REVIEW

Natural autoantibodies and associated B cells in immunityand autoimmunity

KAIISSAR MANNOOR1, YANG XU2, & CHING CHEN1

1Department of Pathology, University of Maryland, Baltimore, Maryland, USA, and 2Central South University Xiangya

School of Medicine, Changsha, China

(Submitted 7 November 2012; accepted 7 November 2012)

AbstractA substantial proportion of circulating antibodies in healthy individuals exhibit self-reactivity. These antibodies, referred to asnatural autoantibodies, are thought to arise naturally without actual antigen stimulation as they are present in human cordblood and in mice housed in germfree conditions and fed an antigen-free diet. Natural autoantibodies are mainly of the IgMclass, unmutated, and typically polyreactive. They provide critical early protection against pathogens, and play important rolesin maintenance of homeostasis and modulation of innate and adaptive immune responses, thereby conferring protection fromrampant autoimmune and inflammatory injuries. In this review, we summarize current information regarding the properties ofnatural autoantibodies and the B cells that produce them, their roles in immunity and autoimmunity, their mechanisms ofaction, and their therapeutic potential.

Keywords: natural antibodies, autoantibodies, B cells, autoimmunity

The properties of natural autoantibodies

In the 1980s, two groups of researchers led by Stratis

Avrameas in France [1–5] and byAbnerNotkins in the

USA [6–9], respectively, published a series of studies

demonstrating the existence of natural autoantibodies

(NAAs). Such antibodies (Abs) are present in cord

blood of newborn humans [10–12] and in newborn

mice [13], as well as in mice housed in germfree

conditions and fed an Ag-free diet [14].

The composition of serum IgM NAA is independent

of external stimulation [15]. Therefore, NAAs are

considered to arise “naturally,” similar to themolecules

of the innate immune system. NAAs are evolutionarily

conserved: they have been found in all jawed

vertebrates, from cartilaginous fish to amphibians,

birds, and mammals [16,17]. NAAs constitute a

substantial proportion of the normal serum Abs [3].

B cells that produce NAAs comprise 15–20% of the

circulating B cells in adults and 50% of the B cells in

cord blood of newborns [11].

A defining feature of NAA is the broad spectrum of

their binding specificities, i.e. each NAA is capable of

binding multiple structurally unrelated antigens such

as proteins, polysaccharides, nucleotides, and phos-

pholipids [4,18–20], many of which are components

of self-constituents. Although polyreactive, the bind-

ing of NAA is selective rather than nonspecific in that

each NAA has its own fine specificity pattern [1,9,20].

The majority of NAA are IgM, but they may also be

IgA or IgG [1,3,4]. The structure of NAA has been

extensively studied. Early studies indicated that the

general biochemical and immunologic features of

NAA are comparable to those of antigen-induced,

monospecific antibodies [21,22]. NAA are usually

encoded by germline Ig variable region genes

with little or no somatic mutation [20,23,24].

However, they use the same spectrum of VH and VL

genes as those used by antigen-induced antibodies

[20,23,25,26].

To understand the cellular origin of the so-called

Group II anti-phosphocholine (PC) antibodies, we

generated a panel of 49 hybridomas from non-

immunized neonatal and adult mice [20]. The

hybridomas were originally selected based on their

Correspondence: Ching Chen, Department of Pathology, University of Maryland, 22 Green St, Baltimore, Maryland 21201, USA.E-mail: qchen@som.umaryland.edu

Autoimmunity, March 2013; 46(2): 138–147q Informa UK, Ltd.ISSN 0891-6934 print/1607-842X onlineDOI: 10.3109/08916934.2012.748753

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Group II anti-PC phenotype, i.e., binding to

PC-protein and p-nitrophenyl-PC (NPPC) but not PC.

Subsequently their binding activities were tested on a

panel of 15 antigens, including self and foreign

proteins, DNA, polysaccharides and various haptens.

Nearly all hybridomas from neonates and more than

50% from adults showed polyreactivity, ranging from

2 to 12 antigens. This is in sharp contrast to the classic

type of antigen-induced Group II anti-PC antibodies

that bind only the immunizing antigen. However, the

polyreactivity is not nonspecific or due to “stickiness”

because each individual monoclonal antibody had a

distinct binding profile and some could distinguish

structurally similar antigens such as DNP and TNP,

indicating a high degree of discrimination.

The structural basis for such diverse, yet seemingly

specific binding is not clear. Does a single antigen-

binding pocket recognize different ligands, perhaps via

ligand-induced conformational changes? Or do differ-

ent antigens bind to different regions outside of

the binding pocket? To test these possibilities, we

performed cross-inhibition assays using a prototypic

polyreactive NAA and four different antigens. We

found that each of the four antigens was able to inhibit

antibody binding to the other antigens [20]. This

indicated that the various antigens were recognized by

the same antigen-binding site of the polyreactiveNAA.

To further understand the structural basis of

polyspecificity, we generated several matching pairs

of NAA and antigen-induced antibodies that were

encoded by the same VH-VL combinations. We found

that the only prominent structural difference between

these two types of antibodies resided in the third

hypervariable region of the heavy chain [VH CDR3]

[20]. To more precisely determine the antibody

sequences that are crucial for polyspecificity, Casali

and colleagues performed a series of elegant experi-

ments using gene assortment, gene shuffling and site-

directed mutagenesis strategies [27,28]. They found

that the main structural correlate for multiple antigen

binding was provided by the VHCDR3 [29]. This was

confirmed by other investigators [30,31]. Crystal

structural analysis of the antibody molecules has

shown that the H chain CDR3 forms the physical

center of the combining site of many antibodies

[32,33] and that it has the highest degree of variation

in amino acid composition and length. It is therefore

likely that the VH CDR3 plays the most important

role in determining the poly/auto reactivity of NAAs.

The B cells that produce NAAs

Mature B cells in adult mice can be divided into three

major populations based on their phenotypes and

anatomic locations: B1, follicular [FO] and marginal

zone [MZ] B cells [34]. In mice, it has been widely

accepted that the CD5þ B1 cells are the major

producers of NAA [35–37]. Indeed, B1 cell-derived

antibodies bear similarity to NAAs in that they often

recognize self-antigens such as phosphatidylcholine

and carbohydrate epitopes on cell membrane glyco-

proteins, as well as common bacterial antigens such as

phosphocholine. In addition to B1 cells, MZ B cells

may also be an important source of NAAs since B cells

with a low level of self-reactivity or with reactivity to

bacterial wall components preferably reside in the

marginal zone [38].

However, there are several differences between

NAAs and B1-derived antibodies. First, NAAs use a

broad spectrum of Ig genes similar to those used by

Ag-induced antibodies [20,23,29] whereas B1 anti-

bodies are encoded by a limited set of VH/VL genes

[39–41], although recent studies showed a less

restricted V gene use by B1 cells [42]. Secondly,

many NAAs have N additions [20,24] whereas B1

antibodies typically lack N nucleotides [43]. Finally,

NAAs bind a large variety of foreign and self antigens

[1,3,20] while B1 antibodies are reactive mostly

toward cell membrane or bacterial wall components

[36]. Using a florescence antigen labeling strategy,

Notkins and colleagues have analyzed B cells that bind

multiple antigens [called polyreactive Ag-binding

B cells or PAB]. They found that PAB cells are widely

distributed in various lymphoid tissues and many of

them are not B1 cells [44]. Using Ig allotype chimeric

mice to identify B1 vs B2 cell-derived antibodies, it

has been shown that ,50% of the serum IgM

was produced by B1 cells and the remainder by

B2 cells [45]. Bone marrow reconstitution studies

have shown that B1 cells are not essential for NAA

regeneration [46].

To directly examine the origin and function of

NAA, we have established an IgH knock-in [KI]

mouse model [47] that expresses a prototypic NAA,

named ppc1-5, that characteristically binds to a

variety of self and foreign Ags, include DNA, actin,

and p-nitrophenyl-phosphocholine, among others.

The ppc1-5 NAA is encoded by a germline VH gene

of the 7183 family and a germline Vll gene. In the

H-only KI mice, the l1 B cells represent NAA-

producing B cells while most of the k B cells are

non-NAA and serve to maintain a relatively normal

and diverse B cell repertoire. Using this model, we

have found that the ppc1-5H/l1 NAA B cells

exhibited a phenotype that was different from that of

B1: they were negative for CD5 and CD43.

In addition, they were not concentrated in the

peritoneal cavity but rather were mainly located in the

follicles of the spleen and lymph nodes, and they were

part of the circulating B cells in the peripheral blood.

These are features of FO B cells. However, the ppc1-

5H/l1 B cells were distinguished from the bulk of FO

cells by their decreased sIgD and increased CD23

expression. Furthermore, these B cells had increased

levels of MHC Class II, CD69 and B7.2, and

decreased level of CD79b [Ig-b chain of the BCR],

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indicating BCR engagement and activation. It is not

clear whether the ppc1-5H/l1 NAAB cells represent a

distinct B cell population or they are a subset of FO

B cells with a distinct activation profile.

Despite their autoreactivity, the ppc1-5/l1 B cells

were not negatively selected, but rather they were

overrepresented in the peripheral lymphoid organs

[47]. The number of ppc1-5H/l1 NAA B cells

increased with maturation, from less than 7% in

the transitional B to nearly 30% in the mature

B compartments in the spleen, demonstrating a

positive selection during peripheral B cell maturation,

likely due to their self-reactivity. This is in agreement

with the seminal work by Hayakawa and colleagues,

who have shown that B1 cells specific for the Thy-1

glycoprotein self-antigen were positively selected by

the antigen [48]. These findings underscore the

functional importance of NAAs and associated B cells.

The role of NAAs and NAA-producing B cells

in immunity

There is substantial evidence that NAA, as an integral

part of innate immunity, are an essential first line of

defense against microbial invasion [49–54]. Such

activity is thought to be largely due to their ability to

recognize a broad spectrum of bacterial antigens,

thereby inhibiting pathogen growth either by direct

neutralization or by enhancing phagocytosis by

macrophages [50,55–57]. The importance of natural

IgM in protective immunity is best demonstrated by

studying mice that lack secreted IgM. These studies

have shown that natural IgM antibodies were required

not only in early effective control of bacterial and viral

infection but also in priming the ensuing IgG

responses [52–54,58,59].

A long-standing question regarding NAAs is

whether they are capable of participating in antigen

specific adaptive immune response, or if their

production is spontaneous. Studies with transgenic

mice have led to conflicting results. B cells producing

anti-phosphocholine natural antibodies could respond

to S. pneumoniae injection [34], whereas B cells

producing innate anti-influenza antibodies did not

participate in a primary immune response [49]. It has

not been determined whether NAAs can participate

in T cell-dependent [TD] memory response. The

ppc1-5H mice represent a particularly useful model to

examine TD response because the KI gene can

undergo somatic hypermutation [SHM] and class

switch recombination [CSR], both of which are

defining features of the TD response. In addition,

the ppc1-5 NAA can bind to a well-studied TD Ag

PC-KLH, thereby guaranteeing an initial engagement

with Ag upon PC-KLH immunization.

Using this model, we have shown that the ppc1-5

NAA B cells could give rise to a quick anti-PC IgMAb

response upon PC-KLH immunization, but they

neither sustained this response nor did they mount a

significant memory IgG response [60]. Immuno-

histologic examination of the spleen revealed many

IgM ppc1-5H/l1 antibody forming cells [AFCs] but

few, if any, IgG ppc1-5H/l1 AFCs. Moreover,

extensive efforts to isolate IgG ppc1-5H/l1 B cells

from memory response by hybridoma generation and

cDNA cloning have demonstrated the extreme paucity

of such B cells. These results point to a checkpoint in

the germinal center that prevents NAA from devel-

oping to high affinity IgG autoantibody production.

This could indicate an inability of these B cells to

receive T cell help in the GC. Alternatively, the ppc1-5

NAA may not be able to improve antigen binding via

SHM or such mutations may be detrimental to the

ppc1-5 antibody function, as shown previously in

the prototypic B1-derived anti-PC antibody T15

[61–64].

The lack of both CSR and SHM in ppc1-5 B cells

during TD immune response indicates a checkpoint

linked to the activation-induced cytidine deaminase

[AID], which is required for both events. AID

expression is induced by activation of B cells through

CD40, BCR and various other signaling pathways

[65]. In addition, AID levels are regulated by multiple

posttranslational factors [66]. As mentioned above,

the ppc1-5 NAA B cells have a unique phenotype and

appear to be chronically activated. In particular, they

have markedly reduced level of Igb [CD79b], which

may impair BCR signaling. We have shown that

ppc1-5 NAA B cells did not respond to anti-CD40

stimulation in vitro with regard to cytokine production

[67]. It is therefore possible that these NAA B cells,

while able to enter GCs, cannot receive or initiate

proper activation signals that are required for

induction of AID expression.

Although CSR and SHM were limited in ppc1-5

NAA B cells, most of the isolated IgG B cell clones

from the KI mice had replaced the ppc1-5 VH with an

endogenous VH via H chain editing [60]. At least

some of these editing events took place in the germinal

centers as demonstrated by the presence of VHppc1-5

double-stranded DNA breaks in the GC B cells and

the expression of RAG1 and RAG2 genes. These

results are consistent with an earlier finding that B1

cells have high levels of receptor editing [68].

Therefore, receptor editing may be an important

mechanism for preventing expression of NAA BCR in

IgG B cells during adaptive immune response.

The role of NAAs and NAA-producing B cells in

autoimmunity

The role of NAAs in autoimmunity is not completely

settled. Many NAAs have reactivity towards con-

served self-components such as DNA, histones,

nucleoproteins, and phospholipids, which are also

the common targets of pathologic autoantibodies in

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autoimmune diseases. This is therefore conceivable

that NAAs could potentially serve as precursors of

pathologic autoantibodies. This notion is supported

by the finding that patients with systemic autoimmune

diseases have increased levels of polyreactive B cells

[69,70]. In addition, examples exist where NAAs can

obtain increased autoreactivity via V gene mutation

and isotype switching [71–73].

In contrast, recent evidence has pointed to a role of

natural IgM in maintaining self-tolerance. Two groups

of investigators independently generated mice

deficient in secreted IgM, but retaining normal

B cell numbers and IgG Abs [74,75]. The sIgM-

deficient mice developed lupus-like autoimmune

symptoms in normal background and had exacerbated

autoimmune disease in the autoimmune MRLlpr

background [76,77], indicating a protective role of

natural IgM. Diaz and colleagues have analyzed AID-

deficient mice, which lack IgG but have increased

levels of natural IgM. They found that development of

lupus nephritis was abrogated in AID deficient

MRL-lpr mice [78] and that the protective effect

could be ascribed to the autoreactive IgM [79].

To directly address the role of NAA and

NAA-producing B-cells in the development of auto-

immune disease, we crossed the ppc1-5 NAA

knock-in gene onto the MRL-lpr background. We

have found that the expression of ppc1-5H in MRL-

lpr mice resulted in near complete protection from

lupus nephritis as shown by prevention of proteinuria

and drastically reduced kidney pathology and immune

complex deposition [67]. Consequently, the mice had

significantly prolonged survival. The ppc1-5 KI mice

had significantly reduced levels of IgG anti-dsDNA

and anti-Sm/RNP antibodies. In addition, there was a

skewing of the IgG subclass profile: while the wild-

type [wt] MRL-lpr mice had high titers of IgG2a and

IgG3, both of which are highly pathogenic in lupus

nephritis, the ppc1-5 KI mice had a predominance of

the least pathogenic IgG1 subclass.

To determine whether the protective effects seen in

ppc1-5 KI mice are, at least in part, due to secreted

NAA per se, we injected the purified ppc1-5 IgM to wt

MRL-lpr mice. We observed a significant reduction in

proteinuria and much improved survival although

these are not as dramatic as seen in ppc1-5 KI mice

[67]. Injection of ppc1-5 IgM also resulted in a

markedly decreased level of anti-Sm/RNP antibodies

while the anti-dsDNA IgG antibody levels were not

significantly affected [unpublished data]. These

findings are consistent with earlier reports from

other investigators that administration of anti-

dsDNA IgM had protective effects in animal models

of SLE [79,80]. Studies from Silverman’s group have

demonstrated a protective role of the T15 anti-PC

natural IgM in collagen-induced arthritis [81].

In humans, it has been found that the level of

anti-dsDNA IgM and the ratio of IgM to IgG

antidsDNA were inversely correlated with the severity

of lupus nephritis in SLE patients [82,83]. Similarly,

higher titers of natural anti-PC IgM were correlated

with low disease activity in SLE patients [84,85]. By

analyzing lupus patient’s sera against a multiplex

autoantigen microarray, it has been found that the

presence of IgM polyreactivity was correlated with

reduced disease severity [86]. These findings demon-

strate a regulatory and protective function of natural

IgM in human autoimmune diseases. However, the

cellular origin of human protective IgM remains to be

identified.

Mechanisms of action

There is strong evidence that natural antibodies with

immune regulatory and protective functions are self-

reactive. However, the exact antigen specificities that

are essential for their function are unclear. Several

reports have demonstrated an association between

anti-dsDNA specificity and the protective effects

[79,80]. However, it has not been thoroughly

investigated whether these anti-dsDNA IgM anti-

bodies are also capable of binding to other antigens, in

particular to epitopes on apoptotic cells. It has been

shown that anti-dsDNA IgG antibodies frequently had

reactivity towards apoptotic cell membrane [87]. The

prototypic B1-derived natural Ab, T15, has specificity

towards PC, an epitope present on bacterial wall, in

oxidized LDL and on apoptotic cell membrane. The

protective function of T15 is dependent on its binding

to apoptotic cells [81]. As described before, the ppc1-5

NAA binds DNA [strongly for ssDNA and weakly for

dsDNA], NPPC, and apoptotic cells [unpublished

data]. Therefore, binding to apoptotic cells may be a

common feature of protective NAAs.

Apoptotic cells are constantly generated in the body

and are a major source of lupus-associated autoanti-

gens [88–90]. Deficiencies in clearance of apoptotic

cells result in autoantibody production and auto-

immune diseases [91–94]. When cells are undergoing

apoptosis, cellular constituents in the nuclei, cyto-

plasm and membranes are fragmented, reorganized

and repacked to form neo-self-Ags [95,96]. These

neo-self-Ags include histones, nucleosomes and

phospholipids [96,97]. Many broadly reactive NAAs

bind these apoptosis-associated Ags, which, in turn,

promotes clearance of apoptotic cells. Indeed, IgM is

required for complement-mediated clearance of

apoptotic cells [98–100]. Mice deficient in secreted

IgM are defective in clearance of apoptotic cells and

have accelerated autoimmune disease [77, 81, 101].

The T15 anti-PC IgM can bind to the PC-containing

neo-self Ag on the apoptotic cell membrane and

subsequently recruit C1q and mannose-binding lectin

[MBL]; this complex interacts with the scavenger

receptors on dendritic cells and macrophages, thereby

enhancing clearance of apoptotic cells [81, 102].

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It has been proposed that the inhibitory effect of

NAAs could be mediated by their anti-idiotypic

activity. For example, IgM NAA can block binding

of autoreactive IgG to their respective autoantigens via

interaction with the V-region of the IgG Abs [103].

The blocking activity is dose-dependent with maximal

inhibition occurring at a specific molar ratio between

the patient’s IgG and a given antiidiotypic IgM. It is

therefore thought that an idiotypic network is

functioning to maintain a low level of IgG auto-

reactivity. This notion is supported by the finding that

anti-idiotypic Abs are constantly present in healthy

individuals and in patients recovered from auto-

immune disease, but they are depleted during active

disease [104, 105].

NAAs can modulate the function of dendritic cells

[DC]. Studies from Silverman’s group have demon-

strated that the T15 Ab, by virtue of binding to

apoptotic cells and recruitment of C1q and MBL, can

suppress TLR-mediated activation and maturation of

DCs [81]. Treatment with T15 Ab inhibited both

in vitro and in vivoDC responses to agonists to TLR-3,

TLR-4, TLR-7 and TLR-9, resulting in decreased

production of IL-6, IL-12, IL-17, TNFa, and a variety

of chemokines. There was also a reduction in the

expression of MHC II, CD40, CD80 and CD86 on

DCs. Therefore, the NAAs not only facilitate removal

of apoptotic cells, but also function as immune

regulators.

B cells that produce NAAs may possess functions

independent of antibodies. Recent studies have

demonstrated diverse functions of B cells in immunity

and autoimmunity. B cells can serve as Ag presenting

cells [106], play a crucial role in T-cell receptor

diversification [107] and in dendritic cell maturation.

They are also essential for secondary lymphoid organ

formation [108]. B cells produce many kinds of

cytokines [109, 110], and, like T-cells, can be divided

into functionally distinct subsets [109]: B effector 1

[Be-1] cells produce cytokines associated with

Th1 response [IFNg and IL-12], and B effector 2

[Be-2] cells make Th2 type cytokines [IL-2, IL-4

and IL-13].

B cells are potent Ag presenting cells [APC] [106].

There is clear evidence that B cells as APC can induce

T cell tolerance. By targeting antigen to B cells, Parker

and colleagues first demonstrated that small resting

B cells, acting as APC, were able to induce T cell

tolerance [111]. Subsequent studies from several

groups confirmed this finding [112–114], and further

demonstrated that tolerance induction was mediated

by up-regulation of negative co-stimulatory molecules

such CTLA-4 and PD-1 on CD4 þ T cells. There are

reports that B cells expressing polyreactive BCRs

could present antigen to T cells but failed to cause

T cell proliferation [115], suggesting tolerance

induction in T cells. By expressing the ppc1-5 NAA

in MRL-lpr mice, we have shown a decreased CD69

expression and an increased CTLA-4 expression on

CD4þT cells [67]. Such changes were not observed

in wt MRL-lpr mice infused with ppc1-5 IgM

[unpublished data]. Therefore, the ppc1-5 NAA

B cells, but not the secreted Ab, can regulate T-cell

function.

Recently, regulatory B cells, called Breg, have been

identified inmice [116–119] and in humans [120, 121].

Using B-cell deficient mouse models or cell transfer

strategies, several groups have demonstrated a

protective and regulatory role of B cells in various

autoimmune and inflammatory conditions, including

experimental autoimmune encephalomyelitis [122, 123],

inflammatory bowel disease [124, 125], collagen

induced arthritis [126, 127], non-obese diabetes

[128], contact hypersensitivity [119], and lupus

[129]. The origin and phenotype of Bregs are not

completely understood. In some experimental systems

Bregs have a marginal zone [MZ] or transitional 2

[T2] B-cell phenotype with expression of CD21 and

CD1d, yet in others they have a B1-like phenotype

with expression of CD5 and CD1d. Nonetheless,

a common feature of these Bregs is the production of

IL-10. Therefore, Bregs are also called B10 cells

[119]. IL-10 is a key regulatory cytokine that inhibits

inflammation mainly by suppressing proinflammatory

cytokine production by innate immune cells [116, 130].

However, the effects of IL-10 depend on the target

cells; it can activate B cells and promote autoantibody

production [130].

Several studies have shown that toll-like receptor

[TLR] signaling can induce regulatory functions

and/or IL-10 production in B cells [131–133].

Other studies have demonstrated a requirement for

BCR and CD40 signaling in the induction of IL-10

producing Breg [126]. It has been postulated that

there are two kinds of regulatory B cells: the “acquired

type” of Bregs are induced by BCR and CD40

stimulation whereas the “innate type” respond to TLR

stimulation [116].

We have found that the ppc1-5 NAA B cells had a

vigorous proliferative response and produced large

amount of IL-10 upon stimulation by CpG [TLR-9

agonist] and, to a lesser extend, by LPS [TLR-4

agaonist] [67]. Therefore, The ppc1-5 NAA B cells

may be examples of Bregs. The NAA-expressing

B cells may be especially responsive to TLR

stimulation because their polyreactive BCRs often

bind TLR ligands such as DNA, RNA and microbial

antigens, which in turn facilitate TLR activation by

dual engagement of BCR and TLR [134, 135].

In addition to B cells, the CD4 þ T cells from

ppc1-5 KImice also producedmore IL-10 than T cells

from wt mice upon anti-CD3 stimulation in the

presence of ppc1-5 B cells [67]. This is in agreement

with a previous report that co-culture of IL-10-

producing B cells with CD4 þ T cells induced

differentiation of CD4 þ IL-10 þ regulatory T cells

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[Tr1 cells] [129]. This was thought to be a mechanism

by which regulatory B cells control autoimmunity.

Some B cell populations can also induce CD3þ

NK1.1þ regulatory T cells [125] and CD4þ

FoxP3þ Tregs [136]. IL-10 and regulatory B cells

can suppress Th1 and Th17 T cells, and influence

Th1/Th2 polarization [116, 129, 132]. Our finding

[67] that the ppc1-5 NAA KI mice had significantly

reduced Th1 type IgG2a and IgG3 Abs and increased

Th2 type IgG1 Abs supports this notion.

In summary, the current data suggest that NAA and

NAA B cells suppress inflammatory and autoimmune

disorders by several mechanisms. The soluble NAAs

can suppress autoimmunity by promoting removal of

apoptotic cells and self-antigens and by modulating

maturation/activation of dendritic cells. The IgM

NAAs may, in some cases, suppress IgG autoantibody

production by antiidiotypic activity. The NAA B cells

can function as tolerogenic APCs by inducing

expression of negative regulators such as CTLA-4

and PD-1. The NAA B cells, upon activation by

TLR ligands, can differentiate to IL-10 producing

regulatory B cells, which, in turn, can induce Tr1

and Treg cells and suppress Th1 and Th17

differentiation (Fig. 1).

Therapeutic Potential

Several studies have shown that infusion of natural

IgM can efficiently ameliorate autoimmune manifes-

tations in mouse models of SLE and collagen-induced

arthritis. These NAAs include antidsDNA [79,80],

anti-PC [81], and poly/autoreative IgM [67]. In

humans, the anti-inflammatory effects of pooled

natural IgG [IVIG] have long been recognized, and

it is now widely used in a number of autoimmune and

inflammatory diseases [137, 138]. More recently, it

has been shown that pooled human IgM [IVIgM], as

well as IgM enriched [75% IgM] and IgM-containing

[12% IgM] preparations have beneficial therapeutic

effects in several experimental autoimmune diseases

and in protecting from graft-versus-host disease in

bone marrow transplant recipients [139–141]. The

effects of IVIgM were stronger on a molar basis than

those of IVIG [139]. An IVIG preparation enriched

for anti-dsDNA anti-idiotypic antibodies [IVIG-ID]

has shown superior therapeutic effects over conven-

tional IVIG in the treatment of murine SLE [142]. It is

therefore conceivable that enriched or engineered

auto/polyreactive IgM preparations may represent a

class of more effective therapeutic antibodies.

NAA

NAA B cell

TLR9 TLR7

TLR4

B cell

IL-10

MΦ

AC

DCAC

Clearance of apoptotic cellsregulation of DC

Suppress inflammationprotect from tissue injury

Suppression ofTh1 and Th17 Induction of Tregs

Decreasedproduction of IgG

T cellnaive

T cellanergy

T cellTreg

APC

APC

Figure 1. Mechanisms by which NAA and NAA B cells suppress inflammation and autoimmunity. When NAA-expressing B cells are

activated by dual engagement of BCR and TLR, they secrete large amount of 1gMNAA and regulatory cytokines such as IL- 10. The soluble

NAA can bind to apoptotic cells (AC) and facilitate their clearance bymacrophages (MF) and dendritic cells (DC). The AC-IgM complex can

modulate DCmaturation and activation. The secreted IL-b can suppress Th1 and Th17 cells and induce Tr1 and Treg cells. The NAA B cells

can also function as tolerogenic APC and induce T cell anergy.

Natural autoantibodies and autoimmunity 143

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The prospect that B cells expressing NAA have

regulatory functions has important therapeutic

implications. We explored this possibility by infusing

NAA-expressing B cells to MRL-lpr mice. Our

preliminary results showed a significant suppression

of autoimmune nephritis in mice treated with

NAA B cells. Recently, regulatory B cells were also

identified in humans [121]. In addition, a substantial

proportion of human circulating B cells express

NAA-like polyspecific BCRs [44]. In the future, a

better knowledge of the properties of human Bregs

could open the door to exploitation of such B cells as

novel therapeutic agents for the treatment of

autoimmune diseases.

Declaration of interest: The authors report no

conflicts of interest. The authors alone are responsible

for the content and writing of the paper.

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