Phosphatidylserine Receptor Research Articles

Tim-4+ cavity-resident macrophages impair anti-tumor CD8+ Tcell immunity.

Immune checkpoint blockade (ICB) has been a remarkable clinical advance for cancer; however, the majority of patients do not respond to ICB therapy. We show that metastatic disease in the pleural and peritoneal cavities is associated with poor clinical outcomes after ICB therapy. Cavity-resident macrophages express high levels of Tim-4, a receptor for phosphatidylserine (PS), and this is associated with reduced numbers of CD8+ Tcells with tumor-reactive features in pleural effusions and peritoneal ascites from patients with cancer. We mechanistically demonstrate that viable and cytotoxic anti-tumor CD8+ Tcells upregulate PS and this renders them susceptible to sequestration away from tumor targets and proliferation suppression by Tim-4+ macrophages. Tim-4 blockade abrogates this sequestration and proliferation suppression and enhances anti-tumor efficacy in models of anti-PD-1 therapy and adoptive Tcell therapy in mice. Thus, Tim-4+ cavity-resident macrophages limit the efficacy of immunotherapies in these microenvironments.

Phosphatidylserine Receptor Enhancement of SARS-CoV-2 Entry: AXL as a Therapeutic Target for COVID-19

Abstract Phosphatidylserine (PS) receptors enhance infection of a wide range of enveloped RNA viruses through virion-associated PS binding that is termed apoptotic mimicry. Here we show that this broadly shared uptake mechanism is utilized by SARS-CoV-2. Under ACE2lo conditions, expression of members of the TIM and TAM families of PS receptors synergized with ACE2 to enhance SARS-CoV-2 infection; however, PS receptors alone did not mediate infection. PS receptors enhanced SARS-CoV-2 binding to the surface of cells. While PS receptors did not interact directly with purified SARS-CoV-2 spike, addition of PS liposomes reduced entry of VSV-SARS-CoV-2 spike pseudovirions, providing evidence that PS/PS receptor interactions are mediating the effect. AXL being abundant on airway cell lines, we evaluated small molecule inhibitors of AXL signaling such as bemcentinib for their ability to inhibit SARS-CoV-2 infection. Bemcentinib robustly inhibited virus infection of some lung cell lines that expressed AXL. This inhibition correlated well with inhibitors that block cathepsin activity, consistent with AXL-mediated uptake of SARS-CoV-2 into the endosomal compartment. RNAseq studies in Vero E6 and A549-hACE2 cells demonstrated that bemcentinib reduced SARS-CoV-2 transcripts dramatically. The ability of bemcentinib to decrease coronavirus infection in vivo was assessed using the betacoronavirus mouse hepatitis virus (MHV). Liver titers and virus load of MHV were significantly inhibited by bemcentinib at day 5 of infection. In total, our findings provide evidence that PS receptors facilitate infection of the pandemic coronavirus SARS-CoV-2 and that inhibition of signaling of the PS receptor AXL has therapeutic potential against SARS-CoV-2.

TIM4 expression by dendritic cells mediates uptake of tumor-associated antigens and anti-tumor responses

Acquisition of cell-associated tumor antigens by type 1 dendritic cells (cDC1) is essential to induce and sustain tumor specific CD8+ T cells via cross-presentation. Here we show that capture and engulfment of cell associated antigens by tissue resident lung cDC1 is inhibited during progression of mouse lung tumors. Mechanistically, loss of phagocytosis is linked to tumor-mediated downregulation of the phosphatidylserine receptor TIM4, that is highly expressed in normal lung resident cDC1. TIM4 receptor blockade and conditional cDC1 deletion impair activation of tumor specific CD8+ T cells and promote tumor progression. In human lung adenocarcinomas, TIM4 transcripts increase the prognostic value of a cDC1 signature and predict responses to PD-1 treatment. Thus, TIM4 on lung resident cDC1 contributes to immune surveillance and its expression is suppressed in advanced tumors.

Hemolysis-associated phosphatidylserine exposure promotes polyclonal plasmablast differentiation.

Antimalarial antibody responses are essential for mediating the clearance of Plasmodium parasite-infected RBCs from infected hosts. However, the rapid appearance of large numbers of plasmablasts in Plasmodium-infected hosts can suppress the development and function of durable humoral immunity. Here, we identify that the formation of plasmablast populations in Plasmodium-infected mice is mechanistically linked to both hemolysis-induced exposure of phosphatidylserine on damaged RBCs and inflammatory cues. We also show that virus and Trypanosoma infections known to trigger hemolytic anemia and high-grade inflammation also induce exuberant plasmablast responses. The induction of hemolysis or administration of RBC membrane ghosts increases plasmablast differentiation. The phosphatidylserine receptor Axl is critical for optimal plasmablast formation, and blocking phosphatidylserine limits plasmablast expansions and reduces Plasmodium parasite burden in vivo. Our findings support that strategies aimed at modulating polyclonal B cell activation and phosphatidylserine exposure may improve immune responses against Plasmodium parasites and potentially other infectious diseases that are associated with anemia.

AXL, an Important Host Factor for DENV and ZIKV Replication.

Flaviviruses, as critically important pathogens, are still major public health problems all over the world. For instance, the evolution of ZIKV led to large-scale outbreaks in the Yap island in 2007. DENV was considered by the World Health Organization (WHO) as one of the 10 threats to global health in 2019. Enveloped viruses hijack a variety of host factors to complete its replication cycle. Phosphatidylserine (PS) receptor, AXL, is considered to be a candidate receptor for flavivirus invasion. In this review, we discuss the molecular structure of ZIKV and DENV, and how they interact with AXL to successfully invade host cells. A more comprehensive understanding of the molecular mechanisms of flavivirus-AXL interaction will provide crucial insights into the virus infection process and the development of anti-flavivirus therapeutics.

Abstract PO032: TIM3 regulation by phosphatidylserine

Abstract Immune responses have been successfully reactivated in cancer by targeting inhibitory receptors such as PD-1 on exhausted T cells to block immune checkpoints. Many patients do not respond, however, and a key challenge in immuno-oncology is to identify and understand new immune-regulatory targets to increase response rates. One other co-inhibitory receptor in T cells is TIM3, which – like PD-1 – is upregulated on exhausted T cells in the tumor microenvironment. TIM3 belongs to a family of phosphatidylserine (PS) receptors with well-documented roles in efferocytosis by phagocytes. However, the role of PS in regulating TIM3 remains unclear. Previous studies have shown that TIM3 can variously enhance or inhibit T cell signaling, though exactly how TIM3 accomplishes this range of activity is unknown. We therefore investigated how TIM3 levels modulate T cell signaling, and how PS in turn influences TIM3 activity. We found that TIM3 promoted T cell activity in a Jurkat T cell model when expressed at high – but not low – levels on the cell surface. We also confirmed the ability of human TIM3 to bind PS with low micromolar affinity using surface plasmon resonance. To investigate how PS affects the ability of TIM3 to promote T cell signaling in this model, we employed several approaches, including adding PS to the cell culture, manipulating TIM3 to reduce or abrogate PS binding (using chimeric receptors and TIM3 mutants), and pharmacological blockade of PS binding using a functional TIM3 antibody. Together, these studies supported a role for PS in regulating TIM3 activity, where decreased or abrogated PS binding prevented TIM3 from promoting T cell signaling. We also sought to understand what changes in T cell receptor (TCR) signaling might account for the TIM3-dependent effects we observed. We found that TIM3 did not impact phosphorylation of downstream components in the TCR signaling cascade. Rather, cells expressing high – but not low – surface levels of TIM3 displayed enhanced phosphorylation of receptor components in the TCR signaling pathway, similar to what has been reported for PD-1. We hypothesize that TIM3 modulates TCR signaling by modifying the localization and/or dwell time of one or more signaling effector proteins (e.g. phosphatases) in a manner dependent on PS engagement. The consequences of this are then determined by surface levels of TIM3. Our results demonstrate that PS does function as a ligand capable of modulating TIM3 activity, and provide new mechanistic insight into how TIM3 modulates TCR signaling. Citation Format: Courtney M. Smith, Alice Li, Nithya Krishnamurthy, Mark A. Lemmon. TIM3 regulation by phosphatidylserine [abstract]. In: Abstracts: AACR Virtual Special Conference: Tumor Immunology and Immunotherapy; 2020 Oct 19-20. Philadelphia (PA): AACR; Cancer Immunol Res 2021;9(2 Suppl):Abstract nr PO032.

Tim4 recognizes carbon nanotubes and mediates phagocytosis leading to granuloma formation

Macrophage recognition and phagocytosis of crystals is critical for the associated fibrosis and cancer. Of note, multi-walled carbon nanotubes (MWCNTs), the highly representative products of nanotechnology, induce macrophage NLRP3 inflammasome activation and cause asbestosis-like pathogenesis. However, it remains largely unknown how macrophages efficiently recognize MWCNTs on their cell surfaces. Here, we identify by a targeted screening of phagocyte receptors the phosphatidylserine receptors Tcell immunoglobulin mucin 4 (Tim4) and Tim1 as the pattern-recognition receptors for carbon crystals. Docking simulation studies reveal spatiotemporally stable interfaces between aromatic residues in the extracellular IgV domain of Tim4 and one-dimensional carbon crystals. Further, CRISPR-Cas9-mediated deletion of Tim4 and Tim1 reveals that Tim4, but not Tim1, critically contributes to the recognition of MWCNTs by peritoneal macrophages and to granuloma development in a mouse model of direct mesothelium exposure to MWCNTs. These results suggest that Tim4 recognizes MWCNTs through aromatic interactions and mediates phagocytosis leading to granulomas.

Phosphatidylethanolamine and Phosphatidylserine Synergize To Enhance GAS6/AXL-Mediated Virus Infection and Efferocytosis.

Phosphatidylserine (PS) receptors mediate clearance of apoptotic cells-efferocytosis-by recognizing the PS exposed on those cells. They also mediate the entry of enveloped viruses by binding PS in the virion membrane. Here, we show that phosphatidylethanolamine (PE) synergizes with PS to enhance PS receptor-mediated efferocytosis and virus entry. The presence of PE on the same surface as PS dramatically enhances recognition of PS by PS-binding proteins such as GAS6, PROS, and TIM1. Liposomes containing both PE and PS bound to GAS6 and were engulfed by AXL-expressing cells much more efficiently than those containing PS alone. Further, infection of AXL-expressing cells by infectious Zika virus or Ebola, Chikungunya, or eastern equine encephalitis pseudoviruses was inhibited with greater efficiency by the liposomes containing both PS and PE compared to a mixture of liposomes separately composed of PS and PE. These data demonstrate that simultaneous recognition of PE and PS maximizes PS receptor-mediated virus entry and efferocytosis and underscore the important contribution of PE in these major biological processes.IMPORTANCE Phosphatidylserine (PS) and phosphatidylethanolamine (PE) are usually sequestered to the inner leaflet of the plasma membrane of the healthy eukaryotic cells. During apoptosis, these phospholipids move to the cell's outer leaflet where they are recognized by so-called PS receptors on surveilling phagocytes. Several pathogenic families of enveloped viruses hijack these PS receptors to gain entry into their target cells. Here, we show that efficiency of these processes is enhanced, namely, PE synergizes with PS to promote PS receptor-mediated virus infection and clearance of apoptotic cells. These findings deepen our understanding of how these fundamental biological processes are executed.

Porcine Reproductive and Respiratory Syndrome Virus Utilizes Viral Apoptotic Mimicry as an Alternative Pathway To Infect Host Cells.

Porcine reproductive and respiratory syndrome (PRRS), caused by PRRS virus (PRRSV), has led to enormous economic losses in global swine industry. Infection by PRRSV is previously shown to be via low pH-dependent clathrin-mediated endocytosis, and CD163 functions as an essential receptor during viral infection. Despite much research focusing on it, PRRSV infection remains to be fully elucidated. In this study, we demonstrated that PRRSV externalized phosphatidylserine (PS) on the envelope as viral apoptotic mimicry and infected host cells through T-cell immunoglobulin and mucin domain (TIM)-induced and CD163-involved macropinocytosis as an alternative pathway. In detail, we identified that PS receptor TIM-1/4 recognized and interacted with PRRSV as viral apoptotic mimicry and subsequently induced macropinocytosis by the downstream Rho GTPases Rac1, cell division control protein 42 (Cdc42), and p21-activated kinase 1 (Pak1). Altogether, these results expand our knowledge of PRRSV infection, which will support implications for the prevention and control of PRRS.IMPORTANCE PRRS has caused huge economic losses to pig farming worldwide. Its causative agent, PRRSV, infects host cells through low pH-dependent clathrin-mediated endocytosis and CD163 is indispensable during the process. Whether there exist alternative infection pathways for PRRSV arouses our interest. Here, we found that PRRSV exposed PS on its envelope and disguised as apoptotic debris. The PS receptor TIM-1/4 recognized PRRSV and induced the downstream signaling pathway to mediate viral infection via CD163-dependent macropinocytosis. The current work deepens our understanding of PRRSV infection and provides clues for the development of drugs and vaccines against the virus.

The Tim gene family in efferocytosis.

One of the key features of the plasma membrane is the asymmetrical distribution of phospholipids across it. Especially, phosphatidylserine (PS) exclusively locates on its inner leaflet. Thus, the exposure of PS on the surface of cells could function as a signal initiating various cellular processes such as phagocytosis of apoptotic cells called efferocytosis, blood clotting, muscle formation, and viral entry. Indeed, PS on apoptotic cells stimulates phagocytes to engulf them and functions as an essential ligand for efferocytosis. Due to the importance of PS in efferocytosis, the existence of the PS receptor had been conceived. However, the PS receptor had not been revealed for a long time. Thus, the first identification of the PS receptor was significant excitement. Tim-4, a member of the T cell immunoglobulin and mucin domain containing family of genes, was one of PS receptors which first identified and received the greatest attention due to its expression in macrophages and relevance to autoimmune and allergic diseases. This review will serve to provide a comprehensive overview of Tim proteins as PS receptors.

Tim4 Recognizes Carbon Nanotubes and Mediates Frustrated Phagocytosis Leading to Granuloma Formation

Frustrated phagocytosis is critical for crystal-associated fibrosis and cancer. Of note, multi-walled carbon nanotubes (MWCNTs), the highly representative products of nanotechnology, induce macrophage NLRP3 inflammasome activation and cause asbestosis-like pathogenesis. However, it remains largely unknown how macrophages efficiently recognize MWCNTs on their cell surfaces. Here we identify by a targeted screening of phagocyte receptors the phosphatidylserine receptors T-cell immunoglobulin mucin 4 (Tim4) and Tim1 as the pattern-recognition receptors for carbon crystals. Docking simulation studies revealed spatiotemporally stable interfaces between aromatic residues in the extracellular IgV domain of Tim4 and one-dimensional carbon crystals. Further, CRISPR/Cas9-mediated deletion of Tim4 and Tim1 revealed that Tim4, but not Tim1, critically contributed to recognition of MWCNTs by peritoneal macrophages and to granuloma development in a mouse model of direct mesothelium exposure to MWCNTs. These results suggest that Tim4 recognizes MWCNTs through aromatic interactions and mediates frustrated phagocytosis leading to granulomas.

Tim-4 functions as a scavenger receptor for phagocytosis of exogenous particles

The phosphatidylserine (PS) receptor Tim-4 mediates phagocytosis of apoptotic cells by binding to PS exposed on the surface of these cells, and thus functions as a PS receptor for apoptotic cells. Some of PS receptors are capable of recognizing other molecules, such as LPS on bacteria, besides PS on apoptotic cells. However, it is unclear whether Tim-4 perceives other molecules like the PS receptors. Here, we report that Tim-4 facilitates the phagocytosis of exogenous particles as well as apoptotic cells. Similar to the process that occurs during Tim-4-mediated efferocytosis, the uptake of exogenous E. coli and S. aureus bioparticles was promoted by overexpression of Tim-4 on phagocytes, whereas phagocytosis of the bioparticles was reduced in Tim-4-deficient cells. A truncation mutant of Tim-4 lacking the cytoplasmic tail promoted phagocytosis of the particles, but a mutant lacking the IgV or the mucin domain failed to enhance phagocytosis. However, expression of Tim-4AAA (a mutant form of Tim-4 that does not bind phosphatidylserine and does not promote efferocytosis) still promoted phagocytosis. Tim-4-mediated phagocytosis was not blocked by expression of the phosphatidylserine-binding protein Anxa5. Furthermore, binding of lipopolysaccharide (LPS), which is found in the outer membrane of Gram-negative bacteria, was higher in Tim-4-overexpressing cells than in Tim-4-deficient cells. In summary, our study suggests that Tim-4 acts as a scavenger receptor and mediates phagocytosis of exogenous particles in a phosphatidylserine-independent manner.

Dual role of phosphatidylserine and its receptors in osteoclastogenesis

Fusion and apoptosis share a breakdown of the membrane phospholipids asymmetry, modes of which are largely unknown in osteoclastogenesis. Here, we investigated the externalization of phosphatidylserine (PS) and its receptors, and their biological functions in osteoclastogenesis. Strong immunoreactivities in vivo for the PS receptors TIM4, BAI1, and STAB2 were observed in the TRAP-positive multinucleated cells in the alveolar bone that was being remodeled around the developing dental follicles in rats. These receptors were significantly upregulated during M-CSF/RANKL-induced in vitro osteoclastogenesis using mouse bone marrow-derived cells. PS externalization in preosteoclasts was increased by the M-CSF/RANKL treatment. Multinucleation of preosteoclasts was markedly inhibited by antibodies against PS and its receptors. Among the investigated lipid transporter proteins, floppases (Abcb4, Abcc5, and Abcg1) were upregulated, whereas flippases (Atp11c and Atp8a1) downregulated during osteoclastogenesis. Preosteoclast fusion was markedly blocked by the ATPase inhibitor Na3VO4 and siRNAs against Abcc5 and Abcg1, revealing the importance of these lipid transporters in PS externalization. Further, the levels of Cd47 and Cd31, don’t-eat-me signal inducers, were increased or sustained in the early phase of osteoclastogenesis, whereas those of AnnexinI and Mfg-e8, eat-me signals inducers, were increased in the late apoptotic phase. In addition, Z-VAD-FMK, a pan caspase inhibitor, had no effect on preosteoclast fusion in the early phase of osteoclastogenesis, whereas Abs against PS, TIM4, and BAI1 decreased osteoclast apoptosis during the late phase. These results suggest that PS externalization is essential for the whole process of osteoclastogenesis and share PS receptors and transporters in the early stage fusion and late stage apoptosis. Therefore, modulation of PS and its receptors could be a useful strategy to develop anti-bone resorptive agents.

Checkpoint Receptor TIGIT Expressed on Tim-1+ B Cells Regulates Tissue Inflammation

Tim-1, a phosphatidylserine receptor expressed on B cells, induces interleukin 10 (IL-10) production by sensing apoptotic cells. Here we show that mice with B cell-specific Tim-1 deletion develop tissue inflammation in multiple organs including spontaneous paralysis with inflammation in the central nervous system (CNS). Transcriptomic analysis demonstrates that besides IL-10, Tim-1+ B cells also differentially express a number of co-inhibitory checkpoint receptors including TIGIT. Mice with B cell-specific TIGIT deletion develop spontaneous paralysis with CNS inflammation, but with limited inflammation in other organs. Our findings suggest that Tim-1+ B cells are essential for maintaining self-tolerance and restraining tissue inflammation, and that Tim-1 signaling-dependent TIGIT expression on B cells is essential for maintaining CNS-specific tolerance. A possible critical role of aryl hydrocarbon receptor (AhR) in regulating the B cell function is discussed, as we find that AhR is among the preferentially expressed transcription factors in Tim-1+ B cells and regulates their TIGIT and IL-10 expression.

Exosome mimicry by a HAVCR1-NPC1 pathway of endosomal fusion mediates hepatitis A virus infection.

Cell-to-cell communication by exosomes controls normal and pathogenic processes1,2. Viruses can spread in exosomes and thereby avoid immune recognition3. While biogenesis, binding, and uptake of exosomes are well-characterized4,5, delivery of exosome cargo into the cytoplasm is poorly understood3. We report that phosphatidylserine receptor HAVCR16,7 and cholesterol transporter NPC18 participate in cargo delivery from exosomes of hepatitis A virus (HAV)-infected cells (exo-HAV) by clathrin-mediated endocytosis. Using CRISPR/Cas9 knockouts we show that these two lipid receptors, which interact in the late endosome9, are necessary for membrane fusion and delivery of RNA from exo-HAVs into the cytoplasm. The HAVCR1/NPC1 pathway, which Ebola virus exploits to infect cells9, mediates HAV infection by exo-HAV indicating that viral infection by this exosome mimicry mechanism does not require an envelope glycoprotein. The luminal viral RNA but not endosomal uncoating of HAV particles (vpHAV) contained in the exosome is mainly responsible for exo-HAV infectivity as assessed by methylene blue-inactivation of non-encapsidated RNA. In contrast, infectivity of vpHAV is pH-independent and requires HAVCR1 or other yet unidentified receptor(s) but not NPC1. Our findings show that envelope glycoprotein-independent fusion mechanisms are shared by exosomes and viruses, and call for a reassessment of the role of envelope glycoproteins in infection.

Biophysical characterization of the adhesion between enveloped virus and phosphatidylserine-binding proteins

Abstract Enveloped viruses are viruses that have a lipid membrane. Many enveloped viruses, such as the emerging virus Ebola, are pathogenic and of great clinical importance. Recent studies have shown that the outer lipid leaflets of viral envelopes contain phosphatidylserine (PS), which serves as an important attachment factor for enveloped virus entry into host cells. The envelope PS interacts with cellular PS receptors in a manner that mimics apoptotic body-PS receptor interactions during the engulfment of dead cell debris by phagocytes. The PS receptors utilized by enveloped viruses for viral internalization are designated phosphatidylserine-mediated virus entry enhancing receptors (PVEERs). While the importance of PVEERs in mediating viral entry has been demonstrated, the biophysical properties of the PVEER-virus interactions, including the detailed kinetics of PVEER-virion-associated PS binding, have not been comprehensively evaluated and compared. Using Ebola virus (EBOV) as the model system, this study investigated and compared the interactions between model Ebola viral particles and PVEER family proteins that are known to mediate enveloped virus attachment and entry. We used single-molecule force spectroscopy to quantify the specific interaction forces between PS or model EBOV viral particles and PVEERs. Based on the dynamic force spectra of various PS-receptor interactions, our results suggest that PS-PVEER interactions are mechanically comparable to antibody-antigen interactions, and that Gas6 and MFG-E8 may generate more force-resistant intersections than TIM-1 and ProteinS. The findings can help explain the biophysical mechanisms underlying the PVEER-mediated entry of enveloped virus into cells.

Effect of phosphatidylserine on cirrhosis-induced hepatic encephalopathy: Response to acute endotoxemia in cirrhotic rats

Background/aimsIn cirrhosis, the levels of proinflammatory cytokines are high in the liver and blood. Endotoxin decreases level of consciousness in cirrhotic rats. Phosphatidylserine exists in the cell membrane structure and is essential for the survival of neurons. Phosphatidylserine receptor is found in phagocytic cells and also activates the signaling of membrane proteins in apoptotic process. Therefore this study was aimed to explore the hypothesis that hepatic encephalopathy is prevented by phosphatidylserine treatment and if so, whether this is associated with altered level of proinflammatory cytokines in the brain. MethodsCirrhosis was induced by surgical ligation of the bile duct in male Wister rats. The groups were treated with phosphatidylserine and saline for 4 weeks. Brain IL6, TNFα and the expression of phosphatidylserine receptor were assessed. Intraperitoneal injections of either saline or lipopolysaccharide (0.1 mg/kg) were administered to each group. Finally, animal behavior, blood ammonia and the expression of toll like receptor 4 were examined in the brain. ResultsCirrhosis in rats was associated with altered expression of toll-like receptor4 in brain cortex and phosphatidylserine treatment increases toll-like receptor4 receptor expression. Phosphatidylserine had anti-inflammatory effect in healthy rats but no effect in cirrhotic rats. Chronic phosphatidylserine treatment decreased blood ammonia in BDL cirrhotic rats treated with lipopolysaccharide. ConclusionThe brain of cirrhotic rat is more susceptible to acute endotoxemia and chronic phosphatidylserine treatment decreases blood ammonia and encephalopathy in cirrhotic rats by encountering endotoxin. Phosphatidylserine may boost immune system against endotoxin.

Transplantation of activated olfactory ensheathing cells by curcumin strengthens regeneration and recovery of function after spinal cord injury in rats

Background aimsThe pro-regeneration capabilities of olfactory ensheathing cells (OECs) remain controversial. However, little is known regarding whether the transplantation of activated OECs by curcumin (CCM) elicits neural regeneration and functional recovery after spinal cord injury (SCI) in rats, and the possible molecular mechanisms have never been investigated. MethodsPrimary OECs were treated with 1μM CCM for 1–3 days. Concomitantly, activated OECs were transplanted into the traumatic spinal cord of Sprague Dawley rats. One to 9 weeks after surgery, the assessment of behavior recovery was made using the Basso, Beattie and Bresnahan (BBB) locomotor scale; electrophysiology tests, such as somatosensory evoked potential (SEP) and motor evoked potential (MEP); and the cylinder test. Pathological study, including hematoxylin and eosin staining and immunofluorescence staining for neurofilaments (NFs), was conducted at 5 weeks post-surgery. In addition, activation profiles of OECs by CCM stimulus were assessed and levels of transglutaminase-2 (TG2) and phosphatidylserine receptor (PSR) in OECs stimulated by CCM were further determined. ResultsCCM remarkably enhanced OEC proliferation, improved cell viability and strengthened secretion of neurotrophins and anti-inflammatory factors. In addition, the levels of TG2 and PSR in CCM-treated OECs were significantly elevated. More importantly, beyond 1 week post-transplantation of CCM-treated OECs into lesioned spinal cord, BBB score and cylinder test score were significantly higher than that seen in the other three groups and a more postponed latent SEP and MEP period was noted. Furthermore, 5 weeks later, numerous, well-arranged NF-positive nerve fibers, lesions with less cavities and reduced levels of pro-inflammatory cytokines were found in activated OEC implantation groups. In addition, the number of NF-positive fibers was significantly improved and the number and area of both cavities and gliotic scars were remarkably decreased compared with the corresponding controls. ConclusionsTransplantation of OECs activated by CCM promotes neural regeneration and functional recovery following SCI, the underlying mechanisms of which are intimately associated with the elevated production of neurotrophic factors and anti-inflammatory factors in OECs stimulated by CCM as well as reduced pro-inflammatory cytokines from the post-contusion spinal cord. In addition, OECs activated by CCM were mediated through TG2 and PSR.

Blocking elevated p38 MAPK restores efferocytosis and inflammatory resolution in the elderly.

Increasing age alters innate immune-mediated responses; however, the mechanisms underpinning these changes in humans are not fully understood. Using a human dermal model of acute inflammation, we found that, although inflammatory onset is similar between young and elderly individuals, the resolution phase was substantially impaired in elderly individuals. This arose from a reduction in T cell immunoglobulin mucin receptor-4 (TIM-4), a phosphatidylserine receptor expressed on macrophages that enables the engulfment of apoptotic bodies, so-called efferocytosis. Reduced TIM-4 in elderly individuals was caused by an elevation in macrophage p38 mitogen-activated protein kinase (MAPK) activity. Administering an orally active p38 inhibitor to elderly individuals rescued TIM-4 expression, cleared apoptotic bodies and restored a macrophage resolution phenotype. Thus, inhibiting p38 in elderly individuals rejuvenated their resolution response to be more similar to that of younger people. This is the first resolution defect identified in humans that has been successfully reversed, thereby highlighting the tractability of targeting pro-resolution biology to treat diseases driven by chronic inflammation.

Tim-4 in Health and Disease: Friend or Foe?

T-cell immunoglobulin and mucin domain containing 4 (Tim-4) is a phosphatidylserine receptor and is selectively expressed on antigen presenting cells. Recently, Tim-4 was reported to be expressed on iNKT cells, B1 cells, and tumor cells, suggesting it has multiple biological functions. In this review, we mainly summarize the expression and regulation of Tim-4 in immune cells including T cells, macrophages, dendritic cells, NKT cells, B cells, and mast cells. The expression of Tim-4 in these cells implies that Tim-4 might participate in immune related diseases. Emerging evidence emphasizes a substantial role for Tim-4 in maintaining homeostasis by regulating various immune responses, including viral infection, allergy, autoimmunity, and tumor immunity. Here, we collectively evaluated the role of Tim-4 in health and diseases. This summary will be extremely useful to fully understand the function of Tim-4 in the pathogenesis of immune related diseases, which would provide novel clues for the diagnosis and treatment of diseases.