Impact of controlling abnormal joint movement on the effectiveness of subsequent exercise intervention in mouse model of early knee osteoarthritis

Abstract

Purpose: Moderate mechanical stress is necessary for preserving the cartilage. Recent evidence from animal models of surgically induced OA indicates that the progression of cartilage degeneration can be delayed by moderate exercise. On the other hand, joint instability induces excessive mechanical stress contributes to OA onset and progression. The anterior cruciate ligament transection (ACL-T) model, which is a commonly used animal model of surgically induced OA. In contrast, the controlled abnormal joint movement (CAJM) model, proposed by (Murata et al. 2017) and (Onitsuka et al. 2018) involves ACL-T followed by another surgical procedure to re-stabilize the joint and reduce joint instability. In the CAJM model, the expression of inflammatory factors and cartilage matrix degrading factors is suppressed, joint instability is reduced, and progression to OA is slower. We hypothesized that exercise intervention is more effective at preventing the degeneration of articular cartilage in knee OA if joint instability is controlled. Methods: This study used 28 adults (12-week-old) ICR male mice. The mice were divided into 4 experimental groups: ACL-T without exercise intervention, CAJM without exercise intervention, ACL-T with exercise intervention (ACL-T/Ex), and CAJM with exercise intervention (CAJM/Ex). For mice assigned to CAJM groups, a 25-gauge needle was used to create bone tunnels in the anterior portion of the proximal tibia and in the posterior portion of the femoral condyle in order to decrease the anterior translation of the tibia onto the femur. Subsequently, 4-0 nylon thread was passed through the bone tunnels to compensate for ACL function and help control the anterior translation of the tibia. To assess knee joint instability, the anterior drawer test was performed using a constant force spring (0.05 kgf; Sanko Spring Co., Ltd., Fukuoka, Japan) and a soft X-ray device (M-60; Softex Co., Ltd., Kanagawa, Japan). Mice allocated to the exercise groups (ACL-T/Ex and CAJM/Ex) were exercised on a rodent treadmill, were exercised on the treadmill at a constant speed of 18 m/min for 30 min/d, 3 d/wk, for 4 weeks. Thin sections (7 μm) were cut in the sagittal plane, stained with safranin-O/fast green, and subjected to histology evaluation to estimate the degree of cartilage damage. Two independent observers (T Kano and KO) per- formed OARSI scoring on a scale of 8 stages (0, 0.5, 1-6), and the average value was retained. To evaluate the expression of IL-1βand MMP-13, we performed immuno- histochemical staining using the avidin-biotinylated enzyme complex method. For analysis, we calculated the ratio between the number of IL-1β- or MMP-13-positive cells and the number of chondrocytes in an articular cartilage area of 10000 μm2 (100 μm × 100 μm). Results: Joint instability was quantified using X-ray radiography with the anterior drawer test (Fig. 1A).Compared with the ACL-T and ACL-T/Ex groups, the CAJM and CAJM/Ex groups had significantly lower anterior displacement of the tibia (p<0.05) (Fig. 1B).In the ACL-T/Ex group, marked surface fibrillation and reduction in cartilage thickness were noted in the posterior part of the articular cartilage, together with a significantly higher OARSI score than that noted for the other 3 groups (p<0.001) (Fig. 2). With regard to inflammation, IL-1β-positive cells were confirmed from the cartilage surface layer to the deep layer in the ACL-T, CAJM, and ACL-T/Ex groups but not in the CAJM/Ex group (Fig. 3A). The percentage of IL-1β- positive cells in the anterior part of the cartilage was significantly higher in the ACL-T and ACL-T/Ex groups than in the CAJM/Ex group, the percentage of IL-1β-positive cells in the posterior part of the cartilage was significantly higher for the ACL-T and ACL-T/Ex groups than for the CAJM/Ex group (p<0.05) (Fig. 3B). Similarly, MMP-13-positive cells were confirmed in the ACL-T and ACL-T/Ex groups, compared with very weak MMP-13 expression in the CAJM and CAJM/Ex groups (Fig. 3A). However, while no significant difference among the four groups was noted in terms of the percentage of MMP-13-positive cells in the anterior part of the cartilage, the percentage of MMP-13-positive cells in the posterior part of the cartilage was significantly higher for the ACL-T/Ex group than for the CAJM/Ex group (p=0.02) (Fig. 3C). Conclusions: Previous studies reported that exercise alone could delay the progress of OA. However, our present Results indicate that the internal environment of the joint strongly modulates the effect of exercise, sometimes completely canceling the benefit. In other words, when prescribing exercise, we need to consider the internal environment of the knee joint.In the clinical setting, these findings may trans- late in the need to consider an intervention to correct abnormal joint movement before prescribing physical exercise in the treatment of OA.View Large Image Figure ViewerDownload Hi-res image Download (PPT)View Large Image Figure ViewerDownload Hi-res image Download (PPT)