Blockade of dengue virus infection and viral cytotoxicity in neuronal cells in vitro and in vivo by targeting endocytic pathways

Abstract

Dengue virus (DENV) infection in neuronal cells was speculated to trigger neuropathy. Herein, we determined the blockade of DENV infection by targeting endocytic pathways in vitro and in vivo. In DENV-infected mouse brains, we previously showed that viral proteins are expressed in neuronal cells around the hippocampus with accompanying neurotoxicity. DENV caused infection, including entry, double-stranded (ds)RNA replication, protein expression, and virus release, followed by cytotoxicity in the mouse neuronal Neuro-2a cell line. Pharmacologically blocking clathrin-mediated endocytosis of the DENV retarded viral replication. Targeting vacuolar-type H+-ATPase (V-ATPase)-based endosomal acidification effectively blocked the DENV replication process, but had no direct effect on viral translation. Blockade of the clathrin- and V-ATPase-based endocytic pathways also attenuated DENV-induced neurotoxicity. Inhibiting endosomal acidification effectively retarded DENV infection, acute viral encephalitis, and mortality. These results demonstrate that clathrin mediated endocytosis of DENV followed by endosomal acidification-dependent viral replication in neuronal cells, which can lead to neurotoxicity.

Anti-TNF-α restricts dengue virus-induced neuropathy

“…According to our findings, DENV infection induces TNF‐α production in brain and TNF‐α is highly correlated with encephalitic progression and the induction of neuronal cell apoptosis. This study, along with our previous works, show that DENV caused neurotoxicity in vivo and in vitro as shown by histopathologic changes and apoptotic staining. We further demonstrated an enhanced effect of TNF‐α on DENV‐induced neurotoxicity in vitro.…”

“…Our previous work showed that DENV infection directly causes neurotoxicity in vitro in neuronal cells . Regarding the proapoptotic role of TNF‐α, we next assessed the neurotoxic effects of TNF‐α under DENV infection in murine neuronal cell line Neuro‐2a.…”

“…Baby hamster kidney (BHK)‐21 cells (CCL10; ATCC) and murine Neuro‐2a cells (CCL131; ATCC), cultured in DMEM (Invitrogen Life Technologies, Rockville, MD), and Aedes albopictus C6/36 cells (CRL1660; ATCC), grown on plastic in RPMI (Invitrogen Life Technologies), were carried out for DENV infection accordingly . DENV2 PL046, a Taiwanese human isolate obtained from the Centers for Disease Control in Taiwan, was propagated in C6/36 cells and viral titers were quantified by plaque assay using the BHK‐21 cells.…”

“…The presence of viral supernatants was evaluated using plaque assays. Protocols for plaque assay were performed according to our previous studies …”

Research advancements in the neurological presentation of flaviviruses

“…55 Apart from the above receptors, JEV can invade human neuronal cells through a caveolin-1-dependent endocytosis pathway ( Figure 2), 56 whereas DENV entry may be mediated by clathrin ( Figure 3). 57 This suggests that flaviviruses have multiple routes to cell infection, rather than a single receptor that allows cell entry. Subsequent to entry, viral acidification-dependent viral replication within nerve cells leads to neurotoxicity.…”