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Costimulatory and coinhibitory mechanisms appear to be involved throughout immune responses to control their specificity and level. Many mechanisms operate; therefore, various theoretical models should be considered complementary rather than competing. One such coinhibitory model, pictured in 1971, involves the crosslinking of antigen receptors with inhibitory Fc receptors by antigen/antibody complexes. This model was prompted by observations that the Fc portion of antibody was required for potent suppression of immune responses by antibody. The signal via the antigen receptor wakes up T or B cells, providing specificity, while costimulators and coinhibitors stimulate or inhibit these awoken cells. The recent observations that administration of monoclonal anti-SARS-CoV-2 spike antibodies in early COVID-19 patients inhibits the induction of clinically damaging autoimmune antibodies suggest they may provide negative Fc signals that are blocked in COVID-19 patients. Furthermore, the reduced ability of SARS-CoV-2 antigen to localize to germinal centres in COVID-19 patients also suggests a block in binding of the Fc of antibody bound to antigen on FcγRIIb of follicular dendritic cells. The distinction between self and foreign is made not only at the beginning of immune responses but also throughout, and involves multiple mechanisms and models. There are past beginnings (history of models) and current and future beginnings for solving serious clinical problems (such as COVID-19) and different types of models used for understanding the complexities of fundamental immunology., (© 2021 The Scandinavian Foundation for Immunology.)

Antibodies are essential components of an adaptive immune response. Immunoglobulin G (IgG) is the most common type of antibody found in circulation and extracellular fluids. Although IgG alone can directly protect the body from infection through the activities of its antigen binding region, the majority of IgG immune functions are mediated via proteins and receptors expressed by specialized cell subsets that bind to the fragment crystallizable (Fc) region of IgG. Fc gamma (γ) receptors (FcγR) belong to a broad family of proteins that presently include classical membrane-bound surface receptors as well as atypical intracellular receptors and cytoplasmic glycoproteins. Among the atypical FcγRs, the neonatal Fc receptor (FcRn) has increasingly gained notoriety given its intimate influence on IgG biology and its ability to also bind to albumin. FcRn functions as a recycling or transcytosis receptor that is responsible for maintaining IgG and albumin in the circulation, and bidirectionally transporting these two ligands across polarized cellular barriers. More recently, it has been appreciated that FcRn acts as an immune receptor by interacting with and facilitating antigen presentation of peptides derived from IgG immune complexes (IC). Here we review FcRn biology and focus on newer advances including how emerging FcRn-targeted therapies may affect the immune responses to IgG and IgG IC.

Autoantibodies, including rheumatoid factor (RF), are an important characteristic of rheumatoid arthritis (RA). Interestingly, several studies reported a correlation between the presence of IgA autoantibodies and worse disease course. We demonstrated previously that triggering the IgA Fc receptor (FcαRI) on neutrophils results in neutrophil recruitment and the release of neutrophil extracellular traps (NETs). Because this can lead to tissue damage, we investigated whether IgA immune complexes in plasma and synovial fluid of RA patients activate neutrophils. RF isotypes were measured with ELISA, and immune complexes were precipitated using polyethylene glycol 6000. Isolated neutrophils were incubated with immune complexes, and activation and release of NETs were determined in the presence or absence of FcαRI-blocking Abs. Plasma and SF of RA patients contained IgM, IgG, and IgA RFs. Patient plasma IgA RF and IgM RF showed a strong correlation. No uptake of IgM and minimal endocytosis of IgG immune complexes by neutrophils was observed, in contrast to avid uptake of IgA complexes. Incubation of neutrophils with immune complexes resulted in the production of reactive oxygen species, as well as the release of NETs, lactoferrin, and chemotactic stimuli. Importantly, activation of neutrophils was reduced when FcαRI was blocked. Neutrophils were activated by IgA immune complexes, which suggests that neutrophils play a role in inducing joint damage in RA patients who have IgA autoantibody complexes, thereby increasing the severity of disease. Blocking FcαRI inhibited neutrophil activation and, as such, may represent an additional attractive novel therapeutic strategy for the treatment of RA., (Copyright © 2016 by The American Association of Immunologists, Inc.)

Human IgGs are increasingly used in the therapy of many different immune and inflammatory diseases, however their mechanism of action still remains unclear in most diseases. To gain insight into the mechanism by which IgGs might also exert their effects on endothelial cells, we tested human IgGs on human umbilical vein endothelial cells (HUVECs). IgGs induced a time-dependent increase in the synthesis and secretion of IgGs, together with a marked angiogenic-like transformation of HUVECs that was maximal after a 20-h incubation. IgGs stimulated IG gene transcription without affecting the process of gene rearrangement, already present in control HUVECs. The mechanism involved the activation of transcription factors with the increased expression of HSP90, HSP70 and inactive MMP-9 responsible for the phenotypic differentiation associated with the most intense IgG synthesis and secretion. However, even a short incubation with IgGs followed by recovery of cells was sufficient to trigger and sustain in time the synthesis and secretion of new IgGs, independently of the angiogenic-like transformation visible only when cells were continuously exposed to IgGs. Under the stimulus of IgGs, specific secretory pathways were also activated in HUVECs together with the expression of FcRn, which was always associated with IgGs of new synthesis, forming complexes that were also secreted. Our results disclose a so far unknown and unexpected mechanism of IgGs on HUVECs that behave as Ig-producing immune cells. Results might have relevance for the effects that IgGs also exert in vivo in physiological conditions., (Copyright © 2016 Elsevier GmbH. All rights reserved.)

Broadly neutralizing antibodies that recognize the conserved hemagglutinin (HA) stalk have emerged as exciting new biotherapeutic tools to combat seasonal and pandemic influenza viruses. Our general understanding of the mechanisms by which stalk-specific antibodies achieve protection is rapidly evolving. It has recently been demonstrated that broadly neutralizing HA stalk-specific IgG antibodies require Fc-Fcγ receptor (FcγR) interactions for optimal protection in vivo Here we examine the neutrophil effector functions induced by stalk-specific antibodies. As the most abundant subset of blood leukocytes, neutrophils represent a critical innate effector cell population and serve an instrumental role in orchestrating downstream adaptive responses to influenza virus infection. Yet, the interplay of HA stalk-specific IgG, Fc-FcγR engagement, and neutrophils has remained largely uncharacterized. Using an in vitro assay to detect the production of reactive oxygen species (ROS), we show that human and mouse monoclonal HA stalk-specific IgG antibodies are able to induce the production of ROS by neutrophils, while HA head-specific antibodies do not. Furthermore, our results indicate that the production of ROS is dependent on Fc receptor (FcR) engagement and phagocytosis. We went on to assess the ability of monoclonal HA stalk-specific IgA antibodies to induce ROS. Consistent with our findings for monoclonal IgGs, only HA stalk-specific IgA antibodies elicited ROS production by neutrophils. This induction is dependent on the engagement of FcαR1. Taken together, our findings describe a novel FcR-dependent effector function induced by HA stalk-specific IgG and IgA antibodies, and importantly, our studies shed light on the mechanisms by which HA stalk-specific antibodies achieve protection., Importance: The present study provides evidence that broadly neutralizing HA stalk-specific antibodies induce downstream Fc-mediated neutrophil effector functions. In addition to their ability to neutralize, this class of antibodies has been shown to rely on Fc-Fc receptor interactions for optimal protection in vivo Curiously, neutralizing antibodies that bind the HA head domain do not require such interactions. Our findings build on these previous observations and provide a more complete picture of the relationship between stalk-specific antibodies and cells of the innate immune compartment. Furthermore, our data suggest that the ability of HA stalk-specific antibodies to mediate Fc-Fc receptor engagement is epitope dependent. Overall, this work will inform the rational design of improved influenza virus vaccines and therapeutics., (Copyright © 2016 Mullarkey et al.)

Innate and adaptive immune responses represent well balanced reactions aimed at resolving microbial infections without causing major collateral damage to the host. Disturbances in this system either due to enhanced activating or decreased inhibitory signaling pathways may lead to excessive immune activation resulting in tissue damage, the induction of autoimmune disease and/or chronic inflammation. On the molecular level this balance is achieved by the integration of inhibitory and activating signals, which are delivered by pairs of activating and inhibitory cell surface receptors expressed on innate and adaptive immune cells. The regulation of immunoglobulin G activity through cellular Fc receptors is a prime example for this type of regulation. This is not only relevant for the regulation of antibody-mediated effector functions through innate immune effector cells but also for the regulation of B cell activation and antibody production itself.

Although both Fcα/μ receptor (Fcα/μR) and polymeric Ig receptor (poly-IgR) are Fc receptors for IgA and IgM and are functionally and genetically related, the expression profile of Fcα/μR is unique. Unlike poly-IgR, Fcα/μR is expressed on marginal zone (MZ) B cells and follicular dendritic cells, suggesting that Fcα/μR plays an important role in humoral immune responses. Fcα/μR mediates endocytosis of the IgM immune complex (IC). Recent research demonstrated that Fcα/μR downregulated retention of the IgM IC with a T-independent (TI) antigen on MZ B cells and follicular dendritic cells due to endocytosis of the IgM IC, suppressing germinal center formation, affinity maturation, and memory B-cell generation in response to TI antigen challenge. In addition, Fcα/μR physically associates with Toll-like receptor 4 (TLR4) and augments TLR4 oligomerization and signaling in MZ B cells upon lipopolysaccharide (LPS) challenge, leading to increased proinflammatory cytokine production by MZ B cells. Thus, Fcα/μR is a unique Fc receptor that is involved in humoral immune responses and inflammation., (© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: The mechanism(s) responsible for acquisition of maternal antibody isotypes other than IgG are not fully understood. This uncertainty is a major reason underlying the continued controversy regarding whether cord blood (CB) IgE originates in the mother or fetus., Objective: To investigate the capacity of maternal IgE to be transported across the placenta in the form of IgG anti-IgE/IgE immune complexes (ICs) and to determine the role of the neonatal Fc receptor (FcRn) in mediating this process., Methods: Maternal and CB serum concentrations of IgE, IgG anti-IgE, and IgG anti-IgE/IgE ICs were determined in a cohort of allergic and non-allergic mother/infant dyads. Madin-Darby canine kidney (MDCK) cells stably transfected with human FcRn were used to study the binding and transcytosis of IgE in the form of IgG anti-IgE/IgE ICs., Results: Maternal and CB serum concentrations of IgG anti-IgE/IgE ICs were highly correlated, regardless of maternal allergic status. IgG anti-IgE/IgE ICs generated in vitro bound strongly to FcRn-expressing MDCK cells and were transcytosed in an FcRn-dependent manner. Conversely, monomeric IgE did not bind to FcRn and was not transcytosed. IgE was detected in solutions of transcytosed IgG anti-IgE/IgE ICs, even though essentially all the IgE remained in complex form. Similarly, the majority of IgE in CB sera was found to be complexed to IgG., Conclusions and Clinical Relevance: These data indicate that human FcRn facilitates the transepithelial transport of IgE in the form of IgG anti-IgE/IgE ICs. They also strongly suggest that the majority of IgE in CB sera is the result of FcRn-mediated transcytosis of maternal-derived IgG anti-IgE/IgE ICs. These findings challenge the widespread perception that maternal IgE does not cross the placenta. Measuring maternal or CB levels of IgG anti-IgE/IgE ICs may be a more accurate predictor of allergic risk., (© 2015 The Authors. Clinical & Experimental Allergy Published by John Wiley & Sons Ltd.)

Given the critical role of Fc gamma receptors (FcγR) as primary targets for autoantibody-mediated effects an important issue is how the FcγR pathway is affected in autoimmune disorders. Here we investigated the FcγR function in monocytes from rheumatoid arthritis (RA) patients in relation to immunoglobulin levels and disease activity. Peripheral blood was obtained from 30 RA patients with clinical acute joint synovitis (active RA), 28 RA patients with no clinical signs of acute joint synovitis (non-active RA) and 34 healthy controls. Prior the functional studies the monocytes were characterized of their FcγRI (CD64), II (CD32), IIb (CD32b) and III (CD16) expression as well as their cell surface bound IgG. The monocytic FcγR function was assessed by binding of human IgG1 and IgG3 immune complexes (IC) and TNF secretion in vitro. IgG anti-citrullinated peptide antibodies (ACPA) were analyzed in the plasma. We found that monocytes from active RA patients had increased levels of FcγRI, II and cell surface IgG concurrently with impaired FcγR function. This was evident by reduced IgG1-IC binding and decreased TNF secretion in response to IgG3-IC. In contrast, monocytes from non-active RA patients displayed a normal FcγR function and had increased FcγRIIb expression together with elevated FcγRI, II and cell surface IgG. The ACPA levels did not differ in active and non-active RA patients but correlated with the monocytic FcγRIII expression in the patients. In conclusion, active RA patients display a dysregulated FcγR function that may represent a novel phenotypic and likely pathogenetic marker for active RA. A disease and FcγR function controlling effect is suggested by the increased inhibitory FcγRIIb in non-active RA., (Copyright © 2014 Elsevier B.V. All rights reserved.)

During pathogen infection, antibodies can be carried into the infected cell, where they are detected by the ubiquitously expressed cytosolic antibody receptor TRIM21. Here we found that recognition of intracellular antibodies by TRIM21 activated immune signaling. TRIM21 catalyzed the formation of Lys63 (K63)-linked ubiquitin chains and stimulated the transcription factor pathways of NF-κB, AP-1, IRF3, IRF5 and IRF7. Activation resulted in the production of proinflammatory cytokines, modulation of natural killer stress ligands and induction of an antiviral state. Intracellular antibody signaling was abrogated by genetic deletion of TRIM21 and was restored by ectopic expression of TRIM21. The sensing of antibodies by TRIM21 was stimulated after infection by DNA or RNA nonenveloped viruses or intracellular bacteria. Thus, the antibody-TRIM21 detection system provides potent, comprehensive activation of the innate immune system independently of known pattern-recognition receptors.

Background: The mechanism(s) responsible for the acquisition of maternal antibody isotypes other than IgG are not fully understood., Objective: To define the ability of the neonatal Fc receptor for IgG uptake (FcRn) to mediate intestinal absorption of IgG(1) anti-IgE/IgE immune complexes., Methods: C57BL/6 allergic ovalbumin (OVA)-immune foster mothers were generated to nurse naïve FcRn(+/-) or FcRn(-/-) progeny. At the time of weaning, serum levels of OVA-specific antibodies and IgG(1) anti-IgE/IgE immune complexes were determined in allergic foster mothers and FcRn(+/+), FcRn(+/-), or FcRn(-/-) breastfed offspring. In separate experiments, FcRn(+/-) or FcRn(-/-) neonatal mice were gavage fed TNP-specific IgE as IgG(1) anti-IgE/IgE immune complexes, IgG(1) isotype control and IgE, or IgE alone. Mice were killed 2 h after feeding to determine serum levels and biological activity of absorbed TNP-specific IgE., Results: As expected, the absorption of maternal OVA-specific IgG(1) in FcRn(-/-) offspring was at levels 10(3) -10(4) less than observed in FcRn(+/+) or FcRn(+/-) offspring. Surprisingly, FcRn expression also influenced the absorption of maternal IgE. OVA-specific IgE was detected in FcRn(+/+) and FcRn(+/-) offspring, but not in FcRn(-/-) offspring. IgG(1) anti-IgE/IgE immune complexes were detected in allergic foster mothers and correlated strongly with levels in FcRn(+/+) and FcRn(+/-) offspring (ρ = 0.88, P < 0.0001). Furthermore, FcRn expression was required for neonatal mice to absorb TNP-specific IgE when fed as IgG(1) anti-IgE/IgE immune complexes. When immune complexes were generated with IgG(1) anti-IgE directed against the Cε4 domain, the absorbed IgE was able to function in antigen-dependent basophil degranulation., Conclusions and Clinical Relevance: These data demonstrate a novel mechanism by which FcRn may facilitate absorption of maternal antibodies other than IgG. These findings are clinically relevant because FcRn mediates the transplacental passage of maternal IgG to the fetus. This raises the possibility that FcRn could mediate the transplacental passage of maternal IgE as IgG anti-IgE/IgE immune complexes., (© 2012 Blackwell Publishing Ltd.)

Modulating the binding affinities to IgE or changing the FcγR binding properties of anti-IgE antibodies offers an opportunity to enhance the therapeutic potential of anti-IgE antibodies, but the influence of increased affinity to IgE or reduced Fc effector function on the pharmacological properties of anti-IgE therapies remains unclear. Our studies were designed to characterize the pharmacokinetics, pharmacodynamics and immune-complex distribution of two high-affinity anti-IgE monoclonal antibodies, high-affinity anti-IgE antibody (HAE) 1 and 2, in mice and monkeys. HAE1, also known as PRO98498, is structurally similar to omalizumab (Xolair®), a humanized anti-IgE IgG1 marketed for the treatment of asthma, but differs by 9 amino acid changes in the complementarity-determining region resulting in a 23-fold improvement in affinity. HAE2 is similar to HAE1, but its Fc region was altered to reduce binding to Fcγ receptors. As expected given the decreased binding to Fcγ receptors, systemic exposure to pre-formed HAE2:IgE complexes in mice was greater (six-fold) and distribution to the liver lower (four-fold) compared with HAE1:IgE complexes. In monkeys, systemic exposure to HAE1 was similar to that previously observed for omalizumab in this species, but required comparatively lower serum drug concentrations to suppress free IgE levels. HAE2 treatment resulted in greater exposure and greater increase of total IgE, relative to HAE1, because of decreased clearance of HAE2:IgE complexes. Overall, these data suggest that increased binding affinity to IgE may provide a more effective therapeutic for asthma patients, and that retaining FcγR binding of the anti-IgE antibody is important for elimination of anti-IgE:IgE complexes.

The Fc-α receptor (FcαR/CD89) is involved in IgA complex formation and may affect the development of IgA nephropathy (IgAN). In this study, we tested the genetic variations of the CD89 gene in relation to disease susceptibility in IgAN and the expression of soluble CD89 (sCD89) in sera of patients with IgAN and in controls. There was a significant difference between the levels of sCD89-IgA complexes, measured by sandwich enzyme-linked immunosorbent assay (ELISA), in 177 patients with IgAN with and without disease progression at the time of first diagnosis. No such difference was found in 42 patients with other renal diseases. The patients with IgAN without disease progression had stable but high levels of sCD89 over 5-15 years of follow-up in contrast to stable but low levels of sCD89 in the disease progression group. Moreover, levels of sCD89 complexes were correlated with one of the five CD89 genetic variants in 212 patients with IgAN and 477 healthy Caucasians; the single-nucleotide polymorphism (SNP) rs11084377 was significantly associated with a lower expression of sCD89. However, no association between CD89 gene polymorphisms and susceptibility to IgAN was detected. Thus, we found an association between the levels of sCD89-IgA complexes in serum and the severity of IgAN, and a possible genetic component in regulating the production or expression of sCD89.

New data suggest that a soluble form of the IgA receptor CD89 may be protective against development of progressive renal injury in IgA nephropathy (IgAN). Understanding how IgA triggers shedding of CD89 from myeloid cell surfaces could help clarify the process of immune complex formation in IgAN, and measurement of this soluble form of CD89 may in the future prove a useful prognostic indicator of end-stage renal disease in this common glomerulonephritis.

Allergic asthma is a chronic lung disease resulting from an inappropriate T helper (Th)-2 response to environmental antigens. Early tolerance induction is an attractive approach for primary prevention of asthma. Here, we found that breastfeeding by antigen-sensitized mothers exposed to antigen aerosols during lactation induced a robust and long-lasting antigen-specific protection from asthma. Protection was more profound and persistent than the one induced by antigen-exposed non-sensitized mothers. Milk from antigen-exposed sensitized mothers contained antigen-immunoglobulin (Ig) G immune complexes that were transferred to the newborn through the neonatal Fc receptor resulting in the induction of antigen-specific FoxP3(+) CD25(+) regulatory T cells. The induction of oral tolerance by milk immune complexes did not require the presence of transforming growth factor-beta in milk in contrast to tolerance induced by milk-borne free antigen. Furthermore, neither the presence of IgA in milk nor the expression of the inhibitory FcgammaRIIb in the newborn was required for tolerance induction. This study provides new insights on the mechanisms of tolerance induction in neonates and highlights that IgG immune complexes found in breast milk are potent inducers of oral tolerance. These observations may pave the way for the identification of key factors for primary prevention of immune-mediated diseases such as asthma.

In intestinal secretions, secretory IgA (SIgA) plays an important sentinel and protective role in the recognition and clearance of enteric pathogens. In addition to serving as a first line of defense, SIgA and SIgA x antigen immune complexes are selectively transported across Peyer's patches to underlying dendritic cells in the mucosa-associated lymphoid tissue, contributing to immune surveillance and immunomodulation. To explain the unexpected transport of immune complexes in face of the large excess of free SIgA in secretions, we postulated that SIgA experiences structural modifications upon antigen binding. To address this issue, we associated specific polymeric IgA and SIgA with antigens of various sizes and complexity (protein toxin, virus, bacterium). Compared with free antibody, we found modified sensitivity of the three antigens assayed after exposure to proteases from intestinal washes. Antigen binding further impacted on the immunoreactivity toward polyclonal antisera specific for the heavy and light chains of the antibody, as a function of the antigen size. These conformational changes promoted binding of the SIgA-based immune complex compared with the free antibody to cellular receptors (Fc alphaRI and polymeric immunoglobulin receptor) expressed on the surface of premyelocytic and epithelial cell lines. These data reveal that antigen recognition by SIgA triggers structural changes that confer to the antibody enhanced receptor binding properties. This identifies immune complexes as particular structural entities integrating the presence of bound antigens and adds to the known function of immune exclusion and mucus anchoring by SIgA.

Background: The obligate intracellular bacterial pathogen Coxiella burnetii causes the zoonosis Q fever. The intracellular niche of C. burnetii has led to the assumption that cell-mediated immunity is the most important immune component for protection against this pathogen. However, passive immunization with immune serum can protect naïve animals from challenge with virulent C. burnetii, indicating a role for antibody (Ab) in protection. The mechanism of this Ab-mediated protection is unknown. Therefore, we conducted a study to determine whether Fc receptors (FcR) or complement contribute to Ab-mediated immunity (AMI) to C. burnetii., Results: Virulent C. burnetii infects and replicates within human dendritic cells (DC) without inducing their maturation or activation. We investigated the effects of Ab opsonized C. burnetii on human monocyte-derived and murine bone marrow-derived DC. Infection of DC with Ab-opsonized C. burnetii resulted in increased expression of maturation markers and inflammatory cytokine production. Bacteria that had been incubated with naïve serum had minimal effect on DC, similar to virulent C. burnetii alone. The effect of Ab opsonized C. burnetii on DC was FcR dependent as evidenced by a reduced response of DC from FcR knockout (FcR k/o) compared to C57Bl/6 (B6) mice. To address the potential role of FcR in Ab-mediated protection in vivo, we compared the response of passively immunized FcR k/o mice to the B6 controls. Interestingly, we found that FcR are not essential for AMI to C. burnetii in vivo. We subsequently examined the role of complement in AMI by passively immunizing and challenging several different strains of complement-deficient mice and found that AMI to C. burnetii is also complement-independent., Conclusion: Despite our data showing FcR-dependent stimulation of DC in vitro, Ab-mediated immunity to C. burnetii in vivo is FcR-independent. We also found that passive immunity to this pathogen is independent of complement.

Tissue damage in autoimmune diseases involves excessive production of reactive oxygen species (ROS) triggered by immune complexes (IC) and neutrophil (PMN) interactions via receptors for the Fc portion of IgG (FcgammaR) and complement receptors (CR). Modulation of both the effector potential of these receptors and ROS generation may be relevant to the maintenance of body homeostasis. In the present study, the modulatory effect of four flavonols (myricetin, quercetin, kaempferol, galangin) on rabbit PMN oxidative metabolism, specifically stimulated via FcgammaR, CR or both classes of receptors, was evaluated by luminol- and lucigenin-dependent chemiluminescence assays. Results showed that flavonol inhibitory effect was not dependent on the cell membrane receptor class stimulated but related to the lipophilicity of the compounds (their apparent partition coefficient values were obtained by high-performance liquid chromatography), and was also inversely related to the number of hydroxyl groups in the flavonol B ring and the ROS-scavenger activity (assessed by the luminol--H2O2--horseradish peroxidase reaction). Under the experimental conditions the flavonols tested were not toxic to PMNs (evaluated by lactate dehydrogenase release and trypan blue exclusion) and did not interfere with IC-induced phagocytosis (evaluated by transmission electron microscopy). Our results suggested that inhibition of IC-stimulated PMNs effector functions by the flavonols tested herein was the result of cooperation of different cellular mechanisms.

Receptors for immunoglobulins [Fc-receptors (FcRs)] are widely expressed throughout the immune system. By binding to the antibody Fc-portion, they provide a link between the specificity of the adaptive immune system and the powerful effector functions triggered by innate immune effector cells. By virtue of coexpression of activating and inhibitory FcRs on the same cell, they set a threshold for immune cell activation by immune complexes (ICs). Besides their involvement in the efferent phase of an immune response, they are also important for modulating adaptive immune responses by regulating B cell and dendritic cell (DC) activation. Deletion of the inhibitory FcR leads to the loss of tolerance in the humoral immune system and the development of autoimmune disease. Uptake of ICs by FcRs on DCs and the concommitant triggering of activating and inhibitory signaling pathways will determine the strength of the initiated T-cell response. Loss of this balanced signaling results in uncontrolled responses that can lead to the damage of healthy tissues and ultimately to the initiation of autoimmune processes. In this chapter, we will discuss how coexpression of different activating and inhibitory receptors on different immune cells of the innate and adaptive immune system modulates cell activity. Moreover, we will focus on exogenous factors that can influence the balanced triggering of activating and inhibitory FcRs, such as the cytokine milieu and the role of differential antibody glycosylation.

IgA nephropathy is the most common form of primary glomerulonephritis worldwide. The basic pathogenesis of this disease is believed to be due to the complex formation between Fc IgA, alphaRI (CD89), and transferrin receptor (CD71), leading to the damage of the glomerulus. However, studying the functional aberration in the human IgA, CD89, and CD71 complex formation in pathogenesis of IgA nephropathy is hard. Here, the author used a new gene ontology technology to predict the molecular function of human IgA, CD89, and CD71 complex. It can be seen that the two main functional aberrations of IgA-CD89 complex might be due to signal transduction and binding activity.

Objective: To study the regulation of Fc receptor expression by immune complexes (ICs) on neutrophils and U937 cells., Methods: IgA ICs, IgG1 ICs, IgG2 ICs, IgG3 ICs, IgG4 ICs, and IgM ICs were incubated with neutrophils or U937 cells for 1 h. Then their surface Fc receptors were stained by anti-Fc gammaR I, anti-Fc gammaR II , anti-Fc gammaR III, and anti-Fc alphaR I monoclonal antibodies and analyzed by fluorescent activated cell sorting (FACS)., Results: IgG1 ICs and IgG3 ICs up-regulated Fc gammaR II and Fc gammaR III on U937 cells, Fc gammaR I and Fc alphaR I on neutrophils. Almost all ICs down-regulated Fc gammaR II on neutrophils., Conclusions: ICs can regulate Fc receptor expression on neutrophils and U937 cells, among which IgG1 ICs and IgG3 ICs are most effective.

Mucosal secretions of the human gastrointestinal, respiratory, and genital tracts contain significant quantities of IgG. The mechanism by which IgG reaches luminal secretions and the function of IgG in these locations are unknown. Here, we find that the human neonatal Fc receptor (FcRn) is the vehicle that transports IgG across the intestinal epithelial barrier into the lumen where the IgG can bind cognate antigen. The FcRn can then recycle the IgG/antigen complex back across the intestinal barrier into the lamina propria for processing by dendritic cells and presentation to CD4(+) T cells in regional organized lymphoid structures. These results explain how IgG is secreted onto mucosal surfaces and scavenges luminal antigens for recognition by the immune system.

In immune complex (IC) diseases, FcR are essential molecules facilitating polymorphonuclear cell (PMN) recruitment and effector functions at the IC site. Although FcR-dependent initial tethering and FcR/integrin-dependent PMN accumulation were postulated, their underlying mechanisms remain unclear. We here addressed potential mechanisms involved in PMN recruitment in acute IC glomerulonephritis (nephrotoxic nephritis). Since some renal cells may be recruited from bone marrow (BM) lineages, reconstitution studies with BM chimeras and PMN transfer between wild-type (WT) and FcR-deficient mice (gamma(-/-)) were performed. Severe glomerular damage was induced in WT and W gamma chimeras (BM from WT to irradiated gamma(-/-)), while it was absent in gamma(-/-) and gamma W chimeras (gamma(-/-) BM to WT). Moreover, WT PMN transfer, but not gamma(-/-) PMN, reconstituted the disease in gamma(-/-), indicating that FcR on resident cells is not a prerequisite for PMN recruitment in this disease. Surprisingly, transferred WT PMN were recruited coincidentally with NF-kappa B activation and TNF-alpha overexpression even in glomeruli with preformed IC (nephrotoxic Ab administered 3 days previously), suggesting that PMN can initially be recruited via its own FcR without previous chemoattractant release. Furthermore, H(2)O(2) inhibition by catalase attenuated the acute WT PMN recruitment and the induction of NF-kappa B and TNF-alpha much more than integrin (CD18) blockade, indicating a role for the respiratory burst before integrin-dependent accumulation. In coculture experiments with IC-stimulated PMN and glomeruli, PMN caused acute glomerular TNF-alpha expression predominantly via FcR-mediated H(2)O(2) production. In conclusion, glomerular IC, even preformed, can cause PMN recruitment and injury through PMN FcR-mediated respiratory burst during initial PMN tethering to IC.

Objective: To explore the Fc receptor-dependent handling of immune complexes (ICs) by the fixed mononuclear phagocytic systems (MPS) in patients with systemic lupus erythematosus (SLE)., Methods: The processing in vivo of soluble model (123)I-hepatitis B/ anti-hepatitis B ICs was studied in 12 healthy subjects and 10 patients with active SLE. ICs that fixed complement poorly were prepared specifically in order to explore Fc receptor-dependent clearance mechanisms. Clearance kinetics and organ uptake were assessed by computer-aided gamma scintigraphy and serial blood sampling., Results: In both patients and controls, the main site of IC clearance was the liver; only 2-6% of injected ICs were taken up in the spleen. The kinetics of initial IC clearance were similar in both groups, but defective hepatic retention of ICs was demonstrated in patients with SLE. At 1 hour, hepatic activity in patients had fallen to 56% of maximum, compared with 74% in controls (P = 0.0002). Precipitation studies performed on serum samples using staphylococcal protein A-Sepharose indicated that antibody-complexed tracer was released from the liver 20-50 minutes after injection., Conclusion: These results indicate that Fc-mediated clearance of ICs is defective in patients with SLE and suggest that ligation of ICs by Fc receptors is critical for their efficient binding and retention by the fixed MPS in the liver.

Immature dendritic cells (DC) sample Ags within nonlymphoid tissues and acquire exogenous proteins/pathogens via scavenger receptors or Ig FcR such as Fc gamma R and Fc epsilon R. IgA is present in a significant proportion among serum Ig and is the main isotype in mucosae, where DC are numerous. We found that a functional Fc alpha R (CD89) was expressed in situ and in vitro on interstitial-type DC but not on Langerhans cell-type DC. Interstitial-type DC expressed CD89 as a 50- to 75-kDa glycoprotein with a 32-kDa protein core, which was down-regulated upon addition of TGF-beta 1. DC, Fc alpha R specifically, bound IgA1 and IgA2. Cross-linking of CD89 on DC triggered endocytosis in time-dependent manner. In addition, internalization of polymeric IgA complexes induced the production of IL-10 and DC activation, as reflected by up-regulation of CD86 costimulatory molecules, class II MHC expression, and increased allostimulatory activity. Therefore, interstitial-type DC may use Fc alpha R-mediated Ag sampling in the subepithelium to check tissue integrity while Langerhans cells inside epithelial layers may neglect IgA immune complexes.

The human IgA Fc receptor (FcalphaR, CD89) plays an important role in host defence against invading pathogens. To study the properties of the receptor, 12 MoAbs, namely, MIP7c, MIP8a, MIP9a, MIP10c, MIP11c, MIP14b, MIP15b, MIP38c, MIP59c, MIP65c, MIP68b and MIP71a, were generated. The inhibitory effects of the antibodies on FcalphaR functions were tested. Three of the antibodies, MIP7c, MIP8a and MIP59c, were able to block up to 90% of soluble FcalphaR binding to IgA-coated beads and 70-80% of neutrophil phagocytosis of IgA immune complexes (IC). MIP8a could also inhibit IgA IC-induced neutrophil lactoferrin release, while cross-linking of FcalphaR with MIP8a and anti-mouse IgG could elicit neutrophil lactoferrin release. However, IgA IC-induced lactoferrin release required both extracellular calcium and magnesium, whereas MIP8a-induced release did not require extracellular magnesium and only partially required extracellular calcium. In addition, the time course of IgA IC-induced lactoferrin release was slow. Lactoferrin was not detectable if the incubation time was less than 0.5 h. In contrast, MIP8a-induced lactoferrin release was fast. Lactoferrin could be detected within 5 min of incubation. Therefore, neutrophil lactoferrin release induced by IgA IC differed from that induced by cross-linking of FcalphaR with MIP8a.

Background: Most intensive investigations on the pathogenesis of IgA nephropathy have focused on the process before IgA deposition and the characteristics of IgA/IgA immune complex (IgA IC), but it still remains uncertain whether mesangial IgA ICs may cause glomerular injuries directly or are only secondary events of another pathological process. To assess the role of IgA ICs in IgA nephropathy, we investigated the characteristics of Fc alpha receptor (Fc alphaR) and FcR gamma chain which is a signalling subunit of FcR in human mesangial cells (MCs)., Methods: Gene expression of Fc alphaR and FcR gamma chain of human cultured MCs was examined by RT-PCR and subsequent Southern blot analyses. Sequence analyses after subcloning were also performed for further confirmation. Expression of Fc alphaR and FcR gamma chain at the protein level and their physical association in MCs were determined by immunoprecipitation after stimulation of the cells with heat-aggregated IgA., Results: Two distinct cDNA products were amplified from each cultured MC line. The sequence of the major product of approximately 900 bp was completely identical to that of Fc alphaR previously described. The smaller product had a 288 bp deletion which corresponded to exon 2 encoding the extracellular domain 2 of Fc alphaR. Gene expression of FcR gamma chain was also confirmed. Furthermore, we proved the physical association of Fc alphaR with the FcR gamma chain by co-immunoprecipitation under stimulation with a high dose of the heat-aggregated IgA., Conclusion: These findings suggested that polymeric IgA and/or IgA IC can directly activate MCs via Fc alphaR associated with the gamma chain. Our data also indicated that phenotypic variations of Fc alphaR occur on MC, such as splicing forms, the chain association and/or the alpha chain expression itself, which may contribute to the pathogenesis of IgA nephropathy.

Monomeric maternal antibodies are transmitted to the fetus, but most immune complexes are trapped in the placental stroma. The receptors responsible for trapping immune complexes appear to be Fc gamma RIa, Fc gamma RIIa, and Fc gamma RIIIa on stromal macrophages, and Fc gamma RII on fetal capillary endothelial cells.

Our previous results showed that neutrophil secondary granule release, indicated by release of lactoferrin, was a slow process when induced by IgA immune complexes (IC) formed in heat-inactivated serum, but became very fast if IgA IC were formed in normal human serum. This phenomenon did not apply to the IC of other Ab isotypes. In this paper, we demonstrate that the fast lactoferrin release is caused by complement, mainly due to the deposition of C3b and iC3b on IgA IC. Either CR1 or CR3 can mediate the response and both receptors have to be blocked to prevent it. Complement also influences FcalphaR-mediated lactoferrin release, in that this is enhanced by the anaphylatoxin peptides, C5a and C5a(desArg). Divalent cations are required for FcalphaR and CR3- but not for CR1-mediated lactoferrin release. Genistein, a protein tyrosine kinase inhibitor, totally inhibits FcalphaR-mediated response, but has little effect on CR1-mediated response. Therefore, it is clear that different pathways of intracellular signaling are utilized. In addition, stimulation through FcalphaR promotes the receptor up-regulation, which is abolished by the presence of EDTA or genistein.

The pectic polysaccharide, bupleuran 2IIb, up-regulates Fc-receptor (FcR) expression on peritoneal macrophages in a dose-dependent manner. The intracellular signal transduction by bupleuran 2IIb leading to the expression of FcR was studied. Neither the protein kinase C (PKC) inhibitor, 1-(5-isoquinolinylsulphonyl)-2-methylpiperazine dihydrochloride, nor the structurally distinct PKC antagonist, calphostin C, inhibited bupleuran 2IIb-induced up-regulation of FcR, whereas two direct activators of PKC, L-alpha-1-oleoyl-2-acetyl-sn-3-glycerol and N-(6-phenylhexyl)-5-chloro-1-naphthalenesulphonamide were unable to up-regulate the expression of FcR. The protein kinase A (PKA) inhibitor, N-[2-(methylamino)ethyl]-5-isoquinolinesulphonamide dihydrochloride also did not inhibit bupleuran 2IIb-induced up-regulation of FcR. Fluorescence image analysis using the calcium-sensitive dye, Fura-2, demonstrated that bupleuran 2IIb induced a rapid increase in intracellular levels of calcium (Ca2+). When macrophages were treated with calcium antagonist, 8-(diethylamino)-octyl-3,4,5-trimethoxybenzoate hydrochloride, bupleuran 2IIb-induced up-regulation of FcR was inhibited in a dose-dependent manner. The bupleuran 2IIb-induced up-regulation of FcR was also blocked by two structurally distinct calmodulin antagonists, trifluoperazine and N-(6-aminohexyl)-5-chloro-1-naphthalenesulphonamide hydrochloride. Furthermore, elevation of intracellular Ca2+ using the calcium ionophore, A23187, led to up-regulation of the FcR expression in a dose-dependent manner. These results suggest that bupleuran 2IIb induces the up-regulation of FcR on macrophages by a mechanism dependent on an increase in intracellular Ca2+ followed by activation of the calmodulin, but not by a PKC or PKA pathway.

We have previously shown that a synthetic peptide corresponding to amino acid residues 21-49 of the first extracellular domain of human CD4 binds immunoglobulins (Ig) and antibody: antigen (Ab:Ag) complexes, and greatly enhances the uptake of aggregated Ig by monocyte/macrophage U937 cells. In this report, we investigated the mechanisms of enhanced uptake, and the contribution of different receptors present on the surface of monocyte/macrophage cells to this phenomenon. Our results indicate that both Fc receptor (FcR) and cell surface CD4 participate in the enhanced uptake of Ig promoted by the synthetic peptide of CD4. The involvement of these two receptors was demonstrated in experiments using monoclonal antibodies to FcR and CD4, as well as monosialoganglioside GM1, a substance known to modulate surface CD4. The participation of CD4 was further confirmed using the CD4 monocyte/macrophage cell line MM-6. Together, the results of these experiments indicate that surface CD4 may cooperate with FcR in handling aggregated Ig and Ab:Ag complexes. The implications of these findings for immunoregulation by Ab:Ag and idiotype:anti-idiotype (Id:anti-Id) complexes, and infection of macrophages by HIV, are discussed.

A study was undertaken to reveal the role of Fc and C3b receptor of mouse peritoneal macrophages (MPM) in the uptake of radiolabelled immune complexes. Large latticed preformed complexes consisting of human serum albumin (HSA)-anti HSA at equivalence (IC-Eq) and with antibody excess (IC-Ab) were observed to be avidly taken up by resident macrophages unlike small size complexes with antigen excess (IC-Ag). Macrophages elicited by thioglycollate (Tg) showed higher IC-binding capacity while IC-elicited MPM showed reduction in the same when compared to the resident cells. However, complement coated complexes were significantly taken up by these IC-elicited macrophages. Uptake studies were further extended to determine the expression of Fc and C3b receptor activity in MPM when elicited with preformed IC. Tg-elicited MPM were observed to bind greater number of IgG-coated erythrocytes (E-IgG) than resident MPM whereas IC-elicited MPM bound E-IgG poorly. When Fc receptors were blocked by in vitro IC treatment, poor binding of complement coated E-IgG [E(IgG)C] was recorded in resident MPM. The present complement medicated rosetting data tends to show enhanced expression of C3b receptors on IC-elicited macrophages.

Binding of glucose oxidase-anti-glucose oxidase complexes (GAG), a model of immune complexes, to macrophages was enhanced by treatment with an acidic pectic polysaccharide, bupleuran 2IIb, from Bupleurum falcatum L. GAG binding to macrophages by bupleuran 2IIb increased in a dose-dependent fashion, and was abolished when the Pronase-treated macrophages were incubated with bupleuran 2IIb. The GAG binding enhancing activity of bupleuran 2IIb was reduced by periodate oxidation but not Pronase digestion of bupleuran 2IIb. When bupleuran 2IIb was digested with endo-polygalacturonase, the resulting enzyme resistant carbohydrate portion showed potent activity. Scatchard analysis indicated enhanced expression of the Fc receptor (FcR) on the surface by the action of bupleuran 2IIb. The enhancement of GAG binding by bupleuran 2IIb was inhibited by the presence of actinomycin D or cycloheximide. Bupleuran-2IIb-stimulated cells showed enhanced expression of both FcRI and FcRII mRNA, which were measured as PCR products. These results suggested that the endo-polygalacturonase resistant carbohydrate portion of bupleuran 2IIb is important for the expression of the activity, and that the activity of bupleuran 2IIb on GAG binding was mediated by receptors for polysaccharide on the cells. The up-regulation of the Fc receptor by bupleuran 2IIb was also suggested to mediate by de novo synthesis of the receptor protein.

Fc gamma receptors are a group of three different receptors with several subtypes. They are widely distributed on many cells of the immune system and contribute to the pathogenesis of immune complex- and autoantibody-mediated diseases such as vasculitis, rheumatoid arthritis, idiopathic thrombocytopenic purpura or autoimmune neutropenia. This review focuses on the structure, distribution and function in Fc gamma receptors and their subtypes.

B lymphocytes and macrophages express closely related immunoglobulin G (IgG) Fc receptors (Fc gamma RII) that differ only in the structures of their cytoplasmic domains. Because of cell type-specific alternative messenger RNA splicing, B-cell Fc gamma RII contains an insertion of 47 amino acids that participates in determining receptor function in these cells. Transfection of an Fc gamma RII-negative B-cell line with complementary DNA's encoding the two splice products and various receptor mutants indicated that the insertion was responsible for preventing both Fc gamma RII-mediated endocytosis and Fc gamma RII-mediated antigen presentation. The insertion was not required for Fc gamma RII to modulate surface immunoglobulin-triggered B-cell activation. Instead, regulation of activation involved a region of the cytoplasmic domain common to both the lymphocyte and macrophage receptor isoforms. In contrast, the insertion did contribute to the formation of caps in response to receptor cross-linking, consistent with suggestions that the lymphocyte but not macrophage form of the receptor can associate with the detergent-insoluble cytoskeleton.

The expression and ligand binding activity of Fc receptors for IgG (Fc gamma R) on guinea-pig peripheral blood polymorphonuclear leukocytes (blood PMN) have been compared with those on casein-elicited peritoneal PMN (exudate PMN). Both the PMN were found to express two distinct types of Fc gamma R, one specific for IgG1 and IgG2 (Fc gamma 1/gamma 2 R) and the other for IgG2 alone (Fc gamma 2 R), when evaluated by their reactivity to monoclonal antibodies (mAb) directed against each type of Fc gamma R. The surface density of Fc gamma 1/gamma 2 R was not significantly different between the two cell types, whereas exudate PMN expressed five times as many Fc gamma 2 R as blood PMN. Moreover, IgG immune complex (IC) binding activities of both the Fc gamma R on blood PMN were markedly low as compared with those on exudate PMN. In addition, blood PMN could not significantly generate superoxide anion (O2-) when exposed to IC. However, these lower activities were improved by protease treatment of the cells or by incubation with platelet activating factor (PAF). Fc gamma 2 R on blood PMN was found to be sensitive to pronase, whereas Fc gamma 1/gamma 2 R was resistant. Pronase-treated blood PMN showed a marked IC binding activity, though they lacked Fc gamma 2 R. This activity was completely blocked by anti-Fc gamma 1/gamma 2 R mAb, indicating that the proteolysis augments the ligand binding capacity of Fc gamma 1/gamma 2 R. In contrast, PAF was found to specifically modulate Fc gamma 2 R. The Fc gamma 2 R expression was significantly increased within 5 min incubation with PAF, whereas that of Fc gamma 1/gamma 2 R was not affected. The cells also exhibited enhanced IgG2-IC binding and subsequent O2- generating activities. These results indicate that both the Fc gamma R on blood PMN are functionally immature and are converted to exhibit intrinsic activities by proteases and PAF; such changes may occur in vivo during exudation and at inflammatory sites.

Granulocyte Fc gamma receptors are important components in the recognition of IgG-coated cells and immune complexes. Two proteins have been identified on resting human granulocytes which function as Fc gamma receptors, Fc gamma RII (CD32) and Fc gamma RIII (CD16). A third protein, Fc gamma RI (CD64), is not constitutively expressed on resting granulocytes, but can be induced by activation with gamma-interferon. We examined the role of these three Fc gamma receptors on human granulocytes in the binding of both IgG-sensitized erythrocytes and soluble oligomeric IgG. In these studies we employed anti-Fc gamma receptor antibodies which complete for the Fc gamma RII and Fc gamma RIII ligand binding sites. Preincubation of granulocytes with saturating concentrations of high-affinity anti-Fc gamma RII monoclonal antibody did not alter the recognition of IgG sensitized human cells by granulocytes. Furthermore, ligand binding studies demonstrated that anti-Fc gamma RII antibody altered neither the number nor the affinity of granulocyte binding sites for human trimeric IgG. In contrast, Fab anti-Fc gamma RIII inhibited the binding of both IgG (anti-D) sensitized human RBCs and IgG sensitized sheep RBCs. Similarly, a reduction in the expression of Fc gamma RIII by treatment with phosphatidyl-inositol specific phospholipase C reduced PMN recognition of IgG-sensitized cells. Also, anti-Fc gamma RIII decreased the number of granulocyte binding sites for human IgG trimer without a change in receptor affinity. Fc gamma RI, which was induced by gamma-IFN, increased granulocyte recognition of both IgG sensitized RBCs and IgG trimer. These data suggest that Fc gamma RIII is the primary Fc gamma receptor on granulocytes which recognizes IgG sensitized RBCs and low molecular weight complexes of IgG. With gamma-interferon activated granulocytes, Fc gamma RI appears to enhance this recognition process.

Fc gamma RIII is one of two Fc gamma R constitutively expressed by human neutrophils. We have prepared a panel of anti-Fc gamma RIII mAb following immunization of mice with Fc gamma RIII purified from human neutrophils. Ten mAb which reacted with neutrophils, NK cells, and monocyte-derived macrophages were produced. Immunohistochemical staining demonstrated that these mAb also identified macrophages in the red pulp of spleen. Competitive cross-inhibition binding assays demonstrated that nine of the ten mAb reacted with a common epitope that is spatially associated with the ligand binding site. These nine mAb blocked the binding of immune complexes to neutrophils by 65 to 90%. In addition, two other anti-CD16 mAb, which also blocked immune complex binding to neutrophils, inhibited the binding of each of these nine mAb to neutrophils. One of the mAb produced here, 214.1, failed to block immune complex binding. In addition to immunoprecipitating the native Fc gamma RIII glycoprotein, mAb 214.1 was capable of immunoprecipitating a 28-kDa polypeptide following deglycosylation of Fc gamma RIII isolated from neutrophils. The results of cross-competition experiments suggest that mAb 214.1 may recognize the epitope identified by mAb BW209/2. Thus mAb 214.1 identifies a polypeptide epitope distinct from the ligand binding site of Fc gamma RIII on neutrophils.

Guinea-pig peritoneal macrophages express two distinct types of Fc receptor for IgG (Fc gamma R): one specific for IgG2 (Fc gamma 2R) and the other for both IgG1 and IgG2 (Fc gamma 1/gamma 2R). When we employed flow cytometry for an assay, we found that the amount of ovalbumin (OA) complex of homologous IgG2 antibody bound on the surface of macrophages rapidly decreased during the phagocytosis in the presence of an excessive amount of the complex. This reduced binding capacity of the cells was gradually restored by incubating the cells in the complex-free medium, which showed that the Fc gamma Rs are consumed during the phagocytosis and again expressed on the cell surface. Flow cytometry with monoclonal anti-Fc gamma 2R Fab' and anti-Fc gamma 1/gamma 2R Fab' revealed that only the Fc gamma R type bound to the immune complex was selectively internalized, whereas another Fc gamma R type unbound persisted on the cell surface during the reaction. In addition, the amount of Fc gamma 1/gamma 2R on the cell surface was found to increase to a greater extent than did that of Fc gamma 2R, when phagocytosis was terminated by the removal of the immune complex. This result suggests that Fc gamma 1/gamma 2R is recruited from some intracellular store. In fact, we were able to demonstrate the existence of the membrane-associated intracellular Fc gamma 1/gamma 2R pool that increases the binding capacity of anti-Fc gamma 1/gamma 2R F(ab')2 by treatment of macrophages with saponin, and by fractionation of homogenized macrophages by sucrose density gradient centrifugation. The different behaviour of these two Fc gamma R type, thus shown, may cause the relatively sustained phagocytosing activity mediated by Fc gamma 1/gamma 2R compared with that caused by Fc gamma 2R; the former continued at least up to 6 hr, while the latter ceased within 2 hr.

Murine B lymphocytes cultured with F(ab')2 anti-mouse mu or delta lost (85%) the capacity to bind antigen-IgG antibody complexes as assessed by flow microfluorometry. Anti-mu-induced loss of binding of complexes was concentration, time, and temperature dependent, reversible, and not due to decreased expression of the receptor because binding of monoclonal anti-Fc gamma R II to B lymphocytes cultured with anti-mu was unaffected. Activation of PKC and elevation of [Ca2+]i obtained by culturing B lymphocytes with the combination of PMA and Ca2+ ionophore induced a similar loss of binding of Cx. Since stimulation of B lymphocytes with anti-mu also activates PKC and elevates [Ca2+]i, these changes may be involved in the anti-mu-induced alterations in the binding of complexes to Fc gamma R II. In contrast to the effects of other activators, LPS caused increased expression (threefold) of B lymphocyte Fc gamma R II as measured by the binding of both complexes and monoclonal anti-Fc gamma R II. Thus, different B lymphocyte activators have distinct effects on Fc gamma R II expression or ligand binding capacity and can thereby affect Fc gamma R II-generated regulatory signals.

Two isotypes of the monocyte/macrophage as well as B cell Fc gamma receptor type II (FcRIIa and FcRIIb2, respectively) mainly differ in the length (76 vs. 44 amino acids) and amino acid sequence of their cytoplasmic domains. Only the eight amino acids just behind the putative transmembrane region are identical. Despite this marked difference, both FcRII mediate endocytosis of immune complexes. To determine the functional significance of the cytoplasmic domains, we expressed truncated FcRIIa and FcRIIb2 in FcR- BHK-21 cells. Mutants of both receptors containing only one amino acid (tail-minus) of the cytoplasmic domain failed to mediate immune complex uptake. The significance of the cytoplasmic domain of the receptors could be further demonstrated using a chimeric FcRIII-FcRIIa construct. Therefore we expressed an FcRIII lacking the hydrophobic carboxyl terminus (containing the putative phosphatidyl - inositol - glycan anchor site) fused inframe to the transmembrane and cytoplasmic domain of the FcRIIa in BHK-21 cells. In contrast to the wild type FcRIII, this chimeric receptor mediated immune complex uptake indistinguishable from that mediated by the FcRIIa. Receptor mutants with relatively short cytoplasmic domains (FcRIIb2: 13, and FcRIIa: 16 amino acids) revealed, that these short amino acid stretches are sufficient to allow reduced receptor-mediated endocytosis of bound ligand. Furthermore, using FcRIIa deletion mutants with a cytoplasmic domain consisting of 62, 46, and 28 amino acids, respectively, we found that the capability of these mutants to mediate immune complex uptake decreased gradually with the truncation of the cytoplasmic tails. Thus, only short amino acid sequences of the cytoplasmic domain are sufficient to enable an, albeit reduced, receptor-mediated endocytosis.

B lymphocytes cultured with LPS show increased expression of Fc gamma R II and increased binding of Ag-IgG complexes (both greater than 200%). In contrast, B lymphocytes cultured with either IL-4 or anti-mu show a marked loss (85-90%) of binding of Ag-IgG complexes that is specific, time and temperature dependent, and reversible. Decreased binding of complexes was not due to decreased expression of the receptor and therefore appears to be due to some form of alteration of the receptor. Based on the observation that the loss of binding of complexes requires protein synthesis, we favor the view that the loss is due to association of Fc gamma R II with another membrane molecule whose expression is induced or increased by IL-4 or anti-mu. Anti-mu induced loss of Fc gamma R II ligand binding capacity does not require cross-linking of surface IgM because the effect can be generated with F(ab') anti-mu. Anti-mu induced loss of Fc gamma R II binding of complexes was substantially prevented by IFN-gamma, whereas IFN-gamma did not reduce the anti-mu caused increase in expression of MHC class II molecules. This result shows that increased expression of the latter molecules can be dissociated from loss of Fc gamma R II ligand binding capacity. A myeloid cell line was identified that constitutively expresses Fc gamma R II binds relatively few complexes. This cell line may be useful in identifying alterations of Fc gamma R II which lead to the loss of binding of complexes. These results indicate that various B lymphocyte activators have different effects on B lymphocyte expression and function, and can thereby affect Fc gamma R II generated regulatory signals.

The purpose of this paper is to provide a histopathologic basis for abnormalities in immune-complex clearance in liver disease. Fc receptors in CCl4-induced liver cirrhosis in rats were studied by applying peroxidase-antiperoxidase immunoglobulin G complex as a ligand to the frozen sections. Intravenous injection of bovine serum albumin-antibovine serum albumin complexes or colloidal carbon was combined with histological staining for endogenous peroxidase, fibronectin, laminin, or a lectin, Bandeiraea simplicifolia agglutinin I. In the cirrhotic process, sinusoidal Fc receptors showed a weakened reactivity to the ligand with focal absence, and the length of the Fc receptor-positive portion of the sinusoids in unit area decreased to about 50% of the normal value in the advanced cirrhosis. Fibronectin and the lectin showed the presence of sinusoids where Fc receptors were absent. The endothelium in Fc receptor-negative areas did not take up either immune complexes or carbon, and Kupffer cells were absent in these areas. A disturbed immune-complex metabolism was thus suggested to occur in association with the defect of sinusoidal Fc receptors in liver cirrhosis. These abnormalities appeared to not be directly related to perisinusoidal laminin deposition, i.e., capillarization of the sinusoid.

A cDNA encoding the human beta Fc gamma receptor II (FcRII) was isolated from a placental cDNA library. Analysis of the predicted amino acid sequence indicates that this receptor is synthesized with a 42-amino acid leader sequence. The mature protein consists of 249 amino acids. The leader sequence and the cytoplasmic domain are strikingly different from the CDw32 antigen but show great homology to the mouse beta 2FcR. RNA blot analysis of human cells using CDw32 and beta FcRII-specific DNA fragments demonstrated one beta FcRII transcript (1.7 kb) in B cells and in HL-60 cells which were induced to differentiate along a monocyte-macrophage pathway by phorbol 12-myristate 13-acetate treatment. Under these conditions the CDw32 transcripts (2.5 and 1.7 kb) are induced to a minor extent in HL-60 cells. In contrast, the 2.5-kb CDw32 transcript is strongly induced in HL-60 cells which have been induced to differentiate into granulocytes by exposure to dimethylsulfoxide. To determine the biological properties of the beta FcRII, we expressed the antigen in FcR- hamster cells. Only immune complexes but not monomeric human IgG were bound significantly. Bound ligand was efficiently internalized within 15 min and it was then found in vesicular structures. Thus the low-affinity beta FcRII is able to internalize ligands without cooperation with any other FcR.

A new photometric microassay for immune complex (IC) binding to macrophages was developed in a homologous system using glucose oxidase-anti-glucose oxidase complexes (GOAGO) as a model for IC clearance in vitro. Thioglycollate-elicited murine peritoneal cells were incubated with GOAGO solution and then cell-associated glucose oxidase activity was measured after the washing and solubilization of the cell membrane in a microtitre plate. GOAGO binding to macrophages was inhibited in the presence of either IgG or its Fc fragments in a dose-dependent manner, while yeast mannan or IgG Fab fragments had no effect. These results indicated that this binding occurred solely via the Fc receptors on the macrophages. The Fc receptors for GOAGO were eliminated by trypsin digestion of the cells. When the macrophages were cultured with LPS or TPA, GOAGO binding was enhanced compared to that of control, whereas carrageenan treatment suppressed GOAGO binding. The present results suggest that this assay may be of value in the measurement of IC clearance and for studying the expression of Fc receptors on macrophages.

Fc receptors in sinusoidal cells and immune complex uptake were studied histologically in D-galactosamine HCl (GalN)-induced liver injury in rats. Kupffer cells and monocytes were distinguished from sinusoidal endothelial cells and from each other by endogenous peroxidase staining. Fc receptors were found along the sinusoidal endothelium throughout the lobules in normal livers. In acute injury caused by 300 or 750 mg/kg of GalN, Fc receptors were preserved within necrotic foci until the foci were infiltrated by inflammatory cells. The endothelial Fc receptor activity altered, as demonstrated by their capacity to bind immune complexes, after GalN injection. The activity decreased from 24 h after injection in the periportal areas in both dose groups, and increased transiently with dose-dependence in the remaining areas. Kupffer cell numbers also showed a transient dose-dependent increase, except in the periphery of lobules where they generally decreased. In chronic injury with 400 mg/kg, Fc receptors were lost and Kupffer cells decreased in the periportal areas. Circulating immune complexes were ingested by Kupffer cells and endothelial cells in normal and injured livers, showing the the same distribution as that of Fc receptors except that the complexes decreased gradually towards the centrilobular zones.