Study On Immune Mechanism of Human Marrow Mesenchymal Stem Cells Adjusting Dendritic Cells for Treatment of Aplastic Anemia

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 5137-5137
Author(s):  
Yang Xiao ◽  
Leqin Zhang
Keyword(s):  
Flow Cytometry ◽  
T Lymphocytes ◽  
Surface Antigen ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Conflicts Of Interest ◽  
Human Peripheral Blood ◽  
Interleukin 4 ◽  
Peripheral Blood Flow ◽  
Inhibiting Effect

Abstract Abstract 5137 Objective (1)To explore whether MSC has inhibiting effect on the proliferation ofAA patients' Tcell;(2)To discuss whether MSC affects T cell's proliferation via adjusting the growth of DCs. Materials and Methods (1) MSC were separated and cultured in vitro. Cell morphology was observed and the cell surface antigen was determined by flow cytometry.(2) Peripheral blood mononuclear cells were extracted from 20 patients suffered AA and then T lymphocytes were separated by nylon fiber column. Flow cytometry was applied to determine the surface antigen and subpopulation of T cell.(3)mononuclear cells were separated from normal human peripheral blood. DCs were prepared under the culture condition of recombination human granulocyte-macrophage colony stimulating factor (GM-CSF) and recombinant human interleukin-4 (IL-4). After acquiring the mature DCs induced by LPS, the phenotype analysis of DCs before and after culture was examined by flow cytometry, respectively. Results (1) After co-culture of MSC and T lymphocytes from AA peripheral blood, flow cytometry showed that the ratio of D8+ in T cells reduced significantly from 38.7% to 29.7 % (p < 0.05), whereas the CD4+ ratio increased from 24.9% to 34.9% significantly (p < 0.05). Meanwhile, ELISA analysis indicated that the concentration of IL-2 and IFN-γ were significantly decreased from 38.9 and 38.5 ng/L to 6.8 and 6.6 ng/L, respectively (p < 0.05). However, IL-4 and IL-10 increased from 2.8 and 2.9 to 5.3 and 8.3 ng/L, respectively (p < 0.05).(2) After the induction of immature DCs by LPS, flow cytometry showed that the expression of CD1a increased from 2.4% to 68.4% in the treatment without MSC, while that of CD14+ decreased from 83.6% to 3.5% (p < 0.05).(3) After the co-culture of mature DCs and MSC, the expression of CD14+ increased from 5.8% to 62.8% when the expression of CD1a, CD83 and CD80 decreased from 48.6%, 60.8% and 50.2% to 30.7%,40.9% and 20.3%, respectively. Conclusions Our study shows that (1)MSC inhibits the proliferation of T lymphocytes from AA p-atients by regulating CD8+ and CD4+; (2)futher study indicates that MSC can inhibit the growth of DCs and reverse the status of DCs from matureness to immatureness; (3)thus suggests the possible mechanism of MSC's inhibiting effect as follows: MSC decreases the liveness by controlling the growth of DCs and further inhibits its proliferation. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Optimizing recovery of frozen human peripheral blood mononuclear cells for flow cytometry

PLoS ONE ◽  
2017 ◽  
Vol 12 (11) ◽  
pp. e0187440 ◽  
Author(s):  
Bo Langhoff Hønge ◽  
Mikkel Steen Petersen ◽  
Rikke Olesen ◽  
Bjarne Kuno Møller ◽  
Christian Erikstrup
Keyword(s):  
Flow Cytometry ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

scholarly journals Defective lymphocyte glycosidases in the macrophage activation cascade of juvenile osteopetrosis

Blood ◽  
1996 ◽  
Vol 88 (4) ◽  
pp. 1473-1478 ◽  
Author(s):  
N Yamamoto ◽  
VR Naraparaju ◽  
PJ Orchard
Keyword(s):  
T Lymphocytes ◽  
B Lymphocytes ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Serum Vitamin ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells ◽  
Beta Galactosidase

Generation of macrophage-activating factor requires a precursor protein, Gc protein (serum vitamin D3-binding protein), as well as participation of beta-galactosidase of inflammation-primed B lymphocytes and sialidase of T lymphocytes. The treatment of human peripheral blood mononuclear cells with an inflammatory lysophospholipid induced beta-galactosidase and sialidase activity of lymphocytes, leading to the generation of macrophage-activating factor and activation of monocytes/macrophages. However, lysophospholipid treatment of peripheral blood mononuclear cells from three infantile patients with osteopetrosis resulted in no significant activation of monocytes/macrophages. The lysophospholipid-inducible beta- galactosidase activity of B lymphocytes as well as that of the sialidase of T lymphocytes was found to be defective in these patients.

Dissecting Macrophage-Derived Microvesicles' Content and Function.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3785-3785
Author(s):  
Noura Ismail ◽  
Kara Batte ◽  
Leni Moldovan ◽  
Clay Marsh ◽  
Melissa Piper
Keyword(s):  
Flow Cytometry ◽  
Surface Antigen ◽  
Peripheral Blood ◽  
Conflicts Of Interest ◽  
Antigen Expression ◽  
Mononuclear Phagocytes ◽  
Gm Csf ◽  
Wide Range ◽  
Surface Antigen Expression ◽  
And Function

Abstract Abstract 3785 Microvesicles (MVs) are small membrane-bound vesicles released under normal homeostatic and stimulatory conditions by a wide variety of cell types. Microvesicles are collectively referred to as exosomes and microparticles which vary in size due to different cellular mechanisms responsible for their production. These microvesicles have a wide range of functions from facilitating communication to regulating cellular growth and differentiation. During their production; microvesicles become enriched in various molecules including proteins and nucleic acids. Previously, we have shown that plasma microvesicles derived from many cell lineages contain microRNAs (miRNAs). We also found that the majority of the peripheral blood microvesicles are derived from platelets while those originating from monocytic cells including macrophages represent the second largest population. Since microvesicles derived from mononuclear phagocytes are a large subpopulation in the plasma; we were interested in understanding their content and function. We hypothesized that the content and/or quantity of macrophage-derived microvesicles could induce the maturation of monocytes. To address our hypothesis, peripheral blood monocytes were treated in vitro for 4hr with GM-CSF; washed and cultured in media devoid of cytokines for 24 h then microvesicles were collected. Flow cytometry and electron)confocal microscropy were used to quantify and visualize microvesicles production. To examine the function of the microvesicles on macrophage maturation, the purified microvesicles were then cultured with freshly isolated monocytes. Macrophage differentiation was determined by cellular adherence using a crystal violet uptake assay and changes in surface antigen expression by flow cytometry. We also examined the genetic changes induced in monocytes incubated with the microvesicles compared to GM-CSF-treated cells. We found that freshly isolated monocytes treated with microvesicles from macrophages acquired phenotypic characteristics of a macrophage such as cellular adherence and surface antigen expression. We also found that treatment of naïve monocytes with the microvesicles induced molecular changes similar to GM-CSF treated monocytes. We found more than 7985 mRNAs that were similarly expressed between the two culture conditions. Notably, we observed the unique expression of 1324 and 1079 genes in the GM-CSF-treated compared to the microvesicle-treated cells, respectively. To begin dissecting the molecules contained in the microvesicles responsible for these changes, we performed mass-spectrometry and miRNA profiling. We observed the expression of miRs-223, -222,-191, -484, -016, -026a, and -155 in GM-CSF-derived microvesicles. Notably, these miRNAs were also expressed in the cells from which the microvesicles were released. We have begun bioinformatics analyses to predict whether the expression of the miRNAs may account for the decrease expression of specific genes in cells treated with the microvesicles that undergo differentiation. Many of the proteins found in the vesicles are important in facilitating protein:protein interactions and nucleic acid binding. Based on our observations; we postulate that microvesicles in areas of inflammation may contribute to the inflammatory response through the maturation of immune cells and activation of cells responsible for tissue repair. Thus, defining key components of this response may identify targets to regulate inflammation. Disclosures: No relevant conflicts of interest to declare.

Low Density Granulocytes in Patients after Allogeneic Hematopoietic Stem Cells Transplantation (allo-HSCT): A Distinct Class of Neutrophils in Systemic Alloimmunity

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 4614-4614
Author(s):  
Ekaterina Mikhaltsova ◽  
Valeri G. Savchenko ◽  
Larisa A. Kuzmina ◽  
Mikhail Drokov ◽  
Vera Vasilyeva ◽  
...  
Keyword(s):  
Lupus Erythematosus ◽  
Peripheral Blood ◽  
Hematological Malignancies ◽  
Mononuclear Cells ◽  
Conflicts Of Interest ◽  
Human Peripheral Blood ◽  
Unrelated Donor ◽  
Low Density ◽  
Hematopoietic Stem

Abstract Introduction It's generally considered that all alloimmune process such as acute graft-versus host disease (aGVHD) after allo-HSCT are mostly controlled by lymphocytes. The role of neutrophils in systemic alloimmunity after allo-HSCT is still illusive. In 1987 a distinct subset of proinflammatory, low-density granulocytes (LDGs) isolated from the peripheral blood mononuclear cell fractions of patients with system lupus erythematosus has been described. There is no LDG's in healthy donors. While the origin and role of LDGs still needs to be fully characterized, we try to describe this population in patients with hematological malignancies after allo-HSCT Patients and methods. Peripheral blood samples were collected in EDTA-tubes before allo-HSCT, on day +30,+60,+90 after allo-HSCT and at day of aGVHD from 47 patients with hematological malignancies (AML=22, ALL n=17, LPD=3, MDS =2; CML=2; 17 with active disease, 30 - in CR) after allo-HSCT (from matched unrelated donor n=34, from matched related donor n=13; MAC = 13, RIC=34). Isolation of mononuclear cells from human peripheral blood was made by standard protocol using Lympholyte®-M Cell Separation Media (Cedarlane Labs). The anti-CD66b-PE (Biolegend, USA) antibodies and FSC/SSC were used to determine LDGs cells as FSChigh \SSChigh \CD66b+. 100000 of cells were analyzed on a BD FACSCanto II (Becton Dickinson, USA). Results. Results of blood evaluation of 47 patients with hematological malignancies, whose blood was examined after allo-HSCT presented in table 1. Conclusion Despite the fact that we don't get significant differences. LDG's detection in allo-HSCT patients need further investigation. Table 1. Incidence of LDG after allo-HSCT in patients with and without aGVHD Table 1. Incidence of LDG after allo-HSCT in patients with and without aGVHD Disclosures No relevant conflicts of interest to declare.

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