Gaucher disease is a lysosomal storage disease, which happens due to mutations in GBA1/Gba1 that encodes the enzyme termed as lysosomal acid β-glucosidase. The major function of this enzyme is to catalyze glucosylceramide (GC) into glucose and ceramide. The deficiency of this enzyme and resultant abnormal accumulation of GC cause altered function of several of the innate and adaptive immune cells. For example, augmented infiltration of T cells contributes to the increased production of pro-inflammatory cytokines, (e.g., IFNγ, TNFα, IL6, IL12p40, IL12p70, IL23, and IL17A/F). This leads to tissue damage in a genetic mouse model (Gba19V/−) of Gaucher disease. The cellular mechanism(s) by which increased tissue infiltration of T cells occurs in this disease is not fully understood. Here, we delineate role of the CXCR3 receptor and its exogenous C-X-C motif chemokine ligand 9 (CXCL9) in induction of increased tissue recruitment of CD4+ T and CD8+ T cells in Gaucher disease. Intracellular FACS staining of macrophages (Mϕs) and dendritic cells (DCs) from Gba19V/− mice showed elevated production of CXCL9. Purified CD4+ T cells and the CD8+ T cells from Gba19V/− mice showed increased expression of CXCR3. Ex vivo and in vivo chemotaxis experiments showed CXCL9 involvement in the recruitment of Gba19V/− T cells. Furthermore, antibody blockade of the CXCL9 receptor (CXCR3) on T cells caused marked reduction in CXCL9- mediated chemotaxis of T cells in Gba19V/− mice. These data implicate abnormalities of the CXCL9-CXCR3 axis leading to enhanced tissue recruitment of T cells in Gaucher disease. Such results provide a rationale for blockade of the CXCL9/CXCR3 axis as potential new therapeutic targets for the treatment of inflammation in Gaucher disease.
GBA1 mutations result in excess storage of glucosylceramide (GC) and the induction of Gaucher disease (GD). GD is frequently associated with elevated levels of pro-inflammatory cytokines and the development of brain inflammation. The mechanisms underlying GC-driven brain inflammation in GD are ill-defined. Recently, we described immune complexes of GC-specific IgG autoantibodies in experimental and clinical GD, which induced massive complement activation and C5a generation. Further, we found that C5a-mediated activation of its cognate C5a receptor 1 (C5aR1) tips the balance between GC formation and degradation, thereby fueling excess GC accumulation and inflammation in visceral tissues in experimental and clinical GD. Previously, the C5a/C5aR1 axis was found to regulate the blood brain barrier integrity in systemic lupus and promote neurodegeneration in Alzheimer’s disease. Here, we determined the production of C5a in the brain of Gba1 D409V/knockout (9V/null) GD-prone mice. C5a production in the brain of 9V/null mice was markedly elevated, when compared to WT control mice. Also, 9V/null mice suffered from massive accumulation of GC in the brain and loss of neurons. To assess the relevance of C5a/C5aR1 axis activation for brain inflammation in GD, we targeted glucocerebrosidase (GCase) with conduritol B epoxide (CBE) in WT and C5aR1−/− mice. Strikingly, CBE-injected WT mice died within 30 days. In contrast, all C5aR1−/− mice survived the 60 days observation window, were protected from CBE-induced accumulation of GC in the brain, showed a marked reduction of microglial cell activation and only a minor loss of neurons. Our data suggest that the C5a/C5aR1 axis is a critical driver of neurodegeneration in experimental GD.
GBA1 mutations lead to defective lysosomal glucocerebrosidase resulting in accumulation of glucosylceramide (GC) in Gaucher disease (GD). Patients with GD have an increased risk to develop B cell lymphomas. The exact mechanistic bases for this propensity remain elusive. Recently, we uncovered formation of GC-specific IgG autoantibodies in Gba1 D409V/knockout (Gba19V/−) mice, which recapitulate features of human GD, and in humans with untreated GD. In vivo formation of IgG-GC immune complexes induced massive complement activation and C5a generation. Importantly, C5a-mediated activation of its cognate C5a receptor 1 (C5aR1) on immune cells enhanced GC synthesis, thereby fueling GC accumulation and excess tissue recruitment and activation of inflammatory myeloid and lymphoid immune cells, leading to visceral tissue damage in GD. Here, the expression of Runt-related transcription factor 1 (RUNX-1) was determined in Gba19V/− mice, to evaluate if C5a/C5aR1 axis activation may control the development of lymphomas in GD. RUNX-1 is a member of the Runt oncogene family linked to hematologic malignancies. We determined RUNX-1 expression in tissue from C5aR1 sufficient (+/+) and deficient (−/−) Gba19V/− mice as well as strain-matched control WT and C5aR1−/− mice. Compared to WT, Gba19V/− mice had increased RUNX-1 expression. Strikingly, RUNX-1 expression was markedly downregulated in C5aR−/−Gba19V/− vs. C5aR1+/+Gba19V/− mice. Our findings suggest that the C5a-C5aR1 axis activation in GD drives RUNX1 expression as a novel mechanism to control the development of hematologic malignancies in GD that may be diminished by targeting the C5aR1 axis in GD.
Gaucher disease (GD) is caused by GBA1 mutations that lead to decreased activity of lysosomal acid β-glucosidase and abnormal tissue accumulation of its substrate, glucosylceramide (GC). Monocyte lineage cells, e.g., macrophages (Mφs) and dendritic cells (DCs) are prominent disease effectors due to their massive accumulation of GC resulting in “Gaucher cells.” Interaction of Gaucher cells and T lymphocytes trigger massive secretion of IFNγ, TNFα, IL6, IL12p40, IL12p70, IL23, and IL17A/F, that leads to the tissue destruction in GD. The exact mechanisms that trigger excess tissue T cells recruitment in GD are still unclear. CXCR3+ T cells are enriched at inflammatory sites in patients with rheumatoid arthritis and multiple sclerosis. Using D409V/null (Gba19V/−) GD mouse model, this study uncovered the role of CXCR3 receptor and its C-X-C Motif Chemokine Ligand 9 (CXCL9) in induction of excess tissue recruitment of T cells in GD. Intracellular FACS staining of Mφs and DCs from Gba19V/− mice showed elevated amounts of CXCL9. CD4+ T and the CD8+ T cells purified from Gba19V/− mice showed increased amounts of CXCR3. Ex vivo and in vivo chemotaxis experiments, evaluated CXCL9’s recruitment of Gba19V/− T cells. Blockade of T-cell CXCL9 receptor using antibodies to CXCR3 caused marked reduction in their CXCL9-mediated chemotaxis in Gba19V/− mice. These data suggest that Gba1 defects and the resultant deficiency of acid β-glucosidase activate CXCL9-CXCR3 axis for enhanced tissue Tells recruitment of T cells in GD. Collectively, our results indicate a critical role for CXCR3 mediated T cell transmigration to sites of inflammation. Molecular targeting of CXCL9/CXCR3 axis may provide needed anti-inflammatory therapies in human patients with GD. Supported by Cincinnati Children’s Hospital Medical Center, 3333 Burnet Avenue, Cincinnati, Ohio 45229, USA
Gaucher disease (GD) is the classical example of lysosomal storage disease, which happens due to mutations in GBA1 as well as the resultant deficiency of glucocerebrosidase (GCase) and the excess tissue accumulation of glucosylceramide (GC). GD patients have shown high risk for developing anemia, which is characterized by Red blood cells (RBCs) and the hemoglobin (Hb) abnormalities. The exact mechanism by which such blood aberrations progress in GD is not clear. We have described immune complexes of GC-specific IgG autoantibodies in GD, which induced massive complement activation and the resulting complement 5a (C5a) and its cognate receptor 1 (C5aR1)-mediated immune inflammation and tissue disruption in GD. Complement activation had been linked to RBCs and/or Hb abnormalities in several diseases, (e.g., atypical hemolytic uremic syndrome, paroxysmal nocturnal hemoglobinuria, systemic lupus erythematosus, malaria, and β-thalassemia). Hence, we hypothesized that the C5a-C5aR1 activation is critical for anemia development in GD. To indorse this theory, blood from C5aR1 deficient (C5aR1−/−) and sufficient (C5aR1+/+) experimental mouse model of GD as well as the background matched control WT and C5aR1−/− mice were measured for RBCs and Hb with uses of automated system and the FACS staining methods. Our data illustrate that C5aR1+/+ mouse model of GD causes increased shortage of RBCs and Hb when compared to control WT or C5aR1−/− strains. Strikingly, C5aR1−/− mouse model of GD were protected from the loss of RBCs and Hb. These findings suggest that C5a-C5aR1 axis is a critical driver of disease processing of anemia in GD. Targeting C5a-C5aR1 axis may stop or slow down anemia development in GD.
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