C57bl 10scnj

C57BL/10ScNJ (TLR4 deficient) and C57BL/10SnJ (WT) mice were purchased from the Jackson Laboratory (Bar Harbor, ME) and housed in a specific pathogen-free environment in conformance with institutional guidelines for animal care. Animal welfare consisted in the inspection a minimum of x3 per week, and any found to be in distress were painlessly euthanized. For experiments, animals were euthanized using CO₂, followed by dissection in a manner incompatible with survival that included severing the great vessels. The experiments were approved by the Institutional Animal Care and Use Committee, Seattle BioMed, WA, USA. All mice were used between 6–9 weeks of age.

Three to four-week-old female C57BL/6J and C57BL/10ScNJ mice were obtained from Jackson Laboratory (Bar Harbor, ME, USA). All mice were housed in sterilized cages and fed autoclaved food and water under specific-pathogen-free, controlled temperature, and controlled photoperiod conditions. Fecal transplants were performed as previously described [15 (link)]. The native microbiota was depleted by treating mice with a single oral dose (20 mg/mouse) of streptomycin 24 h prior to the first fecal transplantation. Fresh fecal pellets from three to four donor mice were collected and placed in 1 mL transfer buffer (pre-reduced sterile phosphate buffered saline containing 0.05% cysteine HCl) on ice. Fecal pellets were homogenized and centrifuged at 800× g for 2 min and the supernatant was collected and diluted (1:3) in transfer buffer. One hundred microliters of diluted fecal supernatant was introduced into recipient mice by oral gavage on six separate occasions, every 48 h.