Tobias Junt

Generation of Gut-Homing IgA-Secreting B Cells by Intestinal Dendritic Cells

Normal intestinal mucosa contains abundant immunoglobulin A (IgA)–secreting cells, which are generated from B cells in gut-associated lymphoid tissues (GALT). We show that dendritic cells (DC) from GALT induce T cell–independent expression of IgA and gut-homing receptors on B cells. GALT-DC–derived retinoic acid (RA) alone conferred gut tropism but could not promote IgA secretion. However, RA potently synergized with GALT-DC–derived interleukin-6 (IL-6) or IL-5 to induce IgA secretion. Consequently, mice deficient in the RA precursor vitamin A lacked IgA-secreting cells in the small intestine. Thus, GALT-DC shape mucosal immunity by modulating B cell migration and effector activity through synergistically acting mediators.

Gut microbiota metabolism of dietary fiber influences allergic airway disease and hematopoiesis

Metabolites from intestinal microbiota are key determinants of host-microbe mutualism and, consequently, the health or disease of the intestinal tract. However, whether such host-microbe crosstalk influences inflammation in peripheral tissues, such as the lung, is poorly understood. We found that dietary fermentable fiber content changed the composition of the gut and lung microbiota, in particular by altering the ratio of Firmicutes to Bacteroidetes. The gut microbiota metabolized the fiber, consequently increasing the concentration of circulating short-chain fatty acids (SCFAs). Mice fed a high-fiber diet had increased circulating levels of SCFAs and were protected against allergic inflammation in the lung, whereas a low-fiber diet decreased levels of SCFAs and increased allergic airway disease. Treatment of mice with the SCFA propionate led to alterations in bone marrow hematopoiesis that were characterized by enhanced generation of macrophage and dendritic cell (DC) precursors and subsequent seeding of the lungs by DCs with high phagocytic capacity but an impaired ability to promote T helper type 2 (TH2) cell effector function. The effects of propionate on allergic inflammation were dependent on G protein–coupled receptor 41 (GPR41, also called free fatty acid receptor 3 or FFAR3), but not GPR43 (also called free fatty acid receptor 2 or FFAR2). Our results show that dietary fermentable fiber and SCFAs can shape the immunological environment in the lung and influence the severity of allergic inflammation.

Subcapsular sinus macrophages in lymph nodes clear lymph-borne viruses and present them to antiviral B cells

Lymph nodes prevent the systemic dissemination of pathogens such as viruses that infect peripheral tissues after penetrating the body’s surface barriers. They are also the staging ground of adaptive immune responses to pathogen-derived antigens. It is unclear how virus particles are cleared from afferent lymph and presented to cognate B cells to induce antibody responses. Here we identify a population of CD11b+CD169+MHCII+ macrophages on the floor of the subcapsular sinus (SCS) and in the medulla of lymph nodes that capture viral particles within minutes after subcutaneous injection. Macrophages in the SCS translocated surface-bound viral particles across the SCS floor and presented them to migrating B cells in the underlying follicles. Selective depletion of these macrophages compromised local viral retention, exacerbated viraemia of the host, and impaired local B-cell activation. These findings indicate that CD169+ macrophages have a dual physiological function. They act as innate ‘flypaper’ by preventing the systemic spread of lymph-borne pathogens and as critical gatekeepers at the lymph–tissue interface that facilitate the recognition of particulate antigens by B cells and initiate humoral immune responses.

Immunosurveillance by Hematopoietic Progenitor Cells Trafficking through Blood, Lymph, and Peripheral Tissues

Constitutive egress of bone marrow (BM)-resident hematopoietic stem and progenitor cells (HSPCs) into the blood is a well-established phenomenon, but the ultimate fate and functional relevance of circulating HSPCs is largely unknown. We show that mouse thoracic duct (TD) lymph contains HSPCs that possess short- and long-term multilineage reconstitution capacity. TD-derived HSPCs originate in the BM, enter the blood, and traffic to multiple peripheral organs, where they reside for at least 36 hr before entering draining lymphatics to return to the blood and, eventually, the BM. HSPC egress from extramedullary tissues into lymph depends on sphingosine-1-phosphate receptors. Migratory HSPCs proliferate within extramedullary tissues and give rise to tissue-resident myeloid cells, preferentially dendritic cells. HSPC differentiation is amplified upon exposure to Toll-like receptor agonists. Thus, HSPCs can survey peripheral organs and can foster the local production of tissue-resident innate immune cells under both steady-state conditions and in response to inflammatory signals.

Dynamic Visualization of Thrombopoiesis Within Bone Marrow

Platelets are generated from megakaryocytes (MKs) in mammalian bone marrow (BM) by mechanisms that remain poorly understood. Here we describe the use of multiphoton intravital microscopy in intact BM to visualize platelet generation in mice. MKs were observed as sessile cells that extended dynamic proplatelet-like protrusions into microvessels. These intravascular extensions appeared to be sheared from their transendothelial stems by flowing blood, resulting in the appearance of proplatelets in peripheral blood. In vitro, proplatelet production from differentiating MKs was enhanced by fluid shear. These results confirm the concept of proplatelet formation in vivo and are consistent with the possibility that blood flow–induced hydrodynamic shear stress is a biophysical determinant of thrombopoiesis.

Lymphoid follicle destruction and immunosuppression after repeated CpG oligodeoxynucleotide administration.

DNA containing unmethylated cytidyl guanosyl (CpG) sequences, which are underrepresented in mammalian genomes but prevalent in prokaryotes, is endocytosed by cells of the innate immune system, including macrophages, monocytes and dendritic cells, and activates a pathway involving Toll-like receptor-9 (TLR9). CpG-containing oligodeoxynucleotides (CpG-ODN) are potent stimulators of innate immunity, and are currently being tested as adjuvants of antimicrobial, antiallergic, anticancer and antiprion immunotherapy. Little is known, however, about the consequences of repeated CpG-ODN administration, which is advocated for some of these applications. Here we report that daily injection of 60 μg CpG-ODN dramatically alters the morphology and functionality of mouse lymphoid organs. By day 7, lymphoid follicles were poorly defined; follicular dendritic cells (FDC) and germinal center B lymphocytes were suppressed. Accordingly, CpG-ODN treatment for ≥7 d strongly reduced primary humoral immune responses and immunoglobulin class switching. By day 20, mice developed multifocal liver necrosis and hemorrhagic ascites. All untoward effects were strictly dependent on CpG and TLR9, as neither the CpG-ODN treatment of Tlr9−/− mice nor the repetitive challenge of wild-type mice with nonstimulatory ODN (AT-ODN) or with the TLR3 agonist polyinosinic:cytidylic acid (polyI:C) were immunotoxic or hepatotoxic.

Restoration of lymphoid organ integrity through the interaction of lymphoid tissue–inducer cells with stroma of the T cell zone

The generation of lymphoid microenvironments in early life depends on the interaction of lymphoid tissue–inducer cells with stromal lymphoid tissue–organizer cells. Whether this cellular interface stays operational in adult secondary lymphoid organs has remained elusive. We show here that during acute infection with lymphocytic choriomeningitis virus, antiviral cytotoxic T cells destroyed infected T cell zone stromal cells, which led to profound disruption of secondary lymphoid organ integrity. Furthermore, the ability of the host to respond to secondary antigens was lost. Restoration of the lymphoid microanatomy was dependent on the proliferative accumulation of lymphoid tissue–inducer cells in secondary lymphoid organs during the acute phase of infection and lymphotoxin α1β2 signaling. Thus, crosstalk between lymphoid tissue–inducer cells and stromal cells is reactivated in adults to maintain secondary lymphoid organ integrity and thereby contributes to the preservation of immunocompetence.

β cells are responsible for CXCR3-mediated T-cell infiltration in insulitis

T cell–mediated loss of insulin-secreting β cells in the islets of Langerhans is the hallmark of type 1 diabetes. The molecular basis for the directed migration of autoreactive T cells leading to insulitis is presently unknown. Here we demonstrate that in response to inflammation, β cells secrete the chemokines CXC ligand 10 and CXC ligand 9, which specifically attract T-effector cells via the CXC chemokine receptor 3. In mice deficient for this receptor, the onset of type 1 diabetes is substantially delayed. Thus, in the absence of known etiological agents, CXC receptor 3 represents a novel target for therapeutic interference early in type 1 diabetes.

Subcapsular sinus macrophages prevent CNS invasion on peripheral infection with a neurotropic virus

Lymph nodes (LNs) capture microorganisms that breach the body’s external barriers and enter draining lymphatics, limiting the systemic spread of pathogens. Recent work has shown that CD11b+CD169+ macrophages, which populate the subcapsular sinus (SCS) of LNs, are critical for the clearance of viruses from the lymph and for initiating antiviral humoral immune responses. Here we show, using vesicular stomatitis virus (VSV), a relative of rabies virus transmitted by insect bites, that SCS macrophages perform a third vital function: they prevent lymph-borne neurotropic viruses from infecting the central nervous system (CNS). On local depletion of LN macrophages, about 60% of mice developed ascending paralysis and died 7–10 days after subcutaneous infection with a small dose of VSV, whereas macrophage-sufficient animals remained asymptomatic and cleared the virus. VSV gained access to the nervous system through peripheral nerves in macrophage-depleted LNs. In contrast, within macrophage-sufficient LNs VSV replicated preferentially in SCS macrophages but not in adjacent nerves. Removal of SCS macrophages did not compromise adaptive immune responses against VSV, but decreased type I interferon (IFN-I) production within infected LNs. VSV-infected macrophages recruited IFN-I-producing plasmacytoid dendritic cells to the SCS and in addition were a major source of IFN-I themselves. Experiments in bone marrow chimaeric mice revealed that IFN-I must act on both haematopoietic and stromal compartments, including the intranodal nerves, to prevent lethal infection with VSV. These results identify SCS macrophages as crucial gatekeepers to the CNS that prevent fatal viral invasion of the nervous system on peripheral infection.

Positioning of follicular dendritic cells within the spleen controls prion neuroinvasion

Peripheral infection is the natural route of transmission in most prion diseases. Peripheral prion infection is followed by rapid prion replication in lymphoid organs, neuroinvasion and progressive neurological disease. Both immune cells and nerves are involved in pathogenesis, but the mechanisms of prion transfer from the immune to the nervous system are unknown. Here we show that ablation of the chemokine receptor CXCR5 juxtaposes follicular dendritic cells (FDCs) to major splenic nerves, and accelerates the transfer of intraperitoneally administered prions into the spinal cord. Neuroinvasion velocity correlated exclusively with the relative locations of FDCs and nerves: transfer of CXCR5-/- bone marrow to wild-type mice induced perineural FDCs and enhanced neuroinvasion, whereas reciprocal transfer to CXCR5-/- mice abolished them and restored normal efficiency of neuroinvasion. Suppression of lymphotoxin signalling depleted FDCs, abolished splenic infectivity, and suppressed acceleration of pathogenesis in CXCR5-/- mice. This suggests that prion neuroimmune transition occurs between FDCs and sympathetic nerves, and relative positioning of FDCs and nerves controls the efficiency of peripheral prion infection.