Polar Infection of Echovirus-30 Causes Differential Barrier Affection and Gene Regulation at the Blood–Cerebrospinal Fluid Barrier

Abstract

Echovirus-30 (E-30) is responsible for the extensive global outbreaks of meningitis in children. To gain access to the central nervous system, E-30 first has to cross the epithelial blood–cerebrospinal fluid barrier. Several meningitis causing bacteria preferentially infect human choroid plexus papilloma (HIBCPP) cells in a polar fashion from the basolateral cell side. Here, we investigated the polar infection of HIBCPP cells with E-30. Both apical and basolateral infections caused a significant decrease in the transepithelial electrical resistance of HIBCPP cells. However, to reach the same impact on the barrier properties, the multiplicity of infection of the apical side had to be higher than that of the basolateral infection. Furthermore, the number of infected cells at respective time-points after basolateral infection was significantly higher compared to apical infection. Cytotoxic effects of E-30 on HIBCPP cells during basolateral infection were observed following prolonged infection and appeared more drastically compared to the apical infection. Gene expression profiles determined by massive analysis of cDNA ends revealed distinct regulation of specific genes depending on the side of HIBCPP cells’ infection. Altogether, our data highlights the polar effects of E-30 infection in a human in vitro model of the blood–cerebrospinal fluid barrier leading to central nervous system inflammation.

Echovirus-30 Infection Alters Host Proteins in Lipid Rafts at the Cerebrospinal Fluid Barrier In Vitro

“…Our previous results suggested that the optimal infection rates of HIBCPP cells with E-30 were obtained when the cells were challenged apically with a MOI (multiplicity of infection) of 20 [ 28 ]. Now, we further analyzed several parameters, such as the transepithelial resistance, the cytotoxic effect of E-30, the number of viral particles per HIBCPP cells, and the percentage of infected HIBCPP cells ( Supplemental Figure S1 ).…”

“…A previous study of our laboratory on E-30 infection of the HIBCPP with MOI 20 revealed a strong impact of the virus at the RNA level [ 28 ]. We compared the differentially expressed genes after E-30 infection in HIBCPPP with the new proteomic results.…”

“…Therefore, to investigate the effect of endocytosis blocking on HIBCPP barrier function, we used the inverted model system to infect HIBCPP cells from the basolateral side [ 8 , 9 ]. As previously published, to induce the same percentage of infected HIBCPP cells with a MOI 20 on the apical side, we used a MOI 0.7 to infect the basolateral side of the HIBCPP cells [ 28 ]. We evaluated the effect on barrier disruption with different blocking drugs, such as Pitstop-2, chlorpromazine, and Dynasore ( Figure 2 ).…”

“…Further, we determined the consequences of inhibiting clathrin-mediated endocytosis for the impact on viability and barrier function of HIBCPP cells in context of viral infection. An MOI of 20 for 24 h was chosen for infection with E-30 on the basis of previous published studies and was well suited for a high infection without inducing cell death to perform the proteomic assay on infected HIBCPP cells [ 28 ]. The infection led to a decrease in barrier properties, a high number of infected cells, and a low cytotoxicity rate.…”

“…Furthermore, a study focused on E-30 strain Bastianni revealed that basolateral infection of the HIBCPP cells has a disruptive effect on tight junction proteins such as ZO-1, but also on adherens junction proteins such as E-cadherin [ 27 ]. In addition, another recent study described the importance of HIBCPP cells polarity for E-30 infection and its impact on the cells at an RNA level [ 28 ].…”