scholarly journals Toll-Like Receptor Homolog CD180 Expression Is Diminished on Natural Autoantibody-Producing B Cells of Patients with Autoimmune CNS Disorders

2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Zsófia Hayden ◽  
Szabina Erdő-Bonyár ◽  
Beáta Bóné ◽  
Noémi Balázs ◽  
Kornélia Bodó ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Peripheral Blood ◽  
Bacterial Infections ◽  
Citrate Synthase ◽  
Memory B Cells ◽  
Igg Antibody ◽  
Autoantibody Production ◽  
B Cell Subsets ◽  
Natural Autoantibody

Purpose. Decreased expression of TLR homolog CD180 in peripheral blood B cells and its potential role in antibody production have been described in autoimmune diseases. Effectiveness of anti-CD20 therapy in neuromyelitis optica spectrum disorder (NMOSD) and multiple sclerosis (MS) strengthens the role of B cells in the pathogenesis. Therefore, we aimed to investigate the CD180 expression of peripheral blood B cell subsets in NMOSD and MS patients and analyze the levels of natural anti-citrate synthase (CS) IgG autoantibodies and IgG antibodies induced by bacterial infections reported to play a role in the pathogenesis of NMOSD or MS. Methods. We analyzed the distribution and CD180 expression of peripheral blood B cell subsets, defined by CD19/CD27/IgD staining, and measured anti-CS IgM/G natural autoantibody and antibacterial IgG serum levels in NMOSD, RRMS, and healthy controls (HC). Results. We found decreased naïve and increased memory B cells in NMOSD compared to MS. Among the investigated four B cell subsets, CD180 expression was exclusively decreased in CD19+CD27+IgD+ nonswitched (NS) memory B cells in both NMOSD and MS compared to HC. Furthermore, the anti-CS IgM natural autoantibody serum level was lower in both NMOSD and MS. In addition, we found a tendency of higher anti-CS IgG natural autoantibody levels only in anti-Chlamydia IgG antibody-positive NMOSD and MS patients. Conclusions. Our results suggest that reduced CD180 expression of NS B cells could contribute to the deficient natural IgM autoantibody production in NMOSD and MS, whereas natural IgG autoantibody levels show an association with antibacterial antibodies.

scholarly journals Increased Frequency of Activated Switched Memory B Cells and Its Association With the Presence of Pulmonary Fibrosis in Diffuse Cutaneous Systemic Sclerosis Patients

2021 ◽  
Vol 12 ◽  
Author(s):  
Diána Simon ◽  
Péter Balogh ◽  
Szabina Erdő-Bonyár ◽  
Katalin Böröcz ◽  
Tünde Minier ◽  
...  
Keyword(s):  
Systemic Sclerosis ◽  
B Cells ◽  
Pulmonary Fibrosis ◽  
B Cell ◽  
Topoisomerase I ◽  
Citrate Synthase ◽  
Severe Form ◽  
Memory B Cells ◽  
Autoantibody Production ◽  
B Cell Subsets

Disease-associated, high-affinity pathological autoantibody production is a well-described consequence of immune dysregulation affecting B cells in systemic sclerosis (SSc), including the distribution of B-cell subsets. We have previously shown that the increased relative frequency of CD19+CD27+IgD− switched memory B cells is associated with the severe form of SSc. This study sought to analyze memory B cell subsets using an extended range of markers for further subdivision based on CD19, IgD, CD27, CD38 and CD95 phenotype, to define relationship between the alterations of memory B cell subsets and the clinical features of SSc. Peripheral blood samples were obtained from 21 SSc patients, including 14 diffuse (dcSSc) and 7 limited (lcSSc) cutaneous SSc patients, with disease duration of 2.7 ( ± 1.6) years. After purification of CD19+ B cells, multiparametric flow cytometry was performed and the frequencies of CD19+IgD−CD27−CD38+ double negative (DN) 1, CD19+IgDloCD27+CD38+ unswitched, CD19+IgD−CD27+CD38+CD95− resting switched and CD19+IgD−CD27+CD38−CD95+ activated switched memory (ASM) B cells were determined, and correlated with clinical features of SSc. The dcSSc patients had a higher frequency of ASM B cells (p = 0.028) compared to lcSSc patients. The percentage of ASM B cells was elevated in anti-Scl-70 (anti-topoisomerase I) antibody positive patients compared to negative patients (p = 0.016). Additionally, the frequency of ASM B cells was also increased in patients with pulmonary fibrosis (p = 0.003) suggesting that patients with severe form of SSc have higher ASM B cell ratios. Furthermore, the ratio of DN1 B cells was decreased (p = 0.029), while the level of anti-citrate synthase IgG natural autoantibody was elevated (p = 0.028) in patients with active disease. Our observations on the increase of ASM B cells in dcSSc and in patients with pulmonary fibrosis may point to the association of this alteration with the severe form of the disease. Functionally the correlation of ASM B cells as effector memory-plasma cell precursors with anti-topoisomerase I antibody positivity could reflect their contribution to pathological autoantibody production, whereas the decrease of memory precursor DN B cells and the increase of anti-citrate synthase IgG autoantibody may have potential significance in the assessment of disease activity.

scholarly journals In VitroMeasles Virus Infection of Human Lymphocyte Subsets Demonstrates High Susceptibility and Permissiveness of both Naive and Memory B Cells

2018 ◽  
Vol 92 (8) ◽  
pp. e00131-18 ◽  
Author(s):  
Brigitta M. Laksono ◽  
Christina Grosserichter-Wagener ◽  
Rory D. de Vries ◽  
Simone A. G. Langeveld ◽  
Maarten D. Brem ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
B Cell ◽  
Immune Suppression ◽  
Peripheral Blood ◽  
Measles Virus ◽  
Memory B Cells ◽  
T And B Cells ◽  
B Cell Subsets

ABSTRACTMeasles is characterized by a transient immune suppression, leading to an increased risk of opportunistic infections. Measles virus (MV) infection of immune cells is mediated by the cellular receptor CD150, expressed by subsets of lymphocytes, dendritic cells, macrophages, and thymocytes. Previous studies showed that human and nonhuman primate memory T cells express higher levels of CD150 than naive cells and are more susceptible to MV infection. However, limited information is available about the CD150 expression and relative susceptibility to MV infection of B-cell subsets. In this study, we assessed the susceptibility and permissiveness of naive and memory T- and B-cell subsets from human peripheral blood or tonsils toin vitroMV infection. Our study demonstrates that naive and memory B cells express CD150, but at lower frequencies than memory T cells. Nevertheless, both naive and memory B cells proved to be highly permissive to MV infection. Furthermore, we assessed the susceptibility and permissiveness of various functionally distinct T and B cells, such as helper T (TH) cell subsets and IgG- and IgA-positive memory B cells, in peripheral blood and tonsils. We demonstrated that TH1TH17 cells and plasma and germinal center B cells were the subsets most susceptible and permissive to MV infection. Our study suggests that both naive and memory B cells, along with several other antigen-experienced lymphocytes, are important target cells of MV infection. Depletion of these cells potentially contributes to the pathogenesis of measles immune suppression.IMPORTANCEMeasles is associated with immune suppression and is often complicated by bacterial pneumonia, otitis media, or gastroenteritis. Measles virus infects antigen-presenting cells and T and B cells, and depletion of these cells may contribute to lymphopenia and immune suppression. Measles has been associated with follicular exhaustion in lymphoid tissues in humans and nonhuman primates, emphasizing the importance of MV infection of B cellsin vivo. However, information on the relative susceptibility of B-cell subsets is scarce. Here, we compared the susceptibility and permissiveness toin vitroMV infection of human naive and memory T- and B-cell subsets isolated from peripheral blood or tonsils. Our results demonstrate that both naive and memory B cells are more permissive to MV infection than T cells. The highest infection levels were detected in plasma cells and germinal center B cells, suggesting that infection and depletion of these populations contribute to reduced host resistance.

Reduced Circulating Memory B-Cells Account for Humoral Immune Defects in Multiple Myeloma, Associated with Infective Risk and Poor Vaccination Responses

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 3393-3393
Author(s):  
Jonathan Carmichael ◽  
Clive R Carter ◽  
Christopher Parrish ◽  
Charlotte Kallmeyer ◽  
Sylvia Feyler ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Bacterial Infections ◽  
Memory B Cells ◽  
Antibody Responses ◽  
Progressive Reduction ◽  
Cell Numbers ◽  
Humoral Immune ◽  
Transitional B Cells ◽  
B Cell Subsets

Abstract Multiple myeloma (MM) is characterized by an increased risk of infection due to the immunosuppressive effect of the disease and conjointly of therapy. Furthermore, there is impaired responses to vaccination to counter the infection risk. The factors that underpin defective B-cell homeostasis and effective humoral immunity are not clear, nor are the extent of the defects. Also, the level of impaired humoral immunity in MGUS is not fully understood. The aim of this study was to delineate the circulating B-cell populations and recall antibody responses in patients with MGUS & MM, compared to age-matched controls, correlating with the responsiveness to vaccinations, incidence of infective complications and concomitant therapy. We performed comprehensive B-cell immunophenotyping by multi-parameter flow cytometry of peripheral blood samples from patients with MGUS (n=16), asymptomatic MM (n=18) and MM (n=108) with a median age of 63 years (range 38-94) comparing them to age-matched controls (n=9). B-cell subsets included naïve (CD19+CD27-), memory (CD19+CD27+; non-switch CD19+IgD+CD27+, switch CD19+IgD-CD27+), transitional (CD19+CD27-CD24hiCD38hi) & regulatory (CD19+CD27+CD24hi) B-cells. Serum uninvolved total IgG, IgM & IgA levels along with vaccine-specific antibody responses were analysed. There is a progressive decrease in the uninvolved immunoglobulin classes with significant reduction in total IgA (p=0.006) and IgM levels (p=0.007) in aMM/MM compared to MGUS & control (Figure 1). When anti-pneumococcal antibodies were measured, only 30% of aMM/MM patients had adequate protective levels compared to 79% of age-matched controls, with 40% of aMM/MM patients with inadequate levels experiencing recurrent respiratory tract infections compared to 25% of aMM/MM patients with adequate proactive antibodies. Patients with MGUS, aMM and MM have lower total B-cell numbers compared to controls (1-way ANOVA p=0.004; Figure 1). The reduction in B-cell numbers were primarily the consequence of reduced memory B-cells (percentage and absolute 1-way ANOVA p<0.0001), noted in both MGUS and aMM/MM but a progressive reduction with increasing disease activity (MGUS>aMM>MM). Furthermore, a correlation with total IgG levels & memory B-cell numbers is evident (r2=-0.053) & progressive reduction in memory B-cell numbers is seen with advancing cycles of therapy. The ratio of switch:non-switch memory B-cells is unaltered (control 1.05, MGUS 0.53, aMM 1.41 & MM 1.49; 1-way ANOVA p=ns). Conversely, there is a compensatory increase in the percentage of transitional B-cells when increasing disease stage is compared to controls (control 7.38% (95%ci 4.9,9.9) vs MGUS 14.0% (95%ci 7.4, 20.7) vs aMM 14.95% (95%ci 8, 21.9); 1-way ANOVA p<0.001) but a reduction is noted in MM (5.82%, 95%ci 4.5,7.2; p<0.0001), primarily being driven by sequential lines of therapy. As a consequence, the ratio of Memory:transitional B-cells is significantly reduced in aMM/MM compared to MGUS & controls (control 10.35, MGUS 20.46, aMM 7.74 & MM 4.57; 1-way ANOVA p=0.006), associated with increasing incidence of bacterial infections. A non-significant correlation is seen between transitional B-cells and total uninvolved immunoglobulin levels and with recall responses to vaccinations. There is a progressive decrease in the CD19+CD27+CD24hi B-cell subset between control and plasma cell dyscrasias (control 20.4% (95%ci 15.5,25.2), MGUS 14.0% (95%ci 7.4, 20.7), aMM 14.95% (95%ci 8, 21.9) & MM 5.82%, 95%ci 4.5,7.2; p<0.0001), primarily being driven by sequential lines of therapy and associated with increased incidence of infection. This study illustrates that patients with myeloma demonstrate reduced total circulating B-cells primarily as a consequence of reduced memory B-cells, associated with reduced immunoglobulin and recall antibody responses. This is associated with increased incidence of bacterial infections and is worsened by sequential exposure to lymphodepleting therapies. Of particular importance is the identified aberration in B-cell subsets seen in MGUS compared with age-matched control, indicative of humoral immune dysregulation highlighting that MGUS may not be an immunologically inert disorder. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

scholarly journals Naive and Effector B-cell Subtypes are Increased in Chronic Rhinosinusitis with Polyps

2018 ◽  
Vol 32 (1) ◽  
pp. 3-6 ◽  
Author(s):  
Dijana Miljkovic ◽  
Alkis Psaltis ◽  
Peter-John Wormald ◽  
Sarah Vreugde
Keyword(s):  
B Cells ◽  
B Cell ◽  
Chronic Rhinosinusitis ◽  
Peripheral Blood ◽  
Nasal Polyposis ◽  
Potential Role ◽  
Memory B Cells ◽  
Live Cells ◽  
B Cell Subsets ◽  
And Control

Background Recent studies demonstrated that B cells and their chemoattractants are elevated in the nasal mucosa of patients with chronic rhinosinusitis (CRS) with nasal polyposis (CRSwNP). However, the presence of naive B cells and of plasmablasts and memory B-cell subsets in the mucosa and periphery of the same patient with CRS is yet to be characterized. Objective Here we sought to quantify naive, plasmablasts, and memory B cells in mucosal tissue and peripheral blood of patients with CRSwNP, patients with CRS without nasal polyps (CRSsNP), and control patients. Methods Polyps, mucosa, and peripheral blood samples were prospectively collected from the patients with CRS and from the non-CRS controls. We used flow cytometry to distinguish among naive, plasmablast, and memory B cells in sinus tissue and peripheral blood. Results A total of 45 patients were recruited for the study. The patients with CRSwNP had significantly increased mucosal B-cell numbers versus the controls (3.39 ± 4.05% versus 0.39 ± 1.05% of live cells; p < 0.01, Kruskal-Wallis test), which included naive B cells (0.61 ± 0.94 versus 0.11 ± 0.24% of live cells; p < 0.03, Kruskal-Wallis test), plasmablasts (0.06 ± 0.26 versus 0.00 ± 0.00% e cells; p < 0.055, Kruskal-Wallis test), and memory B cells (0.62 ± 1.26 versus 0.05 ± 0.15% of live cells; p < 0.02, Kruskal-Wallis test). Conclusion Our study identified increased frequencies of different B-cell subtypes in the mucosa of patients with CRSwNP but not in the peripheral blood. We also found that patients with CRSwNP had significantly increased B-cell subtypes compared with the patients with CRSsNP and the controls. These results implied a potential role for mucosal B cells in the ongoing inflammation in patients with CRSwNP.

scholarly journals A splenic IgM memory subset with antibacterial specificities is sustained from persistent mucosal responses

2018 ◽  
Vol 215 (8) ◽  
pp. 2035-2053 ◽  
Author(s):  
Simon Le Gallou ◽  
Zhicheng Zhou ◽  
Lan-Huong Thai ◽  
Remi Fritzen ◽  
Alba Verge de los Aires ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Bacterial Infections ◽  
Plasma Cells ◽  
Memory B Cells ◽  
Endogenous Retroviruses ◽  
Lymphoid Tissues ◽  
Gut Flora ◽  
Protective Capacity ◽  
B Cell Subsets

To what extent immune responses against the gut flora are compartmentalized within mucosal tissues in homeostatic conditions remains a much-debated issue. We describe here, based on an inducible AID fate-mapping mouse model, that systemic memory B cell subsets, including mainly IgM+ B cells in spleen, together with IgA+ plasma cells in spleen and bone marrow, are generated in mice in the absence of deliberate immunization. While the IgA component appears dependent on the gut flora, IgM memory B cells are still generated in germ-free mice, albeit to a reduced extent. Clonal relationships and renewal kinetics after anti-CD20 treatment reveal that this long-lasting splenic population is mainly sustained by output of B cell clones persisting in mucosal germinal centers. IgM-secreting hybridomas established from splenic IgM memory B cells showed reactivity against various bacterial isolates and endogenous retroviruses. Ongoing activation of B cells in gut-associated lymphoid tissues thus generates a diversified systemic compartment showing long-lasting clonal persistence and protective capacity against systemic bacterial infections.

scholarly journals Multiple Functions of B Cells in the Pathogenesis of Systemic Lupus Erythematosus

2019 ◽  
Vol 20 (23) ◽  
pp. 6021 ◽  
Author(s):  
Kongyang Ma ◽  
Wenhan Du ◽  
Xiaohui Wang ◽  
Shiwen Yuan ◽  
Xiaoyan Cai ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
B Cells ◽  
B Cell ◽  
Lupus Erythematosus ◽  
Therapeutic Interventions ◽  
Systemic Lupus ◽  
Autoantibody Production ◽  
Regulatory B Cell ◽  
Functional Features ◽  
B Cell Subsets

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by excessive autoantibody production and multi-organ involvement. Although the etiology of SLE still remains unclear, recent studies have characterized several pathogenic B cell subsets and regulatory B cell subsets involved in the pathogenesis of SLE. Among pathogenic B cell subsets, age-associated B cells (ABCs) are a newly identified subset of autoreactive B cells with T-bet-dependent transcriptional programs and unique functional features in SLE. Accumulation of T-bet+ CD11c+ ABCs has been observed in SLE patients and lupus mouse models. In addition, innate-like B cells with the autoreactive B cell receptor (BCR) expression and long-lived plasma cells with persistent autoantibody production contribute to the development of SLE. Moreover, several regulatory B cell subsets with immune suppressive functions have been identified, while the impaired inhibitory effects of regulatory B cells have been indicated in SLE. Thus, further elucidation on the functional features of B cell subsets will provide new insights in understanding lupus pathogenesis and lead to novel therapeutic interventions in the treatment of SLE.

scholarly journals Complexity of the human memory B-cell compartment is determined by the versatility of clonal diversification in germinal centers

2015 ◽  
Vol 112 (38) ◽  
pp. E5281-E5289 ◽  
Author(s):  
Bettina Budeus ◽  
Stefanie Schweigle de Reynoso ◽  
Martina Przekopowitz ◽  
Daniel Hoffmann ◽  
Marc Seifert ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Clone ◽  
Human Memory ◽  
Memory B Cells ◽  
Sequencing Analysis ◽  
Cell Compartment ◽  
Memory B Cell ◽  
Cell Clones ◽  
B Cell Subsets

Our knowledge about the clonal composition and intraclonal diversity of the human memory B-cell compartment and the relationship between memory B-cell subsets is still limited, although these are central issues for our understanding of adaptive immunity. We performed a deep sequencing analysis of rearranged immunoglobulin (Ig) heavy chain genes from biological replicates, covering more than 100,000 memory B lymphocytes from two healthy adults. We reveal a highly similar B-cell receptor repertoire among the four main human IgM+ and IgG+ memory B-cell subsets. Strikingly, in both donors, 45% of sequences could be assigned to expanded clones, demonstrating that the human memory B-cell compartment is characterized by many, often very large, B-cell clones. Twenty percent of the clones consisted of class switched and IgM+(IgD+) members, a feature that correlated significantly with clone size. Hence, we provide strong evidence that the vast majority of Ig mutated B cells—including IgM+IgD+CD27+ B cells—are post-germinal center (GC) memory B cells. Clone members showed high intraclonal sequence diversity and high intraclonal versatility in Ig class and IgG subclass composition, with particular patterns of memory B-cell clone generation in GC reactions. In conclusion, GC produce amazingly large, complex, and diverse memory B-cell clones, equipping the human immune system with a versatile and highly diverse compartment of IgM+(IgD+) and class-switched memory B cells.

scholarly journals IgM+IgD+CD27+ B cells are markedly reduced in IRAK-4–, MyD88-, and TIRAP- but not UNC-93B–deficient patients

Blood ◽  
2012 ◽  
Vol 120 (25) ◽  
pp. 4992-5001 ◽  
Author(s):  
Sandra Weller ◽  
Mélanie Bonnet ◽  
Héloïse Delagreverie ◽  
Laura Israel ◽  
Maya Chrabieh ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Signaling Pathways ◽  
Size Distribution ◽  
Heavy Chain ◽  
Peripheral Blood ◽  
Somatic Hypermutation ◽  
Key Factors ◽  
Tlr Signaling ◽  
B Cell Subsets

Abstract We studied the distribution of peripheral B-cell subsets in patients deficient for key factors of the TLR-signaling pathways (MyD88, TIRAP/MAL, IL-1 receptor–associated kinase 4 [IRAK-4], TLR3, UNC-93B, TRIF). All TLRs, except TLR3, which signals through the TRIF adaptor, require MyD88 and IRAK-4 to mediate their function. TLR4 and the TLR2 heterodimers (with TLR1, TLR6, and possibly TLR10) require in addition the adaptor TIRAP, whereas UNC-93B is needed for the proper localization of intracellular TLR3, TLR7, TLR8, and TLR9. We found that IgM+IgD+CD27+ but not switched B cells were strongly reduced in MyD88-, IRAK-4-, and TIRAP-deficient patients. This defect did not appear to be compensated with age. However, somatic hypermutation of Ig genes and heavy-chain CDR3 size distribution of IgM+IgD+CD27+ B cells were not affected in these patients. In contrast, the numbers of IgM+IgD+CD27+ B cells were normal in the absence of TLR3, TRIF, and UNC-93B, suggesting that UNC-93B–dependent TLRs, and notably TLR9, are dispensable for the presence of this subset in peripheral blood. Interestingly, TLR10 was found to be expressed at greater levels in IgM+IgD+CD27+ compared with switched B cells in healthy patients. Hence, we propose a role for TIRAP-dependent TLRs, possibly TLR10 in particular, in the development and/or maintenance of IgM+IgD+CD27+ B cells in humans.

A role for Toll-like receptors in acquired immunity: up-regulation of TLR9 by BCR triggering in naive B cells and constitutive expression in memory B cells

Blood ◽  
2003 ◽  
Vol 101 (11) ◽  
pp. 4500-4504 ◽  
Author(s):  
Nadia L. Bernasconi ◽  
Nobuyuki Onai ◽  
Antonio Lanzavecchia
Keyword(s):  
B Cells ◽  
B Cell ◽  
Constitutive Expression ◽  
Human Memory ◽  
Cell Receptor ◽  
Primary Response ◽  
Memory B Cells ◽  
Toll Like Receptors ◽  
Polyclonal Activation ◽  
B Cell Subsets

Abstract Toll-like receptors (TLRs) are pattern recognition receptors that trigger innate immunity. In this study we investigated the expression of 10 TLRs in human naive and memory B-cell subsets. We report that in human naive B cells most TLRs are expressed at low to undetectable levels, but the expression of TLR9 and TLR10 is rapidly induced following B-cell-receptor (BCR) triggering. In contrast, memory B cells express several TLRs at constitutively high levels. The differential expression of TLR9 correlates with responsiveness to its agonist, CpG DNA. Thus, human memory B cells proliferate and differentiate to immunoglobulin (Ig)–secreting cells in response to CpG, while naive B do so only if simultaneously triggered through the BCR. The BCR-induced expression of TLRs in human naive B cells prevents polyclonal activation in a primary response, because it restricts stimulation to antigen-specific B cells. In contrast, the constitutive expression of TLRs in memory B cells allows polyclonal activation of the entire memory pool. Thus, in human B cells TLRs are downstream of BCR and play a role both in the primary response and in the memory phase.

Different Expression of FcgammaRIIb in Chronic Lymphocytic Leukemia and Human Normal B Lymphocytes

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3134-3134
Author(s):  
Carol Moreno ◽  
Rajendra Damle ◽  
Sonia Jansa ◽  
Gerardo Ferrer ◽  
Pau Abrisqueta ◽  
...  
Keyword(s):  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
B Lymphocytes ◽  
Surface Membrane ◽  
Memory B Cells ◽  
Lymphocytic Leukemia ◽  
Cd38 Expression ◽  
B Cell Subsets

Abstract The Fcgamma receptors (FcγRs) are a family of molecules that modulate immune responses. FcγRIIb is an inhibitory FcγR that bears immunoreceptor tyrosine-based inhibitory motifs which transduce inhibitory signals on coligation with the surface membrane Ig of the B-cell antigen receptor (BCR). The role of FcγRIIb in controlling B cell activation through inhibition of BCR signaling has been extensively studied in animal models. Nevertheless, data on FcγRIIb are scant in human normal and neoplastic B cells, this being due to the lack of a specific antibody for human FcγRIIb. Consequently, there is little information on this receptor in chronic lymphocytic leukemia (CLL). Considering the activated nature of CLL cells and the central role of the BCR in the biology of the disease, studies of FcγRs are warranted. We used a novel specific mAb directly conjugated with Alexa 488 fluorophore that solely reacts with the human FcγRIIb (MacroGenics, Inc.) to investigate the receptors expression on CLL and normal human B cells. The study population included 84 patients with CLL and 24 age- and sex-matched controls. FcγRIIb expression was assessed as the mean fluorescence intensity (MFI) of surface membrane staining. In CLL cells, FcγRIIb was measured on CD19+CD5+ cells in combination with CD38, CD49d or CD69. Normal B cells were immunostained for CD19, CD5, IgD and CD38 expression and B cell subsets: naïve (IgD+CD38−), activated (IgD+CD38+) and memory B cells (IgD−CD38−) were studied for their relative expression of FcγRIIb. FcγRIIb expression was found significantly higher in naïve B cells compared to activated and memory B cells [median MFI: 17420 (11960–21180) vs. 11.140 (7899–16970) and 11.830 (6984–17100); p&lt;0.001]. Significant differences were also observed between CD5− and CD5+ normal B cells. In contrast, FcγRIIb expression was lower in CLL cells than in CD5+ and CD5− normal B lymphocytes [median MFI: 6901(1034–42600), 10180 (5856–14820) and 12120 (7776–16040); p&lt;0.05)]. Interestingly, FcγRIIb expression was variable within individual CLL clones, this being higher in CD38+ and CD49d+ cells than in CD38− and CD49d− cells (p&lt;0.05). Furthermore, the highest density of FcγRIIb was observed on those cells which coexpressed CD38 and CD49d. In contrast, no significant differences were observed between FcγRIIb and the expression of the activation antigen CD69. Although CD69 and CD38 expression was significantly higher on unmutated IGHV cases, no correlation was found between FcγRIIb levels and IGHV mutational status. Similarly, there was no correlation between FcγRIIb and other poor prognostic variables such as ZAP-70 (≥20%), CD38 (≥ 30%) or high risk cytogenetics. Nevertheless, cases with ≥ 30% CD49d+ cells had higher FcγRIIb expression than those with &lt;30% CD49d+ cells (p=0.006). The findings presented in this study suggest a hierarchy of FcγRIIb expression in normal B-cells, CLL cells and their subpopulations: circulating normal CD5− B cells &gt; circulating normal CD5+ B cells &gt; circulating CD5+ CLL B cells. In addition, although FcγRIIb is present on all normal B cell subsets its expression is higher in naïve B cells. Furthermore, in CLL FcγRIIb density is greater in CD38+ and CD49d+ cells within the clone. Although CD49d and FcγRIIb on CLL clones is linked in a direct manner, there is no relationship with FcγRIIb density and IGHV mutations, ZAP-70, CD38 and unfavorable cytogenetic markers. Finally, the relationship between FcγRIIb expression on CLL cells and functional responses to BCR and other receptor-mediated signals deserve further investigation.

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