Sphingosine 1-phosphate Receptor 2 Signaling Suppresses Macrophage Phagocytosis and Impairs Host Defense against Sepsis

2015 ◽  
Vol 123 (2) ◽  
pp. 409-422 ◽  
Author(s):  
JinChao Hou ◽  
QiXing Chen ◽  
Kai Zhang ◽  
BaoLi Cheng ◽  
GuoHao Xie ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mononuclear Cells ◽  
Multiorgan Failure ◽  
Cecal Ligation ◽  
Lung Damage ◽  
Peripheral Blood Mononuclear ◽  
Fluorescent Intensity ◽  
Macrophage Phagocytosis ◽  
Sphingosine 1 Phosphate ◽  
Genetic Deletion

Abstract Background: Sepsis is characterized by an inappropriate systemic inflammatory response and bacteremia that promote multiorgan failure and mortality. Sphingosine 1-phosphate receptor 2 (S1PR2) modulates endotoxin-induced inflammation in endothelium. However, as a highly expressed S1P receptor in macrophages, its role in regulating macrophage response to bacterial infection remains unclear. Methods: Cecal ligation and puncture or intratracheal instillation of Escherichia coli was induced in wild-type or S1pr2-deficient mice. The antibacterial ability of cell-specific S1PR2 was tested in bone marrow reconstitution mice or mice with macrophage-specific deletion. Signaling molecules responsible for S1PR2-mediated phagocytosis were also measured in the bone marrow–derived macrophages. In addition, S1PR2 expression levels and its correlation with severity of sepsis were determined in critically ill patients (n = 25). Results: Both genetic deletion and pharmaceutical inhibition of S1PR2 significantly limited bacterial burden, reduced lung damage, and improved survival (genetic deletion, 0% in S1pr2+/+vs. 78.6% in S1pr2−/−, P < 0.001; pharmaceutical inhibition, 9.1% in vehicle vs. 22.2% in S1PR2 antagonist, P < 0.05). This protection was attributed to the enhanced phagocytic function of S1PR2-deficient macrophages (mean fluorescent intensity, 2035.2 ± 202.1 vs. 407.8 ± 71.6, P < 0.001). Absence of S1PR2 in macrophage inhibits RhoA-dependent cell contraction and promotes IQGAP1-Rac1-dependent lamellipodial protrusion, whose signaling pathways depend on extracellular stimulators. In septic patients, increased S1PR2 levels in peripheral blood mononuclear cells were positively correlated with the severity of sepsis (r = 0.845, P < 0.001). Conclusions: This study implies that S1PR2, as a critical receptor in macrophage, impairs phagocytosis and antimicrobial defense in the pathogenesis of sepsis. Interventions targeting S1PR2 signaling may serve as promising therapeutic approaches for sepsis.

scholarly journals Defective Autophagy in MPN Were Predominantly Detected in Megakaryocytes in Philadelphia Negative (Ph -) Myeloproliferative Neoplasm

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 5386-5386
Author(s):  
Jen-Chin Wang ◽  
Guanfang Shi ◽  
Karan Josan ◽  
Preethi Ramachandran ◽  
Vladimir Gotlieb ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Western Blot ◽  
Cell Signaling ◽  
Mononuclear Cells ◽  
Myeloproliferative Neoplasm ◽  
Beclin 1 ◽  
Positive Staining ◽  
Rt Pcr ◽  
Peripheral Blood Mononuclear ◽  
Cytoplasmic Staining

We previously reported that autophagy was defective in classical Philadelphia negative (Ph -) Myeloproliferative Neoplasm (MPN) by demonstrating increased p62, decreased Beclin-1 by RT-PCR and Western Blot (WB) methods and decreased LC3-II by WB (ASH annual meeting poster 2018) in peripheral blood mononuclear cells. Now, we further performed immunohistochemical staining of these autophagy markers on the bone marrow biopsy specimens. Methods: Formalin-fixed and paraffin-embedded bone marrow samples were stained with primary antibodies against Beclin-1 (mouse monoclonal, Millipore), LC3B (rabbit monoclonal, Cell Signaling Technology), and p62 (mouse monoclonal, Cell Signaling Technology). The tissue sections were analyzed using VENTANA BenchMark Ultra System (Ventana Medical Systems, Inc.) according to the manufacturer's protocol. Patients who were studied included 12 essential thrombocythemia (ET), 10 polycythemia (PV), and 5 myelofibrosis (MF) (including 1 post-ET MF and 1 post-PV MF). Scoring was given based on the degree of cytoplasmic staining. Score 1 included weak cytoplasmic stain, score 2 included moderate cytoplasmic stain and score 3 was given for strong cytoplasmic staining . Results: 1) As shown in Fig1, the predominant cells which stained positive including p62, Beclin-1 , or LC3 B were mostly found on the megakaryoctes, although very few of other cell types were found to have positive staining as well 2) As shown in Table 1, the immunostaining results were in general correlated to the RT-PCR , or western blot findings that a defective autophagy was demonstrated in MPN patients with combined finding of a stronger staining in p62,and weak staining in Beclin-1 and LC3B. Conclusion: We have demonstrated defective autophagy in these Ph (-) MPN, by immunostaining of the bone marrow specimens, and demonstrating positivity for defective autophagy predominantly in megakaryocytes. Since megakaryocytes are the most important cells involved in the pathogenesis of MPN, we propose that defective autophagy in megakaryocytes play an important role in the pathogenesis of these diseases. Disclosures Wang: Incyt: Research Funding.

scholarly journals Reprogramming of bone marrow myeloid progenitor cells in patients with severe coronary artery disease

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Marlies P Noz ◽  
Siroon Bekkering ◽  
Laszlo Groh ◽  
Tim MJ Nielen ◽  
Evert JP Lamfers ◽  
...  
Keyword(s):  
Coronary Artery Disease ◽  
Bone Marrow ◽  
Coronary Artery ◽  
Immune Cells ◽  
Mononuclear Cells ◽  
Immune Cell ◽  
Ex Vivo ◽  
Peripheral Blood Mononuclear ◽  
Artery Disease ◽  
Myeloid Progenitors

Atherosclerosis is the major cause of cardiovascular disease (CVD). Monocyte-derived macrophages are the most abundant immune cells in atherosclerotic plaques. In patients with atherosclerotic CVD, leukocytes have a hyperinflammatory phenotype. We hypothesize that immune cell reprogramming in these patients occurs at the level of myeloid progenitors. We included 13 patients with coronary artery disease due to severe atherosclerosis and 13 subjects without atherosclerosis in an exploratory study. Cytokine production capacity after ex vivo stimulation of peripheral blood mononuclear cells (MNCs) and bone marrow MNCs was higher in patients with atherosclerosis. In BM-MNCs this was associated with increased glycolysis and oxidative phosphorylation. The BM composition was skewed towards myelopoiesis and transcriptome analysis of HSC/GMP cell populations revealed enrichment of neutrophil- and monocyte-related pathways. These results show that in patients with atherosclerosis, activation of innate immune cells occurs at the level of myeloid progenitors, which adds exciting opportunities for novel treatment strategies.

scholarly journals The Human SHIP Gene Is Differentially Expressed in Cell Lineages of the Bone Marrow and Blood

Blood ◽  
1997 ◽  
Vol 89 (6) ◽  
pp. 1876-1885 ◽  
Author(s):  
Susan J. Geier ◽  
Paul A. Algate ◽  
Kristen Carlberg ◽  
Dave Flowers ◽  
Cynthia Friedman ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mononuclear Cells ◽  
Sh2 Domain ◽  
Protein Species ◽  
Peripheral Blood Mononuclear ◽  
Phosphorylated Proteins ◽  
Cd34 Cells ◽  
Negative Effect ◽  
Mrna And Protein Expression ◽  
Macrophage Colony

Abstract The macrophage colony-stimulating factor receptor and several other hematopoietic growth factor receptors induce the tyrosine phosphorylation of a 145- to 150-kD protein in murine cells. We have previously cloned a cDNA for the murine 150-kD protein, SHIP, and found that it encodes a unique signaling intermediate that binds the SHC PTB domain through at least one tyrosine phosphorylated (NPXY) site in the carboxyl-terminal region. SHIP also contains several potential SH3 domain-binding sites, an SH2 domain for binding other tyrosine phosphorylated proteins, and an enzymatic activity that removes the phosphate from the 5 position of phosphatidylinositol 3,4,5-phosphate or from inositol 1,3,4,5-phosphate. SHIP has a negative effect on cell growth and therefore loss or modification may have profound effects on hematopoietic cell development. In this study, we have cloned a cDNA for human SHIP and examined mRNA and protein expression of SHIP and related species in bone marrow and blood cells. Flow cytometry indicates that at least 74% of immature CD34+ cells express SHIP cross-reacting protein species, whereas within the more mature population of CD33+ cells, only 10% of cells have similar expression. The majority of T cells react positively with the anti-SHIP antibodies, but significantly fewer B cells are positive. Immunoblotting detects up to seven different cross-reacting SHIP species, with peripheral blood mononuclear cells exhibiting primarily a 100-kD protein and a CD34+ acute myeloblastic leukemia expressing mainly 130-kD and 145-kD forms of SHIP. Overall, these results indicate that there is an enormous diversity in the size of SHIP or SHIP-related mRNA and protein species. Furthermore, the expression of these protein species changes according to both the developmental stage and differentiated lineage of the mature blood cell.

scholarly journals Proliferation and cytolytic function of anti-CD3 + interleukin-2 stimulated peripheral blood mononuclear cells following bone marrow transplantation

Blood ◽  
1991 ◽  
Vol 78 (5) ◽  
pp. 1286-1291 ◽  
Author(s):  
E Katsanis ◽  
PM Anderson ◽  
AH Filipovich ◽  
DE Hasz ◽  
ML Rich ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Transplantation ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Marrow Transplantation ◽  
Mononuclear Cells ◽  
Interleukin 2 ◽  
Peripheral Blood Mononuclear ◽  
Cytolytic Function ◽  
Blood Mononuclear Cells

Abstract We evaluated the proliferation, cytolytic function, and phenotypic characteristics of anti-CD3 plus interleukin-2 (IL-2) stimulated peripheral blood mononuclear cells (PBMCs) from 44 patients with leukemia or non-Hodgkin's lymphoma (NHL) treated with multiagent chemotherapy or following bone marrow transplantation (BMT). BMT patients had decreased cell growth with only a 1.35 +/- 0.25 (autologous BMT for acute lymphoblastic leukemia [ALL]), 1.24 +/- 0.25 (autologous BMT for NHL), and 0.8 +/- 0.1 (allogeneic BMT for leukemia) mean fold increase by day 5 of culture compared with controls (4.0 +/- 0.4), P less than .001. Anti-CD3 + IL-2 activated cells from patients with ALL and NHL who had received autologous BMT and cells from patients with leukemia who underwent allogeneic BMT were more effective in lysing the natural killer (NK) sensitive target, K562, and the NK- resistant target, Daudi, compared with controls. In contrast, cytolysis of K562 and Daudi by cultured PBMCs from patients with ALL and NHL receiving multi-agent chemotherapy was similar to that of controls. Cultures from BMT recipients had a significant increase in CD16+ (autologous ALL 5.7 +/- 1.5%, P less than .01; autologous NHL 12.4 +/- 3.5%, P less than .001; allogeneic 14.3 +/- 2.9%, P less than .001) and CD56+ cells (autologous ALL 27.6 +/- 12.0%, P less than .01; autologous NHL 39.3 +/- 9.5%, P less than .001; allogeneic 42.7 +/- 7.4%, P less than .001) compared with controls (CD16+ 2.5 +/- 0.4%; CD56+ 6.9 +/- 0.9%). Stimulation of PBMCs with anti-CD3 + IL-2 is effective in generating cells with high cytolytic function post-BMT.

scholarly journals Interleukin-2 production and response to interleukin-2 by peripheral blood mononuclear cells from patients after bone marrow transplantation: II. Patients receiving soybean lectin-separated and T cell-depleted bone marrow

Blood ◽  
1987 ◽  
Vol 70 (5) ◽  
pp. 1595-1603 ◽  
Author(s):  
K Welte ◽  
CA Keever ◽  
J Levick ◽  
MA Bonilla ◽  
VJ Merluzzi ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Interleukin 2 ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

Abstract The ability of peripheral blood mononuclear cells (PBMC) to produce and respond to interleukin-2 (IL-2) was evaluated in 50 recipients of HLA- identical bone marrow (BM) depleted of mature T cells by soybean agglutination and E rosetting (SBA-E-BM). In contrast to our previous findings in recipients of unfractionated marrow, during weeks 3 to 7 post-SBA-E-BM transplantation (BMT), PBMC from the majority of patients spontaneously released IL-2 into the culture medium. This IL-2 was not produced by Leu-11+ natural killer cells, which were found to be predominant in the circulation at this time, but by T11+, T3+, Ia antigen-bearing T cells. The IL-2 production could be enhanced by coculture with host PBMC frozen before transplant but not by stimulation with mitogenic amounts of OKT3 antibody, thus suggesting an in vivo activation of donor T cells or their precursors by host tissue. Spontaneous IL-2 production was inversely proportional to the number of circulating peripheral blood lymphocytes and ceased after 7 to 8 weeks post-SBA-E-BMT in most of the patients. In patients whose cells had ceased to produce IL-2 spontaneously or never produced this cytokine, neither coculture with host cells nor stimulation with OKT3 antibody thereafter induced IL-2 release through the first year posttransplant. Proliferative responses to exogenous IL-2 after stimulation with OKT3 antibody remained abnormal for up to 6 months post-SBA-E-BMT, unlike the responses of PBMC from recipients of conventional BM, which responded normally by 1 month post-BMT. However, the upregulation of IL- 2 receptor expression by exogenous IL-2 was found to be comparable to normal controls when tested as early as 3 weeks post-SBA-E-BMT. Therefore, the immunologic recovery of proliferative responses to IL-2 and the appearance of cells regulating in vivo activation of T cells appear to be more delayed in patients receiving T cell-depleted BMT. Similar to patients receiving conventional BMT, however, the ability to produce IL-2 after mitogenic stimulation remains depressed for up to 1 year after transplantation.

Expression of Programmed Death 1 Ligand in Different Compartments of Chronic Lymphocytic Leukemia

2015 ◽  
Vol 134 (4) ◽  
pp. 255-262 ◽  
Author(s):  
Maciej Grzywnowicz ◽  
Agnieszka Karczmarczyk ◽  
Katarzyna Skorka ◽  
Malgorzata Zajac ◽  
Joanna Zaleska ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Chronic Lymphocytic Leukemia ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Lymphocytic Leukemia ◽  
Strongly Correlated ◽  
Peripheral Blood Mononuclear ◽  
Programmed Death ◽  
Distinct Disease ◽  
Programmed Death 1

Background: The programmed death 1 (PD-1) receptor pathway is responsible for the negative regulation of both T and B lymphocytes upon activation of these cells. There is growing evidence that chronic lymphocytic leukemia (CLL) cells exploit the PD-1 ligand (PD-L1) to resist antitumor immune reactions and maintain their survival by shaping their own microenvironment. Methods: We used a quantitative RT-PCR method to analyze PD-L1 gene expression in bone marrow and peripheral blood mononuclear cells, representing the proliferation and accumulation compartments of CLL. Results: PD-L1 expression was found to be significantly higher in 112 CLL patients than in controls. Levels of PD-L1 expression in bone marrow and peripheral blood were comparable and showed a positive correlation. Furthermore, expression of PD-L1 strongly correlated with expression of PD-1 receptor in mononuclear cells from the same compartment, and was not affected by incubation with immunomodulatory drug thalidomide. Conclusion: PD-L1 expression is shared between CLL cells localized in distinct disease compartments, demonstrating that PD-1/PD-L1 a universal target for therapy.

Human CD8+/TCR−/CD56dim/- Facilitating Cells Facilitate Homing and Engraftment of Human HSC in Vivo

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 4093-4093
Author(s):  
Yiming Huang ◽  
Mary J Elliott ◽  
Thomas Miller ◽  
Deborah R Corbin ◽  
Larry D. Bozulic ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mononuclear Cells ◽  
Mouse Bone Marrow ◽  
Common Procedure ◽  
Peripheral Blood Mononuclear ◽  
Hematopoietic Stem ◽  
Donor Chimerism ◽  
Nsg Mice ◽  
Hsc Transplantation

Abstract Abstract 4093 Hematopoietic stem cell (HSC) transplantation has become a common procedure for treatment of hematopoietic malignancies and autoimmune disease. Despite significant advances in HSC transplantation, the morbidity and mortality of ablative conditioning and graft-versus-host disease (GVHD) remain limitations to application in the clinic. However, these risks can be overcome through less toxic nonmyeloablative conditioning and cell depletion strategies to remove GVHD causing-cells while retaining engraftment enhancing-tolerogeneic cells. We were the first to discover CD8+/TCR− graft facilitating cells (FC) in mouse bone marrow. The addition of as few as 30,000 FC to 10,000 HSC significantly enhances engraftment of HSC in allogeneic recipients without causing GVHD. FC also potently enhance engraftment of limiting numbers of syngeneic HSC. Human CD8+/TCR- FC comprised 1.1% ± 0.27% of total G-CSF-mobilized peripheral blood mononuclear cells (mPBMC). In the CD8+/TCR- FC, 48% of cells expressed CD3ε+, 43% were FoxP3+, 43% were CD11c+, 19% were CD19+, and 30% were HLA-DR+. Approximately 55% of FC are also CD56dim/-, and the remaining population is CD56bright. The morphology of human CD8+/TCR− FC with Wright-Giemsa staining under light microscopy suggested that the human FC population is heterogeneous. Here we evaluated if human FC enhance human HSC or progenitor homing to bone marrow of NOD/SCID/IL-2rγnull (NSG) mouse recipients. CD45+CD34+ HSC and CD8+/TCR−/CD56dim/-FC were sorted from mPBMC. NSG recipients were conditioned with 1100 cGy of total body irradiation (TBI). 24 hours after TBI, 100,000 HSC with or without 300,000 CD8+/TCR−/CD56dim/- FC were transplanted into conditioned NSG recipients. Recipients were euthanized 16 hours after transplantation. Bone marrow was harvested from femurs and tibias of recipients and plated in Colony Forming Culture (CFC) Assays. Recipients of HSC plus FC generated significantly more colony formation (colonies = 110) compared with HSC alone (colonies = 65) (P = 0.011), suggesting that CD8+/TCR−/CD56dim/- FC enhanced homing of HSC or progenitors to bone marrow. To test if human CD8+/TCR−/CD56dim/- FC facilitate engraftment of human HSC in NSG mice, 300,000 CD8+/TCR−/CD56dim/- FC were mixed with 100,000 HSC and transplanted into NSG recipient mice conditioned with 325 cGy TBI. Mice that received HSC alone served as controls. At 30 days after transplantation, PBL typing showed that 34% (10 of 29) recipients of HSC alone engrafted. In contrast, 78% of recipients (n = 23) of HSC plus CD8+/TCR−/CD56dim/- FC engrafted, and donor chimerism in PB was 1.1% ± 0.8% and 4.1% ± 1.3% (P <0.05), respectively. At 6 months after transplantation, NSG recipients of HSC + CD8+/TCR−CD56dim/- FC exhibited persistent donor chimerism in PB (9.1% ± 6% vs. 3.8% ± 3.5%) (P <0.05) and significantly higher levels of donor chimerism in spleen (26.3% ± 11.8% vs. 12.3% ± 9.8%) (P <0.05) and BM (11.6% ± 4.8% vs. 2.9% ± 1.3%) (P <0.05) compared to recipients of HSC alone. Our data indicate that CD8+/TCR−/CD56dim/- FC facilitate homing of human HSC or progenitors and enhance engraftment of human HSC in NSG recipient mice. Disclosures: Bozulic: Regenerex, LLC: Employment. Ildstad:Regenerex, LLC: Equity Ownership.

Solannm lyratum extract affected immune response in normal and leukemia murine animal in vivo

2010 ◽  
Vol 29 (5) ◽  
pp. 359-367 ◽  
Author(s):  
Jai-Sing Yang ◽  
Chia-Chun Wu ◽  
Chao-Lin Kuo ◽  
Chin-Chung Yeh ◽  
Fu-Shin Chueh ◽  
...  
Keyword(s):  
Immune Responses ◽  
Mononuclear Cells ◽  
Nk Cell ◽  
Folk Medicine ◽  
Cell Activity ◽  
Colon Adenocarcinoma ◽  
Target Cells ◽  
Peripheral Blood Mononuclear ◽  
Macrophage Phagocytosis

Solanum lyratum Thunberg (Solanaceae) has been used as a folk medicine for treating liver, lung and esophagus in the Chinese population. Our previous studies have shown that the crude extract of S. lyratum Thunberg (SLE) induced apoptosis in colo 205 human colon adenocarcinoma cells; however, there is no report to show SLE affect immune responses in vivo. In this study, the in vivo effects of SLE on leukemia WEHI-3 cells and immune responses such as phagocytosis and natural killer (NK) cell activity in normal and leukemia mice were investigated. The SLE treatment decreases surface markers of CD3 and Mac-3 in normal and leukemia mice but promoted the cell markers of CD19 and CD11b in normal mice and CD11b in leukemia mice indicating that the precursors of T cells was inhibited and B cells and macrophage were promoted. The SLE treatment promoted the activity of macrophage phagocytosis in the peripheral blood mononuclear cells (PBMC) and peritoneal cells from normal and leukemia mice. The results also showed that NK cells from the normal and leukemia mice after treatment with SLE can kill the YAC-1 target cells. Therefore, the SLE treatment increased macrophage and NK cell activities. These consistent results indicate SLE could be a potent immune responses agent.

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