Cd 1 swiss mice

Mice were housed in animal facilities at Newcastle University, and all procedures were licensed (PPL 70/7960 and PPL PDD4CCF4F) by the UK Home Office under the Animal (Scientific Procedures) Act 1986. GV‐stage oocytes were harvested from ovaries of 8–12‐week‐old CD‐1 Swiss mice (Charles River) in M2 medium (Sigma) containing IBMX (200 µM) and then cultured in IBMX‐free G‐IVF PLUS medium (Vitrolife) for 16 h at 37°C in 6% CO₂, 5% O₂. Oocytes that produced a polar body were placed in activation medium (KSOM [Millipore] plus 5 mM SrCl₂ and 2 mM EGTA) for 1 h. Nocodazole (1 µM), reversine (1 µM), MG‐132 (10 µM), and ICRF‐193 (20 µM) were used at the indicated concentrations.

All animal experiments were performed according to the guidelines for the use and care of laboratory animals and they were authorized by the local Animal Care Committee “Organismo Preposto al Benessere degli Animali” and by Italian Ministry of Health, “Ministero della Salute” (DL 26/2014). Animal studies are reported in compliance with the ARRIVE guidelines (Kilkenny et al., 2010 (link); Curtis et al., 2015 (link)). All appropriate measures were taken to minimize pain or discomfort of animals. Female CD1 Swiss mice (7–12 weeks old) (Charles River Laboratories, Calco, Italy), weighing 25–30 g, were housed five per cage and maintained under standard conditions at our animal facility (12:12 h light–dark cycle, water and food ad libitum, 22–24°C). Intragastric gavage administration was performed using suitable gavage needles (22 gauge, length 3 cm, ball diameter 1.5 mm). All the experimental procedures (induction of colitis, SPX) and euthanasia by CO₂ inhalation were performed between 9 a.m. and 12 a.m.

Virulence of the M1T1-WT, M1T1ΔTyK mutant, and the tyk-complemented GAS strains was determined in the mouse peritonitis model (10–12 mice/group). CD-1 Swiss mice (5 weeks, 20–22 g Charles River Laboratories) were injected intraperitoneally with 5 × 10⁸ CFU of individual GAS strains. Morbidity and mortality in the infected animals were monitored twice daily for 10 days. All animal experiments were performed per the recommendations of the National Institute of Health Guide for the Care and Use of Laboratory Animals. The animal experiment protocol (2008A0222R4) was approved by the Ohio State University Institutional Animal Care and Use Committee (OSU-IACUC). These experiments were once repeated for reproducibility. Percentage survival in each group was combined and statistically evaluated by the Log Rank test (Kaplan–Meier) using GraphPad Prism 6 software. A value of p < 0.05 was treated as a significant difference.