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T cells sense and respond to their local environment at the nanoscale by forming small actin-rich protrusions, called microvilli, which play critical roles in signaling and antigen recognition, particularly at the interface with the antigen presenting cells. However, the mechanism by which microvilli contribute to cell signaling and activation is largely unknown. Here, we present a tunable engineered system that promotes microvilli formation and T cell signaling via physical stimuli. We discovered that nanoporous surfaces favored microvilli formation and markedly altered gene expression in T cells and promoted their activation. Mechanistically, confinement of microvilli inside of nanopores leads to size-dependent sorting of membrane-anchored proteins, specifically segregating CD45 phosphatases and T cell receptors (TCR) from the tip of the protrusions when microvilli are confined in 200-nm pores but not in 400-nm pores. Consequently, formation of TCR nanoclustered hotspots within 200-nm pores allows sustained and augmented signaling that prompts T cell activation even in the absence of TCR agonists. The synergistic combination of mechanical and biochemical signals on porous surfaces presents a straightforward strategy to investigate the role of microvilli in T cell signaling as well as to boost T cell activation and expansion for application in the growing field of adoptive immunotherapy., Competing Interests: The authors declare no competing interest.

Immunotherapy stands out as a powerful and promising therapeutic strategy in the treatment of cancer, infections, and autoimmune diseases. Adoptive immune therapies are usually centered on modified T cells and their specific expansion towards antigen-specific T cells against cancer and other diseases. However, despite their unmatched features, the potential of B cells in immunotherapy is just beginning to be explored. The main role of B cells in the immune response is to secrete antigen-specific antibodies and provide long-term protection against foreign pathogens. They further function as antigen-presenting cells (APCs) and secrete pro- and anti-inflammatory cytokines and thus exert positive and negative regulatory stimuli on other cells involved in the immune response such as T cells. Therefore, while hyperactivation of B cells can cause autoimmunity, their dysfunctions lead to severe immunodeficiencies. Only suitably activated B cells can play an active role in the treatment of cancers, infections, and autoimmune diseases. As a result, studies have focused on B cell-targeted immunotherapies in recent years. For this, the development, functions, interactions with the microenvironment, and clinical importance of B cells should be well understood. In this review, we summarize the main events during B cell activation. From the viewpoint of mechanobiology we discuss the translation of external cues such as surface topology, substrate stiffness, and biochemical signaling into B cell functions. We further dive into current B cell-targeted therapy strategies and their clinical applications. STATEMENT OF SIGNIFICANCE: B cells are proving as a promising tool in the field of immunotherapy. B cells exhibit various functions such as antibody production, antigen presentation or secretion of immune-regulatory factors which can be utilized in the fight against oncological or immunological disorders. In this review we discuss the importance of external mechanobiological cues such as surface topology, substrate stiffness, and biochemical signaling on B cell function. We further summarize B cell-targeted therapy strategies and their clinical applications, as in the context of anti-tumor responses and autoimmune diseases., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Antibodies play an important role in host defense against microorganisms. Besides direct microbicidal activities, antibodies can also provide indirect protection via crosstalk to constituents of the adaptive immune system. Similar to many human chronic viral infections, persistence of Lymphocytic choriomeningitis virus (LCMV) is associated with compromised T- and B-cell responses. The administration of virus-specific non-neutralizing antibodies (nnAbs) prior to LCMV infection protects against the establishment of chronic infection. Here, we show that LCMV-specific nnAbs bind preferentially Ly6C hi inflammatory monocytes (IMs), promote their infection in an Fc-receptor independent way, and support acquisition of APC properties. By constituting additional T-cell priming opportunities, IMs promote early activation of virus-specific CD8 T cells, eventually tipping the balance between T-cell exhaustion and effector cell differentiation, preventing establishment of viral persistence without causing lethal immunopathology. These results document a beneficial role of IMs in avoiding T-cell exhaustion and an Fc-receptor independent protective mechanism provided by LCMV-specific nnAbs against the establishment of chronic infection., (© 2021 The Authors. European Journal of Immunology published by Wiley-VCH GmbH.)

Nucleoprotein (N) is an immunodominant antigen in many enveloped virus infections. While the diagnostic value of anti-N antibodies is clear, their role in immunity is not. This is because while they are non-neutralising, they somehow clear infection by coronavirus, influenza and LCMV in vivo. Here, we show that anti-N immune protection is mediated by the cytosolic Fc receptor and E3 ubiquitin ligase TRIM21. Exploiting LCMV as a model system, we demonstrate that TRIM21 uses anti-N antibodies to target N for cytosolic degradation and generate cytotoxic T cells (CTLs) against N peptide. These CTLs rapidly eliminate N-peptide-displaying cells and drive efficient viral clearance. These results reveal a new mechanism of immune synergy between antibodies and T cells and highlights N as an important vaccine target., (© 2020 The Authors. Published under the terms of the CC BY 4.0 license.)

Fibroblast growth factors (FGFs) play key roles in the pathogenesis of different human diseases, but the cross-talk between FGFs and other cytokines remains largely unexplored. We identified an unexpected antagonistic effect of FGFs on the interferon (IFN) signaling pathway. Genetic or pharmacological inhibition of FGF receptor signaling in keratinocytes promoted the expression of interferon-stimulated genes (ISG) and proteins in vitro and in vivo. Conversely, FGF7 or FGF10 treatment of keratinocytes suppressed ISG expression under homeostatic conditions and in response to IFN or poly(I:C) treatment. FGF-mediated ISG suppression was independent of IFN receptors, occurred at the transcriptional level, and required FGF receptor kinase and proteasomal activity. It is not restricted to keratinocytes and functionally relevant, since FGFs promoted the replication of herpes simplex virus I (HSV-1), lymphocytic choriomeningitis virus, and Zika virus. Most importantly, inhibition of FGFR signaling blocked HSV-1 replication in cultured human keratinocytes and in mice. These results suggest the use of FGFR kinase inhibitors for the treatment of viral infections., (© 2020 The Authors. Published under the terms of the CC BY 4.0 license.)

Protective immunity against intracellular pathogens, including bacteria, usually relies on cellular immunity. However, antibodies are also implicated in mediating protection against intracellular bacteria. In case of airway infection with Legionella pneumophila (Lpn), the causative agent of Legionnaires' disease, pre-existing Lpn-specific antibodies were shown to afford protection within two days of infection. Here we dissected the early kinetics of Ab-mediated protection against airway Lpn infection and observed two kinetically and mechanistically distinct phases of protection by passively administered antibodies. Within the first hour of infection, Lpn-opsonizing antibodies provided almost 10-fold protection in an antibody Fc-dependent, but FcR-independent manner. Later on, by two days post infection, Lpn-specific Ab-mediated protection strictly involved FcγR, Syk kinase activity in alveolar macrophages and induction of reactive oxygen species (ROS). The findings presented here contribute to the understanding of the mechanisms of Ab-mediated control of Lpn infection in actively or passively immunized individuals., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

In lymphopenic mice, T cells become activated and undergo lymphopenia-induced proliferation (LIP). However, not all T cells are equally sensitive to lymphopenia. Several lymphopenia-insensitive T cell clones were described and their non-responsiveness was mainly attributed to clone-specific properties. Here, we provide evidence for an additional, host-dependent mechanism restraining LIP of lymphopenia-insensitive CD4 + T cells. We show that such cells undergo LIP in lymphopenic mice lacking IFN-γ receptor (IFN-γR) expression, a process, which is promoted by the autocrine action of T cell-derived IFN-γ. Additionally, LIP of lymphopenia-insensitive CD4 + T cells requires an intact microflora and is accompanied by the massive accumulation of IL-6 and dendritic cells (DCs). Consistent with these results, IL-6 neutralization and the DC-specific restoration of IFN-γR expression are both sufficient to restrict LIP. Hence, the insensitivity of CD4 + T cells to lymphopenia relies on cell-intrinsic properties and a complex interplay between the commensal microflora, IL-6, IFN-γR + DCs, and T cell-derived IFN-γ.

The adoptive transfer of antigen-specific CD8+ T cells is a promising approach for the treatment of chronic viral and malignant diseases. In order to improve adoptive T cell therapy (ATT) of cancer, recent strategies aim at the antibody-based blockade of immunosuppressive signaling pathways in CD8+ T cells. Alternatively, adjuvant effects of immunostimulatory cytokines might be exploited to improve therapeutic CD8+ T cell responses. For example, Interleukin-7 (IL-7) is a potent growth, activation and survival factor for CD8+ T cells that can be used to improve virus- and tumor-specific CD8+ T cell responses. Although direct IL-7 effects on CD8+ T cells were studied extensively in numerous models, the contribution of IL-7 receptor-competent (IL-7R+) host cells remained unclear. In the current study we provide evidence that CD8+ T cell-mediated tumor rejection in response to recombinant IL-7 (rIL-7) therapy is strictly dependent on IL-7R+ host cells. On the contrary, CD8+ T cell expansion is independent of host IL-7R expression. If, however, rIL-7 therapy and peptide vaccination are combined, host IL-7R signaling is crucial for CD8+ T cell expansion. Unexpectedly, maximum CD8+ T cell expansion relies mainly on IL-7R signaling in non-hematopoietic host cells, similar to the massive accumulation of dendritic cells and granulocytes. In summary, we provide evidence that IL-7R+ host cells are major targets of rIL-7 that modulate therapeutic CD8+ T cell responses and the outcome of rIL-7-assisted ATT. This knowledge may have important implications for the design and optimization of clinical ATT protocols.

In response to primary Ag contact, naive mouse CD8(+) T cells undergo clonal expansion and differentiate into effector T cells. After pathogen clearance, most effector T cells die, and only a small number of memory T cell precursors (TMPs) survive to form a pool of long-lived memory T cells (TMs). Although high- and low-affinity CD8(+) T cell clones are recruited into the primary response, the TM pool consists mainly of high-affinity clones. It remains unclear whether the more efficient expansion of high-affinity clones and/or cell-intrinsic processes exclude low-affinity T cells from the TM pool. In this article, we show that the lack of IFN-γR signaling in CD8(+) T cells promotes TM formation in response to weak, but not strong, TCR agonists. The IFN-γ-sensitive accumulation of TMs correlates with reduced mammalian target of rapamycin activation and the accumulation of long-lived CD62L(hi)Bcl-2(hi)Eomes(hi) TMPs. Reconstitution of mammalian target of rapamycin or IFN-γR signaling is sufficient to block this process. Hence, our data suggest that IFN-γR signaling actively blocks the formation of TMPs responding to weak TCR agonists, thereby promoting the accumulation of high-affinity T cells finally dominating the TM pool., (Copyright © 2015 by The American Association of Immunologists, Inc.)

Identifying the antigens that have the potential to trigger endogenous antitumor responses in an individual cancer patient is likely to enhance the efficacy of cancer immunotherapy, but current methodologies do not efficiently identify such antigens. This study describes what we believe to be a new method of comprehensively identifying candidate tissue antigens that spontaneously cause T cell responses in disease situations. We used the newly developed automated, two-dimensional chromatography system PF2D to fractionate the proteome of human tumor tissues and tested protein fractions for recognition by preexisting tumor-specific CD4+ Th cells and CTLs. Applying this method using mice transgenic for a TCR that recognizes an OVA peptide presented by MHC class I, we demonstrated efficient separation, processing, and cross-presentation to CD8+ T cells by DCs of OVA expressed by the OVA-transfected mouse lymphoma RMA-OVA. Applying this method to human tumor tissues, we identified MUC1 and EGFR as tumor-associated antigens selectively recognized by T cells in patients with head and neck cancer. Finally, in an exemplary patient with a malignant brain tumor, we detected CD4+ and CD8+ T cell responses against two novel antigens, transthyretin and calgranulin B/S100A9, which were expressed in tumor and endothelial cells. The immunogenicity of these antigens was confirmed in 4 of 10 other brain tumor patients. This fast and inexpensive method therefore appears suitable for identifying candidate T cell antigens in various disease situations, such as autoimmune and malignant diseases, without being restricted to expression by a certain cell type or HLA allele.

IFN-gamma regulates multiple processes in the immune system. Although its antimicrobial effector functions are well described, less is known about the mechanisms by which IFN-gamma regulates CD8(+) T cell homeostasis. With the help of adoptive T cell transfers, we show in this study that IFN-gammaR signaling in CD8(+) T cells is dispensable for expansion, contraction, and memory differentiation in response to peptide vaccination. In contrast, host IFN-gammaR signaling counterregulates CD8(+) T cell responses and the generation of effector memory T cell processes, which are partially regulated by CD11b(+) cells. Similar to vaccination-induced proliferation, host IFN-gammaR signaling limits the expansion of naive CD8(+) T cells and their differentiation into effector memory-like T cells in lymphopenic mice. In contrast to peptide vaccination, IFN-gammaR signaling in CD8(+) T cells contributes to memory fate decision in response to lymphopenia, an effect that is fully reversed by high-affinity TCR ligands. In conclusion, we show that host IFN-gammaR signaling controls the magnitude of CD8(+) T cell responses and subsequent memory differentiation under lymphopenic and nonlymphopenic conditions. In contrast, IFN-gammaR signaling in CD8(+) T cells does not affect cell numbers under either condition, but it directs memory fate decision in response to weak TCR ligands.

In earlier studies (Ivanov, Staneva-Stoycheva, 1984) we have found an inhibitory effect of metamyzole (Analgin-Pharmachim) on the contractile responses of electrically stimulated guinea-pig ileum and mouse vas deferens, which is antagonized by naloxone. In the present study an attempt is made to clarify whether these presynaptic effects of metamyzole are due to direct interaction with presynaptic opiate receptors. The method of radioligand binding of /3H/-naloxone to the opiate receptors in a crude membrane fraction of rat brain was used. It was found that in the wide concentration range used (10(-5)-10(-9) M), metamyzole does not compete with /3H/-naloxone for binding to the receptor sites. The biochemical study has shown that the presynaptic opiate receptors do not participate in the mechanism of the presynaptic effects of metamyzole, established in earlier studies.

The antiarrhythmic activity of the compound N-(trans-3-hydroxy-1,2,3,4-tetrahydro-2-naphthyl)-N-(3-oxo-3-phenyl-2-methylpropyl)-piperazine hydrochloride, referred to as P11, is studied on anaesthesized cats and Wistar albino rats, as well as on non-anaesthesized rabbits. Four types of experimental arrhythmia are used--with BaCl2, with chloroform-adrenaline, with strophantine G and with aconitine. The compound P11 is introduced in doses of 0.25 and 0.50 mg/kg intravenously and 10 mg/kg orally. The compound manifests antiarrhythmic activity in all models of experimental arrhythmia used, causing greatest inhibition on the arrhythmia induced by chloroform-adrenaline (in 90 per cent) and with BaCl2 (in 84 per cent). The results obtained are associated with the beta-adrenoblocking and with the membrane-stabilizing action of the compound.

The effect of Analgine in different concentrations on the contractions of electrically stimulated isolated guinea-pig ileum and mouse vas deferens is studied and compared with the effect of morphine on the same isolated organs. Analgine causes morphine-like but weaker concentration-dependent inhibitory effect (IC50 = 5.75 X 10(-6) M) on guinea-pig ileum, which is completely eliminated by naloxon and 4-aminopyridine. Upon simultaneous administration of morphine analgine, potentiation of the effect of morphine is observed, which is also eliminated by naloxon and 4-aminopyridine. Analgine has a weaker inhibitory effect (IC50 = 1 X 10(-4) M) on the contractions of mouse vas deferens, compared with its effect on the ileum. The same is also valid for the independent and combined with analgine effect of morphine, completely eliminated only by 4-aminopyridine, but not by naloxon. In cases of reduced Na+-concentration in the medium, the effect of analgine on the contractions of the ileum is weaker (IC25 = 4.43 X 10(-5) M), while that of morphine is stronger compared with normal conditions. Increased Ca++-concentration is parallelled by a weaker effect of analgine (IC25 = 4.76 X 10(-5) M), unlike the effect of morphine which is potentiated. It is possible to assume some interaction of analgine with a definite type of presynaptic opiate receptors, similar or identical to these receptors which are responsible for the peripheral effects of the narcotic analgesics.

The experiments were carried out on Wistar rats, using a conflict situation consisting in opposition of two motivations: positive (water drinking) and negative (electrical pain). Two newly-synthesized piperazine derivatives with proved psychotropic activity (4P-183 and GP4) were studied and compared with the classical benzodiazepine tranquilizing agent diazepam. Parallel with this, the affinity of these compounds to the benzodiazepine receptor in mouse brain was tested (radioligand binding of 3H-diazepam), as well as the changes in the characteristics of binding (dissociation constant and density of the receptors) after 14-day administration o doses equal to 1/3 LD50 of the two compounds. The results indicate the existence of anxiolytic (anticonflict) activity in both compounds. The fact that the above-mentioned derivatives do not interact with 3H-diazepam (in vitro), changing above all the dissociation constant after chronic administration, suggests predominantly nonspecific type of binding to the receptors. It is possible that their anxiolytic activity is associated with indirect interactions with one of the components of the GABA-benzodiazepine receptor complex, similar to the atypical tranquillizers.