Objectives:
Anterior cruciate ligament (ACL) tears are common knee injuries that initiate biologic cascades that may not be reversed with ACL reconstruction (ACLR). These cascades culminate in post-traumatic osteoarthritis (PTOA) in a relatively high proportion of patients despite successful ACLR. The synovium is often overlooked in ACL injuries, but it is a major regulator of the intraarticular environment and its responses to injuries. Further characterization of patient-specific synovial responses can help in developing strategies to regulate the post-injury joint environment and alter the natural history of the ACL-injured knee. In this study, we aim to characterize the histological changes, MRI appearance, and bioactivity of the human synovium after ACL injuries.
Methods:
Patient Cohort:
Patients with ACL tears were prospectively enrolled (IRB #2009879). Patient demographics, intraoperative findings, and the time from injury to MRI and to ACLR/synovial biopsy (days) were recorded. A total of 31 patients (male=18; female=13; mean age: 25.8±12 y/o) and 61 patients (male=30; female=31; mean age 23.7 y/o) were included in the histologic/MRI and bioactivity analyses, respectively.
Histologic and MRI Characteristic of the Synovial Response to ACL Injury:
Synovial biopsies were obtained during the primary ACLR. Histopathological changes were scored using the Krenn system that evaluated synovial hyperplasia, density of the resident cells, and inflammatory infiltrates (maximum of 9 points; 0-1=no synovitis; 2-4=low-grade synovitis; ≥ 5= high-grade synovitis). A semiquantitative scoring system (MRI OsteoArthritis Knee Score or MOAKS) was used to determine Hoffa’s-synovitis (0-3) and effusion-synovitis (0-3) with a higher score indicative of greater synovitis. A total MOAKS score was calculated by combining the Hoffa-synovitis and effusion-synovitis scores.
Bioactivity of Synovial Tissues after ACL Injury:
Two-4mm explants were created from the synovium and cultured. After 3 days, the media were assessed for inflammation (IL-6, IL-8, MCP-1, MCP-3, GRO-α, MIP-1α, MIP-1β, RANTES, IL-1β, INF-γ, TNF-α, FGF-2, VEGF, PDGF-AA, PGE2) and degradative biomarker levels (MMP-1, MMP-2, MMP-3, MMP-7, MMP-8, MMP-9, MMP-13, total MMP activity, TIMP-1, TIMP-2, TIMP-3, TIMP-4).
Statistics:
The cohorts were divided based on age (<20 y/o or ≥20 y/o), sex, time from injury to ACLR or MRI (<60 days, 60-120 days, > 120 days), and presence of concurrent injuries (meniscus injuries (ACL+MEN), cartilage lesions (ACL+CART), or meniscus and cartilage lesions combined (ACL+MEN+CART)). Two-sample t-Tests were used for age, sex, and time from injury to MRI and MOAKS scores comparisons. A one-way ANOVA with Tukey post-hoc analysis was performed for time from injury to ACLR and concurrent injury analyses. Statistical significance was set at p<0.05.
Results:
Histological Scoring:
The mean Krenn synovitis score was 3.42 ± 0.96. All scores, regardless of time from injury to ACLR, were within the range of low-grade synovitis (Fig. 1). There were no statistical differences in Krenn scores based on time from injury to ACLR (p=0.877). Females had higher scores than males (p=0.03) (Fig. 2). Age (p=0.56) and presence of meniscus (p=0.06) or cartilage lesions (p=0.524) did not result in differences in synovitis severity.
MRI Assessment:
MRI demonstrated a higher total MOAKS scores for the <60-day vs. >120-day group (p<.001) (Fig. 3). Both Hoffa’s-synovitis (p=0.036) and effusion-synovitis (p=<0.001) scores were also higher for the <60-day vs. >120-day group. There were no statistical differences between Total MOAKS score with respect to age (p=0.935), sex (p=0.734), or presence of meniscus (p=0.187) or chondral lesions (p=0.415).
Bioactivity Analyses:
Synovial tissues from patient >20 y/o released higher levels of inflammation (MCP-3, PDGF-AA, MCP-1) and degradative biomarkers (MMP-2, MMP-3, MMP-7, and MMP-13) compared to <20 y/o (p<0.05). Female patients also produced higher levels of various inflammation and degradative proteins compared to males (GRO-α, IL-6, MMP-1, MMP-2, MMP-9, TIMP-3 and TIMP-4) (P<0.05) (Fig. 4). There was also a time dependency on inflammatory and degradative biomarkers. Synovial tissues collected <60-day from injury produced higher levels of MIP-1α, MIP-1β, RANTES, and MMP-8 compared to the >120-day group (P<0.05). When concurrent intraarticular injuries were considered, tissues from ACL+CART injury had lower various inflammatory and degradative biomarkers when compared to ACL-only, ACL+MEN, and ACL+MEN+CART groups (P<0.05).
Conclusions:
The present study suggests that ACL tears result in early low-grade synovitis that persists for months after injury. The MRI data indicates, however, that the level of Hoffa’s synovitis and effusion may subside with time. The MRI data correspond with bioactivity analyses documenting diminishing inflammatory and degradative biomarker production amongst synovial tissues collected from longer duration ACL tears. Patient demographics and injury characteristics may also impact the synovial response in ACL injury. Female patients with ACL injuries were found to have a greater histologic synovitis response and more active synovium with production of inflammatory and degradative biomarkers. These differences could provide the biologic rationale to explain the higher level of pain and lower patient satisfaction reported amongst female patients after ACLR. While our study demonstrated a persistent low-grade synovitis after ACL injury, what is unknown is how ACLR would impact these findings. Understanding this phenomenon is important given the prevalence of morbidity that occurs after ACL injury even after ACLR. Current work in our laboratory with post-operative biomarkers may elucidate the magnitude of the surgery insult and the joint response after surgery.
