Dissecting the Roles of Immune Receptors in MCMV Infection

All in vivo experiments were performed according to UK Home Office guidelines and were performed at Cardiff University (Reference: P7867DADD) in specific pathogen-free/SPF conditions at ambient temperature and humidity. Nogo-A/B^−/− mice^{73 (link)}, and IFITM3-deficient (Ifitm3^−/−) and wt control mice have been described previously^{1 (link)} and were crossed with Myd88^−/− (Jackson Laboratory) or for some experiments Nogo-A/B^−/− mice to generate Ifitm3^−/−/Myd88^−/− and Ifitm3^−/−/Nogo-A/B^−/− mice. Novel strains are available upon request. Age- and sex-matched mice between 7 and 12 weeks of age were used in the experiments. Tlr3^{−/−74 (link)}, Tlr7^{−/−75 (link)} and Tlr9^{−/−76 (link)} mice were a kind gift from Caetano Reis e Sousa (Crick Institute, London). Relevant wild type control mice were all bred inhouse. MCMV (pSM3fr-MCK-2fl BACmid) was grown and titred using 3T3 cells (ATCC, CRL-1658) with a carboxycellulose overlay. Mice were infected via intraperitoneal (i.p.) injection with between 5 × 10⁵ to 2 × 10⁶ PFU MCMV. For Anakinra treatment, mice were injected i.p. with Anakinra (KINERET: Cardiff & Vale NHS Pharmacy) (25 mg/kg) or PBS control on day 0 p.i. For infectious virus quantification form harvested tissue, viral load was determined via plaque assay as previously described^{77 (link)}. All mice were euthanized using carbon dioxide.