Cholinergic B cells: New players in immune regulation and tissue homeostasis

As key players in humoral immunity, B cells play diverse roles in immune responses and disease development. Although early studies focused on the hallmark features of antibody production in B cells, increasing evidence has demonstrated the effector functions of cytokine secretion and antigen presentation by B cells in the development of various human diseases. Recent studies on the interactions of immune cells and nerve systems through neurotransmitters, including acetylcholine (ACh), have revealed novel functions of B cells in immune regulation and tissue homeostasis. Two newly published studies in Nature Immunology and Immunity demonstrated that B cells produce ACh to modulate antiviral inflammatory responses and liver regeneration after injury [1, 2]. These pivotal findings suggest the existence of a new functional subset of ACh-producing B cells, termed “cholinergic B cells”, which are characterized by their ability to synthesize ACh via high expression of choline acetyltransferase (ChAT). Cholinergic B cells span both innate B1 and conventional B2 cells and play a unique role in bridging neuroimmune crosstalk. Thus, elucidation of the functional features of cholinergic B cells will open new avenues for therapeutic intervention in acute inflammatory and regenerative disorders.

It has been reported that vagus nerve-derived ACh suppresses proinflammatory cytokine production by macrophages through a cholinergic anti-inflammatory pathway [3]. Moreover, immune cells, including B cells, produce massive amounts of ACh upon inflammation induction, suggesting neuron-independent production of ACh. ChAT⁺ cholinergic B cells have been shown to regulate local innate but not adaptive immunity in an adoptive transfer model of colitis [4]. Notably, B cells are important sources of ACh in the bone marrow, and cholinergic B cells limit steady-state and emergency hematopoiesis by regulating α7-nicotinic-ACh receptor (α7nAChR)⁺ bone marrow stromal cells [5]. The available evidence also shows that B-cell-derived ACh suppresses acute and chronic cardiovascular inflammation and disease severity by regulating immune cell responses [5]. These findings provide a solid rationale for in-depth investigations on how cholinergic B cells affect pathophysiological processes across diverse organs and tissues. Here, the findings from two newly published papers support the notion that cholinergic B cells are a distinct subset with immunomodulatory functions in viral infections and tissue regeneration.

Cembellin-Prieto et al. elegantly reported that cholinergic B cells are sensitive and critical regulators for the early control of viral infection and tissue damage to the respiratory tract (Fig. 1). Using ChAT-green fluorescent protein (GFP) reporter mice and an influenza A viral infection model, the authors identified B cells, but not T cells, as the primary ACh-producing leukocytes in the respiratory tract at both the baseline and postinfection stages [1]. To elucidate the functional roles of B-cell-derived ACh, B-cell-specific ChAT knockout (ChatBKO) mice lacking ACh production in B cells were infected with influenza virus. Despite the reduced lung viral loads at 1 day postinfection, these ChatBKO mice presented increased epithelial degeneration in the nasal cavities; decreased integrity, function and maturation of the alveolar epithelium; and poorer overall health at 7–10 days postinfection, suggesting that cholinergic B cells regulate lung viral loads at an early stage and restrain excessive tissue inflammation and damage at a later stage. Cholinergic B cells were found in close proximity to macrophages in infected lungs. Mechanistically, B-cell-derived ACh suppressed activation and TNF-α production by α7nAChR-expressing interstitial macrophages but not alveolar macrophages, indicating a spatially restricted regulatory role of cholinergic B cells due to the short half-life and effector distance of ACh. These findings highlight the critical regulatory roles of cholinergic B cells in antiviral inflammation by balancing tissue protection and pathogen control, which may provide new therapeutic targets for respiratory infections such as influenza and COVID-19.

Fig. 1.

Cholinergic B cells in immune regulation and tissue homeostasis. ACh-producing cholinergic B cells participate in liver regeneration, lung viral infection, skin inflammation and bone marrow hematopoiesis. Cholinergic B cells promote liver tissue regeneration by enhancing IL-6 production by Kupper cells and reducing IFNγ production by hepatic CD8 + T cells. During lung viral infection, cholinergic B cells suppress TNF-α expression produced by interstitial macrophages to regulate antiviral inflammatory responses and tissue damage. ACh-producing B cells also inhibit keratinocyte proliferation during skin inflammation. In the bone marrow, B-cell-derived ACh inhibits hematopoiesis by targeting stromal cells

In a parallel study, Modares et al. investigated the roles of B-cell-derived ACh in liver regeneration via a partial hepatectomy (PHX) mouse model [2]. B cells, rather than T cells, rapidly increased ChAT expression after PHX. Notably, ChatBKO mice exhibited high mortality with failed liver regeneration post-PHX, which was rescued by adoptive transfer of wild-type B cells, suggesting that cholinergic B cells respond rapidly to acute liver injury and play an essential role in liver regeneration. The phenotype of impaired liver regeneration in α7nAChR-deficient mice closely resembled that observed in ChatBKO mice, highlighting the indispensable roles of α7nAChR among the ACh receptors in facilitating cholinergic B-cell-mediated liver regeneration. Mechanistically, B-cell-derived ACh targets two α7nAChR-expressing immune cell populations in the liver: Kupffer cells and CD8⁺ T cells. In Kupffer cells, ACh promoted the production of IL-6, a key cytokine for hepatocyte proliferation. In the absence of cholinergic B cells, Kupffer cells exhibit an immunosuppressive phenotype with reduced IL-6 production and increased expression of anti-inflammatory genes. Moreover, ACh suppressed IFNγ production by CD8⁺ T cells to prevent excessive inflammation from damaging the liver. This biaxial regulation that promotes regenerative signals by Kupffer cells and dampens inflammatory signals by T cells underscores the pivotal role of cholinergic B cells in liver repair and regeneration (Fig. 1).

In both viral infection and liver regeneration models, B1 cells constitute one of the dominant populations for ChAT expression at inflammatory sites [1, 2]. Conventional mature B2 cells are important sources of ACh in the spleen, lung, mediastinal lymph nodes and liver, suggesting that ChAT⁺ cholinergic B cells span both the B1 and B2 subsets and exhibit diverse tissue localizations [1, 2]. Transcriptional analysis revealed distinct features of cholinergic B cells. The ChAT⁺ B cells presented marked transcriptional enrichment of genes associated with HIF-1, PI3K-Akt signaling, and axon guidance [2], suggesting the critical involvement of these pathways in the induction and maintenance of the cholinergic B-cell response. In contrast, the ChAT^- B cells presented enriched signatures of antigen processing and presentation, NF-kappa B signaling and chemokine signaling pathways [2]. BCR repertoire analysis revealed that post-PHX ChAT⁺ B cells were significantly more clonal than ChAT^- B-cell counterparts and were enriched in public IgM clones [2]. Consistent with previous findings that B cells produce ACh in a MyD88-dependent manner [4], cholinergic B cells respond to innate signals such as toll-like receptor 4 (TLR4) ligands [1, 2]. The levels of LPS markedly increased in the liver upon PHX, which may account for the rapid ChAT induction in B cells [2]. However, stimulation with both the adrenergic neurotransmitter norepinephrine and BCR induced ChAT expression in splenic B cells in culture [2], indicating that diverse upstream signals can induce cholinergic B cells. Since sympathetic nerves surround the hepatic arteries and portal veins, the roles of sympathetic nerve-derived NE in early cholinergic B-cell responses need further investigation. The presence of TLR ligands and the extensive distribution of sympathetic nerves suggest that cholinergic B cells may reside in a variety of organs and tissues.

In both studies, ChAT-GFP reporter mice and ChatBKO mouse models were used, which provide compelling evidence for B-cell-specific ACh production and support the concept of cholinergic B cells as a functional subset. Two studies highlighted B-cell-derived ACh as a nonneuronal regulatory molecule that acts through α7nAChR, a receptor known for its anti-inflammatory properties [3]. The available data show that α7nAChR mediates its effects across interstitial macrophages in the lung, Kupffer cells and CD8⁺ T cells in the liver during pulmonary viral infection and liver regeneration. The short half-life of ACh and the close contact of cholinergic B cells with target cells suggest a precise and localized regulatory circuit in these tissues. Interestingly, the suppressive effects of cholinergic B cells on TNF-α production by macrophages and IFNγ secretion by CD8 + T cells resemble those of regulatory B cells with immunosuppressive functions. The observed high expression of IL-10 and ACh by B1 cells may suggest developmental similarities between cholinergic B cells and IL-10-producing regulatory B cells. Moreover, LPS stimulation induces the generation of both regulatory B cells and cholinergic B cells. Despite these similarities, cholinergic B cells and regulatory B cells are distinct subsets on the basis of the available evidence. Although a small fraction of ChAT⁺ B cells produce IL-10, adoptive transfer experiments have shown that these cholinergic B cells do not suppress effector CD4 T-cell responses [6]. Although plasma cells have been shown to secrete IL-10 under specific conditions, TLR-induced ChAT expression is inhibited during plasma cell differentiation [1]. Regulatory B cells exhibit strong suppressive effects on many other immune populations. However, the functions of cholinergic B cells extend far beyond immune suppression and include tissue repair, liver regeneration, skin inflammation, hematopoiesis and neuroimmune crosstalk (Fig. 1) [1, 2, 4, 5]. Currently, it remains unclear whether cholinergic B cells and regulatory B cells share phenotypic and transcriptional features. Future studies are needed to elucidate the developmental, transcriptional, phenotypic, metabolic and functional characteristics of cholinergic B cells, especially the identification of key transcription factors defining this unique B-cell subset.

Recently, the characteristics and functions of cholinergic T cells in tumor development have been well documented [7]. The emerging roles of tumor-infiltrating B cells in cancers are gaining increasing attention [8]. B-cell-derived gamma-aminobutyric acid (GABA), an important neurotransmitter, has been reported to suppress tumor progression by regulating macrophage and CD8⁺ T-cell responses [9]. The existence and functional implications of cholinergic B cells in the tumor microenvironment warrant further investigation. B cells are central players in the development of many autoimmune diseases [10]. Cholinergic B cells have been shown to mitigate skin inflammation in an imiquimod-induced autoimmune psoriasis model (Fig. 1) [6]. Future investigations on cholinergic B cells will provide new insights into the underlying pathologic mechanisms and novel therapeutic interventions in autoimmune diseases. It has been proposed that the cholinergic activity of T cells could be targeted to enhance immunotherapy via various strategies [7]. These new findings suggest that cholinergic B cells may also have significant therapeutic potential. Similarly, cholinergic B cells can be targeted by chemicals that increase the level and duration of ACh action and by selective α7nAChR modulators that directly influence cholinergic B-cell functions. In respiratory infections, enhancing cholinergic B-cell activity or activating α7nAChR may reduce excessive tissue inflammation and damage. Pharmacological α7nAChR agonists or acetylcholinesterase inhibitors might be explored as adjunctive therapies for infectious diseases. In liver diseases, increasing cholinergic B-cell signaling may have therapeutic implications for enhancing regeneration with reduced inflammation, which could be applied in liver resection, transplantation, and chronic liver diseases. Currently, translating cholinergic B cells into clinical applications remains a major challenge because of the diverse sources of ACh from nerves and lymphocytes. Moreover, the roles of other ACh receptors, including muscarinic receptors, and their interactions with α7nAChR warrant further investigations to fully map the cholinergic immune network. These findings may pave the way for future clinical trials to validate the efficacy and safety of targeting cholinergic B cells in human diseases.

The characterization of B-cell-derived ACh has led to significant progress in understanding the diverse functions of B cells. The identification of cholinergic B cells adds a new layer of neuroimmune crosstalk to this field. By acting through α7nAChR, cholinergic B cells modulate the inflammatory response and tissue homeostasis during lung infection and liver regeneration. These new findings on the tissue-specific roles of cholinergic B cells reveal the importance of the local inflammatory milieu in shaping immune cell functions, which will facilitate the development of innovative therapies for organ-specific inflammatory and regenerative disorders.

Competing interests

The authors declare that they have no conflicts of interest. Dr. Liwei Lu is an editorial board member of Cellular & Molecular Immunology, but he has not been involved in peer review or decision-making related to the article.