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Inflammation Unrestrained by SIRPα Induces Secondary Hemophagocytic Lymphohistiocytosis Independent of IFN-γ.

ABSTRACT: A hallmark of secondary hemophagocytic lymphohistiocytosis (sHLH), a severe form of cytokine storm syndrome, is the emergence of overactivated macrophages that engulf healthy host blood cells (i.e., hemophagocytosis) and contribute to the dysregulated inflammation-driven pathology. In this study, we show that depleting SIRPα (SIRPα^-/-) in mice during TLR9-driven inflammation exacerbates and accelerates the onset of fulminant sHLH, in which systemic hemophagocytosis, hypercytokinemia, consumptive cytopenias, hyperferritinemia, and other hemophagocytic lymphohistiocytosis hallmarks were apparent. In contrast, mice expressing SIRPα, including those deficient of the SIRPα ligand CD47 (CD47^-/-), do not phenocopy SIRPα deficiency and fail to fully develop sHLH, albeit TLR9-inflamed wild-type and CD47^-/- mice exhibited hemophagocytosis, anemia, and splenomegaly. Although IFN-γ is largely considered a driver of hemophagocytic lymphohistiocytosis pathology, IFN-γ neutralization did not preclude the precipitation of sHLH in TLR9-inflamed SIRPα^-/- mice, whereas macrophage depletion attenuated sHLH in SIRPα^-/- mice. Mechanistic studies confirmed that SIRPα not only restrains macrophages from acquiring a hemophagocytic phenotype but also tempers their proinflammatory cytokine and ferritin secretion by negatively regulating Erk1/2 and p38 activation downstream of TLR9 signaling. In addition to TLR9 agonists, TLR2, TLR3, or TLR4 agonists, as well as TNF-α, IL-6, or IL-17A, but not IFN-γ, similarly induced sHLH in SIRPα^-/- mice but not SIRPα⁺ mice. Collectively, our study suggests that SIRPα plays a previously unappreciated role in sHLH/cytokine storm syndrome pathogenesis by preventing macrophages from becoming both hemophagocytic and hyperactivated under proinflammation.

SUBMITTER: Kidder K

PROVIDER: S-EPMC7658050 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundInflammatory activation of CD8+ T cells can, when left unchecked, drive severe immunopathology. Hyperstimulation of CD8+ T cells through a broad set of triggering signals can precipitate hemophagocytic lymphohistiocytosis (HLH), a life-threatening systemic inflammatory disorder.ObjectiveThe mechanism linking CD8+ T-cell hyperactivation to pathology is controversial, with excessive production of IFN-γ and, more recently, excessive consumption of IL-2, which are proposed as competing hypotheses. We formally tested the proximal mechanistic events of each pathway in a mouse model of HLH.MethodsIn addition to reporting a complete autosomal recessive IFN-γ receptor 1-deficient patient with multiple aspects of HLH pathology, we used the mouse model of perforin (Prf1)KO mice infected with lymphocytic choriomeningitis virus to genetically eliminate either IFN-γ production or CD25 expression and assess the immunologic, hematologic, and physiologic disease measurement.ResultsWe found a striking dichotomy between the mechanistic basis of the hematologic and inflammatory components of CD8+ T cell-mediated pathology. The hematologic features of HLH were completely dependent on IFN-γ production, with complete correction after loss of IFN-γ production without any role for CD8+ T cell-mediated IL-2 consumption. By contrast, the mechanistic contribution of the immunologic features was reversed, with no role for IFN-γ production but substantial correction after reduction of IL-2 consumption by hyperactivated CD8+ T cells. These results were complemented by the characterization of an IFN-γ receptor 1-deficient patients with HLH-like disease, in whom multiple aspects of HLH pathology were observed in the absence of IFN-γ signaling.ConclusionThese results synthesize the competing mechanistic models of HLH pathology into a dichotomous pathogenesis driven through discrete pathways. A holistic model provides a new paradigm for understanding HLH and, more broadly, the consequences of CD8+ T-cell hyperactivation, thereby paving the way for clinical intervention based on the features of HLH in individual patients.

Project description:ObjectiveFamilial hemophagocytic lymphohistiocytosis (FHLH) is a complex cytokine storm syndrome caused by genetic abnormalities rendering CD8+ T cells and natural killer cells incapable of cytolytic killing. In murine models of FHLH, interferon-γ (IFNγ) produced by CD8+ T cells has been identified as a critical mediator of disease, and an IFNγ-blocking antibody (emapalumab) has recently been approved by the Food and Drug Administration. However, development of hemophagocytic lymphohistiocytosis (HLH)/macrophage activation syndrome (MAS) in patients who are genetically unresponsive to IFNγ questions the absolute necessity of IFNγ in driving disease. This study was undertaken to determine the necessity of IFNγ in driving HLH.MethodsIFNγ-/- Prf1-/- mice were infected with lymphocytic choriomeningitis virus (LCMV), and HLH immunopathologic features, including survival, weight loss, cytopenias, cytokine profiles, and immune cell phenotypes, were assessed. Mixed bone marrow chimeras were created to determine the immune cell-intrinsic role of IFNγ receptor signaling. CD8+ T cell depletion and interleukin-33 (IL-33)/ST2 blockade were performed using monoclonal antibodies.ResultsLCMV infection of IFNγ-/- Prf1-/- mice resulted in severe HLH-like disease. CD8+ T cells and the IL-33/ST2 axis remained essential mediators of disease; however, IFNγ-independent HLH immunopathology correlated with a 10-15-fold increase in neutrophilia (P < 0.001) and an altered cytokine milieu dominated by IL-6, IL-1β, and granulocyte-macrophage colony-stimulating factor (GM-CSF) (P < 0.05). Furthermore, IFNγ regulated CD8+ T cell expression of GM-CSF and neutrophil survival.ConclusionIFNγ is not necessary for the development of fulminant HLH, requiring physicians to consider case-by-case treatment strategies. Use of therapies that target upstream activators of CD8+ T cells, such as IL-33/ST2 signaling, may be more universally applicable treatment options that ameliorate both IFNγ-dependent and -independent manifestations of HLH/MAS.

Project description:ObjectiveTo investigate the activation of the IFNγ signaling pathway in monocytes of patients with secondary hemophagocytic lymphohistiocytosis (sHLH)/macrophage activation syndrome (MAS) and to evaluate whether levels of phosphorylated STAT1 represent a biomarker for the identification of patients at early stages of the disease.MethodsFresh whole blood samples from pediatric patients with active sHLH/MAS, not receiving (n=10) and receiving glucocorticoids (n=14) at time of sampling, were prospectively collected. As disease control groups, patients with active systemic juvenile idiopathic arthritis (sJIA) without MAS, patients with sHLH/MAS in remission and patients with other rheumatic diseases were also sampled. Whole blood cells were left unstimulated or stimulated with increasing concentrations of IFNγ for 10 minutes and the intracellular Tyrosine (701)-phosphorylated STAT1 (pSTAT1) levels were evaluated in monocytes by flow cytometry.ResultsMonocytes from untreated sHLH/MAS patients showed significantly higher basal levels of pSTAT1 compared to those observed in monocytes from glucocorticoid-treated sHLH/MAS patients and from all the other disease controls. In addition, a significant increase in responsiveness to IFNγ, as assessed by increased levels of pSTAT1 following ex vivo stimulation, was observed in monocytes from untreated sHLH/MAS patients. pSTAT1 levels in monocytes distinguished patients with sHLH/MAS not treated with glucocorticoids from patients with active sJIA or with other rheumatic diseases [AUC, 0.93; 95% confidence interval 0.85-1.00, p<0.001]. Statistically significant correlations between IFNG mRNA levels in whole blood cells, circulating IFNγ levels and pSTAT1 levels in sHLH/MAS monocytes were found.ConclusionOur data demonstrating higher basal levels of pSTAT1 as well as a hyperreactivity to IFNγ stimulation in monocytes from patients with sHLH/MAS point to perturbations in the activation of downstream IFNγ signaling pathway as a contributor to the hyperinflammation occurring in these patients. Finally, the observation that glucocorticoids affect pSTAT1 levels in vivo, makes it difficult to consider the measurement of pSTAT1 levels as a biomarker to identify patients at early stages of sHLH/MAS in clinical practice.

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Inflammation Unrestrained by SIRPα Induces Secondary Hemophagocytic Lymphohistiocytosis Independent of IFN-γ.

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