Acute Synovitis after Trauma Precedes and is Associated with Osteoarthritis Onset and Progression

Abstract

Osteoarthritis (OA) is a whole-joint disease characterized by cartilage destruction, subchondral bone sclerosis, osteophyte formation, and synovitis. However, it remains unclear which part of the joint undergoes initial pathological changes that drives OA onset and progression. In the present study, we investigated the longitudinal alterations of the entire knee joint using a surgically-induced OA mouse model. Histology analysis showed that synovitis occurred as early as 1 week after destabilization of the medial meniscus (DMM), which preceded the events of cartilage degradation, subchondral sclerosis and osteophyte formation. Importantly, key pro-inflammatory cytokines such as IL-1β, IL-6, TNFα, and Ccl2, major matrix degrading enzymes including Adamts4, Mmp3 and Mmp13, as well as nerve growth factor (NGF), all increased significantly in both synovium and articular cartilage. It is notable that the inductions of these factors in synovium are far more extensive than those in articular cartilage. Results from behavioral tests demonstrated that sensitization of knee joint pain developed after 8 weeks, later than histological and molecular changes. In addition, the nanoindentation modulus of the medial tibiae decreased 4 weeks after DMM surgery, simultaneous with histological OA signs, which is also later than appearance of synovitis. Collectively, our data suggested that synovitis precedes and is associated with OA, and thus synovium may be an important target to intervene in OA treatment.

Keywords: cartilage; osteoarthritis; subchondral bone; synovitis.

Figure 1

Synovitis precedes cartilage degradation and angiopoiesis during the pathogenesis of surgically-induced OA. Knee joints were harvested at different time points of 1, 2, 4, 8, or 12 weeks after surgery as indicated. Hematoxylin-eosin staining results showed synovitis (yellow arrow) in the DMM surgery group one week after surgery, and cartilage degradation (black arrow) and angiopoiesis (black arrowhead) 4 weeks after DMM surgery. Scale bar, 50 μm. n = 5.

Figure 2

Longitudinal histologic analysis of cartilage degradation and subchondral bone sclerosis induced by DMM surgery. (A) Alcian blue/Hematoxylin and Orange G staining of the coronal sections of the knee joints collected at different time points as indicated. Black arrow, cartilage degradation. Green arrow, subchondral bone sclerosis. Scale bar, 50 μm. n = 5. (B) Alcian blue/Hematoxylin Orange G staining of the coronal sections of the joints collected 4 weeks after surgery. Black arrow, cartilage degradation. White arrow, subchondral bone sclerosis in the medial tibiae. Red arrow, subchondral bone loss in the lateral tibiae. L, lateral tibiae; M, medial tibiae. Scale bar, 200 μm. n = 5.

Figure 3

Longitudinal μCT analysis of osteophyte formation in the pathogenesis of surgically-induced OA. Osteophytes (yellow arrow) outgrowth and joint space decrease (white arrow) were evident 8 weeks after DMM surgery, and the phenotype aggravated 12 weeks after DMM surgery. L, lateral; M, medial. Scale bar, 1 mm. n = 7.

Figure 4

Inductions of MMP13 and ColX in medial articular cartilage during OA progression. Immunohistochemistry of MMP13 (A) and ColX (B) of the knee joint sections 4 weeks after DMM surgery. Red arrow, positive IHC signals. It is notable that, in both lateral and medial tibiae of the sham group as well as in the lateral tibiae of the DMM group, MMP13 expressions were weak in articular cartilage and was restricted mainly in deep zone and adjacent to subchondral bone. However, MMP13 expressions increased in medial tibia cartilage and expanded into the middle and superficial zones in the DMM group. Scale bar, 50 μm. *** p < 0.001. **** p < 0.0001. One-way ANOVA followed by Tukey's multiple comparison post-test n = 5.

Figure 5

Long-term changes of nanoindentation modulus of tibia cartilage during OA pathogenesis. Knee joints were harvested at different time points as indicated for nanoindentation tests. Both medial tibiae (A) and lateral tibiae (B) were measured. * p < 0.05. ** p < 0.01. One-way ANOVA followed by Bonferroni's multiple comparison post-test. n = 4.

Figure 6

Longitudinal behavior tests of pain sensitivity induced by DMM surgery. (A) Hot plate testing results showed that DMM surgery decreased latency since 8 weeks after surgery, suggesting development of thermal hyperalgesia. (B and C) Spontaneous rearing activity, displayed as vertical photobeam crossings (B), and ambulation, measured as horizontal photobeam crossings (C), were reduced since 8 weeks after DMM surgery. * p < 0.05. ** p < 0.01. One-way ANOVA followed Bonferroni's multiple comparison post-test. Values are mean ± SD. n = 7.

Figure 7

Early induction of OA-associated factors in synovium. The mRNA expression levels of inflammatory cytokines (IL-1β, IL-6, TNFα and Ccl2), NGF, Runx2, and extracellular matrix degrading enzymes (Adamts4, Adamts5, Adamts7, Adamts12, Mmp13, Mmp3, and Mmp9) in synovium and articular cartilage were measured 3 days after DMM surgery. * p < 0.05. ** p < 0.01. Unpaired Student's t-test. Values are mean ± SD. n = 3.