Myeloid Impairment Contributes to Immunoparesis in Multiple Myeloma but Not MGUS

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1831-1831
Author(s):  
Alessandra Romano ◽  
Nunziatina Parrinello ◽  
Calogero Vetro ◽  
Piera La Cava ◽  
Annalisa Chiarenza ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Flow Cytometry ◽  
T Cell ◽  
T Lymphocytes ◽  
Peripheral Blood ◽  
Healthy Subjects ◽  
Conflicts Of Interest ◽  
Myeloid Suppressor Cells ◽  
Hla Dr ◽  
Healthy Donors

Abstract Abstract 1831 Introduction In Multiple Myeloma (MM), but not in the monoclonal gammopathy of unknown significance (MGUS), the immune function is impaired as consequence of an immunologically hostile microenvironment and cellular defects, including reduction of immuno-surveillance and T-cell immunoparesis. We conducted an study focused on the myeloid compartment in MM, and its role in the progression from MGUS to MM. Methods Between January 2009 and April 2011 peripheral blood obtained from 60 consecutive newly diagnosed MM and 70 MGUS plus 30 healthy subjects was studied for evaluation of myeloid subpopulations and lymphoid paresis. Myeloid dysfunction was evaluated as percentage and absolute count of circulating myeloid suppressor cells (MDSC) in peripheral blood assessed by flow cytometry as follows: im-MDSC (CD34+/CD11b+/CD13+/CD14-/ HLA-DR-/CD45+), neutrophilic-like N-MDSC (CD11b+/CD13+/CD15+/CD14-/HLA-DR-/Lin-) and monocytic-like mo-MDSC (CD14+/HLA-DRlow/-). Myeloid function was evaluated by phagocytic activity using a commercially available kit (Phagotest R). Further, we investigated whether MM-neutrophils were able to induce anergy in T-cells. Neutrophils isolated from healthy donor (N=6), MGUS (N=3) or MM (N=6) peripheral blood (PB) were co-cultured with T-lymphocytes obtained from healthy donors. Expression of markers of activation in response to stimulation with PHA-P for 2 hours was assessed by flow cytometry as antigen density expressed as normalized mean of fluorescence intensity (N-MFI) of CD71 at 48 hours. Results The capability of phagocytosis of in neutrophils and monocytes from MM patients at diagnosis was significantly reduced compared to healthy subjects (p<0.001) and MGUS (p<0.0001). While the mature suppressive N-MDSC subset was not increased in MGUS and MM patients, the mo-MDSC subpopulation showed an increasing trend from healthy donors through MM (p=0.06) and the im-MDSC subset was significantly higher in MM vs healthy (p=0.002) and MGUS (p=0.001). After PHA-P stimulation, expression of CD71 (a marker of activation) in normal T-lymphocytes was increased (2954 ± 240.6 arbitrary units, au), and it was reduced (751.3 ± 30.48 au, p=0.0001) when co-coltured with MM-neutrophils, while no differences were evident in co-colture with MGUS- (2783 ± 206.1 au, p=0.61) or healthy donors-neutrophils (2588 ± 135.4, p=0.38). Conclusion Taken together, our findings suggest that in MM but not in MGUS there is a myeloid cell dysfunction that is correlated to impairment of T- cell arm. These alterations may have a role in the development of MM. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Pomalidomide Promotes the Maturation of Healthy Donors and Multiple Myeloma Patients Derived-Dendritic Cells

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 4702-4702
Author(s):  
Xi Wang ◽  
Feifei Che ◽  
Wanjun Cao ◽  
Zichen Ye ◽  
Hong Zheng ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Dendritic Cells ◽  
Peripheral Blood ◽  
Conflicts Of Interest ◽  
Control Group ◽  
Incubation System ◽  
Hla Dr ◽  
Healthy Donors ◽  
Tnf Α ◽  
Significant Difference

Abstract Abstract: Objective To investigate the effect of pomalidomide on the maturation of monocyte derived dendritic cells (moDCs) from healthy donors (HDs) and multiple myeloma (MM) patients. Method Peripheral blood mononuclear cells (PBMCs) were isolated from the peripheral blood of HDs and MM patients. The generation of moDCs from monocytes in PBMCs was conducted by the incubation of 7 days in a medium consisting of RPMI 1640 medium, 5% human serum, 800U/mL GM-CSF, 500U/mL IL-4, 100U/mL penicillin and 0.1mg/mL streptomycin. During this period, the incubation system was administrated with pomalidomide at 10 µM or 1×PBS as the control group. On the 8 th day, cells were harvested, and the immunophenotyping of cells were analyzed by the flow cytometry. The CD80+CD86+ cell population in total cells is gated as DCs in the FACS analyzing system. Then, the median fluorescence intensity (MFI) of surface markers CD40 and HLA-DR as well as the proportion of CD40+ DCs and HLA-DR+ DCs in total DCs were analyzed respectively. In addition, supernatant from the incubation system with or without pomalidomide administration was collected and detected for the concentration of cytokines IL-12, TNF-α and MIP-1α. Results The proportion of CD80+CD86+ cells in total cells was higher in HD group (n=15) than in MM patient group (n=11), but there was no statistically significant difference (93.49%±6.43% vs 77.04%±27.17%, P=0.094). When analyzing all the HD-derived moDCs (n=15), pomalidomide significantly enhanced the MFI of CD40 (P=0.003) and HLA-DR (P=0.040) on moDCs when compared with the control group. Meanwhile, the proportion of CD40+ DCs (P=0.008) and HLA-DR+ DCs (P=0.032) in total DCs was significantly higher in pomalidomide group than in control group. When analyzing all MM patients-derived moDCs (n=11), pomalidomide significantly enhanced the MFI of CD40 (P=0.047) and HLA-DR (P=0.006) on moDCs when compared with the control group. Meanwhile, the proportion of HLA-DR+ DCs in total DCs was significantly higher in pomalidomide group than in the control group (P=0.000). Moreover, pomalidomide treated HD-derived moDCs (n=8) produced 192% IL-12 (P=0.020), 110% TNF-α (P=0.006) and 112% MIP-1α (P=0.055) of untreated moDCs. However, when analyzing MM patients-derived moDCs (n=10), the expression of IL-12 (P=0.458), TNF-α (P=0.377) and MIP-1α (P=0.248) from moDCs showed no significant difference between pomalidomide group and the control group. Conclusions Pomalidomide at 10 µM can promote the maturation of both HD-derived moDCs and MM patients-derived moDCs. Pomalidomide shows potential to be a DC adjuvant for DC-based therapeutic strategies, such as DC vaccine and DC cell-therapy in MM. Key words: Pomalidomide; Monocyte derived Dendritic Cells; Multiple Myeloma; DC Adjuvant Disclosures No relevant conflicts of interest to declare.

Identification of a new subset of myeloid suppressor cells in peripheral blood of melanoma patients and modulation by GM-CSF-based anti-tumor vaccine

2007 ◽  
Vol 25 (18_suppl) ◽  
pp. 21082-21082
Author(s):  
P. Filipazzi ◽  
R. Valenti ◽  
V. Huber ◽  
M. Iero ◽  
L. Pilla ◽  
...  
Keyword(s):  
T Cell ◽  
Peripheral Blood ◽  
Tumor Vaccine ◽  
Suppressor Cells ◽  
Suppressive Activity ◽  
Gm Csf ◽  
Myeloid Suppressor Cells ◽  
Hla Dr ◽  
Healthy Donors ◽  
Melanoma Patients

21082 Background: Phenotypic and functional features of myeloid suppressor cells (MSC), known to serve as critical regulators of anti-tumor T cell responses in tumor-bearing mice, are still poorly defined in human cancers. Here we analyzed myeloid subsets with suppressive activity present in peripheral blood of metastatic melanoma patients (MM) and evaluated their modulation by a GM-CSF-based anti- tumor vaccine. Methods: Stage IV AJCC MM patients (n=16) vaccinated with autologous tumor-derived heat-shock protein peptide complexes gp96 (HSPPC-96) and low dose GM-CSF provided pre- and post-treatment whole blood samples. Peripheral blood mononuclear cells (PBMC) were analyzed by flow cytometry, separated into cellular subsets and used for in vitro proliferation assays. PBMC from stage- matched melanoma patients (n=12) treated with non-GM-CSF-based vaccines (i.e. HSPPC-96 alone or IFNa/melanoma-derived peptides) or gender and age-matched healthy donors (n=16) were also analyzed for comparison. Results: The lack or low levels of HLA-DR expression was found to identify a CD14+ cell subset with high suppressive activity on lymphocyte proliferation and functions. CD14+HLA-DR-/lo cells were significantly expanded in all MM patients, while undetectable in healthy donors. Suppressive activity was mediated by TGFβ, as suggested by functional experiments with neutralizing specific antibodies. In contrast, no involvement of arginase and iNOS pathways could be detected. CD14+HLA-DR-/lo cells, as well as spontaneous ex- vivo release and plasma levels of TGFβ, were augmented after administration of the HSPPC-96/GM-CSF vaccine. Interestingly, the expansion of suppressive CD14+ monocytes was associated to the inability to mount a significant CD8-mediated T cell response upon vaccination. On the other hand, no quantitative or qualitative enhancement of the CD14+HLA-DR-/lo suppressive cell population was observed in patients receiving a non-GM-CSF based vaccine. Conclusions: CD14+HLA-DR- /lo cells exerting TGFβ-mediated immune suppression may represent a new subset of myeloid suppressive cells, potentially expandable by the administration of GM-CSF-based vaccines in melanoma patients. [Table: see text]

A Population of Suppressive Monocytes Inhibiting T Cell Proliferation and Dendritic Cell Differentiation in Relapsed Non-Hodgkin’s Lymphoma

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 808-808
Author(s):  
Yi Lin ◽  
Peggy A Bulur ◽  
Michael P. Gustafson ◽  
Thomas E. Witzig ◽  
Allan B. Dietz
Keyword(s):  
Cell Proliferation ◽  
T Cell ◽  
Peripheral Blood ◽  
Hodgkin’S Lymphoma ◽  
Hodgkin's Lymphoma ◽  
T Cell Proliferation ◽  
Hla Dr ◽  
Healthy Donors ◽  
Non Hodgkin's Lymphoma ◽  
Kinase Phosphorylation

Abstract Despite advances in treatments for patients with non-Hodgkin’s lymphoma (NHL), the relapse rates remain high and 40% of diffuse large B-cell NHL (DLBCL) patients die of disease. New therapies to augment the host anti-tumor immune response are needed. Reports of graft-versus-lymphoma responses in patients who have received allogeneic hematopoietic cell transplant indicate a role for cellular immunotherapy. However, these patients have variable levels of immunodeficiency which may impact the efficacy of cellular therapy. To study this we first evaluated the cellular immune status of patients with relapsed NHL. Proliferation of peripheral blood mononuclear cells (PBMNC) stimulated with anti-CD3/CD28 beads was reduced by more than 3.5 folds for patients with DLBCL (n = 3) compared to that of age-matched healthy donors (n = 5; p = 0.02). Removal of monocytes from PBMNC by use of anti-CD14 immunomagnetic beads restored proliferation to that of healthy donors. Further, monocytes from these patients were deficient in stimulating allogeneic T cell proliferation by 3 folds compared to monocytes from healthy donors (n = 3 NHL; n = 8 normal; p < 0.01). Peripheral blood from 12 NHL patients (9 DLBCL; 1 grade 3 follicular lymphoma; 2 composite) and 12 age-matched healthy donors were characterized by flow cytometry to determine the phenotype of these suppressive monocytes. There was no difference in the % monocytes in the blood between NHL patients and healthy donors; however, NHL patients had elevated % monocytes with a suppressive phenotype (CD14+HLA-DRneg) compared to normals (NHL 38.9 ± 4.93%; normal 8.3 ± 2.15; p < 0.0001). This phenotype is distinct from other myeloid suppressors (Lin-CD33+HLA-DR-) or non-classical monocytes (CD16+), neither of which was different in numbers between NHL and normal donors. This suggests that the CD14+HLA-DRneg monocytes are responsible for the observed T cell suppression. To further characterize the function of these cells, we cultured purified CD14+ monocytes from NHL and compared their differentiation capacity with those from normal donors. The percentage of CD14+HLA-DRneg monocytes in initial ex vivo culture was inversely correlated with the percentage of pure, mature dendritic cells (mDC) generated with TNF-a and PGE2 as maturation factors (CD80+CD83+; n = 9; p = 0.015). As CpG oligonucleotides are also capable of immune stimulation and have some anti-tumor activity in clinical trials, we investigated the effect of CpG on mDC differentiation in NHL. In healthy donors, maturation of monocytes with CpG yielded highly pure mDC (90.7 ± 2.15%, n = 3). However, preliminary results of mDC yield from monocytes of NHL patients matured with CpG was only 22.6 ± 11.2% (n = 2). This data suggests alternative signaling pathways of these suppressive monocytes. Preliminary analysis of a proteome array for 46 kinase phosphorylation sites from 37 proteins in 2 NHL and 3 healthy donors suggest changes in phosphorylation of 17 protein kinases for CD14+HLA-DRneg compared to CD14+HLA-DR+ cells. Functional correlation of these protein kinase phosphorylation changes is needed to definitively target the pathway characteristic of CD14+HLA-DRneg monocytes. Finally, we have identified a serum-free culture method that can consistently generate highly pure mDC from monocytes of NHL patients (93.0 ± 3.6%, n = 9) and is readily adaptable to good manufacturing practice for clinical use in immunotherapy. Taken together we have described for the first time a population of CD14+HLA-DRneg monocytes that is a significant source of immunosuppression in NHL patients and are beginning to target methods of overcoming this suppression.

Lenalidomide IS Able to Restore Immune SYSTEM IN MULTIPLE MYELOMA PATIENTS.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4909-4909 ◽  
Author(s):  
Annalisa Chiarenza ◽  
Nunziatina Parrinello ◽  
Piera La Cava ◽  
Eleonora Spina ◽  
Daniele Tibullo ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Immune System ◽  
T Cell ◽  
T Lymphocytes ◽  
Mononuclear Cells ◽  
Conflicts Of Interest ◽  
Gradient Centrifugation ◽  
Target Cells ◽  
First Line ◽  
Number Of Patients

Abstract Abstract 4909 LENALIDOMIDE IS ABLE TO RESTORE IMMUNE SYSTEM IN MULTIPLE MYELOMA PATIENTS Annalisa Chiarenza, Nunziatina Parrinello, Piera La Cava, Eleonora Spina, Daniele Tibullo, Cesarina Giallongo, Maide Cavalli, Alessandra Romano, Paolo Fiumara, Giuseppe A. Palumbo, Francesco Di Raimondo Background Multiple myeloma (MM) is a malignant plasma-cell proliferative disorder associated with dysfunctional T-cell responses. The immunomodulatory Thal derivative (IMiD) CC-5013 (lenalidomide) appears to be a promising agent for the treatment of myeloma. Although the exact antitumor mechanism of action of lenalidomide is unknown, a number of mechanisms are postulated to be responsible for it's activity (inhibition of angiogenesis, direct antiproliferative and proapoptotic effects on MM cells, suppression of pro-inflammatory cytokines, modulation of myeloma-stromal cells adhesive interactions). In addition, it has been demonstrated that lenalidomide in vitro is able to enhance T cell proliferation and to promotes ADCC. In this study we evaluated if MM patients have a deficit of T-reg (CD4+, CD25+, and FOXP3+) and of T lymphocytes bearing CD200 (a tolerogenic molecule) and the effect of lenalidomide treatment on these parameters. In addition, we investigated whether lenalidomide could improve ex vivo the ADCC against myeloma cells. Materials and methods Eight patients with previously untreated MM (median age 56 years) were treated with lenalidomide plus dexamethasone as first line therapy. Lenalidomide was given orally 25 mg daily on days 1 to 21 of a 28-day cycle. Dexamethasone was given orally 40 mg daily on days 1, 8, 15, 22 of each cycle. All patients were evaluable for response and toxicity. Peripheral blood mononuclear cells (PBMNc) were obtained from MM patients using density gradient centrifugation (Fycoll) under sterile condictions, at the beginning of treatment and after 4 cycles of therapy. The percentage of T-reg (CD4+CD25+FOXP3+) and the expression of CD200 on T- lymphocytes were evaluated by cytometry. Twelve healthy subjects were used as control. Moreover, PBMNc (effector cells, E) were incubated with MM cells line ARH-77 (target cells, T), previously labelled with CFDA,SE (carboxyfluorescein diacetate, succinimidyl ester) as a tracing fluorescent marker, in culture medium (RPMI-1640, 10%FCS, 1%penicillin/streptomycin) at different concentration (T/E ratio 1:20, 1:40). After 18-24 h co-colture cells were analyzed by flow cytometry and MM plasma cells cytotoxicity was calculated as the percentage of positive CFDA,SE/propidium cells. Myeloma cell viability was determined by tripan blue esclusion and apoptosis was also evaluated using Annexin V/propidium assay. Two MM patients treated in first line with a combination of Velcade, Thalidomide and Dexamethasone (VTD) were used as control and the experiments were performed in duplicate. Results MM patients have a significantly lower rate of CD4+/CD25+/FOXP3+ and CD200+/CD3+ than normal (28,3±14,9/mmc and 37,8±24,7 /mmc vs 79,3±27,8 and 79,5± 48,9)(p=0,0001 and p=0,01 respectively). In our study, lenalidomide treatment resulted in an increase both of Treg cells and T-lymphocytes espressing CD200. This improvement is not statistically significant probably due to the low number of patients examined (tab I). More important, we observed that PBMC derived from patients treated with lenalidomide showed an increase ability to kill a target MM cell line compared to PBMC collected at diagnosis (CFDA,SE/propidium cells 11% vs 68%). This effect was more prominent in patients treated with lenalidomide than in MM patients treated with VTD (CFDA,SE/propidium cells 12% vs 39%), Fig.1. Conclusions Our data emphasize the role of lenalidomide in modulating the endogenous tumor-specific immune response and underline the anti-myeloma activity of these new class of drugs. Disclosures No relevant conflicts of interest to declare.

Morphologic and Phenotypic Features of Concurrent Clonal T-Cell Large Granular Lymphocytic Expansion and Myelodysplastic Syndrome

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2803-2803
Author(s):  
Xiaohui Zhang ◽  
Lynn Moscinski ◽  
John M. Bennett ◽  
Reza Setoodeh ◽  
Deniz Peker ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
T Cell ◽  
Peripheral Blood ◽  
Low Grade ◽  
Gene Rearrangements ◽  
High Grade ◽  
Bone Marrow Cellularity ◽  
Hla Dr ◽  
Marrow Cellularity

Abstract Abstract 2803 Myelodysplastic syndrome (MDS) and T-cell large granular (T-LGL) leukemia are both bone marrow failure disorders. It has been reported in a small number of cases that clonal T-LGL proliferation or leukemia can coincidentally occur with MDS. Also, clonal CD8+/CD57+ effector T cells expansion was detected in as many as 50% of MDS bone marrows [Epling-Burnette, 2007]. How clonal LGL cells that reside in the bone marrow interfere with hematopoiesis remains unclear, particularly in the setting of MDS. We analyzed the clinicopathological features of concomitant MDS and T-LGL, and evaluated bone marrow status for lineage or pan-hypoplasia in these patients. Design: Clinical and pathologic data from patients with a diagnosis of MDS and flow cytometry performed on the peripheral blood between 1/2005 and 12/2009 were reviewed. The concurrent bone marrow biopsies from each patient at the time of flow cytometric analysis were reviewed by two hematopathologists. Bone marrow cellularity, lineage hypoplasia (M:E >5: 1 or <1:2) were documented. Peripheral lymphocyte count and CD3+/CD57+ and CD8+/CD57+ populations by flow cytometry were calculated and T cell gene receptor (TCR) rearrangements were correlated. Results: We performed LGL flow cytometry panel on 76 MDS patients (high grade MDS, n=23; low grade, n=54), as well as TCR gene rearrangements, and identified clonal T-LGL cells in peripheral blood of 37 patients (48.7%), including 15 high grade MDS (40.5%, RAEB-I and RAEB-II), and 22 low grade MDS (59.4%), including RCMD(13), RA(1), RS(1), RCMD-RA(1), RCMD-RS (2), 5q- MDS(1), and MDS unclassifiable(3). The immunophenotype of the T-LGL cells was typically CD3+/CD57+/CD7 dim+/CD5 dim+/CD8+ with variable CD11b,CD11c, CD16, CD56 and HLA-DR. A frequent variant in these MDS patients was CD11b-,CD11c -, CD16+/−, CD56+/−, HLA-DR- and CD62L+.The TCRβ or/and TCRγ gene rearrangements were positive in 35 of the 38 cases (92.1%). The peripheral blood lymphocyte counts were 300–3820 cells/μL (1199±799 cells/μL); the CD3+/CD8+/CD57+ T-LGL cell counts were 30–624 cells/μL (229±154 cells/μL). In comparison, the remaining 39 patients with non-clonal T-LGL included 11 high grade MDS cases, and 28 low grade MDS cases. The peripheral blood lymphocyte counts were 308–2210 cells/μL (1030±461 cells/μL). CD3+/CD57+ cells were 1–425 cells/μL (105±98 cells/μL). There was no identifiable phenotypic features suggestive of clonal T-LGL cells such as dim CD5 and/or dim CD7 with aforementioned aberrant expressions on T-cells, although 7 of the 39 cases had TCRβ or/and TCRγ gene rearrangements. Thirty healthy donors were included for controls with absolute lymphocyte counts of 2136±661 cells/μL and baseline CD3+/CD57+ cells of 162±109 cells/μL. All showed no clonal LGL phenotype and negative TCR gene rearrangements. Since the presence of T-LGL cells may impair bone marrow hematopoiesis, we examined if there are bone marrow status differences between these two groups. All the bone marrows were obtained at diagnosis or not on chemotherapy. The bone marrow cellularity of the MDS patients with clonal T-LGL ranged from <3% to almost 100%, averaging 56%, with 8 cases with dramatic hypocellularity (<3%-20%), while the bone marrow cellularity of the MDS patients without clonal T-LGL ranged from 20% to 90%, averaging 62%, with only 2 cases with mild hypocellularity (20% in 73- and 65-year-old). In addition, among MDS patients with clonal T-LGL cells, 14 of 37 (37.8%; 5 high grade, and 9 low grade) bone marrows had certain lineage hypoplasia, including 3 cases of trilineal hypoplasia, 9 cases of erythroid hypoplasia, and 2 cases of myeloid hypoplasia. In contrast, among 39 MDS patients without T-LGL, there were only 1 bone marrow with trilineal hypoplasia and 3 others with erythroid hypoplasia (10.2%). The difference between the two groups was statistically significant (p=0.004, chi square test). In conclusion, our studies indicate that clonal T-LGL cells expansion is a fairly common finding in high grade as well as low grade MDS. The clonal T-LGL cells have more than one variant immunophenotypes and are typically positive for TCR gene rearrangements. Additionally, we observed that the clonal LGL cells present in MDS bone marrows could be associated with lineage hypoplasia, which, in this respect, might impact clinical treatment. Disclosures: No relevant conflicts of interest to declare.

Recovery Of Myeloid Derived Suppressor Cell Subsets Following Allogeneic Hematopoietic Stem/Progenitor Cell Transplantation

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4617-4617
Author(s):  
Qingdong Guan ◽  
Anna Blankstein ◽  
Anjos Karla ◽  
Oleksandra Synova ◽  
Marie Tulloch ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cell ◽  
Airway Inflammation ◽  
Progenitor Cell ◽  
Hematopoietic Stem ◽  
Post Transplant ◽  
Inflammatory Conditions ◽  
Hla Dr ◽  
Healthy Donors ◽  
Immature Myeloid Cells

Myeloid derived suppressor cells (MDSCs) are a heterogeneous population of immature myeloid cells that expand during many inflammatory conditions and malignancies. MDSCs may play an important role following allogeneic hematopoietic stem/progenitor cell transplant (HSCT). MDSCs suppress T-cell, B-cell and dendritic cell responses by a number of mechanisms, including promoting regulatory T cell expansion and producing soluble mediators such as Arginase 1 (Arg-1) and iNOS. MDSCs are divided into two subsets: monocytic (M-MDSCs) and granulocytic (G-MDSCs). MDSC morphology and function differ in various tissues under different inflammatory conditions. In a murine asthma model, M-MDSCs inhibit airway inflammation, but the other subset of MDSCs exacerbated airway inflammation. In a sepsis model, MDSCs exaggerated inflammation in the early stage, but suppressed inflammation in the later stage of sepsis. As the early post-transplant period is characterized by the rapid expansion of immature myeloid cells, we postulated this time period may also be a time when MDSCs might play a major role in modulating immune recovery post-transplant, and aid in the development of immune regulatory networks potentially important in the pathophysiology of graft-versus-host disease (GVHD). In nine patients undergoing allogeneic HSCT, peripheral blood was drawn on the day prior to the start of conditioning, days +4-5, +7-9, +14-16, +21-23, +27-29 and +80-100 post HSCT. White blood cells were quantified, red cell depleted using HetaSep (Stem Cell Technologies), then stained with fluorescence-labelled antibodies against CD45, CD15, CD14, HLA-DR, CD33 and CD66b and analyzed by flow cytometry for MDSC subsets. The soluble mediators iNOS and Arg-1were evaluated by intracellular staining for iNOS and Arg-1 and analyzed by flow cytometry. Four of the nine patients developed acute GVHD (II-IV) and/or extensive chronic GVHD. Early recovery of CD33+CD14+HLA-DR-/low M-MDSCs and CD33+CD15+CD66b+ G-MDSCs was seen post-transplant. Compared to healthy donors, the percentage of M- and G-MDSCs was increased by 3 weeks post-transplant. Interestingly, the patients who went on to develop GVHD had lower percentage and number of M-MDSCs, but inversely had higher numbers of G-MDSCs by day +27-29 and day +80-100 post-transplant (Fig. 1). When compared with healthy donors, the expression of Arg-1 in G-MDSCs, a measure of activation of MDSCs, was increased in patients pre- and post-HSCT, especially at day +80-100; while there was no difference seen iNOS expression in G-MDSCs (Fig 2). The expression of Arg-1 and iNOS in M-MDSCs was increased pre-transplant but fell by day +80-100 post HSCT (Fig 2). Taken together, our pilot data indicates that both M- and G- MDSCs recover early post HSCT and may contribute to the pathophysiology of GVHD. Patients with lower numbers of M-MDSCs and higher numbers of G-MDSCs at earlier time points post-transplant might be at greater risk for developing GVHD. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Myeloidderived peripheral blood suppressor cells at haematopoietic stem cell mobilisation in multiple myeloma patients

2021 ◽  
Vol 66 (2) ◽  
pp. 218-230
Author(s):  
T. A. Aristova ◽  
E. V. Batorov ◽  
V. V. Sergeevicheva ◽  
S. A. Sizikova ◽  
G. Yu. Ushakova ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Stem Cell ◽  
Partial Response ◽  
Peripheral Blood ◽  
Plasma Cells ◽  
Suppressor Cells ◽  
Complete Response ◽  
Haematopoietic Stem Cell ◽  
Hla Dr ◽  
Healthy Donors

Introduction. Multiple myeloma (MM) is a B-cell malignancy with clonal expansion of plasma cells in bone marrow. Highdose chemotherapy with autologous haematopoietic stem cell transplantation is among main consolidation therapies in MM. Myeloid-derived suppressor cells (MDSCs) are immature myeloid-accompanying cells able to suppress the immune response. The administration of granulocyte colony stimulating factor (G-CSF) to mobilise haematopoietic stem cells (HSCs) increases the MDSC count in peripheral blood (PB).Aim — to study MDSC subsets in PB of remission MM patients and their incidence dynamics at HSC mobilisation.Methods. The study surveyed 35 MM patients prior to and after HSC mobilisation. The counts of granulocytic (G-MDSCs; Lin–HLA-DR–CD33+ CD66b+), monocytic (М-MDSCs; CD14+ HLA-DRlow/–) and early MDSCs (E-MDSCs; Lin–HLA-DR– CD33+ CD66b–) were estimated in flow cytometry.Results. Remission MM patients differed from healthy donors in higher relative counts of G-MDSCs (Lin–HLA-DR– CD33+ CD66b+) and increased relative and absolute counts of М-MDSCs (CD14+ HLA-DRlow/–). М-MDSCs significantly outnumbered G-MDSCs. MDSC subset counts were elevated in complete response (CR) and very good partial response (VGPR), as well as in partial response (PR). Higher relative MDSC counts were associated with greater pretreatment (2–3 lines of chemotherapy). After HSC mobilisation with cyclophosphamide 2–4 g/m2 + G-CSF (filgrastim 5 μg/kg/day), the median relative E-MDSC and M-MDSC counts increased by 2.3 and 2.0 times, respectively, while the relative G-MDSC count raised 46-fold perturbing the MDSC subset balance.Conclusion. Remission MM patients had the increased relative G-MDSC and both relative and absolute M-MDSC counts compared to donors. A greater patient pretreatment was associated with higher relative G-MDSC counts. Treatment response (CR/VGPR vs. PR) was not coupled with MDSC count variation. The G-CSF-induced HSC mobilisation entailed a significant expansion of all three MDSC subsets in PB.

Myeloid-Derived Suppressor Cells in Patients with Hodgkin Lymphoma.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3662-3662
Author(s):  
Nunziatina Parrinello ◽  
Piera La Cava ◽  
Daniele Tibullo ◽  
Cesarina Giallongo ◽  
Orazio Di Bartolo ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Hodgkin Lymphoma ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Conflicts Of Interest ◽  
Suppressor Cells ◽  
Cell Number ◽  
Myeloid Derived Suppressor Cells ◽  
Peripheral Blood Mononuclear ◽  
Hla Dr

Abstract Abstract 3662 Poster Board III-598 Background Immune suppression and angiogenesis are mechanisms key to tumour growth and progression. Myeloid-derived suppressor cells (MDSC) are a heterogeneous population of cells of myeloid origin and include immature macrophages, dendritic cells (DC) and other myeloid cells. In mice are phenotypically characterized as CD11b+Gr-1+ cells, while in human they have an immature phenotype, including lineage negative (Lin-), CD14-, HLA-DR-, CD15+, CD34+, CD11b+, CD33+, and CD13+ cells. MDSC reduce activated T-cell number and inhibit their function through different mechanisms including: L-arginine metabolism, nitric oxide (NO), up-regulation of reactive oxygen species (ROS), and secretion of immunosuppressive cytokines. MDSC also promote tumor-dependent angiogenesis as well as tumor metastasis. Their accumulation has been described in patients affected by some solid tumors but information on haematological neoplasms are lacking. Our study investigated by flow cytometry the presence of MSDC in the peripheral blood of patients affected by Hodgkin Lymphoma (HL). Methods We studied 14 patients with HL at diagnosis and 10 age-matched healthy controls (HC). Peripheral blood mononuclear cells were stained with the following monoclonal antibodies:CD11b, CD13, CD14, CD34, CD45, for 20 minutes at room temperature. After lysing red cells, cells were analyzed by flow cytometry. Results we observed a increased number of MDSC (CD11b+,CD13+,CD34+,CD14-, CD45+) in the peripheral blood of patients with HL compared to HC (13,37 ± 17,77 ×109/l vs 1,45± 0,98 ×109/l, p=0,0007). We also found that patients with advanced-stage Hodgkin disease (III and IV) have higher number of MDSC, compared to patients stage I and II (p= 0,04). Conclusion These data suggest a role for myeloid-derived suppressor cells in promoting tumor cell proliferation in hodgkin lymphoma. Disclosures: No relevant conflicts of interest to declare.

Identification of a New Subset of Myeloid Suppressor Cells in Peripheral Blood of Melanoma Patients With Modulation by a Granulocyte-Macrophage Colony-Stimulation Factor–Based Antitumor Vaccine

2007 ◽  
Vol 25 (18) ◽  
pp. 2546-2553 ◽  
Author(s):  
Paola Filipazzi ◽  
Roberta Valenti ◽  
Veronica Huber ◽  
Lorenzo Pilla ◽  
Paola Canese ◽  
...  
Keyword(s):  
Metastatic Melanoma ◽  
Peripheral Blood ◽  
Suppressor Cells ◽  
Suppressive Activity ◽  
Gm Csf ◽  
Myeloid Suppressor Cells ◽  
Hla Dr ◽  
Healthy Donors ◽  
Melanoma Patients ◽  
Macrophage Colony

Purpose Phenotypic and functional features of myeloid suppressor cells (MSC), which are known to serve as critical regulators of antitumor T-cell responses in tumor-bearing mice, are still poorly defined in human cancers. Here, we analyzed myeloid subsets with suppressive activity present in peripheral blood of metastatic melanoma patients and evaluated their modulation by a granulocyte-macrophage colony-stimulating factor (GM-CSF) –based antitumor vaccine. Patients and Methods Stage IV metastatic melanoma patients (n = 16) vaccinated with autologous tumor-derived heat shock protein peptide complex gp96 (HSPPC-96) and low-dose GM-CSF provided pre- and post-treatment whole blood specimens. Peripheral-blood mononuclear cells (PBMCs) were analyzed by flow cytometry, separated into cellular subsets, and used for in vitro proliferation assays. PBMCs from stage-matched metastatic melanoma patients (n = 12) treated with non–GM-CSF-based vaccines (ie, HSPPC-96 alone or interferon alfa/melanoma–derived peptides) or sex- and age-matched healthy donors (n = 16) were also analyzed for comparison. Results The lack of or low HLA-DR expression was found to identify a CD14+ cell subset highly suppressive of lymphocyte functions. CD14+HLA-DR–/lo cells were significantly expanded in all metastatic melanoma patients, whereas they were undetectable in healthy donors. Suppressive activity was mediated by transforming growth factor beta (TGF-β), whereas no involvement of the arginase and inducible nitric oxide synthase pathways could be detected. CD14+HLA-DR–/lo cells, as well as spontaneous ex vivo release and plasma levels of TGF-β, were augmented after administration of the HSPPC-96/GM-CSF vaccine. No enhancement of the CD14+-mediated suppressive activity was found in patients receiving non–GM-CSF-based vaccines. Conclusion CD14+HLA-DR–/lo cells exerting TGF-β–mediated immune suppression represent a new subset of MSC potentially expandable by the administration of GM-CSF–based vaccines in metastatic melanoma patients.

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.

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