Arthritis was induced in DBA/1 mice, as described previously (5557). In brief, bovine type II collagen was purified from articular cartilage and dissolved in 0.1 M acetic acid. Male DBA/1 mice (Harlan UK) received subcutaneous 100 μl injections of 200 μg of bovine type II collagen in complete Freund’s adjuvant (BD Biosciences) at the base of the tail and on the flank. After immunization, the mice were monitored daily for arthritis. Once an animal showed signs of arthritis, it was randomly assigned to a treatment group and monitored daily. Animals received GT14829 (10 mg/kg per day) or GT13072 (1, 5, or 10 mg/kg per day) intraperitoneally. Etanercept-treated animals received four doses of 100 μl (10 mg/kg) of etanercept (Pfizer) intraperitoneally on days 1, 3, 6, and 8. The experimental design is depicted in Figs. 1B and 5A. All mice with CIA were treated until day 10 of arthritis. On day 10, some animals were culled, whereas others in remission were maintained until day 18 before being culled. In both cases, peripheral blood, lymph nodes, and paws were taken. Arthritis severity was scored by an experienced nonblinded investigator as follows: 0 = normal, 1 = slight swelling and/or erythema, 2 = pronounced swelling, and 3 = ankylosis. All four limbs were scored, giving a maximum possible score of 12 per animal.

Hind paws of CIA animals on day 10 were fixed in paraformaldehyde and decalcified with EDTA. After being embedded in paraffin, the paws were sectioned and stained with hematoxylin and eosin. Each joint was scored by a researcher blinded to the study as follows: 0 = normal, 1 = cell infiltration with no signs of joint erosion, 2 = inflammation with the presence or erosions limited to discrete foci, and 3 = severe and extensive joint erosion with loss of architecture. Three hind paw joints (proximal interphalangeal, metatarsal phalangeal, and tarsal metatarsal) were scored and a mean was calculated, giving a maximum possible score of three per animal. All procedures were approved by the Clinical Medicine Ethical Review Board and the UK Home Office.

Note: The content above has been extracted from a research article, so it may not display correctly.

Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.

We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.