Fast type I interferon response protects astrocytes from flavivirus infection and virus-induced cytopathic effects

Lindqvist, Richard; Mundt, Filip; Gilthorpe, Jonathan D.; Wölfel, Silke; Gekara, Nelson O.; Kröger, Andrea; Överby, Anna K.

Supernatants from virus-infected astrocytes were inactivated using β-propiolactone (β-PL) (Acros Organics, 269040050), diluted in water to 0.96 % and used at a final concentration of 0.05 %, incubated at 4°C for 16 h followed by 2 h incubation at 37°C for hydrolysis, performed in plates (VWR, 734-2325) to avoid acidification of the samples [47, 48] . The cells (MEFs, astrocytes, or neurons) were seeded in 96-well plates (Greiner CELLSTAR®) and treated with β-PL-inactivated supernatants or serially diluted IFNαB/D [49] (a kind gift of Peter Stäheli, Virology, University of Freiburg, Freiburg, Germany). The cells were infected 24 h post treatment with VSV-eGFP (multiplicity of infection (MOI) 0.01) (MEFs) or TBEV (astrocytes or neurons) and fixed with 4 % formaldehyde at 16 h post infection (hpi). The number of infected cells was determined using a TROPHOS Plate RUNNER HD®, and the IFN levels were quantified using a standard curve (calculated from 1, 5, 10, 50, 100, and 500 U of IFNαB/D).