The antiphospholipid syndrome (APS) is characterized by recurrent fetal loss, vascular thrombosis, and thrombocytopenia occurring in the presence of antiphospholipid (aPL) antibodies. The pathogenesis of fetal loss and tissue injury in APS is incompletely understood, but is thought to involve platelet and endothelial cell activation as well as procoagulant effects of aPL antibodies acting directly on clotting pathway components. Recent studies have shown that uncontrolled complement activation in the placenta leads to fetal death in utero. We hypothesized that aPL antibodies activate complement in the placenta, generating split products that mediate placental injury and lead to fetal loss and growth retardation. To test this hypothesis, we used a murine model of APS in which pregnant mice are injected with human IgG containing aPL antibodies. We found that inhibition of the complement cascade in vivo, using the C3 convertase inhibitor complement receptor 1–related gene/protein y (Crry)-Ig, blocks fetal loss and growth retardation. Furthermore, mice deficient in complement C3 were resistant to fetal injury induced by aPL antibodies. While antigenic epitopes recognized by aPL antibodies are important in the pathogenesis of APS, our data show that in vivo complement activation is required for aPL antibody-induced fetal loss and growth retardation.

In the healthy colon, intestinal epithelial cells (IEC) form a physical barrier separating the myriad of gut Ags from the cells of the immune system. Simultaneously, IEC use several mechanisms to actively maintain immunologic tolerance to nonpathogenic Ags, including commensal bacteria. However, during inflammatory bowel disease (IBD), the line of defense provided by IEC is breached, resulting in uncontrolled immune responses. As IEC are a principal mediator of immune responses in the gut, we were interested in discerning the gene expression pattern of IEC during development and progression of IBD. Laser capture microdissection and microarray analysis were combined to identify the LY6 superfamily as strongly up-regulated genes in inflamed IEC of the colon in two models of murine colitis. Surface expression of LY6A and LY6C on IEC is induced by several cytokines present within the colitic gut, including IL-22 and IFN-γ. Furthermore, cross-linking of LY6C results in production of a number of chemokines which are known to be involved in the immunopathogenesis of IBD. Increased chemokine production was cholesterol dependent, suggesting a role for lipid raft structures in the mechanism. As such, LY6 molecules represent novel targets to down-regulate chemokine expression in the colon and limit subsequent inflammation associated with IBD.

Background-We recently suggested that many anticardiolipin antibodies bind only to oxidized cardiolipin (OxCL) and/or to OxCL-␤ 2 -glycoprotein 1 (␤ 2 GP1) adducts but not to a "reduced" cardiolipin that is unable to undergo oxidation. To test this hypothesis, we investigated 24 sera, 4 protein A-purified IgG fractions, and 3 human monoclonal antibodies that were all isolated from patients with antiphospholipid antibody syndrome (APS); testing was also performed in 7 controls. Two monoclonal antibodies (IS3 and IS4) were selected for binding to CL and one was selected for binding to ␤ 2 GP1 (LJB8). Methods and Results-By chemiluminescent immunoassay, all APS sera samples bound only to OxCL and not to reduced CL, and the binding was inhibited Ͼ95% by OxCL but not reduced CL. All purified IgG fractions bound to ␤ 2 GP1 but only when the ␤ 2 GP1 was plated on microtiter wells coated with OxCL. All 3 monoclonal antibodies bound only to OxCL. On Western blots, IS4 and LJB8 bound to ␤ 2 GP1 as well as to delipidated apoB of oxidized LDL but not to native apoB. IS3 also bound to oxidized apoB on Western blot. Covalent modification of ␤ 2 GP1 with oxidation products of CL made it more antigenic for APS serum samples, for purified IgG fractions, and for the monoclonal antibodies. Conclusions-These data support the hypothesis that oxidation of CL is needed to generate epitopes for many anticardiolipin antibodies and that some of these epitopes are covalent adducts of OxCL with ␤ 2 GP1 or apoB.

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Vimentin is an intermediate filament protein with diverse roles in health and disease far beyond its structural functions. Exosomes or small extracellular vesicles (sEVs) are key mediators for intercellular communication, contributing to tissue homeostasis and the progression of various diseases, especially the metastasis of cancers. In this study, we evaluated a novel vimentin-binding compound (R491) for its anti-cancer activities and its roles in cancer exosome release. The compound R491 induced a rapid and reversible intracellular vacuolization in various types of cancer cells. This phenotype did not result in an inhibition of cancer cell growth, which was consistent with our finding from a protein array that R491 did not reduce levels of major oncoproteins in cancer cells. Morphological and quantitative analyses on the intracellular vacuoles and extracellular exosomes revealed that in response to R491 treatment, the exosomes released from the cells were significantly reduced, while the exosomes retained as intra-luminal vesicles inside the cells were subsequently degraded. Vim+/− cells had lower amounts of vimentin and accordingly, lower amounts of both the retained and the released exosomes than Vim+/+ cells had, while the vimentin-binding compound R491 inhibited only the release of exosomes. Further functional tests showed that R491 significantly reduced the migration and invasion of cancer cells in vitro and decreased the amount of exosome in the blood in mice. Our study suggests that vimentin promotes exosome release, and small-molecule compounds that target vimentin are able to both block cancer exosome release and reduce cancer cell motility, and therefore could have potential applications for inhibiting cancer invasive growth.

With the Delta variant currently fueling a resurgence of new infections in the fully vaccinated population, developing an effective therapeutic drug is especially critical and urgent in fighting COVID-19. In contrast to the many efforts to repurpose existing drugs or address only one aspect of COVID-19, we are developing a novel agent with first-in-class mechanisms of action that address both the viral infection and the overactive immune system in the pathogenesis of the disease.

Little is known of the genetic factors that may contribute to the development of chronic idiopathic thrombocytopenic purpura (cITP). We have previously shown that a developmentally regulated Vh gene (Humhv3005) is absent in 10/41 (24%) of patients with systemic lupus erythematosus while it is absent in only 7/88 (8%) of normal controls. This finding suggests that a homozygous deletion of an Ig variable (V) gene may alter the immune system and thus predispose the host to an autoimmune disorder. We have analyzed the same gene in 44 patients with cITP and found that Humhv3005 and like genes were absent in a higher percentage of patients (14 of 44, 31.8%) than they were absent in either normals (7/88, 8%, p = 0.002) or thrombocytopenic patients without cITP (6/53, 11.3%, p = 0.042); the hv3005 deletion frequency in the latter group did not differ from that in normals (P = 0.74). These data suggest that deletions of Humhv3005 and/or highly homologous Vh genes may predispose individuals to the development of cITP, and may contribute toward production of pathogenic antiplatelet antibodies.

Although serum transfer studies implicate IgG anti-platelet autoantibodies in the premature platelet destruction of idiopathic thrombocytopenic purpura (ITP), many characteristics of these putative pathogenic autoantibodies remain unclear. The inability to obtain relevant monoclonal autoantibodies from patients has prevented their molecular, genetic and functional studies as a homogenous population. We have generated a monoclonal IgG anti-platelet alpha IIb beta 3 autoantibody (termed G1) from an ITP patient. G1 binds human platelets (both resting and activated) and purified alpha IIb beta 3 with a Kd of 1.57 x 10(-8) M. G1 utilizes VH4 and V lambda 2 genes. The G1 VH region apparently has a 30 nucleotide insertion in its second complementarity determining region (CDR). Notably, somatic CDR insertion in the VH region has been observed only in one IgG rheumatoid factor, and not in any characterized polyreactive human autoantibodies reported in the literature. Combined these data suggest G1 may be a disease-relevant autoantibody. Further generation and study of monoclonal IgG anti-platelet antibodies are warranted to determine the significance of such unusual autoantibodies in the immunopathogenesis of chronic ITP.