Andrey A. Kruglov

Nonredundant Function of Soluble LTα3 Produced by Innate Lymphoid Cells in Intestinal Homeostasis

Command and Control Innate lymphoid cells are vital for the development of gut-associated lymphoid tissues, maintenance of the epithelial barrier, and protection against intestinal microbes; their dysfunction can promote immune pathology. Immunoglobulin A (IgA) production is important for maintenance of the gut epithelial barrier and the composition of the gut microbiota. Through the generation of knockout mouse models, Kruglov et al. (p. 1243) were able to distinguish how soluble and membrane-bound lymphotoxins expressed by innate lymphoid cells in the gut specifically regulate IgA production and thereby control gut microbiota composition. Soluble lymphotoxin plays a paracrine role in controlling immunoglobulin A responses and regulating gut microbiota. Immunoglobulin A (IgA) production at mucosal surfaces contributes to protection against pathogens and controls intestinal microbiota composition. However, mechanisms regulating IgA induction are not completely defined. We show that soluble lymphotoxin α (sLTα3) produced by RORγt+ innate lymphoid cells (ILCs) controls T cell–dependent IgA induction in the lamina propria via regulation of T cell homing to the gut. By contrast, membrane-bound lymphotoxin β (LTα1β2) produced by RORγt+ ILCs is critical for T cell–independent IgA induction in the lamina propria via control of dendritic cell functions. Ablation of LTα in RORγt+ cells abrogated IgA production in the gut and altered microbiota composition. Thus, soluble and membrane-bound lymphotoxins produced by ILCs distinctly organize adaptive immune responses in the gut and control commensal microbiota composition.

Eosinophils promote generation and maintenance of immunoglobulin-A-expressing plasma cells and contribute to gut immune homeostasis.

Although in normal lamina propria (LP) large numbers of eosinophils are present, little is known about their role in mucosal immunity at steady state. Here we show that eosinophils are needed to maintain immune homeostasis in gut-associated tissues. By using eosinophil-deficient ΔdblGATA-1 and PHIL mice or an eosinophil-specific depletion model, we found a reduction in immunoglobulin A(+) (IgA(+)) plasma cell numbers and in secreted IgA. Eosinophil-deficient mice also showed defects in the intestinal mucous shield and alterations in microbiota composition in the gut lumen. In addition, TGF-β-dependent events including class switching to IgA in Peyers patches (PP), the formation of CD103(+) T cells including Foxp3(+) regulatory (Treg), and also CD103(+) dendritic cells were disturbed. In vitro cultures showed that eosinophils produce factors that promote T-independent IgA class switching. Our findings show that eosinophils are important players for immune homeostasis in gut-associated tissues and add to data suggesting that eosinophils can promote tissue integrity.

Commensal microbiota influence systemic autoimmune responses

Antinuclear antibodies are a hallmark feature of generalized autoimmune diseases, including systemic lupus erythematosus and systemic sclerosis. However, the processes underlying the loss of tolerance against nuclear self‐constituents remain largely unresolved. Using mice deficient in lymphotoxin and Hox11, we report that approximately 25% of mice lacking secondary lymphoid organs spontaneously develop specific antinuclear antibodies. Interestingly, we find this phenotype is not caused by a defect in central tolerance. Rather, cell‐specific deletion and in vivo lymphotoxin blockade link these systemic autoimmune responses to the formation of gut‐associated lymphoid tissue in the neonatal period of life. We further demonstrate antinuclear antibody production is influenced by the presence of commensal gut flora, in particular increased colonization with segmented filamentous bacteria, and IL‐17 receptor signaling. Together, these data indicate that neonatal colonization of gut microbiota influences generalized autoimmunity in adult life.

Pathogenic and Protective Functions of TNF in Neuroinflammation Are Defined by Its Expression in T Lymphocytes and Myeloid Cells

TNF displays pathogenic activities in many autoimmune disorders. However, anti-TNF therapy in multiple sclerosis patients failed because of poorly understood reasons. We used a panel of gene-targeted mice that allowed cell-type specific ablation of TNF to uncover pathogenic and protective contributions of this cytokine during autoimmune disease of the CNS. T cells and myeloid cells were found to be critical cellular sources of TNF during experimental autoimmune encephalomyelitis (EAE). TNF produced by myeloid cells accelerated the onset of disease by regulation of chemokine expression in the CNS, driving the recruitment of inflammatory cells into the target organ. TNF produced by T cells exacerbated the damage to the CNS during EAE by regulating infiltration of inflammatory myeloid cells into the CNS. In secondary lymphoid organs, TNF expressed by myeloid cells and T cells acted in synergy to dampen IL-12p40 and IL-6 production by APCs, subsequently inhibiting the development of encephalitogenic T cell responses of Th1 and Th17 types. This dual role of TNF during EAE (protective in lymphoid organs and pathogenic in CNS) suggests that global TNF blockade might be inefficient in multiple sclerosis patients because augmented autoreactive T cell development in lymphoid tissues might overwhelm the beneficial effects resulting from TNF inhibition in the CNS.

Cellular sources of pathogenic and protective TNF and experimental strategies based on utilization of TNF humanized mice

Over the years, tumor necrosis factor (TNF) has been implicated in the pathogenesis of various inflammatory conditions and TNF antagonists are highly efficient in treatment of multiple autoimmune diseases. However, it has been shown that various cellular sources of TNF exhibit distinct and non-redundant functions that can be either deleterious or beneficial. This suggests that systemic TNF blockade, in addition to neutralization of pathogenic TNF, may abrogate its protective functions, resulting in adverse effects. Here we review the data on cellular sources of pathogenic and protective TNF and then discuss an experimental system based on humanized mice to study the role of cell-type specific TNF ablation during various disease models for development of cell-type specific TNF blockade.

Tumor necrosis factor, lymphotoxin and cancer

Initially TNF has been discovered as an anti‐tumor factor, but it is now considered as one of the universal effectors of innate signaling implicating its key role in host defense and inflammation. Other physiological functions of TNF are primarily linked to organization of lymphoid tissues. TNF can exert deleterious effects on the organism when its local or systemic concentrations exceed certain levels. This is the main reason for the failure of TNF therapy in oncology. Moreover, in certain experimental models TNF to TNFRp55 signaling axis was found to play a pro‐tumorigenic role. On the other hand, anti‐TNF therapy proved to be beneficial in rheumatic and other autoimmune diseases. Taking into consideration the pivotal function of TNF in the immune system, it is obvious that such therapy cannot be entirely free of adverse effects including suppression of host defense and, possibly, predisposition to lymphomas. Lymphotoxins alpha and beta are the two related cytokines that exist in distinct trimeric forms which can signal through TNFR I and TNFR II, as well LTbetaR receptors, depending on the composition of the trimer. These signals have important functions in the development and homeostasis of the immune system. Importantly, there is a recently uncovered link between the LTalpha/LTbeta to LTbetaR signaling axis and cancer. Here we review the current status of the field with the focus on one particular issue: are TNF and lymphotoxins intrinsically anti‐cancer or pro‐tumorigenic.

Structural Relationship of the Lipid A Acyl Groups to Activation of Murine Toll-Like Receptor 4 by Lipopolysaccharides from Pathogenic Strains of Burkholderia mallei, Acinetobacter baumannii, and Pseudomonas aeruginosa

Toll-like receptor 4 (TLR4) is required for activation of innate immunity upon recognition of lipopolysaccharide (LPS) of Gram-negative bacteria. The ability of TLR4 to respond to a particular LPS species is important since insufficient activation may not prevent bacterial growth while excessive immune reaction may lead to immunopathology associated with sepsis. Here, we investigated the biological activity of LPS from Burkholderia mallei that causes glanders, and from the two well-known opportunistic pathogens Acinetobacter baumannii and Pseudomonas aeruginosa (causative agents of nosocomial infections). For each bacterial strain, R-form LPS preparations were purified by hydrophobic chromatography and the chemical structure of lipid A, an LPS structural component, was elucidated by HR-MALDI-TOF mass spectrometry. The biological activity of LPS samples was evaluated by their ability to induce production of proinflammatory cytokines, such as IL-6 and TNF, by bone marrow-derived macrophages. Our results demonstrate direct correlation between the biological activity of LPS from these pathogenic bacteria and the extent of their lipid A acylation.

Control of Mycobacterial Infections in Mice Expressing Human Tumor Necrosis Factor (TNF) but Not Mouse TNF

ABSTRACT Tumor necrosis factor (TNF) is an important cytokine for host defense against pathogens but is also associated with the development of human immunopathologies. TNF blockade effectively ameliorates many chronic inflammatory conditions but compromises host immunity to tuberculosis. The search for novel, more specific human TNF blockers requires the development of a reliable animal model. We used a novel mouse model with complete replacement of the mouse TNF gene by its human ortholog (human TNF [huTNF] knock-in [KI] mice) to determine resistance to Mycobacterium bovis BCG and M. tuberculosis infections and to investigate whether TNF inhibitors in clinical use reduce host immunity. Our results show that macrophages from huTNF KI mice responded to BCG and lipopolysaccharide similarly to wild-type macrophages by NF-κB activation and cytokine production. While TNF-deficient mice rapidly succumbed to mycobacterial infection, huTNF KI mice survived, controlling the bacterial burden and activating bactericidal mechanisms. Administration of TNF-neutralizing biologics disrupted the control of mycobacterial infection in huTNF KI mice, leading to an increased bacterial burden and hyperinflammation. Thus, our findings demonstrate that human TNF can functionally replace murine TNF in vivo, providing mycobacterial resistance that could be compromised by TNF neutralization. This new animal model will be helpful for the testing of specific biologics neutralizing human TNF.

Modern anti-cytokine therapy of autoimmune diseases

The emergence of genetically engineered biological agents opened new prospects in the treatment of autoimmune and inflammatory diseases. Cytokines responsible for regulation of a wide range of processes during development of the normal immune response are among the most successful therapeutic targets. Studies carried out in recent decades and accompanied by rapid development of biotechnology have promoted establishing in detail the role and place of cytokines in autoimmune and inflammatory pathologies. Nevertheless, mechanisms that underlie anti-cytokine therapy are still not fully understood. This review examines the role of such cytokines as TNF, IL-1, and IL-6 in the development of inflammatory processes and the action mechanisms of their inhibitors.

Deficiency of the B Cell-Activating Factor Receptor Results in Limited CD169+ Macrophage Function during Viral Infection

ABSTRACT The B cell-activating factor (BAFF) is critical for B cell development and humoral immunity in mice and humans. While the role of BAFF in B cells has been widely described, its role in innate immunity remains unknown. Using BAFF receptor (BAFFR)-deficient mice, we characterized BAFFR-related innate and adaptive immune functions following infection with vesicular stomatitis virus (VSV) and lymphocytic choriomeningitis virus (LCMV). We identified a critical role for BAFFR signaling in the generation and maintenance of the CD169+ macrophage compartment. Consequently, Baffr − / − mice exhibited limited induction of innate type I interferon production after viral infection. Lack of BAFFR signaling reduced virus amplification and presentation following viral infection, resulting in highly reduced antiviral adaptive immune responses. As a consequence, BAFFR-deficient mice showed exacerbated and fatal disease after viral infection. Mechanistically, transient lack of B cells in Baffr − / − animals resulted in limited lymphotoxin expression, which is critical for maintenance of CD169+ cells. In conclusion, BAFFR signaling affects both innate and adaptive immune activation during viral infections. IMPORTANCE Viruses cause acute and chronic infections in humans resulting in millions of deaths every year. Innate immunity is critical for the outcome of a viral infection. Innate type I interferon production can limit viral replication, while adaptive immune priming by innate immune cells induces pathogen-specific immunity with long-term protection. Here, we show that BAFFR deficiency not only perturbed B cells, but also resulted in limited CD169+ macrophages. These macrophages are critical in amplifying viral particles to trigger type I interferon production and initiate adaptive immune priming. Consequently, BAFFR deficiency resulted in reduced enforced viral replication, limited type I interferon production, and reduced adaptive immunity compared to BAFFR-competent controls. As a result, BAFFR-deficient mice were predisposed to fatal viral infections. Thus, BAFFR expression is critical for innate immune activation and antiviral immunity.