Scavenger Receptor Ligands Research Articles

SREC-I, a Type F Scavenger Receptor, Is an Endocytic Receptor for Calreticulin

Calreticulin and gp96 (GRP94) traffic associated peptides into the major histocompatibility complex class-I cross-presentation pathway of antigen-presenting cells (APCs). Efficient accession of the cross-presentation pathway requires APC receptor-mediated endocytosis of the chaperone/peptide complexes. Previously, scavenger receptor class-A (SRA) was shown to play a substantial role in trafficking gp96 and calreticulin into macrophages, accounting for half of total receptor-mediated uptake. However, the scavenger receptor ligand fucoidin competed the chaperone uptake beyond that accounted for by SRA, indicating that another scavenger receptor(s) may also contribute. Consistent with this hypothesis, we showed that the residual calreticulin uptake into SRA(-/-) macrophages is competed by the scavenger receptor ligand acetylated low density lipoprotein (LDL). We now report that an additional scavenger receptor, SREC-I (scavenger receptor expressed by endothelial cell-I), mediates the endocytosis of calreticulin and gp96. Ectopic expression of SREC-I in Chinese hamster ovary cells yielded chaperone recognition and uptake, and these processes were competed by the inhibitory ligands fucoidin and acetylated (Ac)LDL. Although AcLDL competes for the chaperone interactions with SRA and SREC, we showed that not all of the scavenger receptors, which bind AcLDL, bind calreticulin or gp96. The overexpression of SREC-I in macrophages increased chaperone endocytosis, indicating that SREC-I functions in APCs and that the cytosolic components necessary for the endocytosis of SREC-I and its cargo are present and not limiting in APCs. These data identify a novel class of ligands for SREC-I and provide insight into the mechanisms by which APCs and potentially endothelial cells traffic chaperone/antigen complexes.

Stabilin-1 and stabilin-2 are both directed into the early endocytic pathway in hepatic sinusoidal endothelium via interactions with clathrin/AP-2, independent of ligand binding

Liver sinusoidal endothelial cells (LSECs) mediate clearance of hyaluronan (HA) and scavenger receptor ligands, for example, advanced glycation end product (AGE)-modified proteins and oxidized lipids from the circulation. We recently cloned stabilin-1 and -2, two members of a novel family of transmembrane proteins expressed in LSECs. By using primary LSECs and HEK293 cells separately expressing either stabilin, we have investigated their roles in the early events of endocytosis with respect to localization, ligand-binding properties, and associations with clathrin and adaptor protein (AP)-2. Both stabilins were present at the cell surface, although surface levels of stabilin-1 were limited. In addition, stabilins were present in early endosomal antigen (EEA)-1+ organelles colocalizing with endocytosed AGE-modified bovine serum albumin (BSA). Treating cells with monensin further pronounced this distribution. Recombinant stabilin-2, but not recombinant stabilin-1, bound HA and the scavenger receptor ligands AGE-modified BSA, formaldehyde-treated BSA, and collagen N-terminal propeptides. In LSECs, both stabilins were associated with clathrin and AP-2, but not with each other. These interactions did not change upon addition of exogenous HA, suggesting that stabilins are constitutively internalized. In conclusion, hepatic stabilins are both present in the early endocytic pathway, associating with clathrin/AP-2, but whereas stabilin-2 has a clear scavenging profile, stabilin-1 does not recognize these ligands.

Nobiletin, a citrus flavonoid isolated from tangerines, selectively inhibits class A scavenger receptor-mediated metabolism of acetylated LDL by mouse macrophages

Flavonoids are a class of chemically related polyphenols that are nearly ubiquitous in nature. Of the more-than 4000 flavonoids thus identified, citrus fruit-derived flavonoids are suggested to have an inverse association with the occurrence of coronary heart disease via their ability to reduce plasma cholesterol concentrations. Our current studies examined whether citrus flavonoids possess an additional antiatherogenic effect by modulating macrophage metabolism of the specific class A scavenger receptor (SR-A) ligand, acetylated LDL (acLDL). In this study, both acLDL-metabolism and SR-A expression by cultured murine J774A.1 macrophages was examined following 24 h pretreatment (100 μM) with the flavonoids: naringenin (from grapefruit), hesperetin (from oranges), and tangeretin and nobiletin (from tangerines). Of these flavonoids, only nobiletin inhibited (50–72%) acLDL metabolism as measured by both cellular cholesterol ester mass and [ 3H]oleate incorporation into cholesterol esters. This nobiletin-mediated effect was specific for SR-A and not a global effect on lipoprotein metabolism by the macrophage, as all four citrus flavonoids significantly reduce the metabolism of beta-VLDL, which is primarily taken up by macrophages via the LDL receptor. Nevertheless, nobiletin did not affect SR-A protein expression, as measured by Western blot analysis, nor was cell surface expression of SR-A affected as measured by 4 °C binding studies using [ 125I]acLDL. In conclusion, our findings suggest that in addition to reducing plasma cholesterol concentrations, nobiletin may prevent atherosclerosis at the level of the vascular wall by inhibiting macrophage foam-cell formation.

Receptor mediated endocytosis of retinal pigment epithelial cell

To evaluate the possibility and mechanism of the turnover of the negatively charged macromolecule by retinal pigment epithelial (RPE) cells. Cultured porcine RPE eye cups were incubated with fluorescence labeled formaldehyde treated serum albumin (F-FSA), a classical ligand for scavenger receptors. The endocytosis of F-FSA by RPE cells under different conditions was evaluated systematically by fluorescent microscopy and electron microscopy. The amount of F-FSA ingested by RPE cells depends on the incubation time and the concentration of ligand. Similar ligands compete for the binding site of RPE cells. Small vesicles containing F-FSA are noticed in the cytosol of RPE cells as demonstrated by electron microscopy. The RPE cells can effectively ingest negatively charged macromolecules, possibly by scavenger receptor mediated endocytosis.

Chemokines Generally Exhibit Scavenger Receptor Activity through Their Receptor-binding Domain

Chemokines are a family of cytokines that induce directed migration of various types of leukocytes through specific interactions with a group of seven transmembrane receptors. Scavenger receptors are a heterogenous family of transmembrane molecules that commonly bind and uptake oxidized low density lipoprotein and bacteria. Here, we show that not only CXC chemokine 16 (CXCL16)/SR-PSOX, a transmembrane chemokine with scavenger receptor activity, but also 12 out of 15 chemokines examined efficiently bound scavenger receptor ligands in competition with cells expressing their specific chemokine receptors. Furthermore both the chemotactic and scavenger receptor activities of SR-PSOX/CXCL16 were similarly impaired in a series of mutants altered in the chemokine domain, indicating that SR-PSOX/CXCL16 binds scavenger receptor ligands as well as CXCR6 using highly overlapping binding motifs. Taken together, chemokines generally have scavenger receptor-like activity through their receptor-binding domain, suggesting a close evolutionary relationship between chemokines and scavenger receptors.

Macrophage-Targeted Photodynamic Therapy

Photodynamic therapy (PDT) uses light activatable molecules that after illumination produce reactive oxygen species and unwanted tissue destruction. PDT has dual selectivity due to control of light delivery and to some extent selective photosensitizer (PS) accumulation in tumors or other diseased tissue, additional targeted selectivity of PS for disease is necessary. The delivery of drugs to selected lesions can be enhanced by the preparation of targeted macromolecular conjugates that employ cell type specific targeting by ligand-receptor recognition. Macrophages and monocytes express a scavenger-receptor that is a high-capacity route for delivering molecules into endocytic compartments in a cell-type specific manner. We have shown that by attaching PS to scavenger-receptor ligands it is possible to get three logs of selective cell killing in macrophages while leaving non-macrophage cells unharmed. The capability to selectively kill macrophages has applications in treating cancer and in the detection and therapy of vulnerable atherosclerotic plaque and possibly for autoimmune disease and some infections.

Induction of intracellular cytokine production in human monocytes/macrophages stimulated with ligands of pattern recognition receptors.

This study addressed the role of the pattern recognition receptors (PRR), which recognize different molecular structures present on microorganisms, apoptotic, senescent and tumor cells, in the stimulation of human monocyte and monocyte-derived macrophages (MDM) for the production of intracellular cytokines. Monocytes and MDM were stimulated with different ligands of scavenger receptors (SR) and mannose receptor (MR). Production of intracellular cytokines: tumor necrosis factor alpha (TNF alpha), interleukin 10 and 12 (IL-10, IL-12) was determined by flow cytometry following staining with anti-cytokine monoclonal antibodies (mAbs). The ligands of SR type A: fucoidan, polyguanylic acid (polyG), chemically modified low density lipoproteins (LDL), ligands of SR-B: native and chemically modified LDL, and ligand of mannose receptor (MR)-mannan induced strong expression of intracellular TNF alpha and weaker IL-10 in monocytes, while phosphatidylserine (PdS) was without effect. IL-12 was stimulated only by fucoidan and polyG. The induction of cytokine m-RNA generally followed the pattern and the magnitude of intracellular cytokine production. In MDM, intracellular TNF alpha and IL-12 expression was induced by mannan, native and modified LDL, but not other ligands. Expression of IL-10 was less pronounced and occurred following stimulation with fucoidan, polyG and modified, but not native, LDL. These results suggest that some PRR ligands may be involved in activation of monocytes/MDM for the production of mainly proinflammatory cytokines (TNF alpha, IL-12) implicating their role in the response to microbial and tumor invasion.

Macrophage endocytosis of superparamagnetic iron oxide nanoparticles: mechanisms and comparison of ferumoxides and ferumoxtran-10.

Superparamagnetic iron oxides (SPIO) used as magnetic resonance (MR) contrast agents undergo specific uptake by macrophages. The purpose of this study was first to determine the mechanism of macrophage uptake for Ferumoxides by using competition experiments with specific ligands of scavenger receptors SR-A (I/II) and second, to evaluate and compare the internalization of 2 different contrast agents, Ferumoxides (SPIO) and Ferumoxtran-10 (USPIO: ultrasmall superparamagnetic iron oxide) using macrophages obtained by chemical activation of human monocytic cells. Ferumoxides and Ferumoxtran-10 are 2 MR contrast agents, composed of dextran-coated iron oxide nanoparticles. The endocytosis pathway of Ferumoxides was studied using competition experiments on mouse peritoneal macrophages in the presence of specific ligands of scavenger receptors SR-A (types I and II): polyinosinic acid and fucoidan. In vitro assays using THP-1 (human promonocyte) cells activated into macrophages were performed in the presence of the 2 superparamagnetic nanoparticles. The cellular uptake was determined by measuring the iron content using ICP-AES (inductively coupled plasma-atomic emission spectrometry) and by Prussian blue staining. In the presence of polyinosinic acid or fucoidan, the endocytosis of Ferumoxides by mouse peritoneal macrophages was inhibited. This inhibition was obtained using 10 microg/mL of scavenger receptor ligands at a concentration of 62.5 microg Fe/mL of SPIO, and a dose-dependent relationship was observed. Without competitors, the percentage of uptake of Ferumoxides by mouse peritoneal macrophages ranged between 3 and 8%. On the human activated monocyte THP-1 cell assay, Ferumoxides underwent a higher macrophage uptake (between 1.1 and 3%) compared with Ferumoxtran-10 (between 0.03 and 0.12%). This difference is attributed to the larger size of Ferumoxides nanoparticles. Competition experiments indicate that the cellular uptake of Ferumoxides involves scavenger receptor SR-A-mediated endocytosis. The comparison between Ferumoxides and Ferumoxtran-10 confirms that macrophage uptake of iron oxide nanoparticles depends mainly on the size of these contrast agents.

Internalization of OspA in rsCD14 complex and aggregated forms

Although the spirochetal protein OspA is capable of stimulating immune cells in a CD14- and TLR2-dependent manner, little is known about how TLR2 receptor complex ligands, such as OspA, are handled by the cell once delivered. We examine here the internalization of the fluorescently derivatized forms of both the full length OspA lipoprotein delivered as a recombinant soluble CD14 (rsCD14) complex and the corresponding lipohexapeptide given to the cells as an aggregate. Both forms of OspA are internalized in a similar manner to acetylated low density lipoprotein (AcLDL), a scavenger receptor ligand. Acetylated low density lipoprotein is capable of competing for internalization with OspA even when OspA is delivered as a rsCD14 complex. We observe co-localization of OspA with lysosomes but not with the Golgi complex. These phenomena are similar between RAW264.7 macrophages and endothelial cells but change drastically when the cells are deprived of serum. Upon serum starvation, OspA shows some localization to the Golgi apparatus whereas the lipohexapeptide remains on the cell surface. Inhibition of internalization of OspA via treatment with cytochalasin D or of the lipohexapeptide via serum starvation does not interfere with TNF induction activity, consistent with signalling from the cell surface.

Cutting edge: SR-PSOX/CXC chemokine ligand 16 mediates bacterial phagocytosis by APCs through its chemokine domain.

SR-PSOX and CXC chemokine ligand (CXCL)16, which were originally identified as a scavenger receptor and a transmembrane-type chemokine, respectively, are indicated to be identical. In this study, we demonstrate that membrane-bound SR-PSOX/CXCL16 mediates adhesion and phagocytosis of both Gram-negative and Gram-positive bacteria. Importantly, our prepared anti-SR-PSOX mAb, which suppressed chemotactic activity of SR-PSOX, significantly inhibited bacterial phagocytosis by human APCs including dendritic cells. Various scavenger receptor ligands inhibited the bacterial phagocytosis of SR-PSOX. In addition, the recognition specificity for bacteria was determined by only the chemokine domain of SR-PSOX/CXCL16. Thus, SR-PSOX/CXCL16 may play an important role in facilitating uptake of various pathogens and chemotaxis of T and NKT cells by APCs through its chemokine domain.

T cell proliferative responses to malondialdehyde–acetaldehyde haptenated protein are scavenger receptor mediated

Malondialdehyde–acetaldehyde (MAA) haptenated proteins have been described in disease processes related to prolonged oxidative stress (via malondialdehyde production), such as alcohol liver disease (ALD), non-alcoholic non-steatohepatitis (NASH) and atherosclerosis. Experimentally, high titer IgG1 antibody responses are seen after immunization without adjuvant; however, T cell proliferative responses and the role of scavenger receptors in this immunogenicity has not previously been described. In this study, T cell proliferative responses to the carrier protein, but not the MAA hapten itself, were identified in vitro. Moreover, these T proliferative responses were inhibited when MAA-hen egg lysozyme (HEL) was co-immunized with excess scavenger receptor ligand polyG (poly-guanylic acid), implicating the role of (a) scavenger receptor(s) in initiating the T helper cell response. Activated B cells were unable to process and present MAA-HEL preferentially to T cells, while thioglycollate-elicited (but not Con A-elicited) macrophages and dendritic cells (DC) did so with ∼32-fold less MAA-HEL than native antigen necessary to initiate equal proliferative responses. While this preferential processing and presentation may be related to several factors, preferential binding of MAA haptenated proteins mediated by scavenger receptors may be one mechanism. IL-4 was absent from the supernatants of T proliferative assays despite a strong IgG1 response in vivo, although the TH2 cytokines IL-6 and IL-10 were expressed. Since the modification of proteins by the MAA have previously been shown to occur after ethanol consumption in vivo, the ability of MAA haptens to experimentally enhance immune responses, specifically humoral and T cell responses, may represent mechanisms by which autoimmune phenomena found in ALD occur.

Identification of a scavenger receptor homologue on nonspecific cytotoxic cells and evidence for binding to oligodeoxyguanosine

In mammals scavenger receptors (SR) are expressed by monocytic-macrophage lineage cells and B-cells. Studies of various teleost species have indirectly demonstrated the presence of SR receptors on phagocytic or endothelial cells by showing the uptake of SR ligands (i.e. derivatised (acetylated) lipoproteins) by these cells. In the present study, nonspecific cytotoxic cells (NCC) were examined for membrane expression of an SR-like protein. Approximately 15–25% of purified NCC expressed scavenger receptor class A (SR-A) demonstrated by binding by a monoclonal (2F8) specific for mouse SR-A (types I, II). Flow cytometric analysis determined that SR binding cells had the same size and ‘side scatter’ characteristics as NCC. Two colour flow analysis of NCC demonstrated that only a subset of NCC expressed the SR-A-like protein and non-NCC were SR-A negative. Membrane expression of SR on NCC was confirmed by fluorescence microscopy. Analysis of the tissue distribution of SR bearing cells demonstrated that in both catfish and tilapia, SR-A was expressed by NCC in the peripheral blood, spleen and anterior kidney. Experiments were also done to determine if the ligands known to bind mammalian SR-A had a similar specificity for the teleost receptor. Cold competition binding experiments determined that anti-SR-A antibody competed with and reduced biotinylated polyguanosine 20-mer binding to NCC by approximately 40%. Two other types of ligands known to bind (mammalian) SR-A (i.e. polyvinyl sulphate and dextran sulphate) likewise decreased anti-SR-A antibody binding to NCC by 40%. These studies for the first time demonstrated that NCC express the teleost orthologue of mammalian SR-A, suggesting that NCC may participate in physiologic regulation of lipid metabolism in addition to functions of innate immunity.

A role for class A scavenger receptor in dendritic cell nibbling from live cells.

Monocyte-derived dendritic cells (DC) possess the unique capacity to capture Ag from live cells through intimate cell contact, a process referred to as nibbling. We sought to define the receptor(s) mediating DC nibbling. Uptake of fluorescently labeled plasma membrane from live cells by DC was inhibited by protease treatment and by a panel of polyanionic ligands, implicating scavenger receptors (SR) in this process. Differential expression of SR on DC and macrophages correlated with the capacity to acquire membrane from live cells. Internalized membrane colocalized with SR ligand and entered the endosomal pathway. DC very efficiently acquired and internalized gp100 tumor Ag expressed at the surface of viable adenocarcinoma cells via recombinant adenoviral infection. Cross-presentation of gp100 by DC to MHC class I-restricted T cells was inhibited by polyanionic SR ligand and an Ab to type A SR (SR-A), whereas Ab to the class B SR CD36, which mediates uptake of apoptotic cells, induced no inhibition. DC capture of fluorescently labeled membrane from live cells was partially blocked by SR-A-specific Ab, suggesting that other SR may also be contributing to nibbling. DC maturation resulted in a switch in expression from type II SR-A (SR-AII) to the SR-AI splice variant. Finally, SR-A was identified on interdigitating DC isolated from monkey lymph nodes. These findings define a novel role for SR-A, and suggest that Ag uptake from live cells by DC may be important in the generation of immunity and in the maintenance of peripheral tolerance in vivo.

ケモカイン活性とスカベンジャー受容体活性を有するＳＲ‐ＰＳＯＸ／ＣＸＣＬ１６の自然免疫と獲得免疫における役割解析

SR-PSOX and CXCL16, which were originally identified as a scavenger receptor and a transmembrane-type chemokine, respectively, are strikingly proved to be essentially identical. Here, we demonstrate that, while soluble SR-PSOX/CXCL16 is chemotactic for CXCR6-expressing cells, membrane-bound SR-PSOX/CXCL16 mediates adhesion and phagocytosis of both Gram-negative and Gram-positive bacteria. Importantly, our prepared anti- SR/PSOX monoclonal antibody, which suppressed chemotactic activity of SR-PSOX, significantly inhibited bacterial phagocytosis by antigen-presenting cells including dendritic cells. Various scavenger receptor ligands inhibited not only scavenger receptor activity but also chemotactic activities of SR-PSOX. In addition, both activities were similarly impaired in a series of mutants in which one basic amino acid in the chemokine domain of SRPSOX was replaced by alanine. Thus, SR-PSOX/CXCL16 was shown to play a role in both innate and adaptive immunity by mediating phagocytosis of bacteria as well as recruitment of CXCR6-expressing T cells by antigenpresenting cells through the same chemokine domain.

Oxidative tyrosylation of high density lipoproteins impairs cholesterol efflux from mouse J774 macrophages: role of scavenger receptors, classes A and B.

Studies were designed to test whether tyrosylation of high-density lipoprotein (HDL(T)) modifies its metabolic features. HDL(T) was less effective than native HDL in promoting cholesterol efflux from J774-AI macrophages. Cell association with fluorescent HDL(T)-apolipoprotein and the uptake of HDL(T)-[(3)H]cholesteryl hexadecyl ether were enhanced by 50% in comparison with native HDL. In addition, neutral cholesterol ester hydrolase (nCEH) activity in J774-AI, which controls the hydrolysis of cholesteryl ester stores to provide free cholesterol for cellular release, declined in the presence of HDL(T). In vitro displacement experiments revealed the ability of HDL(T) to compete with oxidized and acetylated LDL, known as ligands of scavenger receptor (SR) class B type I/II. Similarly, treatment with a blocking antibody to SR-BI/II reduced the cell association of HDL(T) and native HDL by 50%. The addition of polyinosinic acid, an inhibitor of SR class A, reduced the cell association of HDL(T) without affecting that of native HDL. These findings provide evidence that HDL(T) can compete with modified LDL, bind SR-BI/BII and internalize cholesterol ester. Furthermore, the impaired capacity of HDL(T) in promoting cholesterol efflux from J774-AI was accompanied by diminished nCEH and enhanced recognition by SR-AI/II, which appears to involve the transport of cholesterol into cells.

Advanced glycation end products (AGEs) lower the clearance function of hepatic scavenger endothelial cells (SEC)

Pre-incubation of cultured scavenger endothelial cells (SEC) with the scavenger receptor (SR) ligand advanced glycation end product (AGE)-modified bovine serum albumin (AGE-BSA) for 3 h reduced subsequent endocytosis of radioiodinated SR ligands AGE-BSA, formaldehyde-treated BSA (FSA) and 125I-oxidatively modified low-density lipoprotein (OxLDL) by 51%, 58% and 32%, respectively. Endocytosis of non-SR ligands, radioiodinated collagen, mannan or heat-aggregated γ-globulin was not affected by preexposure of SEC to AGE-BSA. Pre-incubation with FSA and other ligands different than AGE-BSA did not affect subsequent endocytosis of ligands for any receptor. Subcellular fractionation showed that pre-incubation with AGE-BSA delayed the transport of SR ligands along the endocytic pathway. Quantitative immuno-EM suggested that endocytosis of AGE-BSA in SEC lowered the amount of a newly described SR for AGE-BSA. Endocytic activity reduced by pre-incubation with AGE-BSA could be restored by incubating the cells for 6 h in AGE-free medium. Receptor restoration was inhibited by the protein synthesis inhibitor cycloheximide. We conclude from these findings that endocytosis of AGE-BSA leads to a loss of SR, and delayed intracellular transport in SEC. De novo synthesis of the receptor is required to restore normal endocytic activity.

SREC-II, a New Member of the Scavenger Receptor Type F Family, Trans-interacts with SREC-I through Its Extracellular Domain

The scavenger receptor expressed by endothelial cells (SREC) with an extremely large cytoplasmic domain, was originally identified in a human endothelial cell line. In this study, we have cloned a second isoform named SREC-II and shown that there is a heterophilic interaction between SREC-I and -II at their extracellular domains. The cDNA for murine SREC-II encodes an 834-amino acid protein with 35% homology to SREC-I. Similar to SREC-I, SREC-II contains multiple epidermal growth factor-like repeats in its extracellular domain. However, in contrast to SREC-I, SREC-II had little activity to internalize modified low density lipoproteins (LDL). A Northern blot analysis revealed a tissue expression pattern of SREC-II similar to that of SREC-I with predominant expression in human heart, lung, ovary, and placenta. Mouse fibroblast L cells with no tendency to associate showed noticeable aggregation when SREC-I was overexpressed in these cells, whereas overexpression of SREC-II caused only slight aggregation. Remarkably, intense aggregation was observed when SREC-I-expressing cells were mixed with those expressing SREC-II. Deletion of almost all of the cytoplasmic receptor domain had no effect on the receptor expression and cell aggregation, indicating that solely the extracellular domain is involved in cell aggregation. The association of SREC-I and -II was effectively suppressed by the presence of scavenger receptor ligands such as acetylated LDL and oxidized LDL. These findings suggest that SREC-I and -II show weak cell-cell interaction by their extracellular domains (termed homophilic trans-interaction) but display strong heterophilic trans-interaction through the extracellular epidermal growth factor-like repeat domains.

Advanced glycation end products impair the scavenger function of rat hepatic sinusoidal endothelial cells.

We have previously reported that advanced glycation end products are eliminated from the circulation mainly by scavenger receptor-mediated uptake in hepatic sinusoidal endothelial cells. Our experiments showed that the degradation of AGE-modified protein after endocytosis in hepatic sinusoidal endothelial cells occurs slowly compared with that of other scavenger receptor ligands. The aim of this study was to investigate further the mechanism whereby AGE-modified protein affects the important scavenger function of hepatic sinusoidal endothelial cells. Primary cultures of hepatic sinusoidal endothelial cells were pre-incubated with unlabelled ligand, unbound ligand was washed off, and the endocytic capacity was measured by addition of radiolabelled ligand, and immune electron microscopy. Pre-incubation with unlabelled AGE-modified bovine serum albumin reduced subsequent endocytosis of radiolabelled scavenger receptor ligands AGE-modified bovine serum albumin, formaldehyde-treated serum albumin, oxidized low density lipoprotein and acetylated low density lipoprotein by 50, 56, 32 and 20%, respectively. Non-scavenger receptor-mediated endocytosis was not affected by pre-exposure to AGE-modified protein. Pre-incubation with a number of non-AGE-ligands did not affect subsequent endocytosis via any of the major endocytosis receptors in hepatic sinusoidal endothelial cells. Incubation in fresh medium for 6 h after pre-exposure to AGE-modified protein almost completely restored normal scavenger receptor-mediated endocytic activity. Quantitative immune electron microscopy showed that the amount of a newly described scavenger receptor for AGE-modified protein is reduced after pre-incubation with AGE-modified protein. Subcellular fractionation showed that pre-incubation with AGE-modified protein delays intracellular transport of scavenger receptor ligands. Endocytosis of AGE-modified protein leads to loss of scavenger receptors and delayed intracellular transport in hepatic sinusoidal endothelial cells.

Metabolism of oxidized and chemically modified low density lipoproteins in rainbow trout—clearance via scavenger receptors

Oxidative modifications of low density lipoprotein (LDL) convert LDL into a ligand recognized by a variety of scavenger receptors (SR) in mammals. This oxidized LDL (oxLDL) activate several cell types, and have been shown to induce expression of a variety of genes in mammals. Lipoproteins of poikilothermic animals like salmonid fishes contain high levels of polyunsaturated fatty acids susceptible to oxidative modifications. We have investigated, and found trout LDL to be susceptible to oxidation in the presence of Cu 2+. When oxidized or acetylated trout LDL was injected intravenously, the clearance rate was increased compared to that of native LDL. Modified LDL was taken up almost exclusively in the kidney, whereas native LDL was also taken up in the liver. Uptake of both oxLDL and acetylated LDL in the kidney was significantly inhibited by lipoteichoic acid (LTA) and formaldehyde treated BSA (fBSA), both of which are known ligands of SR.

Scavenger receptor-specific allergen delivery elicits IFN-γ-dominated immunity and directs established TH2-dominated responses to a nonallergic phenotype

Background: Immunotherapeutic approaches to allergy consist of reliably changing allergen-specific TH2 immunity associated with secretion of IL-4, IL-5, and IL-13, along with IgE antibodies in atopic individuals to TH1 immunity. Our earlier data show that targeting of protein antigens to antigen-presenting cells (APCs), such as macrophages, by means of scavenger receptors (SRs) results in a pronounced TH1 immunity. Here we demonstrate a novel experimental approach for the conversion of TH2 immunity to TH1 immunity by using SR delivery of allergens. Objectives: We sought to show that targeting of allergens by means of SRs to APCs triggers TH1 immunity and that an established TH2 immunity to the Der p 1-immunodominant peptide 111-139 (p1, 111-139) can be modulated to a nonallergic TH1 phenotype. Methods: Analysis of the T cell-derived cytokines IL-4, IL-5, IL-13, and IFN-γ in response to p1, 111-139 in C57BL/6 mice 7 to 42 days after immunization, measurement of specific antibody responses, eosinophilic infiltrate, and skin hypersensitivity in response to allergen challenge constitute the parameters of in vivo immunity. Results: We show that p1, 111-139, when delivered to APCs by means of SR, elicits a TH1-dominant immunity. If it is delivered to APCs either after chemical coupling to SR ligands or by means of mere coadsorption on alum in the presence of an SR ligand, the established TH2 immunity can be modified to TH1 immunity. Conclusions: SR-mediated delivery of allergens has immunotherapeutic potential that may be usable in atopic individuals. (J Allergy Clin Immunol 2002;109:321-8.)