Abstract
Infections with Herpes simplex virus can cause severe ocular diseases and encephalitis. The present study aimed
to investigate potential inhibitors of fusion between HSV-1 and the cellular membrane
of the host cell. Fusion and entry of HSV-1 into the host cell is mimicked by a virus-free
eukaryotic cell culture system by co-expression of the HSV-1 glycoproteins gD, gH,
gL, and gB in presence of a gD receptor, resulting in excessive membrane fusion and
polykaryocyte formation. A microscopic read-out was used for the screening of potential
inhibitors, whereas luminometric quantification of cell-cell fusion was used in a
reporter fusion assay. HSV-1 gB was tagged at its C-terminus with mCherry to express
mCherry-gB in both assay systems for the visualization of the polykaryocyte formation.
Reporter protein expression of SEAP was regulated by a Tet-On 3 G system. The saponin
mixture aescin was identified as the specific inhibitor (IC50 7.4 µM, CC50
24.3 µM, SI 3.3) of membrane fusion. A plaque reduction assay on Vero cells reduced
HSV-1 entry into cells and HSV-1 cell-to-cell spread significantly; 15 µM aescin decreased
relative plaque counts to 41%, and 10 µM aescin resulted in a residual plaque size
of 11% (HSV-1 17 syn+) and 2% (HSV-1 ANG path). Release of the HSV-1 progeny virus was reduced by one log
step in the presence of 15 µM aescin. Virus particle integrity was mainly unaffected.
Analytical investigation of aescin by UHPLC-MS revealed aescin IA and -IB and isoaescin
IA and -IB as the main compounds with different functional activities. Aescin IA had
the lowest IC50, the highest CC50, and an SI of > 4.6.
Keywords
Aescin -
Aesculus hippocastanum
- Sapindaceae -
Herpes simplex virus - HSV-1 - membrane fusion - plaque reduction - triterpene saponins
