Date Published: November 20, 2014
Publisher: Public Library of Science
Author(s): Tsung-Ting Tsai, Yi-Jui Chuang, Yee-Shin Lin, Chih-Peng Chang, Shu-Wen Wan, Sheng-Hsiang Lin, Chia-Ling Chen, Chiou-Feng Lin, Aravinda M. de Silva. http://doi.org/10.1371/journal.pntd.0003320
Abstract: BackgroundInterleukin (IL)-10 levels are increased in dengue virus (DENV)-infected patients with severe disorders. A hypothetical intrinsic pathway has been proposed for the IL-10 response during antibody-dependent enhancement (ADE) of DENV infection; however, the mechanisms of IL-10 regulation remain unclear.Principle FindingWe found that DENV infection and/or attachment was sufficient to induce increased expression of IL-10 and its downstream regulator suppressor of cytokine signaling 3 in human monocytic THP-1 cells and human peripheral blood monocytes. IL-10 production was controlled by activation of cyclic adenosine monophosphate response element-binding (CREB), primarily through protein kinase A (PKA)- and phosphoinositide 3-kinase (PI3K)/PKB-regulated pathways, with PKA activation acting upstream of PI3K/PKB. DENV infection also caused glycogen synthase kinase (GSK)-3β inactivation in a PKA/PI3K/PKB-regulated manner, and inhibition of GSK-3β significantly increased DENV-induced IL-10 production following CREB activation. Pharmacological inhibition of spleen tyrosine kinase (Syk) activity significantly decreased DENV-induced IL-10 production, whereas silencing Syk-associated C-type lectin domain family 5 member A caused a partial inhibition. ADE of DENV infection greatly increased IL-10 expression by enhancing Syk-regulated PI3K/PKB/GSK-3β/CREB signaling. We also found that viral load, but not serotype, affected the IL-10 response. Finally, modulation of IL-10 expression could affect DENV replication.SignificanceThese results demonstrate that, in monocytes, IL-10 production is regulated by ADE through both an extrinsic and an intrinsic pathway, all involving a Syk-regulated PI3K/PKB/GSK-3β/CREB pathway, and both of which impact viral replication.
Partial Text: Four serotypes of dengue virus (DENV) – a mosquito-borne human pathogen belonging to the family Flaviviridae and the genus Flavivirus – infect an estimated 50 million people annually and cause a spectrum of illnesses, ranging from mild dengue fever (DF) to the more severe dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) . However, it is unclear which antiviral strategies are most appropriate for treating DENV progression, as many aspects of DENV pathogenesis remain controversial, including viral load, virulence, cytotoxicity, the nature of the immune response, autoimmunity , , and the potential effects of common diseases such as allergies, diabetes, and hypertension , . There are no licensed antiviral drugs for DENV treatment. Administration of chloroquine (a 9-aminoquinoline) exerts direct antiviral effects by inhibiting the pH-dependent steps of flavivirus replication, although this drug is failed to inhibit the duration of viremia and antigenemia in DENV patients . Balapiravir (4′-azidocytidine) is developed for the treatment of chronic hepatitis C Virus infection by a nucleoside analogue of RNA-dependent RNA polymerase; however, this drug does not alter the kinetics of viremia and NS1 antigenemia in DENV patients . During the early acute phase of DENV infection, oral prednisolone is not related to prolongation of viremia or other pathogenic effects . A recent trial showing that the α-glucosidase inhibitor celgosivir (6-O butanoyl prodrug of castanospermine) has antiviral activity by modulating the host’s unfolded protein response, but it does not significantly reduce viral load or fever burden in DENV patients . The development of a DENV vaccine would represent a powerful new tool for preventing DENV infection. Although a safe vaccine is not yet available, a number of candidate vaccines and strategies for strengthening vaccine efficiency are under active investigation , , .
Both the physiological and the pathogenic roles of IL-10 are immunosupressive. IL-10 not only suppresses inflammation during immune resolution but also affects pathogen clearance and helps alleviate immunopathology . In particular, during microbial infection, IL-10 plays an essential role in relieving IFN-γ- and TNF-α-mediated immunopathology ,  as well as IFNs-mediated antiviral responses. In addition to the canonical extrinsic ADE pathway, which facilitates DENV virus-cell interactions, an intrinsic ADE pathway exists that may play a role during persistent viral infections by inducing IL-10-mediated immune suppression, particularly on IFNs responses , –. ADE of DENV infection requires the Fcγ receptor , but the molecular regulation of Fcγ receptor signaling for IL-10 expression remains unclear; therefore, we chose to further investigate this pathway in monocytes. As summarized in Figure 8, combined with the previous studies which ADE facilitates IL-10 production , –, we speculated that ADE of DENV infection and/or attachment induces not only the Fcγ receptor/Syk-facilitated (i.e., intrinsic) pathway but also the Fcγ receptor/CLEC5A partly/Syk-mediated (i.e., extrinsic) pathway, which both lead to PKA/PI3K/PKB activation, followed by CREB-mediated IL-10 production. In this study, we did not exclude the involvement of CLEC5A/Syk signaling for the intrinsic pathway of IL-10 production under ADE. In addition, we demonstrated that PKB phosphorylates GSK-3β and decreases its activity, enhancing CREB stability and inducing IL-10 production, consistent with previous studies –, , . However, the regulation of PKA by DENV receptors and effectors requires further investigation. Following IL-10 induction, SOCS3 is upregulated and may facilitate attenuation of T cell responses  and induction of IFN resistance to enhance viral replication , –. By interfering nitric oxide generation, which is regulated by STAT1/IRF1 signaling and confers anti-DENV activity, IL-10 may cause immunosuppression through SOCS3 expression , . Based on our findings, IL-10 appears to play a permissive role with respect to DENV pathogenesis, and regulating IL-10 production may therefore provide cellular protection against DENV infection, even under ADE conditions. However, this hypothesis needs to be approved in vivo by using an appropriate animal model.