Phenotypic Analysis of CD19+ Subsets In Monoclonal Gammopathy Patients

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 4998-4998
Author(s):  
Lucie Kovarova ◽  
Pavla Zarbochova ◽  
Tamara Varmuzova ◽  
Ivana Buresova ◽  
Karthick Raja Muthu Raja ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Healthy Volunteers ◽  
Peripheral Blood ◽  
Monoclonal Gammopathy ◽  
Plasma Cells ◽  
Memory B Cells ◽  
Phenotypic Analysis ◽  
Transitional B Cells ◽  
Significant Difference

Abstract Abstract 4998 Background. Monoclonal gammopathy of undetermined significance (MGUS) is the most common plasma cell disorder which can eventually progress into malignant multiple myeloma (MM). Plasma cells (PCs) are the terminal stadium of B cells differentiation, but it is still unclear which population is the source of pathological PCs with malignant transformation and which population is involved in and may give rise to clonogenic myeloma stem cells. Aims. Phenotypic analysis of CD19+ cell subpopulations in monoclonal gammopathy patients and healthy volunteers to asses their frequency and to find differences on cellular level. Methods. Total of 38 samples was analyzed (16 newly diagnosed untreated MM patients, 12 untreated MGUS persons and 10 healthy donors). CD19+ cells were analyzed for surface expression of CD24, CD27, CD38, and IgD by 5-colors immunophenotyping. Subpopulations of pre-plasma cells consist of transitional B cells (CD24+CD38+), naïve B cells (CD38-IgD+), activated B cells (CD38+IgD+), preGC B cells (CD38++IgD+) and memory B cells (CD38-/+IgD-). These were evaluated in whole lysed peripheral blood together with circulating plasmablast/plasma cells (CD38++IgD-). Bone marrow of MGUS and MM patients was analyzed for number of transitional, immature and memory B cells. Results. Flow cytometric analysis shown no statistical difference when compared number of transitional B cells (1.8%; 3.0% and 1.2%) and activated B cells (54.6%; 62.1% and 45.5%) in peripheral blood of healthy volunteers, MGUS and MM patients, respectively. There was found lower number of circulating plasmablast/plasma cells in peripheral blood of healthy volunteers than in MGUS (1.0% vs. 1.7%; p<0.01), but there was no statistically significant difference for MM (1.7%) when compared to others. The highest number of peripheral naive B cells was found in healthy volunteers (21.4%; p<0.001) and the highest number of peripheral memory B cells was found in MM patients (32.9%; p<0.01) when compared to other groups. There was found also higher number of peripheral preGC B cells in MGUS and MM patients (2.7% vs. 1.6% vs. 1.3%; p<0.05) than in healthy volunteers, respectively. Although numbers of transitional and immature B cells in bone marrow were different for MGUS and MM, the only statistically significant difference was found in number of memory B cells (25.4% for MGUS vs. 11.9% for MM; p<0.01). Summary/Conclusions. Our result showed differences in CD19+ subsets when compared peripheral blood of healthy volunteers and monoclonal gammopathy patients as well as in bone marrow of monoclonal gammopathy group. These differences could be a sign of ongoing changes in B cells of monoclonal gammopathy patients. Further analysis will be also focused on changes at DNA level to confirm clonality of selected subpopulations and to find possible myeloma stem cells source. Supported by GACR 301/09/P457, GACR GAP304/10/1395, MSMT LC06027, MSM0021622434, IGA 10408-3, IGA 10406-3. Disclosures: Hajek: Janssen-Cilag: Honoraria; Celgene: Honoraria; Merck, Sharp, and Dohme: Honoraria.

Rituximab Infusion after Allogeneic HCT Delays B Cell Reconstitution and Alloimmunity.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2982-2982
Author(s):  
Bita Sahaf ◽  
Kartoosh Heydari ◽  
George Chen ◽  
David Miklos
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Peripheral Blood ◽  
Plasma Cells ◽  
Memory B Cells ◽  
Influenza B ◽  
Post Transplant ◽  
Allogeneic Hct ◽  
Lymphoid Progenitors

Abstract B cells are implicated in the pathophysiology of chronic graft-vs-host disease (cGVHD) and anti-B cell rituximab is effective cGVHD therapy. We have treated 31 MCL and CLL patients with a nonmyeloablative transplant preparative regimen consisting of total lymphoid irradiation (TLI, 80 cGy in 10 fractions, days -11 to -1) and anti-thymocyte globulin (ATG, 1.5mg/kg/day, days -11 to -7, total 7.5mg/kg) followed by rituximab 375mg/m2 on days 56, 63, 70, and 77 after transplant. Primary GVHD prophylaxis was mycophenolic acid and cyclosporine tapered off by 6 months. Thus far, two patients with MCL have died of disease progression before rituximab infusion and the remaining 29 are alive. Here we study B cell reconstitution in 12 patients with more than one year of follow-up. We used Hi-D FACS technology to distinguish common lymphoid progenitors (CD34+CD117+), early B cell progenitors (CD34+CD10+CD19+), pre B cells (CD3−CD19+CD10+CD34−), immature B cells (CD3−CD19+CD10+CD20+CD5−IgM+), mature and memory B cells (CD3−CD19+CD20+CD27+), and plasma cells (CD138+CD38+). Peripheral B cells (CD19+CD20+) remained undetectable 6 and 9 months after transplant. Peripheral blood CD19+ cells were first detected in 3/9 patients at 1 year and 6/6 patients at 1.5 years. The majority of recovering peripheral blood B cells expressed a memory phenotype (CD19+CD27+, n–=6). Bone marrow aspirates collected 180 and 365 days post transplant showed CD34+CD117+ lymphoid progenitors (n=4) are increased after rituximab and then decline from 13–20% of cells 180 days post transplant to 3–8% of cells 365 days post transplant. Control patients transplanted using a TLI-ATG regimen without rituximab infusion show a lymphoid progenitor cell frequency of 4–7% (n=3). CD19+CD10+ immature progenitor B cells accumulated after rituximab, comprised 3–6% of lymphoid cells in the bone marrow 90 and 180 days after HCT (n=4), and with time were replaced by mature B cells lacking CD10 expression. IgM and/or IgD expressing mature cells (that usually express CD20) were rarely detected in bone marrow until peripheral CD19+ B cell recovery. As expected the frequency of CD19+CD27+ mature memory B cells was very low at 0.3–1.5% (n=4). Finally, CD38+CD138+ plasma cells accounted for 0.5–2% of bone marrow before and after rituximab. In summary, B cells recover from increased proportion of lymphoid progenitors with reconstitution recapitulating B cell ontogeny. No adverse infusion events occurred with rituximab and infectious complications reflected usual transplant incidence including CMV and VZV reactivation, influenza B, aspergillus and pseudomonal bacteremia. Plasma IgG levels increased from the patient’s peritransplant baseline to 110% at 9 months, 158% at 12 months, and 124% at 18months. At 6, 9, and 12 months, EBV titer was 76%, 104%, and 103% relative to pretransplant patient titers demonstrating protective antibodies are maintained despite rituximab therapy presumably secreted from long-lived CD20 negative plasma cells. Thus far, no allogeneic antibody responses have developed in the five male with female donors against H-Y antigens and suggest post-HCT rituximab prevent or diminish allogeneic B cell responses. This first trial of rituximab treatment 2 months after allogeneic HCT was well tolerated, patients maintained protective humoral immunity, and peripheral blood B cells reconstituted 12–18 months after transplant.

scholarly journals Reduced peripheral blood regulatory B cell levels are not associated with the Expanded Disability Status Scale score in multiple sclerosis

2018 ◽  
Vol 46 (9) ◽  
pp. 3970-3978 ◽  
Author(s):  
Shujun Guo ◽  
Qingqing Chen ◽  
Xiaoli Liang ◽  
Mimi Mu ◽  
Jing He ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
B Cells ◽  
Peripheral Blood ◽  
Plasma Cells ◽  
Serum Levels ◽  
Memory B Cells ◽  
Healthy Controls ◽  
Expanded Disability Status Scale ◽  
Disability Status ◽  
Edss Score

Objective To investigate levels of regulatory B (Breg) cells, plasma cells, and memory B cells in the peripheral blood, and interleukin (IL)-10 in the serum of multiple sclerosis (MS) patients, and to determine the correlation between Breg cell levels and the Expanded Disability Status Scale (EDSS) score. Methods Levels of Breg cells, plasma cells, and memory B cells in the peripheral blood of 12 MS patients were measured using flow cytometry. IL-10 serum levels were measured by enzyme-linked immunosorbent assay. The correlation between Breg cell levels and MS EDSS score was measured using Pearson’s correlation coefficient. Results Compared with healthy controls, MS patients had decreased levels of CD19+CD24hiCD38hi Breg cells in their peripheral blood and reduced serum levels of IL-10; however, the ratios of CD19+CD27hiCD38hi plasma cells and CD19+CD27+CD24hi memory B cells to total B cells did not differ significantly between healthy controls and MS patients. CD19+CD24hiCD38hi Breg cell levels in the peripheral blood of MS patients were not significantly correlated with MS EDSS score. Conclusion Peripheral blood CD19+CD24hiCD38hi Breg cell levels and serum IL-10 levels were reduced in MS patients compared with controls, but Breg cell levels were not correlated with MS EDSS score.

scholarly journals The Normal Counterpart of IgD Myeloma Cells in Germinal Center Displays Extensively Mutated IgVH Gene, Cμ–Cδ Switch, and λ Light Chain Expression

1998 ◽  
Vol 187 (8) ◽  
pp. 1169-1178 ◽  
Author(s):  
Christophe Arpin ◽  
Odette de Bouteiller ◽  
Diane Razanajaona ◽  
Isabelle Fugier-Vivier ◽  
Francine Brière ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Light Chain ◽  
Germinal Center ◽  
Plasma Cells ◽  
Precursor Cells ◽  
Memory B Cells ◽  
Hematopoietic Stem ◽  
Myeloma Cells ◽  
Λ Light Chain

Human myeloma are incurable hematologic cancers of immunoglobulin-secreting plasma cells in bone marrow. Although malignant plasma cells can be almost eradicated from the patient's bone marrow by chemotherapy, drug-resistant myeloma precursor cells persist in an apparently cryptic compartment. Controversy exists as to whether myeloma precursor cells are hematopoietic stem cells, pre–B cells, germinal center (GC) B cells, circulating memory cells, or plasma blasts. This situation reflects what has been a general problem in cancer research for years: how to compare a tumor with its normal counterpart. Although several studies have demonstrated somatically mutated immunoglobulin variable region genes in multiple myeloma, it is unclear if myeloma cells are derived from GCs or post-GC memory B cells. Immunoglobulin (Ig)D-secreting myeloma have two unique immunoglobulin features, including a biased λ light chain expression and a Cμ–Cδ isotype switch. Using surface markers, we have previously isolated a population of surface IgM−IgD+CD38+ GC B cells that carry the most impressive somatic mutation in their IgV genes. Here we show that this population of GC B cells displays the two molecular features of IgD-secreting myeloma cells: a biased λ light chain expression and a Cμ–Cδ isotype switch. The demonstration of these peculiar GC B cells to differentiate into IgD-secreting plasma cells but not memory B cells both in vivo and in vitro suggests that IgD-secreting plasma and myeloma cells are derived from GCs.

scholarly journals Abnormal Microenvironment of Bone Marrow B Lymphocytes in Patients with Primary Immune Thrombocytopenia

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 3759-3759
Author(s):  
Tianshu Yu ◽  
Ming Hou ◽  
Xinguang Liu ◽  
Panpan Han
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Lymphocytes ◽  
Chemokine Receptors ◽  
Immune Thrombocytopenia ◽  
Plasma Cells ◽  
Healthy Donors ◽  
Significant Difference ◽  
Primary Immune Thrombocytopenia ◽  
Development And Differentiation

Abstract BACKGROUNDS: Primary immune thrombocytopenia (ITP) is an autoimmune disorder characterized by peripheral blood platelet count<100x10^9/L and increased risk of bleeding, but its exact pathogenesis remains unclear. Previous researches mostly emphasized on periphral blood, in order to elucidate the pathogenesis and provide new strategies for treatment, a series of studies about bone marrow B cells were carried out in our investigation. METHODS: 5ml bone marrow blood samples from 11 newly diagnosed ITP patients and 7 allo-HSCT healthy donors were collected into heparin anticoagulant tubes , bone marrow mononuclear cells (BMMCs) were separated with Ficoll density gradient-centrifugation in two hours. Different subsets of B lymphocytes were determined by multicolor flow cytometry including naive B cells, total memory B cells , plasma cells and regulatory B cells (Bregs) ,as well as some chemokine receptors of B cells (CXCR5, BAFFR, BCMA, TACI) . RNA were extracted from BMMCs using Trizol reagent, transcription factors related to development and differentiation of B cells (PRDM1, Pax5, IRF-4, XBP1) were detected by real-time PCR. RESULTS: The percentage of B cells (CD19+) in bone marrow lymphocytes in ITP patients was significantly lower than that in healthy donors (6.16±0.74% vs. 17.28±2.43%, P < 0.0001). The proportion of naive B (CD19+CD27-) in B cells was also lower compared with normal controls (59.11±7.60% vs. 81.58±4.00%, P=0.041), and the proportion of memory B (CD19+CD27+) was higher(40.17±7.67% vs. 18.01±3.89%, P=0.045),but there was no significant difference in plasma cells (CD19-CD138+). Besides,there was a decrease of Breg (CD19+CD24highCD38high) in ITP patients compared with healthy donors (20.33±5.05% vs. 57.98±9.76% , P = 0.008), and its percentage of total lymphocytes was also significantly lower unsurprisingly (1.38±0.38% vs. 12.23±2.88%, P < 0.001). The level of BAFFR in mature B cells was elevated in ITP patients (80.72±4.53% vs. 45.81±8.49%, P = 0.002). However, no significant difference was observed in other three chemokine receptors. All four Transcription factors related to B cell development and differentiation was not found to be significantly different between ITP patients and healthy controls. CONCLUSIONS: Our results showed that memory B cells which represent the active form were increased, and Bregs which mediate immune tolerance were much more decreased in ITP patients. As BAFFR is the only specific receptor of B cell activating factor(BAFF), its elevated expression suggested the BAFF-BAFFR system enhanced chemotactic function of B cells in ITP patients. All those results indicated that the bone marrow B cells in patients with ITP were in a state of immune overstimulation, this may potentially constitute a novel therapeutic target. Figure. Figure. Disclosures No relevant conflicts of interest to declare.

scholarly journals Generation of Plasma Cells From Peripheral Blood Memory B Cells: Synergistic Effect of Interleukin-10 and CD27/CD70 Interaction

Blood ◽  
1998 ◽  
Vol 91 (1) ◽  
pp. 173-180 ◽  
Author(s):  
Kazunaga Agematsu ◽  
Haruo Nagumo ◽  
Yumiko Oguchi ◽  
Takayuki Nakazawa ◽  
Keitaro Fukushima ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
B Cells ◽  
Peripheral Blood ◽  
Plasma Cells ◽  
Interleukin 10 ◽  
Memory B Cells ◽  
The Other ◽  
Cell Pool ◽  
Activation System ◽  
Antibody Secreting Cells

B cells can differentiate into the antibody-secreting cells, plasma cells, whereas the crucial signals that positively control the entry into the pathway to plasma cells have been unclear. Triggering via CD27 by CD27 ligand (CD70) on purified peripheral blood B cells yielded an increase in the number of plasma cells in the presence of interleukin-10 (IL-10). Differentiation into plasma cells by a combination of IL-10 and CD70 transfectants occurred in CD27+ B cells but not in CD27− B cells. Moreover, addition of IL-2 to the IL-10 and CD70-transfect activation system greatly induced differentiation into plasma cells. In the presence of only IL-2, IL-4, or IL-6, CD70 transfectants did not promote differentiation into plasma cells. On the other hand, CD40 signaling increased the expansion of a B-cell pool from peripheral blood B cells primarily activated by IL-2, IL-10, and anti-CD40 monoclonal antibody (MoAb). Finally, CD27 signaling also rescued B cells from IL-10–mediated apoptosis. These data demonstrate that CD27 ligand (CD70) is a key molecule to prevent the IL-10–mediated promotion of apoptosis and to direct the differentiation of CD27+ memory B cells toward plasma cells in cooperation with IL-10.

scholarly journals Memory persistence and differentiation into antibody-secreting cells accompanied by positive selection in longitudinal BCR repertoires

2022 ◽  
Author(s):  
Artem I. Mikelov ◽  
Evgeniia I. Alekseeva ◽  
Ekaterina A. Komech ◽  
Dmitriy B. Staroverov ◽  
Maria A. Turchaninova ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Peripheral Blood ◽  
Plasma Cells ◽  
Inflammatory Diseases ◽  
Affinity Maturation ◽  
Memory B Cells ◽  
Immune Memory ◽  
Memory B Cell ◽  
Antibody Secreting Cells

B-cell mediated immune memory holds both plasticity and conservatism to respond to new challenges and repeated infections. Here, we analyze the dynamics of immunoglobulin heavy chain (IGH) repertoires of memory B cells, plasmablasts and plasma cells sampled several times during one year from peripheral blood of volunteers without severe inflammatory diseases. We reveal a high degree of clonal persistence in individual memory B-cell subsets with inter-individual convergence in memory and antibody-secreting cells (ASCs). Clonotypes in ASCs demonstrate clonal relatedness to memory B cells and are transient in peripheral blood. Two clusters of expanded clonal lineages displayed different prevalence of memory B cells, isotypes, and persistence. Phylogenetic analysis revealed signs of reactivation of persisting memory B cell-enriched clonal lineages, accompanied by new rounds of affinity maturation during proliferation to ASCs. Negative selection contributes to both, persisting and reactivated lineages, saving functionality and specificity of BCRs to protect from the current and future pathogens.

scholarly journals An apoptosis-dependent checkpoint for autoimmunity in memory B and plasma cells

2020 ◽  
Vol 117 (40) ◽  
pp. 24957-24963 ◽  
Author(s):  
Christian T. Mayer ◽  
Jan P. Nieke ◽  
Anna Gazumyan ◽  
Melissa Cipolla ◽  
Qiao Wang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Lymphocytes ◽  
Antinuclear Antibody ◽  
Plasma Cells ◽  
Memory B Cells ◽  
Measurable Effect ◽  
Gene Diversification ◽  
Shed Light ◽  
Antibody Gene

B lymphocytes acquire self-reactivity as an unavoidable byproduct of antibody gene diversification in the bone marrow and in germinal centers (GCs). Autoreactive B cells emerging from the bone marrow are silenced in a series of well-defined checkpoints, but less is known about how self-reactivity that develops by somatic mutation in GCs is controlled. Here, we report the existence of an apoptosis-dependent tolerance checkpoint in post-GC B cells. Whereas defective GC B cell apoptosis has no measurable effect on autoantibody development, disruption of post-GC apoptosis results in accumulation of autoreactive memory B cells and plasma cells, antinuclear antibody production, and autoimmunity. The data presented shed light on mechanisms that regulate immune tolerance and the development of autoantibodies.

Delayed Redistribution of CD27, CD40 and CD80 Positive B Cells and the Impaired In Vitro Immunoglobulin G Production in Patients with Non-Hodgkin Lymphoma Treated with Rituximab.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 938-938
Author(s):  
Mitsufumi Nishio ◽  
Katsuya Fujimoto ◽  
Satoshi Yamamoto ◽  
Toshiya Sakai ◽  
Kohki Kumano ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Plasma Cells ◽  
Memory B Cells ◽  
Healthy Controls ◽  
Immunoglobulin Production ◽  
Significant Difference ◽  
Healthy Control ◽  
Delayed Recovery

Abstract Rituximab (RT) has been proven to be very effective in depleting normal and malignant B lymphocytes in vivo and it is widely used for the treatment of B cell malignancies, particularly B cell non-Hodgkin’s lymphoma (NHL). RT alone does not appear to cause severe hypogammaglobulinemia according to initial clinical trials. However, recent studies revealed that patients who received RT as an adjuvant to stem cell transplantation (SCT) demonstrated an increased risk of developing severe hypogammaglobulinemia. We have found such hypogammaglobulinemia to be due to the delayed recovery of CD27 positive memory B cells and an impaired isotype expression. (Nishio et al. Eur J Haematol, 2006). This finding suggests that RT can influence not only the quantity, but also the quality of B-cell redistribution. Nevertheless, to our knowledge, precisely how the B-cell repertoire regenerates after anti-CD20-mediated transient B-cell depletion in patients with NHL remains to be elucidated. To clarify this, we performed a phenotypical analysis of B cells. A total of 22 patients with NHL who received RT combined with autologous SCT (n=17) or CHOP (n=5) were evaluated to identify their immunophenotype. The median period after the last administration of RT was 33.5 months (range from 12 to 56 months). We investigated the expression of various markers, including CD27, CD38, CD40, CD80, CD86 and CD95 on B cells by immunofluorescence staining with a flowcytometry analysis. A statistically significant difference was noted in three of the six surface antigens when the expressions of those antigens were compared with those in the healthy control populations (N=14). The most striking differences we found was the expression levels of CD27. The healthy control group had a much higher expression of CD27 in comparison to those of the patients treated with RT (28.1±14.1% vs 8.2±6.1%, p&lt;0.001). In addition, significant differences in the expression of CD40 and CD80 were also noted. While the positive rates of CD80 and CD40 on B cells from healthy controls were 21.5±10.8% and 80.5±16.7%, those of patients treated with RT were 9.9±6.9% and 49.7±33.5%, respectively (p&lt;0.01 and p&lt;0.05). Since CD40-CD40L and CD80-CD28 pathways between B and T cells are necessary for the development of CD27 positive polyclonal B-cell activation and immunoglobulin production, we hypothesized that the B cells from patients treated with RT thus had a reduced ability to differentiate into plasma cells and immunoglobulin production in vitro. To test this hypothesis, we purified the B cells from ten patients with NHL treated with RT and then cultured them upon the engagement of immunoglobulin receptor and CD40 in the presence of IL-2 and IL-10. After eight days of stimulation, the supernatants of the culture were harvested and the concentrations of immunoglobulin were measured by ELISA. As a result, the IgG production was found to be significantly impaired in patients with NHL in comparison to those from the healthy controls. The observation of a delayed recovery of the memory B cells with an abnormal cell marker expression and function demonstrates that naive B cells may therefore be responsible for their failure to differentiate into plasma cells after RT therapy.

Sign in / Sign up

Export Citation Format

Share Document