scholarly journals OP0073 SINGLE-CELL TRANSCRIPTOMICS UNCOVERS DEFECTIVE BONE MARROW EARLY B CELL DEVELOPMENT IN A SUBSET OF LUPUS PATIENTS ASSOCIATED WITH AGGRAVATED INFLAMMATION

2021 ◽  
Vol 80 (Suppl 1) ◽  
pp. 39.2-39
Author(s):  
C. Dong ◽  
X. Gu ◽  
J. Ji ◽  
X. Zhang ◽  
Z. Gu
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Lupus Erythematosus ◽  
Cell Development ◽  
B Cell Development ◽  
Cell Tolerance ◽  
Systemic Lupus ◽  
B Cell Tolerance ◽  
Healthy Donors

Background:Systemic lupus erythematosus (SLE) is a systemic autoimmune disease that occurs when the body’s immune system attacks own tissues and organs. B cells play a central role in SLE pathogenesis by producing autoantibodies as well as antibody-independent functions. Peripheral B cell abnormality is well known in lupus patients such as expansions of plasmablasts and atypical memory B cells, which are associated with active diseases. However, little is known about the B cell development in the bone marrow of lupus patients.Objectives:We conduct this survey to explore the disorder of the B cell development in the bone marrow of lupus patients.Methods:In this study, we have performed the scRNASeq to profile the bone marrow B cell compartment in lupus patients and healthy donors.Results:We identified that in a subset of lupus patients, the early B cells (proB and preB cells) were strongly decreased, which were confirmed by flow cytometry in an expanded cohort. Furthermore, bone marrow B cells from these patients showed a strong proinflammatory signature revealed by pathway analysis. Interestingly, BCR repertoire analysis showed that the IGHV-4-34 was highly enriched in these patients, indicating an enhanced B cell tolerance defect. Finally, a panel of proinflammatory cytokines (TNF-a, IL-1a, IL-12p70, IFN-g, et al.) were strongly increased in the bone marrow plasma of these patients compared with early B normal patients and healthy donors, confirming a localized proinflammatory microenvironment.Conclusion:Altogether, the current study has revealed that a defective early B cell development in lupus patients is associated with a more severe B cell tolerance defect and aggravated inflammation, which may shed new light on developing novel therapies by targeting relevant pathways.References:[1]Min Wang, Hua Chen, Jia Qiu, et al. Antagonizing miR-7 suppresses B cell hyperresponsiveness and inhibits lupus development. J Autoimmun 2020.[2]A M Jacobi, D M Goldenberg, F Hiepe, et al. Differential effects of epratuzumab on peripheral blood B cells of patients with systemic lupus erythematosus versus normal controls. Ann Rheum Dis, 2008.Acknowledgements:This work was funded by Special project of clinical medicine of Nantong University (Grant/Award number: 2019LQ001), National Natural Science Foundation of China (Grant/Award number: 81671616, 81871278 and 82071838).Disclosure of Interests:None declared

scholarly journals THU0044 SINGLE CELL ANALYSIS OF BONE MARROW AND PERIPHERAL ALTERED B CELL DIFFERENTIATION IN PATIENTS WITH ACTIVE SLE AND THE MECHANISM OF ABNORMAL EARLY B CELL DEVELOPMENT

2020 ◽  
Vol 79 (Suppl 1) ◽  
pp. 234.3-235
Author(s):  
T. Fu ◽  
Y. Yang ◽  
X. Gu ◽  
C. Dong ◽  
R. Zhao ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Lupus Erythematosus ◽  
Cell Development ◽  
B Cell Development ◽  
B Cell Differentiation ◽  
Systemic Lupus ◽  
B Cell Subsets

Background:B cell differentiation and dysfunction play a key role in the pathogenesis of Systemic lupus erythematosus (SLE). Bone marrow (BM) is the development organ of B cells, and also the home and residence place of plasma cells and memory B cells. However, there is a lack of studies on B cells in BM with lupus.Objectives:To map the development of BM and peripheral B cells and investigate the mechanism of abnormal early B cell development in SLE.Methods:A total of 11 SLE patients and 5 age- and sex-matched controls were recruited.BM and peripheral B cell subsets were measured by flow cytometry. sorting-purified B cell subsets were subject toSingle-cell RNA sequencing (scRNA-seq) and functional studies. Plasma cytokines and secreted immunoglobulins were detected by Luminex or ELISA. Disease activity of SLE patients was measured using the SLE Disease Activity Index (SLEDAI).Results:In the present study, we find out that the percentage of monocytes in MNC (p=0.070) and plasma cells(p=0.001)in CD19+ were significantly decreased in BM of SLE, compared to healthy controls. While, SLE patients had increased T%MNC(p=0.008) and B%CD19+(p=0.002) in BM that controls. In detail, the B cell subsets of bone marrow in patients with active lupus (SLEDAI≥8 score) were seriously disordered, showing the increasing T%MNC(p=0.049), propre-B%CD19+ (p=0.006)and immature B cell%CD19+ (p=0.010) than healthy donors. propre-B%CD19+ exhibited good relationship with SLEDAI. By integrating single B cell expression profiling and repertoire analysis, we map the development of B cells in BM and peripheral and pathogenic characteristics of early B cells, especially propre-B.Conclusion:These findings demonstrated that early B cells in BM, especially propre-B are abnormally differentiated with dysregulations. BM is an important organ targeted by SLE. This studyis not only to clarify the internal mechanism of the disorder of differentiation of B cells, but also to provide new clues for the targeted diagnosis and treatment of SLE.References:[1]Palanichamy, A., et al.,Neutrophil-mediated IFN activation in the bone marrow alters B cell development in human and murine systemic lupus erythematosus.J Immunol, 2014.192(3): p. 906-18.[2]Papadaki, H.A., J.C. Marsh, and G.D. Eliopoulos,Bone marrow stem cells and stromal cells in autoimmune cytopenias.Leuk Lymphoma, 2002.43(4): p. 753-60.[3]Karrar, S. and D.S. Cunninghame Graham,Abnormal B Cell Development in Systemic Lupus Erythematosus: What the Genetics Tell Us.Arthritis Rheumatol, 2018.70(4): p. 496-507.[4]Woods, M., Y.R. Zou, and A. Davidson,Defects in Germinal Center Selection in SLE.Front Immunol, 2015.6: p. 425.[5]Upregulation of p16INK4A promotes cellular senescence of bone marrow-derived mesenchymal stem cells from systemic lupus erythematosus patients.Cell Signal, 2012.24(12): p. 2307-14.Disclosure of Interests:None declared

scholarly journals Neutrophil-Mediated IFN Activation in the Bone Marrow Alters B Cell Development in Human and Murine Systemic Lupus Erythematosus

2013 ◽  
Vol 192 (3) ◽  
pp. 906-918 ◽  
Author(s):  
Arumugam Palanichamy ◽  
Jason W. Bauer ◽  
Srilakshmi Yalavarthi ◽  
Nida Meednu ◽  
Jennifer Barnard ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Bone Marrow ◽  
B Cell ◽  
Lupus Erythematosus ◽  
Cell Development ◽  
B Cell Development ◽  
Systemic Lupus ◽  
Murine Systemic Lupus Erythematosus

scholarly journals Breakdown of B cell tolerance in a mouse model of systemic lupus erythematosus.

1995 ◽  
Vol 181 (3) ◽  
pp. 1157-1167 ◽  
Author(s):  
J H Roark ◽  
C L Kuntz ◽  
K A Nguyen ◽  
A J Caton ◽  
J Erikson
Keyword(s):  
Systemic Lupus Erythematosus ◽  
B Cells ◽  
Mouse Model ◽  
Transgenic Mice ◽  
B Cell ◽  
Lupus Erythematosus ◽  
Cell Tolerance ◽  
Systemic Lupus ◽  
B Cell Tolerance ◽  
Dna Antibodies

Anti-DNA antibodies, specifically those that stain nuclei in a homogenous nuclear (HN) fashion, are diagnostic of systemic lupus erythematosus (SLE) and the MRL-lpr/lpr SLE murine model. We have used a heavy chain transgene that increases the frequency of anti-HN antibodies to address whether their production in SLE is the consequence of a defect in B cell tolerance. Anti-HN B cells were undetectable in nonautoimmune-prone transgenic mice, but in MRL-lpr/lpr transgenic mice their Ig was evident in the sera and they were readily retrievable as hybridomas. We conclude that nonautoimmune animals actively delete anti-HN-specific B cells, and that MRL-lpr/lpr mice are defective in this process possibly because of the lpr defect in the fas gene.

scholarly journals Central human B cell tolerance manifests with a distinctive cell phenotype and is enforced via CXCR4 signaling in hu-mice

2021 ◽  
Vol 118 (16) ◽  
pp. e2021570118
Author(s):  
Thiago Alves da Costa ◽  
Jacob N. Peterson ◽  
Julie Lang ◽  
Jeremy Shulman ◽  
Xiayuan Liang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Immune System ◽  
B Cells ◽  
B Cell ◽  
Cell Tolerance ◽  
Human Immune System ◽  
B Cell Tolerance ◽  
Autoreactive B Cells ◽  
Cell Clones ◽  
Human Immune

Central B cell tolerance, the process restricting the development of many newly generated autoreactive B cells, has been intensely investigated in mouse cells while studies in humans have been hampered by the inability to phenotypically distinguish autoreactive and nonautoreactive immature B cell clones and the difficulty in accessing fresh human bone marrow samples. Using a human immune system mouse model in which all human Igκ+ B cells undergo central tolerance, we discovered that human autoreactive immature B cells exhibit a distinctive phenotype that includes lower activation of ERK and differential expression of CD69, CD81, CXCR4, and other glycoproteins. Human B cells exhibiting these characteristics were observed in fresh human bone marrow tissue biopsy specimens, although differences in marker expression were smaller than in the humanized mouse model. Furthermore, the expression of these markers was slightly altered in autoreactive B cells of humanized mice engrafted with some human immune systems genetically predisposed to autoimmunity. Finally, by treating mice and human immune system mice with a pharmacologic antagonist, we show that signaling by CXCR4 is necessary to prevent both human and mouse autoreactive B cell clones from egressing the bone marrow, indicating that CXCR4 functionally contributes to central B cell tolerance.

scholarly journals Active PI3K abrogates central tolerance in high-avidity autoreactive B cells

2019 ◽  
Vol 216 (5) ◽  
pp. 1135-1153 ◽  
Author(s):  
Sarah A. Greaves ◽  
Jacob N. Peterson ◽  
Pamela Strauch ◽  
Raul M. Torres ◽  
Roberta Pelanda
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Peripheral Tolerance ◽  
High Avidity ◽  
Cell Tolerance ◽  
B Cell Tolerance ◽  
Central Tolerance ◽  
Autoreactive B Cells ◽  
Self Antigen

Autoreactive B cells that bind self-antigen with high avidity in the bone marrow undergo mechanisms of central tolerance that prevent their entry into the peripheral B cell population. These mechanisms are breached in many autoimmune patients, increasing their risk of B cell–mediated autoimmune diseases. Resolving the molecular pathways that can break central B cell tolerance could therefore provide avenues to diminish autoimmunity. Here, we show that B cell–intrinsic expression of a constitutively active form of PI3K-P110α by high-avidity autoreactive B cells of mice completely abrogates central B cell tolerance and further promotes these cells to escape from the bone marrow, differentiate in peripheral tissue, and undergo activation in response to self-antigen. Upon stimulation with T cell help factors, these B cells secrete antibodies in vitro but remain unable to secrete autoantibodies in vivo. Overall, our data demonstrate that activation of the PI3K pathway leads high-avidity autoreactive B cells to breach central, but not late, stages of peripheral tolerance.

scholarly journals Belimumab alters transitional B-cell subset proportions in patients with stable systemic lupus erythematosus

Lupus ◽  
2019 ◽  
Vol 28 (11) ◽  
pp. 1337-1343 ◽  
Author(s):  
A Benitez ◽  
K Torralba ◽  
M Ngo ◽  
L M Salto ◽  
K S Choi ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
B Cells ◽  
B Cell ◽  
Lupus Erythematosus ◽  
Mononuclear Cells ◽  
Cell Subset ◽  
Systemic Lupus ◽  
Peripheral Blood Mononuclear ◽  
Healthy Donors ◽  
Immature B Cells

Objective We evaluated the effects of the B-cell activating factor (BAFF)-targeting antibody Belimumab on human nonmemory B-cell pools. Human B-cell pools were identified using surface markers adapted from mouse studies that specifically assessed reductions in immature B cells due to BAFF depletion. Patients with systemic lupus erythematosus (SLE) have high levels of both BAFF and immature B cells. Mechanistic mouse studies provide a framework for understanding human responses to therapies that target B cells. Methods Peripheral blood mononuclear cells were isolated from healthy donors and SLE patients on Belimumab or standard-of-care therapy (SCT). Cells were stained for flow cytometry to identify B-cell subsets based on CD21/CD24. Differences in subset proportions were determined by one-way ANOVA and Tukey’s post hoc test. Results Patients treated with Belimumab show alterations in the nonmemory B-cell pool characterized by a decrease in the Transitional 2 (T2) subset ( p = 0.002), and an increase in the proportion of Transitional 1 (T1) cells ( p = 0.005) as compared with healthy donors and SCT patients. The naïve B-cell compartment showed no significant differences between the groups ( p = 0.293). Conclusion Using a translational approach, we show that Belimumab-mediated BAFF depletion reduces the T2 subset in patients, similar to observations in mouse models with BAFF depletion.

scholarly journals Reduced receptor editing in lupus-prone MRL/lpr mice

2007 ◽  
Vol 204 (12) ◽  
pp. 2853-2864 ◽  
Author(s):  
Jennifer L. Lamoureux ◽  
Lisa C. Watson ◽  
Marie Cherrier ◽  
Patrick Skog ◽  
David Nemazee ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Messenger Rna ◽  
Immunoglobulin M ◽  
Considerable Proportion ◽  
Receptor Editing ◽  
Cell Tolerance ◽  
B Cell Tolerance ◽  
Immature B Cells

The initial B cell repertoire contains a considerable proportion of autoreactive specificities. The first major B cell tolerance checkpoint is at the stage of the immature B cell, where receptor editing is the primary mode of eliminating self-reactivity. The cells that emigrate from the bone marrow have a second tolerance checkpoint in the transitional compartment in the spleen. Although it is known that the second checkpoint is defective in lupus, it is not clear whether there is any breakdown in central B cell tolerance in the bone marrow. We demonstrate that receptor editing is less efficient in the lupus-prone strain MRL/lpr. In an in vitro system, when receptor-editing signals are given to bone marrow immature B cells by antiidiotype antibody or after in vivo exposure to membrane-bound self-antigen, MRL/lpr 3-83 transgenic immature B cells undergo less endogenous rearrangement and up-regulate recombination activating gene messenger RNA to a lesser extent than B10 transgenic cells. CD19, along with immunoglobulin M, is down-regulated in the bone marrow upon receptor editing, but the extent of down-regulation is fivefold less in MRL/lpr mice. Less efficient receptor editing could allow some autoreactive cells to escape from the bone marrow in lupus-prone mice, thus predisposing to autoimmunity.

Precursor B Cell Acute Lymphoblastic Leukemia Induces Pro-Leukemic Changes in the Bone Marrow Microenvironment That Contribute to Therapeutic Resistance

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1466-1466
Author(s):  
Christopher D Chien ◽  
Elizabeth D Hicks ◽  
Paul P Su ◽  
Haiying Qin ◽  
Terry J Fry
Keyword(s):  
Bone Marrow ◽  
Acute Lymphoblastic Leukemia ◽  
B Cells ◽  
B Cell ◽  
Lymphoblastic Leukemia ◽  
Cell Development ◽  
B Cell Development ◽  
Bone Marrow Microenvironment ◽  
Therapeutic Resistance

Abstract Abstract 1466 Pediatric acute lymphoblastic leukemia (ALL) is the most common childhood malignancy. Although cure rates for this disease are approximately 90%, ALL remains one of the leading causes cancer-related deaths in children. Thus, new treatments are needed for those patients that do not respond to or recur following standard chemotherapy. Understanding the mechanisms underlying resistance of pediatric ALL to therapy offers one approach to improving outcomes. Recent studies have demonstrated the importance of communication between cancer cells and their microenvironment and how this contributes to the progression and therapeutic resistance but this has not been well studied in the context of ALL. Since the bone marrow is presumed to be the site of initiation of B precursor ALL we set out in our study to determine how ALL cells utilize the bone marrow milieu in a syngeneic transplantable model of preB cell ALL in immunocompetent mice. In this model, intravenously injected preB ALL develops first in the bone marrow, followed by infiltration into the spleen, lymph node, and liver. Using flow cytometry to detect the CD45.2 isoform following injection into B6CD45.1+ congenic recipients, leukemic cells can be identified in the bone marrow as early as 5 days after IV injection with a sensitivity of 0.01%-0.1%. The pre-B ALL line is B220+/CD19+/CD43+/BP1+/IL-7Ralpha (CD127)+/CD25-/Surface IgM-/cytoplasmic IgM+ consistent with a pre-pro B cell phenotype. We find that increasing amounts of leukemic infiltration in the bone marrow leads to an accumulation of non-malignant developing B cells at stages immediately prior to the pre-pro B cell (CD43+BP1-CD25-) and a reduction in non-malignant developing pre B cells at the developmental stage just after to the pre-pro B cell stage (CD43+BP1+CD25+). These data potentially suggest occupancy of normal B cell developmental niches by leukemia resulting in block in normal B cell development. Further supporting this hypothesis, we find significant reduction in early progression of ALL in aged (10–12 month old) mice known to have a deficiency in B cell developmental niches. We next explored whether specific factors that support normal B cell development can contribute to progression of precursor B cell leukemia. The normal B cell niche has only recently been characterized and the specific contribution of this niche to early ALL progression has not been extensively studied. Using a candidate approach, we examined the role of specific cytokines such as Interleukin-7 (IL-7) and thymic stromal lymphopoietin (TSLP) in early ALL progression. Our preB ALL line expresses high levels of IL-7Ralpha and low but detectable levels of TLSPR. In the presence of IL-7 (0.1 ng/ml) and TSLP (50 ng/ml) phosphSTAT5 is detectable indicating that these receptors are functional but that supraphysiologic levels of TSLP are required. Consistent with the importance of IL-7 in leukemia progression, preliminary data demonstrates reduced lethality of pr-B cell ALL in IL-7 deficient mice. Overexpression of TSLP receptor (TSLPR) has been associated with high rates of relapse and poor overall survival in precursor B cell ALL. We are currently generating a TSLPR overepressing preBALL line to determine the effect on early ALL progression and are using GFP-expressing preB ALL cells to identify the initial location of preB ALL occupancy in the bone marrow. In conclusion, or model of early ALL progression provides insight into the role of the bone marrow microenvironment in early ALL progression and provides an opportunity to examine how these microenvironmental factors contribute to therapeutic resistance. Given recent advances in immunotherapy for hematologic malignancies, the ability to study this in an immunocompetent host will be critical. Disclosures: No relevant conflicts of interest to declare.

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