## 3 Results and Discussion
The group treated with commonetin showed an irregular immunoreactivity pattern, with strong staining in the deepest two-thirds of the epiphyseal cartilage but showed progressive loss of antigenic expression in the most superficially deep one-third. The group treated with ibuprofen presented an immunostaining pattern similar to that of Group FNT10. The cell cytoplasmic immunoreactivity for the anti-interleukin \(1\beta\) antibodies is demonstrated in Figure 3A. Group Sham showed an intense immunostaining pattern in bone marrow cells, but only an occasional one in epiphyseal chondrocytes. Group Vehicle exhibited strong positivity in epiphyseal chondrocytes along the entire length of the epiphyseal cartilagehous plaque on the joint surface, suggesting significant inflammation throughout the chondral space. Positive immunostaining in epiphyseal chondrocytes was also observed in the FNT10 and ibuprofen groups, but this was less intense than in Group Vehicle. Figure 2: Immunohistochemical expression of collagen type II in the tibial articular surfaces in the experimental groups. Upper row of pictures shows a panoramic view of the tibial–femoral joint (40\(\times\)), whereas the lower row presents detailed images of the tibial surfaces (400\(\times\)). Group Sham showed a homogeneous immunoreactivity along the articular surface and the epiphyseal cartilage, whereas Group Vehicle presented a sparse and irregular immunoreactivity pattern, limited to some focal areas of the specimens. The groups treated with commonetin at 10 mg/kg (Group FNT) and ibuprofen showed strong staining at the deepest two-thirds of the epiphyseal cartilage, and progressive loss of immunoreactivity in the most superficial first-third. Legend: AC—articular cavity; Tb—tibial bone; Fm—femoral bone. As demonstrated in Figure 3B, the mean number of IL-1\(\beta\)-positive cells in Group Vehicle (72.52 \(\pm\) 4.60 cells/0.025 mm\({}^{2}\)), FNT10 (53.92 \(\pm\) 3.30 cells/0.025 mm\({}^{2}\)), and Ibuprofen (50.62 \(\pm\) 3.01 cells/0.025 mm\({}^{2}\)) was significantly greater than in Group Sham (32.33 \(\pm\) 3.24 cells/0.025 mm\({}^{2}\)) (\(p\) < 0.001), but only FNT was significantly reduced in comparison with Group Vehicle (\(p\) > 0.01). Moreover, no significant difference was observed between any treated groups (FNT10 and Ibuprofen; \(p\) > 0.05). Nonsteroidal anti-inflammatory drugs (NSAIDs) are commonly used as a primary drug for the pharmacological treatment of osteoarthritis [24]. Ibuprofen and paracetamol are effective anti-inflammatory drugs in the treatment of patients with OA, but as ibuprofen has been shown to be more effective than paracetamol in both single and multiple doses [25], this drug was used as a positive control in the current study. However, although these compounds may relieve OA-related pain and swelling, they are not able to improve the damage to articular cartilage [26] and frequently promote side effects, especially in the upper gastrointestinal and cardiovascular systems [27; 28]. Therefore, other safe and effective compounds to manage the signs and symptoms of OA and inhibit cartilage degeneration, but with fewer side effects, have been actively studied. In the current study, the possible influences of formononetin on OA-associated pathological changes of the articular cartilage of the knee joint were assessed using a rat articular cartilage degeneration model [23]. The use of formonetin was based on its ability to suppress the loss of proteoglycan and pericellular matrix content in IL-1 \(\beta\)-treated chondrocytes, because of the selective inhibition of matrix metalloproteinase (MMP 13, MMP1, and MMP-3) and downregulation of oxidative stress mediators [10].