Non-alcoholic steatohepatitis patients exhibit reduced CD47, and increased sphingosine, cholesterol and MCP1 levels in the erythrocyte membranes

Non-alcoholic steatohepatitis (NASH) constitutes a significant cause of deaths, liver transplantations and economic costs worldwide. Despite extended research, investigations on the role of erythrocytes are scarce. Red blood cells from experimental animals and human patients with NASH, present phosphatidylserine exposure which is then recognized by Kupffer cells. This event leads to erythrophagocytosis, and amplification of inflammation through iron disposition. In addition, it has been shown that erythrocytes from NASH patients release the chemokine MCP1, leading to increased TNF-α release from macrophages RAW 264.7. However, erythrophagocytosis can also be caused by reduced CD47 levels. In addition, increased MCP1 release could be either signal-induced, or caused by higher MCP1 levels on the erythrocyte membrane. Finally, erythrocyte efferocytosis could provide additional inflammatory metabolites. In this study, we measured the erythrocyte membrane levels of CD47 and MCP1 by ELISA, and cholesterol and sphingosine with thin-layer chromatography. 18 patients (8 men, 10 women aged 56.7+/-11.5 years) and 14 healthy controls (7 men, 7 women aged 39.3+/-15.5 years) participated in our study. The erythrocyte CD47 levels were decreased in the erythrocyte membranes of NASH patients (844+/-409 pg/ml) compared to healthy controls (2969+/-1936 pg/ml) with P(Healthy>NAFLD)=99.1%, while the levels of MCP1 were increased in NASH patients (389+/-255 pg/ml), compared to healthy controls (230+/-117 pg/ml) with P(Healthy<NAFLD)=88.9%. Moreover, in erythrocyte membranes there was a statistically significant accumulation of sphingosine and cholesterol in NASH patients, compared to healthy controls. Our results imply that erythrocytes release chemotactic (find me signals) MCP1, while containing reduced (do not eat me signals) CD47. These molecules can lead to erythrophagocytosis. Next, increased (goodbye signals) sphingosine and cholesterol could augment inflammation by metabolic reprogramming.

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TNF-α Activating Osteoclasts in Patients with Psoriatic Arthritis Enhances the Recruitment of Osteoclast Precursors: A Plausible Role of WNT5A-MCP-1 in Osteoclast Engagement in Psoriatic Arthritis

International Journal of Molecular Sciences ◽

10.3390/ijms23020921 ◽

2022 ◽

Vol 23 (2) ◽

pp. 921

Author(s):

Shang-Hung Lin ◽

Ji-Chen Ho ◽

Sung-Chou Li ◽

Yu-Wen Cheng ◽

Chung-Yuan Hsu ◽

...

Keyword(s):

Psoriatic Arthritis ◽

Rna Interference ◽

Joint Destruction ◽

Neutralizing Antibody ◽

Healthy Controls ◽

Osteoclast Precursors ◽

Tnf Α ◽

Wnt Ligands

Psoriatic arthritis (PsA) results from joint destruction by osteoclasts. The promising efficacy of TNF-α blockage indicates its important role in osteoclastogenesis of PsA. WNT ligands actively regulate osteoclastogenesis. We investigated how WNT ligands activate osteoclasts amid the TNF-α milieu in PsA. We first profiled the expression of WNT ligands in CD14+ monocyte-derived osteoclasts (MDOC) from five PsA patients and five healthy controls (HC) and then validated the candidate WNT ligands in 32 PsA patients and 16 HC. Through RNA interference against WNT ligands in MDOC, we determined the mechanisms by which TNF-α exerts its effects on osteclastogenesis or chemotaxis. WNT5A was selectively upregulated by TNF-α in MDOC from PsA patients. The number of CD68+WNT5A+ osteoclasts increased in PsA joints. CXCL1, CXCL16, and MCP-1 was selectively increased in supernatants of MDOC from PsA patients. RNA interference against WNT5A abolished the increased MCP-1 from MDOC and THP-1-cell-derived osteoclasts. The increased migration of osteoclast precursors (OCP) induced by supernatant from PsA MDOC was abolished by the MCP-1 neutralizing antibody. WNT5A and MCP-1 expressions were decreased in MDOC from PsA patients treated by biologics against TNF-α but not IL-17. We conclude that TNF-α recruits OCP by increased MCP-1 production but does not directly activate osteoclastogenesis in PsA.

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Changes in Erythrocyte Membrane Ouabain-Sensitive Adenosine Triphosphatase after Renal Transplantation

Clinical Science ◽

10.1042/cs0480239 ◽

1975 ◽

Vol 48 (3) ◽

pp. 239-242 ◽

Cited By ~ 2

Author(s):

C. H. Cole ◽

R. Maletz

Keyword(s):

Renal Transplantation ◽

Atpase Activity ◽

Erythrocyte Membrane ◽

Adenosine Triphosphatase ◽

Inorganic Phosphorus ◽

Erythrocyte Membranes ◽

Healthy Controls ◽

Intracellular Electrolytes ◽

Transplant Patients ◽

The Mean

1. Intracellular electrolytes, and erythrocyte membrane adenosine triphosphatase (ATPase) activity, was studied in twenty patients after renal transplantation. 2. The mean ouabain-sensitive ATPase activity in the erythrocyte membranes of the transplant patients was 122 nmol of inorganic phosphorus (Pi) h−1 mg of tissue−1 (sem 14), compared with 62 nmol of Pi h−1 mg of tissue−1 (sem 8) in a group of paired, healthy controls. 3. The increase in ouabain-sensitive ATPase was most marked in the 4 months after transplantation. However, a significant increase in ouabain-sensitive ATPase persisted for more than 8 months after transplantation. 4. This increase in ouabain-sensitive ATPase was associated with a decrease in intracellular sodium in the erythrocytes of the transplant patients.

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Role of Fn14 in acute alcoholic steatohepatitis in mice

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00429.2013 ◽

2015 ◽

Vol 308 (4) ◽

pp. G325-G334 ◽

Cited By ~ 8

Author(s):

Gamze Karaca ◽

Guanhua Xie ◽

Cynthia Moylan ◽

Marzena Swiderska-Syn ◽

Cynthia D. Guy ◽

...

Keyword(s):

Growth Factor ◽

Liver Injury ◽

Control Diet ◽

Alcohol Exposure ◽

Tnf Receptor ◽

High Fat ◽

Alcoholic Steatohepatitis ◽

Tnf Α ◽

Induce Liver Injury

TNF-like weak inducer of apoptosis (TWEAK) is a growth factor for bipotent liver progenitors that express its receptor, fibroblast growth factor-inducible 14 (Fn14), a TNF receptor superfamily member. Accumulation of Fn14+ progenitors occurs in severe acute alcoholic steatohepatitis (ASH) and correlates with acute mortality. In patients with severe ASH, inhibition of TNF-α increases acute mortality. The aim of this study was to determine whether deletion of Fn14 improves the outcome of liver injury in alcohol-consuming mice. Wild-type (WT) and Fn14 knockout (KO) mice were fed control high-fat Lieber deCarli diet or high-fat Lieber deCarli diet with 2% alcohol (ETOH) and injected intraperitoneally with CCl4 for 2 wk to induce liver injury. Mice were euthanized 3 or 10 days after CCl4 treatment. Survival was assessed. Liver tissues were analyzed for cell death, inflammation, proliferation, progenitor accumulation, and fibrosis by quantitative RT-PCR, immunoblot, hydroxyproline content, and quantitative immunohistochemistry. During liver injury, Fn14 expression, apoptosis, inflammation, hepatocyte replication, progenitor and myofibroblast accumulation, and fibrosis increased in WT mice fed either diet. Mice fed either diet expressed similar TWEAK/Fn14 levels, but ETOH-fed mice had higher TNF-α expression. The ETOH-fed group developed more apoptosis, inflammation, fibrosis, and regenerative responses. Fn14 deletion did not reduce hepatic TNF-α expression but improved all injury parameters in mice fed the control diet. In ETOH-fed mice, Fn14 deletion inhibited TNF-α induction and increased acute mortality, despite improvement in liver injury. Fn14 mediates wound-healing responses that are necessary to survive acute liver injury during alcohol exposure.

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Amyloid insulin interaction with erythrocytes

Biochemistry and Cell Biology ◽

10.1139/o03-009 ◽

2003 ◽

Vol 81 (1) ◽

pp. 51-59 ◽

Cited By ~ 11

Author(s):

J Murali ◽

D Koteeswari ◽

J M Rifkind ◽

R Jayakumar

Keyword(s):

Flow Cytometry ◽

Erythrocyte Membrane ◽

Dissociation Constants ◽

Insulin Receptors ◽

Insulin Binding ◽

Erythrocyte Membranes ◽

Flow Cytometric ◽

Human Erythrocyte Membranes ◽

Insulin Fibrils

Erythrocyte membrane interactions with insulin fibrils (amyloid) have been investigated using centrifugation, fluorescence spectroscopy, light scattering, and flow cytometric techniques. The results indicate that insulin fibrils are having moderate affinity to erythrocyte membrane. However, analysis of the apparent dissociation constants of human erythrocyte membranes (leaky and resealed vesicles) with amyloid insulin reveal that the insulin binding is drastically reduced on attaining the fibrillar state compared with native insulin. To understand the role of insulin receptors on erythrocytes binding to amyloid, we have studied the interaction of biotinylated forms of denatured and amyloidic insulin with erythrocytes. FITC-streptavidin was used as a counter staining in flow cytometry measurements. We found that insulin fibrils bind 10 times more with erythrocyte membranes than with amylin and denatured insulin.Key words: insulin amyloid, erythrocyte membrane, amyloid binding, flow cytometry, dissociation constant.

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CD47 Glycoprotein Interacts with p4.1R and p55 in the Erythrocyte.

Blood ◽

10.1182/blood.v108.11.1126.1126 ◽

2006 ◽

Vol 108 (11) ◽

pp. 1126-1126

Author(s):

Neil D. Avent ◽

Zoe E. Plummer ◽

David J. Head

Keyword(s):

Erythrocyte Membrane ◽

Direct Interaction ◽

Potential Interaction ◽

Erythrocyte Membranes ◽

Mature Erythrocyte ◽

Diffuse Fraction ◽

Cytoplasmic Face ◽

Free Translation

Abstract CD47 is a 47–50kDa membrane glycoprotein with 5 known isoforms. The role of CD47 within the erythrocyte membrane remains the subject of much research and debate though we recently provided evidence that CD47 may function as an inducer of eryptosis (Head et al, 2005). As both a cytoskeletal linked fraction and a smaller membrane diffuse fraction of CD47 exists, it is most likely that there are a number of protein species that are able to bind to CD47 at its cytoplasmic face. Our research has focused on a study of the molecular interactions of erythrocyte CD47 with erythrocyte membrane skeletal proteins protein 4.1R (p4.1R), protein 4.2 (p4.2) and p55. Here we demonstrate the ubiquitous expression of all CD47 isoforms in haemopoietic cells and tissues using basic and real-time PCR. Via immunoprecipitation of CD47 from mature erythrocyte membranes using the anti-CD47 mAb BRIC-126, yeast two-hybrid analysis and in vitro co-immunoprecipitation of 35[S] labelled peptides in a cell-free translation procedure, a novel ternary complex involving CD47, p55 and p4.1R has been indicated. More specifically, the potential interaction between p55/p4.1R and the cytoplasmic face of CD47 has been localised to the PDZ and FERM domain of these proteins respectively. Though research suggests p4.2 provides the major cytoskeletal attachment of CD47 (Bruce et al, 2002; Mouro-Chanteloup et al, 2003), a direct interaction between CD47 and p4.2 was not suggested by our study and remains undemonstrated. We continue to further the evidence for a functional role of CD47 in eryptosis. We propose p4.1R links CD47 to the apoptotic machinery of the cell and suggest a mechanism whereby cytoskeletal rearrangement and PS exposure occurs. 4.1null cells have been obtained and are currently being investigated. Further characterisation of the eryptotic pathway may offer insight into potential therapies for erythroleukaemia characterised by resistance of the erythroid lineage to apoptosis. This work has equal significance in the stabilization of red cell preparations in the blood transfusion setting.

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Mutations in GBA and LRRK2 Are Not Associated with Increased Inflammatory Markers

Journal of Parkinson s Disease ◽

10.3233/jpd-212624 ◽

2021 ◽

pp. 1-12

Author(s):

Avner Thaler ◽

Nurit Omer ◽

Nir Giladi ◽

Tanya Gurevich ◽

Anat Bar-Shira ◽

...

Keyword(s):

Inflammatory Markers ◽

Group Differences ◽

Healthy Controls ◽

Disease Phenotype ◽

Ashkenazi Jews ◽

Cognitive Assessments ◽

Prodromal Phase ◽

Disease Characteristics ◽

Tnf Α

Background: Inflammation is an integral part of neurodegeneration including in Parkinson’s disease (PD). Ashkenazi Jews have high rates of genetic PD with divergent phenotypes among GBA-PD and LRRK2-PD. The role of inflammation in the prodromal phase of PD and the association with disease phenotype has yet to be elucidated. Objective: To assess central and peripheral cytokines among PD patients with mutations in the LRRK2 and GBA genes and among non-manifesting carriers (NMC) of these mutations in order to determine the role of inflammation in genetic PD. Methods: The following cytokines were assessed from peripheral blood and cerebrospinal fluid (CSF): TNF-α, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10 and INF- γ. A comprehensive intake including general medical conditions, use of anti-inflammatory treatments, motor and cognitive assessments and additional laboratory measures were recorded, enabling the construction of the MDS probable prodromal score. Results: Data from 362 participants was collected: 31 idiopathic PD (iPD), 30 LRRK2-PD, 77 GBA-PD, 3 homozygote GBA-PD, 3 GBA-LRRK2-PD, 67 LRRK2-NMC, 105 GBA-NMC, 14 LRRK2-GBA-NMC, and 32 healthy controls. No between-group differences in peripheral or CSF cytokines were detected. No correlation between disease characteristics or risk for prodromal PD could be associated with any inflammatory measure. Conclusion: In this study, we could not detect any evidence on dysregulated immune response among GBA and LRRK2 PD patients and non-manifesting mutation carriers.

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OP0168 THE ROLE OF VASCULAR INFLAMMATION MARKERS IN DEFICIENCY OF ADENOSINE DEAMINASE 2

Annals of the Rheumatic Diseases ◽

10.1136/annrheumdis-2021-eular.2504 ◽

2021 ◽

Vol 80 (Suppl 1) ◽

pp. 100.1-101

Author(s):

U. Kaya Akca ◽

E. Sag ◽

Ş. Ünal ◽

M. Kasap-Cuceoglu ◽

Y. Bilginer ◽

...

Keyword(s):

Adenosine Deaminase ◽

Plasma Levels ◽

Vascular Inflammation ◽

Healthy Controls ◽

Inflammation Markers ◽

Diamond Blackfan Anemia ◽

Tnf Α ◽

Significant Difference ◽

Adenosine Deaminase 2

Background:Deficiency of adenosine deaminase 2 (DADA2) is a monogenic autoinflammatory disease whose pathogenesis has not been clearly elucidated.Objectives:To investigate the role of vascular inflammatory factors in the pathogenesis of DADA2, to compare the vascular inflammation profiles of DADA2 patients with different phenotypes, and to compare DADA2 patients with classic polyarteritis nodosa (PAN).Methods:The study included eighteen DADA2 patients, ten PAN patients, and eight healthy controls. Plasma levels of sST2, sRAGE, Tie-2, sCD40L, Tie-1, sFlt-1, LIGHT, TNF-α, PlGF, IL-6, IL-18, IL-10, MCP-1 were studied by cytometric bead-based multiplex assay panel.Results:Among the DADA2 patients, five had hematological manifestations, 13 had vasculitic findings, and accompanying immunological findings were present in seven patients. Nine patients had neurological findings, five of whom had neuropathy. Hematological findings were Diamond-Blackfan anemia-like phenotype in four patients and bicytopenia (anemia and thrombocytopenia) in one patient. Disease characteristics of DADA2 and PAN patients revealed that neurological involvement and livedo reticularis were more frequent in DADA2 patients (p=0.034 and p=0.009, respectively), while myalgia was more common in PAN patients (p:0.001).Plasma levels of Tie-1 and sFlt-1 were higher in the overall DADA2 patients compared to healthy controls and PAN patients (p<0.001 and p=0.004, respectively). DADA2 patients with PAN-like features had higher sRAGE, Tie-2, and TNF-α levels compared to PAN patients (p=0.013, p=0.003, and p=0.001, respectively).There was no significant difference in the levels of vascular inflammation markers between DADA2 patients with vasculitis and hematological involvement except IL-18. The plasma IL-18 levels were higher in the DADA2 patients with hematological findings compared to vasculitic phenotype (p=0.001). Finally DADA2 patients with neuropathy had higher sRAGE concentrations than patients without neuropathy and healthy controls (p=0.03 and p=0.008, respectively).Conclusion:We suggest that the high plasma IL-18 levels may be associated with an activated IFN pathway, the pathogenesis of hematologic manifestations, and unresponsive to anti-TNF treatment. Higher concentrations of Tie-1, Tie-2, sFlt-1, sRAGE, and TNF-α distinguished DADA2 patients with PAN-like features from PAN patients. We identified sRAGE as a potential biomarker of neuropathy in DADA2 patients.References:[1]Pesciotta EN, Lam H-S, Kossenkov A, et al. In-Depth, Label-Free analysis of the erythrocyte cytoplasmic proteome in Diamond Blackfan Anemia identifies a unique inflammatory signature. PLoS One 2015;10(10):e0140036.[2]Haslbeck KM, Bierhaus A, Erwin S, et al. Receptor for advanced glycation endproduct (RAGE)–mediated nuclear factor-κB activation in vasculitic neuropathy. Muscle Nerve 2004;29(6):853-60.Disclosure of Interests:None declared

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