Junín Virus Infection of Human Hematopoietic Progenitors Impairs In Vitro Proplatelet Formation and Platelet Release via a Bystander Effect Involving Type I IFN Signaling

Abstract

Argentine hemorrhagic fever (AHF) is an endemo-epidemic disease caused by Junín virus (JUNV), a member of the arenaviridae family. Although a recently introduced live attenuated vaccine has proven to be effective, AHF remains a potentially lethal infection. Like in other viral hemorrhagic fevers (VHF), AHF patients present with fever and hemorrhagic complications. Although the causes of the bleeding are poorly understood, impaired hemostasis, endothelial cell dysfunction and low platelet counts have been described. Thrombocytopenia is a common feature in VHF syndromes, and it is a major sign for its diagnosis. However, the underlying pathogenic mechanism has not yet been elucidated. We hypothesized that thrombocytopenia results from a viral-triggered alteration of the megakaryo/thrombopoiesis process. Therefore, we evaluated the impact of JUNV on megakaryopoiesis using an in vitro model of human CD34+ cells stimulated with thrombopoietin. Our results showed that CD34+ cells are infected with JUNV in a restricted fashion. Infection was transferrin receptor 1 (TfR1)-dependent and the surface expression of TfR1 was higher in infected cultures, suggesting a novel arenaviral dissemination strategy in hematopoietic progenitor cells. Although proliferation, survival, and commitment in JUNV-infected cultures were normal, viral infection impaired thrombopoiesis by decreasing in vitro proplatelet formation, platelet release, and P-selectin externalization via a bystander effect. The decrease in platelet release was also TfR1-dependent, mimicked by poly(I:C), and type I interferon (IFN α/β) was implicated as a key paracrine mediator. Among the relevant molecules studied, only the transcription factor NF-E2 showed a moderate decrease in expression in megakaryocytes from either infected cultures or after type I IFN treatment. Moreover, type I IFN-treated megakaryocytes presented ultrastructural abnormalities resembling the reported thrombocytopenic NF-E2−/− mouse phenotype. Our study introduces a potential mechanism for thrombocytopenia in VHF and other diseases associated with increased bone marrow type I IFN levels.

Endothelial Cell Permeability and Adherens Junction Disruption Induced by Junín Virus Infection

“…Circulating IFNs have also been previously shown to play an important role in arenavirus-mediated diseases, with increased levels correlated to increased disease severity 5,[48][49][50] In this study, we were unable to detect IFN-α, -β, or -γ in human endothelial cells infected with JUNV. Although this result indicates that the cells themselves are not responding to infection with JUNV by expressing IFN, we cannot rule out the influence of immune cells and other intercellular signaling events that occur in vivo in response to JUNV infection; in vivo studies will be critical in addressing this issue.…”

CCL5 derived from platelets increases megakaryocyte proplatelet formation

“…Intriguingly, platelet releasate increased MK proplatelet production 47% ( Figure 1B-C). [14][15][16] This novel and unexpected finding prompted further exploration. Previously, we observed that platelets release agonist-dependent factors and cytokines including abundant amounts of CCL5.…”

Platelet production proceeds independently of the intrinsic and extrinsic apoptosis pathways

“…Perhaps, in the context of a viral infection, high levels of type I interferons in vivo sensitize megakaryocytes to apoptotic death via a related mechanism. Given their pleiotropic nature, however, it seems likely that type I interferons also affect megakaryocytes indirectly, and this is supported by work involving Junín virus, the cause of Argentine haemorrhagic fever 62 . Studies utilizing mice carrying a megakaryocyte-specific deletion of IFNAR will be required to resolve the cell intrinsic versus cell extrinsic effects of type I interferons on the megakaryocyte lineage.…”