Role of scavenger receptors in the pathophysiology of chronic liver diseases.

Scavenger receptors comprise a large family of structurally diverse proteins that are involved in many homeostatic functions. They recognize a wide range of ligands, from pathogen-associated molecular patterns (PAMPs) to endogenous, as well as modified host-derived molecules (DAMPs). The liver deals with blood micro-organisms and DAMPs released from injured organs, thus performing vital metabolic and clearance functions that require the uptake of nutrients and toxins. Many liver cell types, including hepatocytes and Kupffer cells, express scavenger receptors that play key roles in hepatitis C virus entry, lipid uptake, and macrophage activation, among others. Chronic liver disease causes high morbidity and mortality worldwide. Hepatitis virus infection, alcohol abuse, and non-alcoholic fatty liver are the main etiologies associated with this disease. In this context, continuous inflammation as a result of liver damage leads to hepatic fibrosis, which frequently brings about cirrhosis and ultimately hepatocellular carcinoma. In this review, we will summarize the role of scavenger receptors in the pathophysiology of chronic liver diseases. We will also emphasize their potential as biomarkers of advanced liver disease, including cirrhosis and cancer.

Interleukin-23 receptor expressing γδ T cells locally promote early atherosclerotic lesion formation and plaque necrosis in mice.

AIMS: Atherosclerosis is a chronic inflammatory disease of the vessel wall controlled by local and systemic immune responses. The role of interleukin-23 receptor (IL-23R), expressed in adaptive immune cells (mainly T-helper 17 cells) and γδ T cells, in atherosclerosis is only incompletely understood. Here, we investigated the vascular cell types expressing IL-23R and addressed the function of IL-23R and γδ T cells in atherosclerosis. METHODS AND RESULTS: IL-23R+ cells were frequently found in the aortic root in contrast to the aorta in low-density lipoprotein receptor deficient IL-23R reporter mice (Ldlr-/-Il23rgfp/+), and mostly identified as γδ T cells that express IL-17 and GM-CSF. scRNA-seq confirmed γδ T cells as the main cell type expressing Il23r and Il17a in the aorta. Ldlr-/-Il23rgfp/gfp mice deficient in IL-23R showed a loss of IL-23R+ cells in the vasculature, and had reduced atherosclerotic lesion formation in the aortic root compared to Ldlr-/- controls after 6 weeks of high-fat diet feeding. In contrast, Ldlr-/-Tcrδ-/- mice lacking all γδ T cells displayed unaltered early atherosclerotic lesion formation compared to Ldlr-/- mice. In both HFD-fed Ldlr-/-Il23rgfp/gfp and Ldlr-/-Tcrδ-/- mice a reduction in the plaque necrotic core area was noted as well as an expansion of splenic regulatory T cells. In vitro, exposure of bone marrow-derived macrophages to both IL-17A and GM-CSF induced cell necrosis, and necroptotic RIP3K and MLKL expression, as well as inflammatory mediators. CONCLUSIONS: IL-23R+ γδ T cells are predominantly found in the aortic root rather than the aorta and promote early atherosclerotic lesion formation, plaque necrosis, and inflammation at this site. Targeting IL-23R may thus be explored as a therapeutic approach to mitigate atherosclerotic lesion development.

CD5L Promotes M2 Macrophage Polarization through Autophagy-Mediated Upregulation of ID3.

CD5L (CD5 molecule-like) is a secreted glycoprotein that controls key mechanisms in inflammatory responses, with involvement in processes such as infection, atherosclerosis, and cancer. In macrophages, CD5L promotes an anti-inflammatory cytokine profile in response to TLR activation. In the present study, we questioned whether CD5L is able to influence human macrophage plasticity, and drive its polarization toward any specific phenotype. We compared CD5L-induced phenotypic and functional changes to those caused by IFN/LPS, IL4, and IL10 in human monocytes. Phenotypic markers were quantified by RT-qPCR and flow cytometry, and a mathematical algorithm was built for their analysis. Moreover, we compared ROS production, phagocytic capacity, and inflammatory responses to LPS. CD5L drove cells toward a polarization similar to that induced by IL10. Furthermore, IL10- and CD5L-treated macrophages showed increased LC3-II content and colocalization with acidic compartments, thereby pointing to the enhancement of autophagy-dependent processes. Accordingly, siRNA targeting ATG7 in THP1 cells blocked CD5L-induced CD163 and Mer tyrosine kinase mRNA and efferocytosis. In these cells, gene expression profiling and validation indicated the upregulation of the transcription factor ID3 by CD5L through ATG7. In agreement, ID3 silencing reversed polarization by CD5L. Our data point to a significant contribution of CD5L-mediated autophagy to the induction of ID3 and provide the first evidence that CD5L drives macrophage polarization.

Deletion of Batf3-dependent antigen-presenting cells does not affect atherosclerotic lesion formation in mice.

Atherosclerosis is the main underlying cause for cardiovascular events such as myocardial infarction and stroke and its development might be influenced by immune cells. Dendritic cells (DCs) bridge innate and adaptive immune responses by presenting antigens to T cells and releasing a variety of cytokines. Several subsets of DCs can be discriminated that engage specific transcriptional pathways for their development. Basic leucine zipper transcription factor ATF-like 3 (Batf3) is required for the development of classical CD8α+ and CD103+ DCs. By crossing mice deficient in Batf3 with atherosclerosis-prone low density lipoprotein receptor (Ldlr-/-)-deficient mice we here aimed to further address the contribution of Batf3-dependent CD8α+ and CD103+ antigen-presenting cells to atherosclerosis. We demonstrate that deficiency in Batf3 entailed mild effects on the immune response in the spleen but did not alter atherosclerotic lesion formation in the aorta or aortic root, nor affected plaque phenotype in low density lipoprotein receptor-deficient mice fed a high fat diet. We thus provide evidence that Batf3-dependent antigen-presenting cells do not have a prominent role in atherosclerosis.

Relevance of MICA antibodies in acute humoral rejection in renal transplant patients.

The presence of MICA antibodies was examined in eleven patients diagnosed with AHR. MICA typing was performed in both recipients and donors. Sera were collected sequentially: pre-transplant, at the AHR episode and at follow-up. Sera from 30 patients with functioning graft were also analysed. A stable MICA()008 transfected cell line was used as target to identify MICA antibodies. MICA antibodies were not detected pre-transplant nor post-transplant in patients receiving a compatible graft. MICA antibodies were detected post-transplant AHR in patients receiving an incompatible graft. The persistence of MICA antibodies was associated with chronic graft dysfunction in 3 of 4 patients in this series; although it was not always associated with the graft loss in treated AHR. None of the 30 patients in the control group with long-term functioning grafts showed antibodies to MICA()008. This report provides some insights of the relevance of MICA antibodies in AHR.

Clinical factors influencing T-cell receptor excision circle (TRECs) counts following allogeneic stem cell transplantation in adults.

To ascertain the clinical factors involved in T-cell reconstitution after allogeneic stem cell transplantation (SCT), we evaluated serial assessments of lymphocyte subsets by flow cytometry and TRECs levels by quantitative PCR in 83 adult patients. Patient age >25 years, unrelated donor, CMV infection and acute graft-versus-host disease (GVHD) adversely affected CD3(+) and CD8(+) T-cell recovery after SCT (p < 0.05). TRECs were low or undetectable during the first months after transplant and progressively increased thereafter. However, median TRECs of patients did never achieve normal values compared to healthy donors (median follow-up 9 months, range 2-42). Presence and severity of chronic GVHD significantly affected TRECs counts: patients with chronic GVHD had lower TRECs than patients without GVHD at 9, 12 and 24 months after SCT (p = 0.002, p = 0.022, p = 0.015). Patients with limited chronic GVHD had higher TRECs compared to patients with extensive GVHD (p = 0.018). No relationship was observed between fungal or bacterial infections and TRECs. Nonetheless, CMV infection was associated with lower TRECs (p = 0.032). Our data support the concept that adult thymus contributes with a slow but continuous production of thymic T cells to immune reconstitution after SCT. Chronic GVHD is the main factor associated to a delay in TRECs counts recovery.

Reduced-intensity conditioning regimen preserves thymic function in the early period after hematopoietic stem cell transplantation.

OBJECTIVE: To compare T-cell reconstitution in two groups of patients submitted to allogeneic stem cell transplantation (SCT): those receiving reduced-intensity conditioning (RIC, n = 24) and those receiving myeloablative conditioning (MA, n = 27). METHODS: Fifty-one consecutive patients undergoing SCT were evaluated. Serial assessments of lymphocyte subsets and T cell receptor excision circles (TRECs) levels were performed using multiparametric flow cytometry and real-time PCR, respectively. RESULTS: During the first 6 months posttransplant, total and naïve CD4(+) T cell counts were higher after RIC-SCT than after MA-SCT (total CD4(+): p = 0.04, p = 0.08, and p = 0.058; naïve CD4(+): p = 0.14, p = 0.05, and p = 0.01 at 1, 3, and 6 months, respectively). In both groups of patients, TRECs levels were low or undetectable in the first 3 months after SCT and progressively increased during the study. However, a higher proportion of patients with detectable levels of TRECs was observed in RIC-SCT at 1 and 3 months and more patients in this group reached normal levels of TRECs at 6 months post-SCT. In the multivariate analysis, including factors such as type of donor (sibling vs unrelated), dose of CD34(+) cells infused with the graft, patient age, and graft-vs-host disease (GVHD), the most important factor influencing TRECs recovery in the early period after SCT was the type of conditioning regimen. CONCLUSIONS: In this study, the pattern of immune reconstitution after RIC-SCT was different from that of MA-SCT and was characterized by higher posttransplant naïve CD4(+) T cell counts and TRECs levels in the early period after transplant.

The human CD5L/AIM-CD36 axis: A novel autophagy inducer in macrophages that modulates inflammatory responses.

CD5L (CD5 molecule-like) is a secreted glycoprotein that participates in host response to bacterial infection. CD5L influences the monocyte inflammatory response to the bacterial surface molecules lipopolysaccharide (LPS) and lipoteichoic acid (LTA) by inhibiting TNF secretion. Here we studied the intracellular events that lead to macrophage TNF inhibition by human CD5L. To accomplish this goal, we performed functional analyses with human monocytic THP1 macrophages, as well as with peripheral blood monocytes. Inhibition of phosphatidylinositol 3-kinase (PtdIns3K) reversed the inhibitory effect of CD5L on TNF secretion. Among the various PtdIns3K isoforms, our results indicated that CD5L activates PtdIns3K (whose catalytic subunit is termed PIK3C3), a key modulator involved in autophagy. Further analysis revealed a concomitant enhancement of autophagy markers such as cellular LC3-II content, increased LC3 puncta, as well as LC3-LysoTracker Red colocalization. Moreover, electron microscopy showed an increased presence of cytosolic autophagosomes in THP1 macrophages overexpressing CD5L. Besides preventing TNF secretion, CD5L also inhibited IL1B and enhanced IL10 secretion. This macrophage anti-inflammatory pattern of CD5L was reverted upon silencing of autophagy protein ATG7 by siRNA transfection. Additional siRNA experiments in THP1 macrophages indicated that the induction of autophagy mechanisms by CD5L was achieved through cell-surface scavenger receptor CD36, a multiligand receptor expressed in a wide variety of cell types. Our data represent the first evidence that CD36 is involved in autophagy and point to a significant contribution of the CD5L-CD36 axis to the induction of macrophage autophagy.

Human scavenger protein AIM increases foam cell formation and CD36-mediated oxLDL uptake.

AIM is expressed by macrophages in response to agonists of the nuclear receptors LXR/RXR. In mice, it acts as an atherogenic factor by protecting macrophages from the apoptotic effects of oxidized lipids. In humans, it is detected in atherosclerotic lesions, but no role related to atherosclerosis has been reported. This study aimed to investigate whether the role of hAIM extends beyond inhibiting oxidized lipid-induced apoptosis. To accomplish this goal, functional analysis with human monocytic THP1 cells and macrophages differentiated from peripheral blood monocytes were performed. It was found that hAIM reduced oxLDL-induced macrophage apoptosis and increased macrophage adhesion to endothelial ICAM-1 by enhancing LFA-1 expression. Furthermore, hAIM increased foam cell formation, as shown by Oil Red O and Nile Red staining, as well as quantification of cholesterol content. This was not a result of decreased reverse cholesterol transport, as hAIM did not affect the efflux significantly from [(3)H] Cholesterol-laden macrophages driven by plasma, apoA-I, or HDL2 acceptors. Rather, flow cytometry studies indicated that hAIM increased macrophage endocytosis of fluorescent oxLDL, which correlated with an increase in the expression of the oxLDLR CD36. Moreover, hAIM bound to oxLDL in ELISA and enhanced the capacity of HEK-293 cells expressing CD36 to endocytose oxLDL, as studied using immunofluorescence microscopy, suggesting that hAIM serves to facilitate CD36-mediated uptake of oxLDL. Our data represent the first evidence that hAIM is involved in macrophage survival, adhesion, and foam cell formation and suggest a significant contribution to atherosclerosis-related mechanisms in the macrophage.

The scavenger protein apoptosis inhibitor of macrophages (AIM) potentiates the antimicrobial response against Mycobacterium tuberculosis by enhancing autophagy.

Apoptosis inhibitor of macrophages (AIM), a scavenger protein secreted by tissue macrophages, is transcriptionally regulated by the nuclear receptor Liver X Receptor (LXR) and Retinoid X Receptor (RXR) heterodimer. Given that LXR exerts a protective immune response against M. tuberculosis, here we analyzed whether AIM is involved in this response. In an experimental murine model of tuberculosis, AIM serum levels peaked dramatically early after infection with M. tuberculosis, providing an in vivo biological link to the disease. We therefore studied the participation of AIM in macrophage response to M. tuberculosis in vitro. For this purpose, we used the H37Rv strain to infect THP-1 macrophages transfected to stably express AIM, thereby increasing infected macrophage survival. Furthermore, the expression of this protein enlarged foam cell formation by enhancing intracellular lipid content. Phagocytosis assays with FITC-labeled M. tuberculosis bacilli indicated that this protein was not involved in bacterial uptake; however, AIM expression decreased the number of intracellular cfus by up to 70% in bacterial killing assays, suggesting that AIM enhances macrophage mycobactericidal activity. Accordingly, M. tuberculosis-infected AIM-expressing cells upregulated the production of reactive oxygen species. Moreover, real-time PCR analysis showed increased mRNA levels of the antimicrobial peptides cathelicidin and defensin 4B. These increases were concomitant with greater cellular concentrations of the autophagy-related molecules Beclin 1 and LC3II, as well as enhanced acidification of mycobacterial phagosomes and LC3 co-localization. In summary, our data support the notion that AIM contributes to key macrophage responses to M. tuberculosis.