Rafael Bragado

Activin A skews macrophage polarization by promoting a proinflammatory phenotype and inhibiting the acquisition of anti-inflammatory macrophage markers

M-CSF favors the generation of folate receptor β-positive (FRβ⁺), IL-10-producing, immunosuppressive, M2-polarized macrophages [M2 (M-CSF)], whereas GM-CSF promotes a proinflammatory, M1-polarized phenotype [M1 (GM-CSF)]. In the present study, we found that activin A was preferentially released by M1 (GM-CSF) macrophages, impaired the acquisition of FRβ and other M2 (M-CSF)-specific markers, down-modulated the LPS-induced release of IL-10, and mediated the tumor cell growth-inhibitory activity of M1 (GM-CSF) macrophages, in which Smad2/3 is constitutively phosphorylated. The contribution of activin A to M1 (GM-CSF) macrophage polarization was evidenced by the capacity of a blocking anti-activin A antibody to reduce M1 (GM-CSF) polarization markers expression while enhancing FRβ and other M2 (M-CSF) markers mRNA levels. Moreover, an inhibitor of activin receptor-like kinase 4/5/7 (ALK4/5/7 or SB431542) promoted M2 (M-CSF) marker expression but limited the acquisition of M1 (GM-CSF) polarization markers, suggesting a role for Smad2/3 activation in macrophage polarization. In agreement with these results, expression of activin A and M2 (M-CSF)-specific markers was oppositely regulated by tumor ascites. Therefore, activin A contributes to the proinflammatory macrophage polarization triggered by GM-CSF and limits the acquisition of the anti-inflammatory phenotype in a Smad2-dependent manner. Our results demonstrate that activin A-initiated Smad signaling skews macrophage polarization toward the acquisition of a proinflammatory phenotype.

Serotonin Skews Human Macrophage Polarization through HTR2B and HTR7

Besides its role as a neurotransmitter, serotonin (5-hydroxytryptamine, 5HT) regulates inflammation and tissue repair via a set of receptors (5HT1–7) whose pattern of expression varies among cell lineages. Considering the importance of macrophage polarization plasticity for inflammatory responses and tissue repair, we evaluated whether 5HT modulates human macrophage polarization. 5HT inhibited the LPS-induced release of proinflammatory cytokines without affecting IL-10 production, upregulated the expression of M2 polarization–associated genes (SERPINB2, THBS1, STAB1, COL23A1), and reduced the expression of M1-associated genes (INHBA, CCR2, MMP12, SERPINE1, CD1B, ALDH1A2). Whereas only 5HT7 mediated the inhibitory action of 5HT on the release of proinflammatory cytokines, both 5HT2B and 5HT7 receptors mediated the pro-M2 skewing effect of 5HT. In fact, blockade of both receptors during in vitro monocyte-to-macrophage differentiation preferentially modulated the acquisition of M2 polarization markers. 5HT2B was found to be preferentially expressed by anti-inflammatory M2(M-CSF) macrophages and was detected in vivo in liver Kupffer cells and in tumor-associated macrophages. Therefore, 5HT modulates macrophage polarization and contributes to the maintenance of an anti-inflammatory state via 5HT2B and 5HT7, whose identification as functionally relevant markers for anti-inflammatory/homeostatic human M2 macrophages suggests their potential therapeutic value in inflammatory pathologies.

Chemical modification of carboxyl groups in human fcγ fragment—II. Location of acidic residues involved in complement activation

Abstract The location of the acidic residues on human Fc γ 1 which are responsible for the loss of complement-activating capacity upon chemical modification has been established. As many as 13 residues get modified at the point of functional inactivation, of which only 6 are located in the C γ 2 region. Residues Glu (269) and Glu (318) are very reactive and may be extensively modified without significantly affecting anticomplementary capacity. Both Glu (333) and Asp (249) appear to be partially labelled. They are possibly involved in intramolecular ionic pairs (Deisenhofer, 1981), so that their contribution to complement activation is considered dubious. Extensive modification of Glu (258) and Glu (293) adds to the more rapid one of Glu (269) and Glu (318) resulting in abrogation of anticomplementary capacity. The data support the idea that a charge component may be an essential part of the mechanism of complement activation. They also help to define the nature of the Fc effector site that mediates this activity.

Intravenous immunoglobulin promotes antitumor responses by modulating macrophage polarization.

Intravenous Igs (IVIg) therapy is widely used as an immunomodulatory strategy in inflammatory pathologies and is suggested to promote cancer regression. Because progression of tumors depends on their ability to redirect the polarization state of tumor-associated macrophages (from M1/immunogenic/proinflammatory to M2/anti-inflammatory), we have evaluated whether IVIg limits tumor progression and dissemination through modulation of macrophage polarization. In vitro, IVIg inhibited proinflammatory cytokine production from M1 macrophages and induced a M2-to-M1 polarization switch on human and murine M2 macrophages. In vivo, IVIg modified the polarization of tumor-associated myeloid cells in a Fcεr1γ chain–dependent manner, modulated cytokine blood levels in tumor-bearing animals, and impaired tumor progression via FcγRIII (CD16), FcγRIV, and FcRγ engagement, the latter two effects being macrophage mediated. Therefore, IVIg immunomodulatory activity is dependent on the polarization state of the responding macrophages, and its ability to trigger a M2-to-M1 macrophage polarization switch might be therapeutically useful in cancer, in which proinflammatory or immunogenic functions should be promoted.

The effect of the chemical modification in human Fcγ fragment on protein A and Fc receptor binding

The role of acidic side-chains on Fc gamma fragment in granulocyte receptor binding and in S. aureus protein A binding has been investigated by means of chemical modification. Alteration of a restricted number of carboxyl groups after 5 min of reaction is sufficient to abrogate the capacity of Fc to inhibit EA rosette formation by human neutrophils. More limited modification, which affects mainly the most exposed acidic chains, does not change receptor binding activity. In contrast, the interaction with protein A is largely unaffected, even under reaction conditions which are able to induce significant changes in the circular dichroism spectrum of Fc fragment. The results suggest that some acidic groups on Fc may be involved in the interaction with neutrophil receptor and that the binding to protein A and Fc receptor involves different sites.

Structure and Immune Recognition of HLA-B27 Antigens: Implications for Disease Association

HLA-B27 is strongly associated with susceptibility to ankylosing spondylitis and other spondyloarthropathies. Structural analysis of this antigen has revealed the existence of multiple variants, or subtypes, in human populations. The structural microheterogeneity of these subtypes deeply affects allospecific T cell recognition and most of it occurs at an spatial cluster within the peptide binding groove of the molecule. Many polymorphic residues whose combination is unique to HLA-B27 but is conserved among subtypes are clustered in a spatially separated site of the groove from that where most subtype polymorphism occurs. Site-directed mutagenesis and DNA-mediated gene transfer has been used to show that the positions that are polymorphic among subtypes are highly relevant for modulating T cell recognition, so that immunologically silent changes do not occur. These studies have also revealed an extremely high clonotypic diversity in the alloreactive response against HLA-B27. The structural basis for this diversity has been examined by sequencing the clonotypic T cell receptors. The analysis shows a certain bias in V beta gene segment usage, as well as other recurrent structural motives, among T cell receptor beta chains from HLA-B27-specific cytotoxic T cell clones.

Structural Analysis of the Polymorphism of HLA Class I Antigens

The exceptional polymorphism of HLA class I antigens is relevant to immunologists because (a) HLA class I molecules are able to elicit strong antibody and cytolytic T lymphocyte (CTL) responses upon allogeneic stimulation, such as in allografting; (b) T cell receptors on the surface of CTLs and their precursors are restricted in their capacity for antigen recognition of virus-infected or otherwise modified cells, by the associative recognition of self-HLA class I molecules; and (c) it appears that some HLA class I specificities are more efficient than others in restricting T cell recognition of a given foreign antigen. A definition of the structural features of HLA polymorphism seems essential for establishing the molecular basis of the function of HLA class I molecules.

Human Fcγ Fragment: Location of Acidic Residues Involved in Complement Activation

Classical activation of the complement system is initiated by the binding of C1 to immune complexes or aggregated IgG1. It is generally assumed that the binding site for the complement subcomponent C1q resides on the Cγ2 domain. However, a convincing picture of the precise topography and chemical nature of the complement binding site is still lacking.