Early events in dengue virus infection inducing cytokine storm: The dynamic interplay of pattern-recognition receptors, inflammasome activation, and biphasic NF-κB and STAT1-dependent inflammatory responses in human mononuclear phagocytes.

Abstract

A Cytokine storm is critical in severe dengue, significantly contributing to disrupted endothelial integrity, plasma leakage, and haemorrhage manifestations in affected patients. Various reports have demonstrated that mononuclear phagocytes, including monocytes, dendritic cells, and macrophages, are target cells of DENV infection. They contribute to viral spread into tissues and promote robust inflammatory responses and immunopathology. However, it remains unclear whether the early events of DENV infection play a role in triggering cytokine storms in infected mononuclear phagocytes. To address this knowledge gap, we conducted a comprehensive analysis of the transcriptomic profile of in vitro DENV-2-infected human monocyte-derived macrophages (MDMs) based on the kinetics of viral replication through a standard growth curve. To verify the accuracy of our approach, we used RT-qPCR, ELISA, and transcriptomic data from in vitro DENV-2-infected monocyte-derived dendritic cells (MDDCs) and monocytes obtained from acute dengue patients. RNA-Seq analysis revealed dynamic changes in the transcriptional profile of DENV-2-infected MDMs throughout the viral growth curve. Two waves of differentially expressed genes were observed: the first occurred during the eclipse period of viral replication (3 to 5.5 h.p.i) and was associated with the induction of NF-kB-dependent pro-inflammatory factors, while the second wave at 24 h.p.i coincided with peaks of DENV-2 replication and induction of both NF-kB- and STAT1-dependent pro-inflammatory responses. Additionally, DENV-2 infection promoted the dynamic activation of Toll-like receptors, RIG-like receptors, inflammasomes, and inflammatory pathways, triggering innate pro-inflammatory and antiviral responses. A robust multi-type IFN-dependent antiviral response was also observed at the late stage of infection. A similar transcriptomic profile was found in DENV-2-infected MDDCs and monocyte subsets from acute dengue patients, further confirming the reliability of our in vitro model of DENV-infected MDMs. Together, results suggest that recognizing viral PAMPs during the eclipse period of DENV-2 infection promotes a robust NF-kB-dependent pro-inflammatory response in MDMs. In addition, at later stages of infection, recognizing structural DENV-PAMPs and/or viral replication intermediates induces both NF-kB- and STAT1-dependent pro-inflammatory responses, leading to a cytokine storm. These findings highlight the critical role of monocytes, macrophages, and dendritic cells in detecting DENV infection and triggering a cytokine storm in vitro and in vivo. This suggests that these cell populations could be potential targets for future immunotherapies to modulate the inflammatory response to DENV infection.