To the editor:

We read with great interest the article by Isnardi et al1  wherein the authors showed very similar phenotypic and genotypic features of CD21−/lo B cells as previously reported.2  In view of the polyreactivity, increased autoreactivity, and strongly reduced calcium response and proliferative capacity of CD21−/lo B cells—reminiscent of anergic B cells in mouse models—the authors propose that CD21−/lo B cells represent anergic human B cells.1  Moreover, they showed that the majority of highly HEp2-reactive CD21−/lo B-cell clones express kappa (κ) light chains and suggested that the receptor editing using lambda (λ) light chains might not be induced in those clones serving as a mechanism to silence autoreactive B cells. We would like to suggest a different interpretation of the origin of CD21low B cells.

Anergy in CD21low B cells in patients with common variable immunodeficiency (CVID) is not complete. In the original model of hen egg lysozyme–transgenic mice,3  anergy in B cells represents a mechanism of central tolerance caused by B cell receptor (BCR)–induced hyporesponsiveness, which is associated with down-regulated BCR, poor calcium response, defective up-regulation of CD86, failure of antibody production, and impaired proliferation.3,4  Although poor Ca++ response and impaired proliferation are both features that CD21low B cells share with anergic mouse B cells, CD21low B cells of CVID patients have not down-regulated their surface BCR (Figure 1A-B) and are able to up-regulate activation markers like CD86 (unpublished data) as well as ICOS-L, TACI, and Fas surface expression upon BCR stimulation.1  Even more importantly, CD21low B cells produce higher levels of IgM than naive B cells after stimulation in vitro.2  In addition, a comparison of gene-expression profiles of mouse anergic B cells3,4  and human CD21low B cells2  showed only little overlap: only 3 (11.5%) of 26 genes describing the signature of anergic mouse B cells were identically expressed in CD21low B cells. In contrast, these cells closely resemble2  the recently characterized tissue memory-like B cells in tonsils5  and circulating “exhausted” CD21low B cells in HIV patients, suggesting rather a mechanism of activation-driven peripheral exhaustion underlying this anergic phenotype.6  This would also better explain the accumulation of CD21low B cells in the bronchoalveolar fluid of CVID patients or the synovial fluid of rheumatoid arthritis patients.2 

Figure 1

IgM surface expression and light chain usage in CD21low B cells. (A) Representative FACS plots for the expression of indicated B-cell markers in B cells of CVID patients (n = 10). CD19+IgM+ B cells were gated on CD21low and CD21+ naive B cells, according to CD38 and CD21 expression, and IgM surface expression was analyzed as the mean fluorescence intensity (MFI) in CD21+ and CD21low B-cell subpopulations, respectively. (B) The diagram shows no significant difference in the cell-surface expression of IgM between CD21+ and CD21low B cells in CVID. (C) Representative FACS plots demonstrate the expression of IgM and λ light chain on CD21low and naive B cells of CVID patients (n = 4). (D) The diagram shows a significantly decreased usage of κ-light chains in CD21low B cells of CVID patients compared with naive B cells of the same individuals.

Figure 1

IgM surface expression and light chain usage in CD21low B cells. (A) Representative FACS plots for the expression of indicated B-cell markers in B cells of CVID patients (n = 10). CD19+IgM+ B cells were gated on CD21low and CD21+ naive B cells, according to CD38 and CD21 expression, and IgM surface expression was analyzed as the mean fluorescence intensity (MFI) in CD21+ and CD21low B-cell subpopulations, respectively. (B) The diagram shows no significant difference in the cell-surface expression of IgM between CD21+ and CD21low B cells in CVID. (C) Representative FACS plots demonstrate the expression of IgM and λ light chain on CD21low and naive B cells of CVID patients (n = 4). (D) The diagram shows a significantly decreased usage of κ-light chains in CD21low B cells of CVID patients compared with naive B cells of the same individuals.

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In addition, in our cohort of CVID patients we cannot confirm a defect in secondary editing and λ light chain usage. We actually found that CD21low B cells preferentially express λ-chains compared with the naive B cells of CVID patients, which was also the case in 1 of the 3 CVID samples shown in Figure 2D of Isnardi and colleagues' article.1  In our experience the κ/λ ratio in CD21+ naive B cells (1.24 ± 0.03; n = 4) of CVID patients was within the normal range (1.0-2.1), whereas CD21low B cells showed a significantly decreased usage of κ-chains (0.69 ± 0.10; n = 4; P = .0019) compared with naive B cells of the same patients (Figure 1C-D). In addition, our analysis of κ recombination excision circles (KREC)7  had revealed a more extensive proliferative history of CD21low B cells compared with naive B cells.2  We appreciate the beautiful demonstration of increased autoimmune reactivity of CD21low B cells by Isnardi et al,1  but our data do not support an editing defect in central B-cell selection as the underlying etiology.

According to our data it seems more likely that CD21low B cells of CVID patients are related to tissue-like memory B cells in tonsils of immunologically healthy individuals5  and therefore resemble tissue associated innate-like B cells, which reveal partial anergy in vitro due to their “exhausted” preactivation state in vivo.

Acknowledgments: We thank Dr Hermann Eibel for his valuable discussions.

This research was funded by the German Research Foundation (DFG) grant SFB620 (project C1), the 7th framework program of the European Union grant Nr. HEALTH-F2-2008-201549, and the Federal Ministry of Education and Research (BMBF 01 EO 0803).

Contribution: M.R., S.G., B.K., and M.S. performed research and analyzed and interpreted data; H.-H.P. and K.W. designed the study; and M.R. and K.W. cowrote the letter.

Conflict-of-interest disclosure: The authors declare no competing financial interests.

Correspondence: Klaus Warnatz, MD, Centre of Chronic Immunodeficiency, Division of Rheumatology and Clinical Immunology, University Medical Center Freiburg, Breisacher Str 117, D-79106 Freiburg, Germany; e-mail: klaus.warnatz@uniklinik-freiburg.de.

1
Isnardi
 
I
Ng
 
YS
Menard
 
L
, et al. 
Complement receptor 2/CD21- human naive B cells contain mostly autoreactive unresponsive clones.
Blood
2010
, vol. 
115
 
24
(pg. 
5026
-
5036
)
2
Rakhmanov
 
M
Keller
 
B
Gutenberger
 
S
, et al. 
Circulating CD21low B cells in common variable immunodeficiency resemble tissue homing, innate-like B cells.
Proc Natl Acad Sci U S A
2009
, vol. 
106
 
32
(pg. 
13451
-
13456
)
3
Glynne
 
R
Akkaraju
 
S
Healy
 
JI
Rayner
 
J
Goodnow
 
CC
Mack
 
DH
How self-tolerance and the immunosuppressive drug FK506 prevent B-cell mitogenesis.
Nature
2000
, vol. 
403
 
6770
(pg. 
672
-
676
)
4
Glynne
 
R
Ghandour
 
G
Rayner
 
J
Mack
 
DH
Goodnow
 
CC
B-lymphocyte quiescence, tolerance and activation as viewed by global gene expression profiling on microarrays.
Immunol Rev
2000
, vol. 
176
 (pg. 
216
-
246
)
5
Ehrhardt
 
GR
Hijikata
 
A
Kitamura
 
H
Ohara
 
O
Wang
 
JY
Cooper
 
MD
Discriminating gene expression profiles of memory B cell subpopulations.
J Exp Med
2008
, vol. 
205
 
8
(pg. 
1807
-
1817
)
6
Moir
 
S
Ho
 
J
Malaspina
 
A
, et al. 
Evidence for HIV-associated B cell exhaustion in a dysfunctional memory B cell compartment in HIV-infected viremic individuals.
J Exp Med
2008
, vol. 
205
 
8
(pg. 
1797
-
1805
)
7
van Zelm
 
MC
Szczepanski
 
T
van der Burg
 
M
van Dongen
 
JJ
Replication history of B lymphocytes reveals homeostatic proliferation and extensive antigen-induced B cell expansion.
J Exp Med
2007
, vol. 
204
 
3
(pg. 
645
-
655
)

Author notes

M.R. and S.G. contributed equally.