We studied suppressor potential of myeloid-derived suppressor cells (MDSC) in multiple myeloma patients, including before and after mobilization of hematopoietic stem cells (HSC), by evaluating the ...expression of arginase-1 (Arg1), indolamine-2,3-dioxygenase (IDO), and PD-L1 in MDSC subsets. The study included 20 multiple myeloma patients in remission, 5 patients with progression, as well as 10 sex-and age-matched healthy donors. The expression of Arg1, IDO, and PD-L1 in circulating granulocytic MDSC (G-MDSC, Lin
–
HLA-DR
–
CD33
+
CD66b
+
), monocytic MDSC (M-MDSC, CD14
+
HLA-DR
low/–
), and early-stage MDSC (E-MDSC, Lin
–
HLA-DR
–
CD33
+
CD66b
–
) was evaluated by flow cytometry. Multiple myeloma patients in remission were characterized by reduced expression of Arg1 in M-MDSC in comparison with donors. The expression of Arg1 in M-MDSC depended on the number of induction therapy lines performed and was significantly lower in patients who received ⩾2 lines and responded with remission. Patients with multiple myeloma progression (resistant to therapy) showed significantly increased expression of Arg1 and PD-L1 in M-MDSC, as well as increased expression of Arg1 in E-MDSC. After G-CSF-induced mobilization of HSC, the content of circulating Arg1-expressing M-MDSC increased significantly. Considering the presence of MDSC in apheresis products, MDSC suppressive activity is discussed as a factor affecting the outcomes of autologous HSC transplantation in multiple myeloma patients.
The avoidance of immune surveillance by malignant plasma cells (PCs) in multiple myeloma (MM) is mediated by different mechanisms, among which an induction of T cell exhaustion and expansion of ...myeloid-derived suppressor cells (MDSCs) appear to play substantial roles, but it is still a lack of data on possible MDSC-mediated induction of T cell exhaustion. The aim of the present work was to evaluate possible relationship between frequencies of MM PCs, MDSCs and phenotypically exhausted PD-1
+
and TIM-3
+
T cells in bone marrow (BM) samples and peripheral blood (PB) of MM patients at various disease stages. Peripheral blood (n = 88) and BM samples (n = 56) were obtained from MM patients (newly diagnosed (n = 6), patients in remission (n = 71) and with progressive disease (n = 11)). Frequencies of T cells expressing checkpoint receptors PD-1 and TIM-3, polymorphonuclear MDSCs (PMN-MDSCs, Lin
-
CD14
-
HLA-DR
-
CD33
+
CD15
+
/CD66b
+
), monocyte MDSCs (M-MDSCs, CD14
+
HLA-DR
low/-
), early MDSCs (E-MDSCs, Lin
-
HLA-DR
-
CD33
+
CD15
-
/CD66b
-
), and MM PCs (CD45
dim
CD38
+
CD138
+
CD56
+
CD19
-
CD117
+
CD27
-
CD81
-
) were assessed with flow cytometry. Circulating and BM-resident PD-1
+
/TIM-3
+
T cell subsets, BM E-MDSCs, as soon as MM PCs and serum beta2-microglobulin (B2-M) levels were gradually increased in patients at different stages. Despite that, there were no associations between the markers of tumor load and the studied cell subsets. In patients in remission, BM PMN-MDSCs negatively correlated with CD4
+
T cells, CD4
+
PD-1
+
and CD8
+
TIM-3
+
T cell subsets; there were positive correlations between BM E-MDSCs and CD4
+
PD-1
+
TIM-3
+
cells and PB M-MDSCs and CD8
+
PD-1
+
and (as a trend) CD8
+
TIM-3
+
T cells. We found no associations for the samples of patients at diagnosis and with progression. We can conclude that a possible mutual influence of malignant PCs, MDSCs and PD-1
+
/TIM-3
+
T cells is nonlinear, especially during a manifest tumor growth at diagnosis and progression. The detected negative correlations between resident PMN- MDSCs and T cell subsets might be associated with MDSC suppressive function, affecting both predominantly activated PD-1
+
cells and exhausted TIM-3
+
subsets. The positive correlations between BM E-MDSCs and CD4+PD-1
+
TIM-3
+
cell subset and circulating M-MDSCs and PD-1
+
and TIM-3
+
CD8
+
T cells might confirm an ability of MDSCs to induce T cell exhaustion.
Myeloid-derived suppressor cells (MDSCs) play an important role in the immune response regulation in many pathologies, primarily in malignant tumors, but their role in the hematopoietic stem cell ...engraftment and the hematopoietic recovery after high-dose chemotherapy and autologous stem cell transplantation remains practically unexplored. This study is aimed at studying the correlation between the number of MDSC subpopulations and blood parameters at the stage of hematopoietic recovery after high-dose chemotherapy and autologous hematopoietic stem cell transplantation in patients with multiple myeloma (MM). Circulating MDSCs were assessed at the stage of leukopenia recovery (absolute leukocyte count in peripheral blood (PB) > 1 x 10
9
/L) by flow cytometry. The number of transplanted CD34
+
CD45
+
hematopoietic stem cells was 4.38 x 10
6
/kg (IQR (3.1—5.6) x 10
6
/kg). The duration of recovery from leukopenia varied from 8 to 18 days (Me 12 days). The number of MDSCs at the engraftment was not associated with the number of CD34
+
cells/kg in the graft. The relative number of monocytic MDSCs (M-MDSCs, CD14
+
HLA-DR
low/-
) directly correlated with the number of monocytes at the stage of recovery from leukopenia (R = 0.417, p = 0.002). Granulocytic MDSCs (PMN-MDSCs, Lin
-
HLA-DR
-
CD33
+
CD66b
+
) were characterized by an inverse correlation with the number of monocytes (R = -0.493, p = 0.0003) while the association with the absolute number of neutrophils was weak (R = 0.273, p = 0.048). The number of lymphocytes at the stage of recovery from leukopenia had an inverse correlation with PMN-MDSCs (R = -0.347, p = 0.014) and did not correlate with M-MDSCs. When analyzing the duration of leukopenia, an inverse correlation with this indicator was revealed for the percentage and absolute number of M-MDSCs (R = -0.347, p = 0.018 and R = -0.469, p = 0.0008, respectively). Multiple regression analysis showed dependence of the lymphopenia duration on the proportion of circulating M-MDSCs (p = 0.014) and the number of transplanted CD34
+
cells/kg (p = 0.032). According to the data of multivariate analysis of variance, the number of transplanted CD34
+
cells/kg and the number of M-MDSCs were significant factors for the duration of leukopenia. At the same time, such clinical parameters as the depth of response and minimal residual disease status before high-dose chemotherapy and hematopoietic stem cell transplantation, as well as the MM stage, did not affect the duration of hematopoietic recovery. Thus, the obtained results indicate the association of a higher number of M-MDSCs with a shorter duration of leukopenia after high-dose chemotherapy with autologous stem cell transplantation and indicate a positive role of M-MDSCs in hematopoietic recovery in the early post-transplant period in patients with MM.
T cells expressing checkpoint receptors PD-1, TIM-3 etc., are potential targets for monoclonal antibody immunotherapy in multiple myeloma (MM). However, checkpoint expressing T cell compartment ...includes different subsets, and their dysregulation following anti-checkpoint therapy can lead to the development of adverse events.
The aim of this study
was to evaluate activation markers – homeostatic cytokine receptors and transcription factors expressed by PD-1and TIM-3-positive T cells.
Material and Methods.
Relative counts of circulating PD-1and/or TIM-3-positive and negative T cells expressing common ɣ-chain cytokine receptors CD25, CD122, CD127, phosphorylated STAT5, and transcription factor EOMES associated with T cell exhaustion were studied using flow cytometry in 17 healthy donors, 22 MM patients with remission and 7 MM patients with progressive disease.
Results.
T cells expressing PD-1 and/or TIM-3 inhibitory checkpoint receptors in MM patients consisted of CD25+EOMESactivated cells, CD4+CD25+CD127-FOXP3+ regulatory T cells (Treg), CD4+CD25-EOMES+ dysfunctional cells. CD25+ T cells from healthy donors and MM patients, regardless of the expression of the studied checkpoint receptors, were EOMES-negative. No such association was found for CD122 and CD127 cytokine receptors. EOMES is a marker of T cell exhaustion for CD4+ T cells, but not for CD8+ T cells, in which it is more associated with activation. The proportion of CD4+ Tregs among circulating PD-1+ and TIM-3+ T cells was relatively low. A higher content of cytokine receptors in the population of TIM-3+ T cells may indicate the predominant involvement of TIM-3 in the control of homeostatic proliferation of mature T cells under lymphopenic conditions, while the expression of PD-1 may be more associated with the regulation of activation through T cell receptor. PD-1+ and/or TIM-3+ levels of activated, dysfunctional, and regulatory T cells can potentially be used to predict the safety and efficacy of targeted immunotherapy.
Maternal adaptation of the immune system aimed at limiting the immune response to fetal antigens is a necessary condition for a successful pregnancy. It involves various mechanisms (Th1/Th2 ...switching, Treg expansion, induction of anergy and apoptosis of T lymphocytes, development of T cell depletion) that are induced through the ligation of inhibitory receptors. Accordingly, the expression of inhibitory receptors on T cells, including PD-1, CTLA-4, and Tim-3 molecules, may reflect the effectiveness of immune adaptation and its impairment in pregnancy pathology. Preeclampsia (PE), the pathogenesis of which is associated with the impairments of immunological tolerance is a major complication of pregnancy. Accordingly, changes in the expression of inhibitory receptors on T cells may be biomarkers of abnormal gestation and potential therapeutic targets. The aim of this work was to study the expression of inhibitory molecules on peripheral blood T cells in women with PE. The study recruited 29 pregnant women with PE and 36 women with uncomplicated pregnancies in the second half of pregnancy. Pregnant women of the study groups were comparable in terms of gestational age, number of pregnancies and parity of childbirth. The control group consisted of 28 fertile women with children. Relative content of CD8
+
PD-1
+
, CD8
+
CTLA-4
+
, CD8
+
TIM-3
+
, CD8
+
PD-1
+
TIM-3
+
, CD4
+
PD-1
+
, CD4
+
CTLA-4
+
, CD4
+
TIM-3
+
, CD4
+
PD-1
+
TIM-3
+
T cells in blood were analyzed by flow cytometry. It has been shown that uncomplicated pregnancy is associated with increased expression of PD-1 and Tim-3 T cells, which is manifested by an increase in the relative content of CD4
+
Tim-3
+
, CD8
+
PD-1
+
and PD-1
+
Tim-3
+
T lymphocytes . In PE, on the contrary, there is a reduction in the expression of PD-1 and Tim-3 by T cells, in particular, a decrease in the proportion of CD4
+
Tim-3
+
and CD8
+
PD-1
+
cells; the absence of elevated levels in PD-1
+
Tim-3
+
cells (compared to uncomplicated gestation) and an increase in CTLA-4
+
cells within CD4
+
lymphocytes. Changes in the expression of inhibitory receptors are associated with the severity of PE. A decrease in CD4
+
Tim-3
+
and CD8
+
PD-1
+
T cells is most typical for patients with moderate PE, and an increase in CD4
+
CTLA-4T cells for pregnant women with severe PE. The relationship between changes in the expression of inhibitory molecules and the onset of PE has also been demonstrated. A distinctive feature of early PE is a decrease in the proportion of CD8
+
CTLA-4
+
cells and a more pronounced increase in CD4
+
CTLA-4
+
cells, while late PE is associated with a decrease in CD4
+
PD-1
+
cells and a more pronounced decrease in CD4
+
Tim-3
+
cells. The results obtained indicate a changes in the expression of CTLA-4, PD-1 and Tim-3 molecules on circulating T cells in pregnant women with PE and the association of these changes with the severity and the onset of PE manifestation.
Introduction
. Myeloid-derived suppressor cells (MDSCs) play an important role in restriction of the immune response and are associated with a poor prognosis in cancer. Mobilization of hematopoietic ...stem cells (HSCs) before high-dose chemotherapy (HCT) with autologous HSC transplantation (auto-HSCT) is accompanied by a signifcant increase in MDSC counts in peripheral blood and apheresis product of multiple myeloma (MM) patients. However, quantitative changes of these cells at the post-transplant and their role at the immune recovery remain unexplored.
The study was aimed
to analyze the dynamics of circulating MDSC counts and the expression of suppressor molecule arginase-1 in patients with MM in the frst 12 months after HCT and auto-HSCT and evaluate association between MDSCs and transplantation outcomes.
Material and Methods
. The study included 44 MM patients who underwent HCT and auto-HSCT. The relative number of granulocytic MDSCs (G-MDSCs), monocytic MDSCs (M-MDSCs), and early-stage MDSCs (E-MDSCs), as well as the expression of arginase-1 in each of MDSC subsets was evaluated by fow cytometry in patient peripheral blood samples.
Results. At the engraftment (day +12 – +16, leukocytes >1×10
9
/l), M-MDSC relative count was increased (p
U
=0.038), as well as the relative (p
U
=0.003) and absolute (p
U
˂
0.0001) counts of G-MDSCs, decreasing after 6 months down to pre-transplant values (рU=0.007, рU=0.024 and рU=0.02, respectively) and remaining at the same level at the 12-month follow-up period. The absolute count of E-MDSCs by the time of the engraftment decreased transiently (p
U
=0.004 vs before HCT), gradually recovering by 12-month follow-up (p
U
=0.032 vs day +12 – +16). The remission within 12 months in the group with G-MDSCs˂0.17 % at the engraftment was observed in 67 ± 11 % of patients, with G-MDSCs >0.17 % – in 94 ± 6 % of patients (p=0.049). During the 12-month post-transplant, the number of M-MDSCs expressing arginase-1 has been increasing, with a tendency to lower values at the engraftment in patients with early MM relapse (p
U
=0.09).
Conclusion
. The association of early MM relapse after auto-HSCT with the lower count of G-MDSCs and the lower count of arginase-1
+
M-MDSCs at the engraftment suggests that MDSCs is involved in the restriction of homeostatic proliferation as a factor for more effective immune recovery.
Background
. Inhibitory receptors and their ligands (also called checkpoint molecules) are important feedback regulators of the immune response. However, their role in immunological adaptation during ...pregnancy remains poorly understood.
The aim of the study
was to evaluate the level of checkpoint molecule (PD-1, CTLA-4, Tim-3) expression in peripheral T cells in pregnant women compared with fertile non-pregnant women.
Materials and methods
. The study included 36 women in the second half of pregnancy without pregnancy complications, 12 of whom had extragenital pathology. The control group consisted of 28 age-matched fertile non-pregnant women. The proportion of CD8
+
PD-1
+
, CD8
+
TIM-3
+
, CD8
+
PD-1
+
TIM-3
+
, CD4
+
PD-1
+
, CD4
+
TIM-3
+
, and CD4
+
PD-1
+
TIM-3
+
was evaluated by flow cytometry using the corresponding monoclonal antibodies (BD Biosciences, USA).
Results.
The proportion of CD4+Tim-3+ and CD8+PD-1+ Т cells and CD4+ and CD8+ Т lymphocytes co-expressing PD-1 and Tim-3 in the peripheral blood of pregnant women was statistically significantly higher than in non-pregnant women. An increase in CD4+Tim-3+ and CD8+PD-1+ T cells was observed both in pregnant women with and without extragenital pathology. However, pregnant women with extragenital pathology were characterized by a higher CD8+PD-1+ count and a smaller number of CD8+Tim-3+ cells, as well as by a lack of an increase in PD-1+Tim-3+ T cells typical of pregnant women. The number of comorbidities was directly correlated with the proportion of CD8+PD-1+ lymphocytes and inversely correlated with the proportion of CD8+Tim-3+ and CD4+ PD-1+Tim-3+ cells. In addition, the expression of checkpoint molecules was associated with gestational age (a direct correlation was found with the proportion of CD8+Tim-3+, CD4+PD-1+Tim-3+, and CD8+PD-1+Tim-3+ cells) and to a lesser extent – with the age of pregnant women (an inverse relationship was found with the proportion of CD8+Tim-3+ cells).
Conclusion.
Pregnant women in the second half of pregnancy are characterized by increased expression of PD-1 and Tim-3 molecules in peripheral T cells. At the same time, concomitant extragenital pathology affects the expression of these molecules.
Expansion of myeloid-derived suppressor cells (MDSCs) due to impaired differentiation of myeloid progenitor cells under conditions of inflammation was described in a number of autoimmune diseases, ...including rheumatoid arthritis, systemic lupus erythematosus, and type 1 diabetes mellitus. Studying the role of MDSCs in ankylosing spondylitis is an important issue, given that increased concentration of proinflammatory mediators in this pathology can also cause myelopoiesis disorders. The aim of present work was to study the quantitative content of MDSC subpopulations in patients with different clinical phenotypes and activity of AS. 37 patients, including 10 patients without peripheral skeletal lesions (axial form) and 27 patients with simultaneous lesions of spine and peripheral joints (peripheral form) were recruited into the study. The control group consisted of 32 age/sex-related healthy donors. Evaluation of granulocytic (LinHLA-DRCD33+CD66b+; G-MDSC), monocytic (CD14+HLA-DRlow/-; M-MDSC) and early-stage MDSCs (LinHLA-DRCD33+CD66b- ; E-MDSC) was performed using corresponding antibodies (BD Biosciences, USA) in the population of peripheral blood mononuclear cells by flow cytometry. In general, the AS patients were characterized by an increased relative and absolute amount of M-MDSC (p = 0.00002 and p = 0.00003, respectively) and G-MDSC (p = 0.0002 and p = 0.0006, respectively). Patient gender, age, and HLA-B27 expression did not significantly affect the content of these cells in peripheral blood. An increase in the median values of M-MDSC was detected both in patients with axial (Ме 5.0 (3.2-6.3) versus 2.4 (1.7-3.5) %; p = 0.001) and peripheral form (Ме 5.0 (3.0-7.0) versus 2.4 (1.7-3.5) %; p = 0.0002) AS. At the same time, the G-MDSC expansion was observed only in patients with involvement of peripheral joints (Ме 0.16 (0.07-0.3) % versus 0.05 (0.04-0.09) %; p = 0.0001). The relative contents of E-MDSC, M-MDSC and G-MDSC in the axial form of AS was in direct correlation with the activity of the disease (R = 0.58, p = 0.02; R = 0.73, p = 0.08 and R = 0.65 p = 0.04, respectively). This relationship was not observed in peripheral form of AS. The data obtained suggest a potential involvement of MDSCs in pathogenesis and phenotypic heterogeneity of AS. Simultaneously, the revealed direct correlation between the MDSC contents and the disease activity suggests a decrease in suppressive activity and/or appearance of pro-inflammatory activity in MDSC, thus requiring further research in the field.
The immunomodulatory activity of vascular endothelial growth factors (VEGFs) reveals a new role of neoangiogenesis in tumor development. Most of VEGF effects on T cells are mediated through the ...VEGF-R2 receptors. Placental growth factor (PlGF) belongs to the VEGFs family and is a selective ligand for VEGF-R1. In order to study the role of VEGF-R1-signaling in the regulation of T-cell functions, the effect of PlGF on the proliferation of donor T cell has been investigated. PlGF has been shown to inhibit the proliferation of T-lymphocytes in cultures of anti-CD3-stimulated mononuclear cells in a wide dose range, suppressing the proliferative response of both CD4 + and CD8 + T cells. The suppressive effect of PlGF was mediated through the direct interaction with VEGFR-1 on T-cells that was evidenced by the expression of VEGFR-1 by T-lymphocytes (especially after their activation) and by blocking the suppressive effect of PlGF with neutralizing anti-VEGFR-1 antibodies. Given the increased levels of PlGF in many tumors, this factor may play an important role in immunomodulation during tumor growth, mediating its effect through the VEGFR-1 signaling pathway.
Membrane TNFα (mTNFα) is expressed on many immune cell types and performs various biological functions. Dendritic cells (DC) of high-grade glioma patients exhibit impaired cytotoxic activity against ...TNFα-sensitive HEp-2 tumor cells. The mechanisms leading to the impairment of the TNFα- dependent tumoricidal activity of DC and the possibility of regulating the cytotoxic activity of DC mediated by the TNFα/TNF-R1 signaling pathway have been studied. The study was conducted on healthy donors and patients with newly diagnosed high-grade glioma. DC were generated by culturing the plastic-adherent peripheral blood mononuclear cell fraction in the presence of GM-CSF and interferon-α (IFN-DC). It was shown that the impairment of the cytotoxic activity of patient IFN-DC was associated with a low number of DC expressing mTNFα and a low level of TNFα mRNA expression in DC. IFN-DC of patients exhibited a tendency of high activity of the TNFα-converting enzyme (TACE), which accomplishes shedding of mTNFα from the cell membrane. An increased number of IFN-DC with mTNFα caused by TACE blocking enhanced cytotoxic activity of the patient’s IFN-DC against HEp-2 cells. It was established that exogenous interleukin-2 and extracellular DNA are up-regulators of the mTNFα expression on IFN-DC of the patients, but their effects are mediated by different mechanisms.
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