Porcine circovirus 2 (PCV2) increases the expression of endothelial adhesion/junction molecules

Abstract

Porcine circovirus type 2 (PCV2) is the primary causative agent of porcine circovirus disease, a complex multisystem syndrome in domestic pigs. Despite the significant economic losses caused by porcine circovirus disease, the mechanisms of pathogenesis underlying the clinical findings remain largely unclear. As various reports have highlighted the potential key role of vascular lesions in the pathogenesis of porcine circovirus disease, the aim of this work was to investigate effects of PCV2 infection on vascular endothelial cells, focusing on cell viability and expression of adhesion/junction molecules. PCV2 infection reduced endothelial cell viability, while viral infection did not affected the viability of several other classical cell lines. Also, PCV2 infection in endothelial cells displayed a dual/biphasic effect: initially, infection increased ICAM-1 expression, which can favor leukocyte recruitment and emigration to tissues and possibly inducing characteristic porcine circovirus disease inflammatory lesions; then, secondarily, infection caused an increase in zonula occludens 1 tight junction protein (ZO-1) expression, which in turn can result in difficulties for cell traffic across the endothelium and a potential impairment the immune response in peripheral tissues. These virus-induced endothelial changes could directly impact the inflammatory process of porcine circovirus disease and associated vascular/immune system disturbances. Data suggest that, among the wide range of effects induced by PCV2 on the host, endothelial modulation can be a pivotal process which can help to explain PCV2 pathogenesis in some porcine circovirus disease presentations.

Keywords: Cell adhesion; Infectious diseases; Pathogenesis; Swine; Viral infection.

Fig. 1

Viability of PCV2-infected cells assessed by flow cytometry. EAhy926, PK-15, Vero and HEK293 cell lines and primary cell culture of swine kidney (ST) were infected with 2 log₁₀ virus copies and cell viability assessed 72 h post-infection. Results are expressed as mean ± SEM of twelve biological replicates of uninfected control cells (open bars) and PCV2-infected cells (black bars). Statistical analysis was performed using Student's t-test for unpaired samples, comparing infected and non-infected cells (*p < 0.05).

Fig. 2

ICAM-1 expression in EAhy926 endothelial cells. ICAM-1 (CD54) expression was assessed by flow cytometry after infection of EAhy926 cells with 2 log₁₀ virus copies. Results are expressed as mean ± SEM of nine biological replicates. Statistical analysis was performed using the one-way analysis of variance (ANOVA) followed by Tukey post hoc test. Different letters indicate statistically significant differences (p < 0.05).

Fig. 3

ZO-1 expression in EAhy926 endothelial cells. ZO-1 expression was assessed by flow cytometry after EAhy926 cells infection with 2 log₁₀ virus copies. Panel A, number of cells expressing ZO-1. Panel B, expression of ZO-1 molecules/cell by mean cell fluorescence intensity. Results are expressed as mean ± SEM of nine biological replicates. Statistical analysis was performed using the one-way analysis of variance (ANOVA) followed by Tukey post hoc test. Different letters indicate statistically significant differences (p < 0.05).