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The relevance of collagen degradation and of articular instability as features of OA, as observed in the clinical setting, leading to histopathological alterations that highly correlate with those described for human OA [], make the i.a.

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The signal intensity depended on the paw area in contact with the platform and increased with the pressure applied by the paw.

Random frames of the videos were analysed: three pairs of frames (one for each hind paw) with the animal walking and three frames with the animal standing still.

A model that comprises both the reproduction of the initiating events and joint tissue pathology observed in human OA as well as the induction of relevant nociceptive responses that mimic patients’ main complaints, such as increased nociception due to movement and loading on the affected joint, would certainly be a clinically-relevant model for the study of OA pain.

The intra-articular (i.a.) injection of collagenase is an established model of OA that has been predominantly used to study the mechanisms underlying structural joint damage [].

We also tested the efficacy of morphine in reversing behavioural changes with the goal of validating them as being nociception-related, as well as the efficacy of a local anaesthetic intra-articularly injected in the knee, in order to demonstrate that the nociceptive behaviours observed are indeed originated in the knee joint.

Furthermore, since there seemed to be an inflammatory reaction occurring after the injection of collagenase, we also tested the effect of the non-steroidal anti-inflammatory drug (NSAID) diclofenac in reversing the nociceptive changes observed.

One week after injection of 500 U collagenase, swelling of the injected knee and inflammation of the synovial membrane were also observed, indicating the occurrence of an early inflammatory reaction.

Behavioural changes induced by the 500 U dose of collagenase were overall effectively reversed by morphine and lidocaine. TRPV1 expression increased six weeks after 500 U collagenase injection.

For each hind paw, the number and intensity of pixels above a defined threshold were quantified (Image J 1.37, []) to determine the total paw print intensity (mean pixel intensity × number of pixels), allowing the comparison of the area/pressure applied by each paw.

Results were expressed as the percentage of the total ipsilateral paw print intensity (%TIPPI) in the total intensity of both paw prints.

Experimental procedures were performed in accordance with the ethical guidelines for the study of experimental pain in conscious animals [ Under brief isoflurane anaesthesia (5% isoflurane for anaesthesia induction, 2% for maintenance), an intra-articular injection was performed with the use of a Hamilton syringe, with a 26 G needle inserted through the patellar ligament into the joint space of the left knee.

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