Increased Age-Related B-Cells in Patients with Aplastic Anemia
Authors: Solomou Elena E. , Plakoula Eva , Symeonidis Argiris , Galanopoulos Athanasios , Vyniou Athina , Nikolaos Giannakoulas , Palasopoulou Maria D. , Verigou Evgenia , Vassilopoulos George , Gogos Haralambos , Kattamis Antonis , Liossis Stamatis-Nick 
Published: October 2019

Abstract
Increased Age-Related B-cells in Patients with Aplastic Anemia
Introduction:
Aplastic anemia is a rare disease characterized by immune dysregulation. T cells in aplastic anemia are characterized by various intrinsic defects leading to increased IFN-g levels and Fas-mediated apoptosis of hematopoietic stem cells. We and others, have previously shown that the transcription factor T-bet is over-expressed in T cells from patients with aplastic anemia. Recently it was shown that a subpopulation of B cells also express T-bet; these cells are characterized as age-related B cells (ABCs) and express high levels of CD11c and CD19, they are CD21 negative and express T-bet. These T-bet+ ABCs are found increased in patients with autoimmune diseases (i.e. systemic lupus erythematosus, rheumatoid arthritis, and multiple sclerosis). Stimulation of B cells with antigens, Toll-like receptors and IFN-g leads to the formation of ABCs, which in turn "talk" to the T cells and stimulate them. Stimulation of T cells leads to IFN-g production, and this IFN-g may lead to further induction of T-bet expressing B cells (ABCs).
Specific aim: In this study we wanted to examine the expression of ABCs in patients with aplastic anemia. We isolated peripheral blood mononuclear cells (PBMCs) from patients with aplastic anemia (n=10) and eight healthy, age- matched controls. Written informed consent was obtained from all study subjects. Cells were stained with the surface markers CD11c, CD19 and CD21, and subsequently analyzed using flow cytometry. An anti-Tbet antibody was also used after cell permeabilization. The CD11c-high, CD19 positive, CD21 negative, T-bet positive population represent the ABCs.
Results: Patients with aplastic anemia at presentation showed increased numbers of circulating ABCs compared to healthy controls (29,08±1,62% vs 4,06 ±0,4 % respectively, p<0.001).(Fig. 1). Patients who responded to treatment showed comparable levels of ABCs to those of healthy individuals (5,93±0,6% vs 4,06 ±0,4 % respectively). Non-responders had statistically significant increased levels of ABCs compared to healthy controls (p<0,001). Also, non-responders had ABCs levels similar to those of aplastic anemia patients at presentation (29,08±1,62% vs 31,7 ±2,97 % respectively, p=0,44).
Conclusion: Although the number of patients analyzed is limited, our preliminary results suggest that aplastic anemia patients show expanded ABCs compared to age-matched control subjects, and these numbers are related to disease status. Further analysis of a larger pool of subjects is underway along with the examination of the specific transcription factor Bcl-6, that is implicated in T-bet expression in ABCs. These results will reveal the role of ABCs in the immune pathogenesis of aplastic anemia.
Figure 1. Figure 1.

Disclosures
Kattamis: CELGENE: Consultancy, Honoraria; Vifor Pharma: Consultancy; Novartis: Consultancy, Honoraria; ApoPharma: Honoraria.