The authors used conditioned media from DENV-infected cells that had been UV-irradiated to inactivate virions (CMDV) in order to isolate the effect of secreted viral proteins and host soluble factors on endothelial cells. This in vitro paradigm preserves viral proteins and host mediators while removing productive infection, allowing study of paracrine effects on endothelial phenotype, barrier function and transcriptional programs.
Using a combination of targeted gene expression assays and global transcriptomics, the study compared untreated endothelial cells, CMDV-treated cells, and cells treated with TGF-β as a positive control for Endothelial-to-Mesenchymal Transition (EndMT). Computational analyses including a non-directed asynchronous network model (NDAM-CMDV) were applied to transcriptomic datasets to infer network topology and identify key regulatory nodes.
Exposure to CMDV produced consistent shifts in lineage-associated transcripts. The study reported upregulation of mesenchymal-associated genes, notably SNA1 and CDH2, alongside downregulation of endothelial junction genes OCLN and CDH5. These changes align with loss or weakening of cell–cell junctions and acquisition of migratory features observed in prior CMDV studies.
The pattern is compatible with a partial or progressive EndMT rather than a binary conversion: endothelial markers are repressed while selected mesenchymal transcripts are induced. The authors emphasize that EndMT-like behavior can include multiple intermediate states rather than a full phenotypic switch.
Unbiased transcriptome profiling revealed a biphasic temporal pattern in CMDV-treated endothelial cells. An early, transient pro-inflammatory program was detected shortly after exposure, followed at later times by induction of genes associated with tissue repair and extracellular matrix (ECM) remodeling.
The authors interpret these two phases in the context of dengue clinical kinetics: the inflammatory phase corresponds to the period of high viremia and fever, while the repair/remodeling phase appears later, during defervescence when viremia is declining. This temporal separation provides a molecular rationale for why severe dengue (SD), characterized by vascular leakage and hemorrhage, often appears during defervescence despite falling circulating virus.
Transcriptomic results therefore link soluble DENV-induced factors to both immediate endothelial activation and a delayed program that may weaken barrier integrity through ECM remodeling and junction repression.
To extract potential regulatory drivers from the transcriptomic data the investigators constructed a non-directed asynchronous network model (NDAM-CMDV). This computational approach integrated differentially expressed genes into a network and evaluated node centrality and perturbation effects.
The NDAM-CMDV model highlighted IL6 and FN1 as central nodes within the CMDV response network. Based on network topology and perturbation analysis, the model predicts that these nodes occupy influential positions linking the early inflammatory response to later ECM and remodeling programs that support endothelial trans-differentiation.
The modeling output does not itself prove causality in vivo, but it provides testable hypotheses and prioritizes molecular targets for follow-up functional studies aimed at modulating CMDV-driven endothelial dysfunction.
Findings from molecular profiling and modeling support a scenario in which host soluble factors induced by DENV infection can drive endothelial changes that contribute to vascular leakage and the clinical features of SD. Key implications are:
CMDV recapitulates endothelial junction weakening and partial adoption of mesenchymal features, consistent with EndMT-like processes that increase permeability.
The identified biphasic transcriptional response offers a mechanistic link between patient-stage timing (high viremia and fever versus defervescence) and the emergence of severe vascular phenotypes.
IL6 and FN1 emerge from network analysis as candidate central mediators connecting inflammation to matrix remodeling and endothelial plasticity; these molecules may warrant targeted experimental validation as potential modulators of CMDV-induced dysfunction.
The authors note that the model and data suggest possible therapeutic avenues but do not provide clinical intervention data. Experimental validation in physiologically relevant models and in vivo correlation would be required before translating these network-derived targets into interventions.
The study states that supporting data and code are publicly accessible. Raw microscopy data are available from a figshare repository. RNA-seq sequence data were deposited in the NCBI BioProject (accession PRJNA1331842). The CMDV non-directed network model code is published on GitHub. These resources permit independent reanalysis and further computational experimentation by other groups.
In vitro exposure of endothelial cells to DENV-conditioned media (CMDV) triggers a temporally separable response characterized by an initial inflammatory wave and a later tissue-repair/matrix-remodeling program. CMDV induces repression of junctional endothelial genes (OCLN, CDH5) and induction of mesenchymal markers (SNA1, CDH2), consistent with partial EndMT and increased permeability. Computational NDAM-CMDV modeling nominates IL6 and FN1 as central network nodes linking inflammation and remodeling, offering prioritized hypotheses for experimental validation in the context of severe dengue pathogenesis. The study's datasets and model code are publicly available to support reproducibility and follow-up work.