. Serum sCD25 and sCD163 were measured by ELISA in 29 patients with SARS-CoV-2, aged between 2 months and 16 years (13 had COVID-19 and 16 had multisystem inflammatory syndrome in children (MIS-C)), in comparison to 30 age- and sex-matched healthy control children and 10 patients with HLH. Levels of these markers were re-measured in 21 patients with SARS-CoV-2 who were followed up 3 months after recovery. Patients with SARS-CoV-2 had significantly higher serum sCD25 and sCD163 than healthy control children (P < 0.001). SARS-CoV-2 patients had significantly higher sCD25 than patients with HLH (P < 0.05). Serum sCD25 was a good differentiating marker between patients with SARS-CoV-2 and HLH. Although there was a significant decrease of serum sCD25 and sCD163 of the 21 SARS-CoV-2 patients who were followed up, these levels were still significantly higher than the healthy controls levels (P < 0.001). Conclusion: Serum sCD25 and sCD163 levels were up-regulated in SARS-CoV-2 patients. Serum sCD25 was a good differentiating marker between SARS-CoV-2 and HLH. This initial report requires further studies, on large scales, to investigate the relationship between SARS-CoV-2 and both sCD25 and sCD163, including the disease severity and outcome. The therapeutic role of sCD25 and sCD163 antagonists should also be studied in SARS-CoV-2 patients.What is Known:• Similar to hemophagocytic lymphohistiocytosis (HLH), some patients with COVID-19 have cytokine storm due to excessive pro-inflammatory host response.• Serum soluble interleukin-2 receptor (sCD25) and soluble CD163 (sCD163) are potential diagnostic biomarkers for HLH. Monitoring of serum sCD25 and sCD163 levels can also help in guiding the treatment.What is New:• Serum sCD25 and sCD163 levels are up-regulated in patients with COVID-19, including patients presenting with multisystem inflammatory syndrome in children (MIS-C).• Serum sCD25 is a good differentiating marker between SARS-CoV-2 and HLH.