From the tiny mouse knee sections, it was not possible to restrict the analysis only to articular cartilage.
Immunohistochemistry analysis of mouse knee sections of surgically induced OA and sham control (with Osteoarthritis Research Society International scores of 5 and 0, respectively) revealed that phosphorylated LKB1 and AMPKα were concomitantly decreased in mouse OA knee cartilage, compared with the sham control.
Mouse knee sections were incubated with rabbit anti-mouse -GCP2 (Biorbyt), -CXCR2 (R&D), -Col X (Abcam) or anti-Col II (Chemicon), followed by chicken anti-rabbit Alexa Fluor 488 or chicken anti-rabbit Alexa Fluor 594 secondary antibodies (Life Technologies).
Human cartilage explant and decalcified mouse knee joint sections were deparaffinised and the subsequent steps including blocking and pepsin antigen retrieval were performed as previously described.
Mouse knee cartilage sections were pretreated with hyaluronidase (2 mg/ml) for 1 hour before being treated with 3% (vol/vol) H2O2 for 10 minutes.
Mouse knee-joint sections were stained with both Safranin O-fast green and Alcian blue-hematoxylin to allow visualization of cartilage tissue.
Histologic evaluation was performed on sagittal sections of mouse knee joint.
Sections from archival paraffin blocks of male C57BL6 mouse knee joints with either AIA (7 and 28 days after induction) or saline injection from a previous study [ 16] were prepared at the same time as serial sections from the mouse knee joints with surgically-induced OA.
Images of non-injured mouse knee (left knee, B) and 2 months after surgical induction of osteoarthritis (DMM model; right knee, A) were acquired ex vivo at 80 kVp, 500 μA and with a pixel size of 35 μm.
Freshly dissected mouse knee joints were fixed overnight in TissuFix (Chaptec, Montreal, Quebec, Canada), decalcified for 1.5 h in RDO Rapid Decalcifier (Apex Engineering, Plainfield, Illinois, USA), further fixed in TissuFix overnight, followed by embedding in paraffin and sectioning.
