Senescence affects endothelial cells susceptibility to dengue virus infection

Abstract

Alteration in the endothelium leading to increased vascular permeability contributes to plasma leakage seen in dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). An earlier study showed that senescent endothelial cells (ECs) altered the ECs permeability. Here we investigated the susceptibility of senescing human umbilical vein endothelial cells (HUVECs) to dengue virus infection and determined if dengue virus infection induces HUVECs senescence. Our results suggest that DENV type-2 (DENV-2) foci forming unit (FFU) and extracellular virus RNA copy number were reduced by at least 35% and 85% in infection of the intermediate young and early senescent HUVECs, respectively, in comparison to infection of young HUVECs. No to low infectivity was recovered from infection of late senescent HUVECs. DENV infection also increases the percentage of HUVECs expressing senescence-associated (SA)-β-gal, cells arrested at the G2/M phase or 4N DNA content stage and cells with enlarged morphology, indicative of senescing cells. Alteration of HUVECs morphology was recorded using impedance-based real-time cell analysis system following DENV-2 infection. These results suggest that senescing HUVECs do not support DENV infection and DENV infection induces HUVECs senescence. The finding highlights the possible role of induction of senescence in DENV infection of the endothelial cells.

Keywords: Dengue virus, Endothelium permeability.; HUVECs, Endothelial cells; Senescence.

Figure 1

SA-β-gal staining and cell cycle population distribution of mock- and DENV-2-infected HUVECs at each passage number. (A) SA-β-gal positive cells which stained as blue-turquoise precipitate in cells. (B) DNA content in 15, 000 cells at the different phase during doublings. The relative percentages of cells in G0/G1, S and G2/M population of the cell cycle were estimated from the propidium iodide (PI) histograms. (C) The percentage of the SA-β-gal positive cells. The data shown are mean ± S.D. from two independent experiments with '*', p < 0.001 represent significance difference.

Figure 2

Dynamic assessment of HUVECs proliferation and morphological changes of both mock- and DENV-2-infected at each passage number. Growth kinetics of (A) young, (B) intermediate young and (C) early senescent HUVECs which captures proliferation, morphological changes and induction of apoptosis events was measured. The data for cell index was expressed as mean ± S.D. from two independent experiments.