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. Author manuscript; available in PMC: 2026 Mar 1.
Published in final edited form as: J Orthop Res. 2024 Nov 25;43(3):692–702. doi: 10.1002/jor.26022

Time-dependent changes in medial meniscus kinematics and attachment strength after anterior root injury and repair in a large animal model

Kyle D Meadows 1, John M Peloquin 1, Madeline Boyes 2, Brendan D Stoeckl 3,4,5, Jamie Benson 1, Sonia Bansal 1, David R Steinberg 3,4,5, Miltiadis H Zgonis 3,4, Thomas P Schaer 2,5, Robert L Mauck 3,4,5, Dawn M Elliott 1
PMCID: PMC12032575  NIHMSID: NIHMS2071018  PMID: 39584751

Abstract

This study investigated joint kinematics and attachment tensile mechanics following resection of the medial meniscus anterior attachment. A secondary objective investigated the repair of the attachment. Yucatan minipigs underwent unilateral surgery for either Injury (en bloc) resection of the anterior attachment of the insertional ligament, (a portion of the cranial medial meniscotibial ligament) or Repair (immediate repair with a suture anchor), with the contralateral knee as Intact control. Evaluation at 6 weeks and 6 months included joint kinematics measured from MRI acquired under knee compression and tensile testing of the attachment. Injury resulted in large levels of meniscus extrusion, despite the development of a fibrovascular scar. At 6 weeks, the meniscus extruded 1.95 mm more than Intact; at 6 months, this extrusion was reduced to 0.77 mm. Under an applied 1× body weight load, the meniscus further extruded and was not different with treatment or time. During attachment tensile testing, elongation was 0.6 mm for Intact, following Injury, elongation was 2.7 mm at 6 weeks and was partially restored to 1.5 mm at 6 months. Despite this, the cartilage wear worsened over time. Repair was inadequate to avoid the extrusion or cartilage wear seen in the injury group at 6 weeks, so it was not continued for the 6-month group. This study demonstrates that while meniscus injury is useful to study cartilage degeneration, a holistic consideration of the role of the meniscus itself, including its changing material properties and its impact on joint mechanics during injury, repair, and rehabilitation, are key factors contributing to overall joint health.

Keywords: biomechanics, functional imaging, kinematics and kinetics, meniscus, surgical repair

1 |. INTRODUCTION

The meniscus plays a critical role in the knee in transmitting and distributing load, maintaining low friction, and protecting the articular cartilage (Figure 1). Both meniscus root tears and full thickness radial tears unload the meniscus fibers, which leads to extrusion of the meniscus from the joint space, altering joint mechanics and articular cartilage loading, and ultimately leading to initiation and progression of osteoarthritis (OA).1–6 While it is well established that meniscus tears are linked to higher rates of OA, OA is often not diagnosed until later stages, when pain develops. Therefore, the field has turned to large animal models to study mechanisms for the initiation and progression of OA that arise due to altered joint loading after a meniscus injury.

FIGURE 1.

FIGURE 1

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Joint loading schematic featuring the complex loading state of the meniscus and the role of the attachment in preventing extrusion.

Destabilization of the medial meniscus, or DMM, is an established animal model used to mimic root tears, alter joint mechanics, and induce OA.7–9 Using the Yucatan mini pig, which has similar physiology and joint loading to humans,10–12 our team has recently developed a porcine DMM model of OA where we sought to link noninvasive MRI measures to destructive tissue measures so that findings could be translated to human OA development.7,8 We found joint and tissue level changes in the knee (note: this paper will use the knee for the quadruped stifle/femorotibial joint) as early as 1-month post-surgery with varied trajectories (resolution, persistence, or worsening) up to 6 months post-surgery. At the joint level, loading was assessed via Tekscan pressure sensors, and the results showed that DMM created stress concentrations on the cartilage at 1 month that resolved at later time points. Abrasions and fissuring of the cartilage were visible via India Ink staining at 1 month, worsened by 3 months, and were unresolved at 6 months.7,8 Notably, a fibrovascular scar formed at the site of the resected root and showed increased tissue organization (healing) over the 6-month period. This raised the possibility of a gradual re-establishment of meniscus load transfer and load distribution and cessation of further breakdown of the articular cartilage. However, in this prior work, the fibrovascular scar was not mechanically tested to determine its capacity to maintain the meniscus’ position under load. It is, therefore, uncertain if the fibrovascular scar developed mechanical properties sufficient to re-establish meniscus load transfer and whether joint kinematics were restored.

Therefore, the primary objective of this study was to use this DMM model to investigate the joint kinematics and attachment tensile mechanics following meniscus Injury compared to Intact control. To slow or prevent functional reattachment of the fibrovascular scar, in the present work, we resected en bloc a 3–5 mm length of the anterior medial meniscus attachment rather than using a simple cut.9 We hypothesized that the Injury group would display meniscus extrusion at 6 weeks that would partially recover at 6 months due to fibrovascular scar formation and remodeling.

Re-establishment of meniscus loading to prevent stress concentrations on the cartilage and prevent long-term catastrophic damage motivates the importance of early repair post-injury.13 We recently studied whether suture repair returned the meniscus to its intact position in a cadaveric model.14 We found that acute repair partially restored joint kinematics compared to the intact condition, however, it was unknown whether the joint mechanics would continue to improve over time following repair.

Therefore, our second objective was to explore the capability of a suture anchor repair to restore intact joint kinematics in our porcine DMM model. We hypothesized that the suture anchor repair would prevent the meniscus extrusion seen in the Injury case by restoring the meniscus to its Intact position and loading state.

2 |. METHODS

This study received ethics approval from the Institutional Animal Care and Use Committee of the University of Pennsylvania. A total of 20 male castrated Yucatan minipigs (Sinclair Bioresources, Auxvasse, MO, age 12–18 months on the day of surgery, weight 56 ± 12 kg) underwent unilateral mini-arthrotomy of the knee (femorotibial) joint as reported previously.7,8 After surgery, animals were allowed unrestricted exercise within their 8’×5’ pens and were evaluated daily for well-being by veterinary surgeons throughout the study. Postsurgical time points were 6 weeks and 6 months. Additionally, the Acute effect of treatment was assessed in cadaveric knees. For the Acute group, experimental conditions were applied serially on the same knee, with some of the outcomes previously published.14

Our primary objective was to evaluate the effect of Injury (n =8 at 6 weeks, n = 8 at 6 months), which was performed via a complete transection and en bloc removal of a section of the anterior medial meniscus ligamentous attachment (insertional ligament) with a No. 11 scalpel blade to create a 3–5 mm gap immediately adjacent to its osseous insertion.9 In quadrupeds the insertional ligaments are often referred to as cranial/caudal meniscotibial ligaments, not to be confused with the coronary ligament, which is also known as the (posterior) meniscotibial ligament. In the minipig, the body of the medial meniscus is tightly attached to the capsule throughout its peripheral border, leading to restricted mobility. Due to these surrounding structures, transection of the anterior medial meniscus attachment did not create a free-floating meniscus. A probe was used to confirm that the attachment was fully detached. Our secondary objective was to evaluate the effect of Repair (n = 4 at 6 weeks), which was performed by first transecting the cranial medial meniscotibial ligament with a scalpel immediately adjacent to its osseous insertion. Next, a 4.5 PushLock suture anchor (Arthrex, Inc.) was placed in the tibia using standard surgical instrumentation, and the ligament was immediately re-attached to its insertion site using a vertical mattress suture technique.14 The en bloc attachment removal in the Injury group was performed to minimize healing, while the attachment transection and immediate reattachment in the Repair group was performed to maximize healing. As will be presented and discussed, the Repair did not restore joint kinematics or prevent cartilage wear at 6 weeks, so it was not continued for the 6-month group. Contralateral legs were left undisturbed and served as Intact controls (n = 12).

2.1 |. Gait analysis

After 2–4 weeks postoperative, all animals were regularly evaluated for gait abnormalities by veterinary surgeons using standard of practice gait evaluation at a walk and a trot, noting any deviations in gait (such as winging or paddling), failure to land squarely on all four feet and the unnatural shifting of weight from one limb to another looking for signs, such as shortening of the stride, irregular foot placement, head bobbing, stiffness, and weight shifting. Using a modified lameness scale ranging from zero to five, with zero being no perceptible lameness, and five being most extreme: 0: Lameness not perceptible under any circumstances; 1: Lameness is difficult to observe and is not consistently apparent, regardless of circumstances; 2: Lameness is difficult to observe at a walk or when trotting in a straight line but becomes more consistent when i.e., circling; 3: Lameness is consistently observable at a trot under all circumstances; 4: Lameness is obvious at a walk; 5: Lameness produces minimal weight bearing in motion and/or at rest or a complete inability to move.15

In addition, gait analysis was performed on four animals from the 6-month Injury group using the GAITFour® walkway system.16,17 The pigs walked across a sensor-embedded mat, and the contact pressure of each hoof-strike was recorded. Pigs were habituated to the walkway before surgery to the point where they willingly trotted down the walkway to receive a treat. A model that normalized by velocity and trial number was used to measure differences between the Injury and Intact sides.

2.2 |. Joint loading and magnetic resonance imaging

The joint loading preparation and experimental design followed the same protocols as previously described.14 Briefly, the hindlimbs were disarticulated, and the femur and tibia long bones were cut near the knee joint, preserving the joint capsule and all ligaments. The femur and tibia were then potted using Ortho-Jet acrylic resin (polymethyl methacrylate, Lang Dental), secured in Delrin cups, and placed in an MRI-compatible loading device at 30° of flexion (Figure 2). This angle is consistent with the porcine joint angle during standing. Fluoroscopy was used during potting to ensure the tibia and femur were aligned parallel with the central axis of the Delrin cups and that both femoral condyles were equally in contact with the tibia.

FIGURE 2.

FIGURE 2

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Loading setup for (A) knee joint and (B–E) anterior medial meniscus attachment. (A) The knee joint was potted in PMMA, placed in an MRI compatible loading frame, loaded to the desired force in an ElectroForce 3510 test system, and then the Lock Nuts were tightened to hold the loaded joint in position for transport to the MRI facility. (B) In the Intact group, the medial meniscus anterior attachment (arrow) was prepared for testing by removal of the meniscus mid-body and the infrapatellar fat pad. (C) In the Injury group, the medial meniscus anterior attachment (arrow) was prepared for tension testing in the same way, except that because the scar and fat pad were structurally integrated, this mass was divided where their boundary normally would be (dotted line). (D) Tension was applied to the insertional ligament by clamping the tibia to the base of an Instron 5943 test frame, such the ligament was vertically aligned, and suturing the meniscus horn to a PTFE rod attached to the frame’s crosshead. (E) Attachment and fiducial markers. Elongation of the attachment was calculated as the displacement of the ink markers at the attachment–meniscus transition (rectangle) relative to the nails driven into the tibia (circles).

To noninvasively measure joint kinematics, MRI was acquired at two loading states: Low Load (44.5 N compression, to ensure the joint was in contact) and High load (510 N compression, which represents approximately one body weight). For the Acute samples, a High Load of 850 N was used for all treatments.14 The load was applied using a mechanical testing system, and once the target load was reached, the nuts on the threaded rod were tightened, resulting in complete load transfer to the MRI-compatible loading frame and locking of the displacements in place (Figure 2A). The loading frame was transferred to the MRI facility, and MRI was acquired 30 min after load application. A T1 vibe MR sequence was used to achieve high-resolution images with good contrast: repetition time = 10 ms, echo time = 3.45 ms, voxel size = 0.2 × 0.2 × 0.2 mm, runtime = 18 min. After MRI, joints were wrapped in PBS-soaked gauze, vacuum sealed, and frozen until dissection. Stress relaxation occurs after locking in the loaded position, which we previously quantified for the Acute timepoint as 65% relaxation from the initial load to the end of imaging, and this was not different between treatment groups.14 Stress relaxation is unavoidable with the time required for this assay but would not affect study outcomes since the treatment group did not affect the magnitude of relaxation.

2.3 |. Magnetic resonance image analysis

Meniscus motion (kinematics) was calculated from MR images comparing between Low Load and High Load (Figure 3), as previously described.14 First, all MR images were resliced into a tibia-aligned RAS+ (right, anterior, and superior positive) coordinate system using the Low Load images. Next, in the Low Load images, the tibia was segmented using an active contour semi-automatic segmentation tool (ITK-SNAP,18). Next, the corresponding High Load image was aligned to the Low Load coordinate system by rigid registration of the tibia, using the segmentation as a registration mask.19–21 The outer boundary of the medial meniscus was manually labeled in 3D by following a line halfway between the superior and inferior rims (3D Slicer22). The initial position at Low Load was standardized across joints by using the medial intercondylar tubercle as the origin. The motion from Low Load to High Load (i.e., displacement vector) was calculated by matching points at the same % arc length along the meniscus boundary lines and subtracting the positions of each point. Meniscus position and motion were analyzed for the whole meniscus by averaging displacement along the entire boundary line and, for regional analysis, averaging displacement in the anterior region (AR), mid-body region (MR), and the posterior region (PR).

FIGURE 3.

FIGURE 3

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Example Low Load and High Load MRIs from the Injury group. The mid-coronal slice, halfway between the meniscal horns, is shown.

2.4 |. Macroscopic cartilage assessment

Joints were dissected by severing the collateral ligaments, and the central portions of the medial and lateral menisci were removed. Images were then taken of the cartilaginous tibial plateau surface. The tibial plateau was then painted with India ink and wiped clean with wet gauze. Cartilage wear was quantified as the percentage of the tibial plateau area that retained ink, with the ink-stained regions outlined by hand.9,23,24

2.5 |. Meniscal horn attachment tensile testing

After joint loading and MRI, the anterior medial meniscus attachment was tested in tension to measure its capacity to restrain medial translation of the meniscus. To allow the insertional ligament to move freely, the infrapatellar fat pad was removed. In the Intact joints, the boundary between the insertional ligament and the fat pad was easily identified and dissected (Figure 2B). In the Injury joints, the scar and the fat pad formed an integrated mass, which was cut where their boundary would have been in an Intact joint (Figure 2C). The tibia was clamped to the base of an Instron 5943 universal testing system, and the remnant of meniscus on the end of the attachment was connected to the test system’s crosshead using a running stitch of 3 loops of 2/0 braided silk suture (Fine Science Tools, Foster City, CA; cat. no. 18020–20) (Figure 2D,E). To allow measurement of tissue displacement from the video of the tensile test, three nails were driven into the tibia, and Sharpie ink marks were placed on the attachment (Figure 2E). A preload of 0.65 N was applied, followed by 10 cycles of tension between 0.65 N and 65 N at a displacement rate of 0.1 mm/s. The 65 N force matches the estimated tension in an Intact attachment under High Load, based on another study.25 Using data from the 10th cycle of the force-displacement response, the attachment’s elongation, which reflects its ability to hold the meniscus in place under load, was measured as the displacement of the ink markers on the attachment–meniscus transition relative to the tibial nails. Elongation at the peak force and tangent stiffness were measured.

2.6 |. Statistical analysis

All outcome measures were compared with a two-way ANOVA, when possible, with one factor being the treatment (Intact, Injury, Repair) and the other factor being time point (Acute, 6 weeks, 6 months). Some outcome measures did not include every factor level, and, in these instances, a one-way ANOVA was used to compare treatments within a time point, as noted in the results when applicable. When significance was met, a post hoc Tukey HSD was used to test for the effect of treatment or time point. Pearson correlations were performed to identify relationships between outcome measures. Significance was set at p < 0.05.

3 |. RESULTS

3.1 |. Surgical details and postsurgical recovery

All surgeries were performed without complication. No animals were removed from the study early; all reached their scheduled date of sacrifice (6 weeks or 6 months). Immediately postoperative, all animals were toe-touching lame (grade 5) on the operated leg (Injury and Repair group) for 1–3 weeks. Over time, the lameness resolved to no visible lameness appreciable at a walk or trot in any of the animals in the 6-week group. Operated animals of the Injured cohort in the 6-month group showed shortened cranial stride of the operated limb with a subtle unloading in the stance phase observed as an asymmetry of the pelvis at a trot (hip hike; grade 3). At the time of sacrifice, of the four animals in the Injured cohort of the 6-month group that qualified for walkway analysis, the total pressure index percentage was significantly lower on the right (operated) hind limb when compared to the left hind limb, when normalized for velocity and trial number (model adjusted effect: −0.5, 95% CI: [−0.9, −0.1], p = 0.005). Similarly, the total scale pressure (the sum of peak pressure values recorded from each activated sensor by a hoof during mat contact) showed significantly less pressure in the right hind limb compared with the left hind limb (model adjusted effect: −1.4, 95% CI: [−2.4, −0.5], p = 0.004).

3.2 |. Effect of injury

3.2.1 |. Meniscus motion

When under load, the meniscus shifted medially out of the joint space in the Injury group, as seen qualitatively in axial plane displacements of the outer boundary outlines (Figure 4A). The anterior portion of the meniscus had more motion than the posterior, as the posterior attachment was still intact, and the meniscus pivoted outward around this restraint.

FIGURE 4.

FIGURE 4

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(A) Representative meniscus High Load position in axial plane at 6 weeks, showing outline of tibial plateau (blue), Intact (black), and Injury (red); scale bar = 5 mm. With Injury, the loaded meniscus moved outward medially. (B) Average meniscus initial position at Low Load (dots) and motion from Low Load to High Load (arrows) at 6 weeks (light color) and 6 months (dark color). At 6 weeks, the Injury initial position was 1.95 mm outward medially compared to Intact (*p < 0.05), and at High Load was 1.62 mm outward medial compared to Intact (^p < 0.05). At six months, the Injury initial position partially recovered and was 0.77 mm outward medially (not significantly different from Intact or 6 weeks Injury). The motion from Low Load to High Load was not different with treatment or time. C–H describes the values and standard deviation for these positions and displacements by their anterior and medial components.

The average meniscus initial position at Low Load and motion from Low Load to High Load is shown in Figure 4B (dots) for the whole meniscus, and their components in the anterior and medial directions are shown in Figure 4C,D. With Injury, the 6-week initial position was 1.95 mm outward (medial) compared to Intact (p < 0.05, Figures 4B,D. At 6 months post Injury, the initial meniscus position in the medial direction partially recovered to 0.77 mm (n.s. from Intact and 6-week Injury), suggesting the fibrovascular scar developed enough tension to partially anchor the anterior meniscus in place. The mean 6-month Injury initial position also shifted 0.43 m posteriorly, but this was quite variable (n.s. with respect to any other group) (Figure 4B,C).

The displacement from Low Load to High Load was primarily in the anterior direction (mean 0.76 ± 0.56 mm across all treatments and times) with some medial motion (mean 0.32 ± 0.34 mm across all treatments and times) (Figure 4B arrows; Figure 4E,F). Displacement did not significantly differ with respect to treatment or time, but the final position of the meniscus under High Load was significantly further outward (medial) at 6 weeks (p < 0.05) (Figure 4B,H) due to the aforementioned change in the initial position. Thus, while DMM compromised the initial position (under a small tare load), the amount of additional motion with full loading was not affected.

3.3 |. Attachment testing

The intact attachments and fibrovascular scars were tested in tension designed to match the load experienced by an intact attachment under High Load. The peak elongation of the Intact attachment was 0.6 ± 0.2 mm at 6 weeks and 6 months (Figure 5). The peak elongation of the Injury attachment was greater: 2.7 ± 0.6 mm at 6 weeks and 1.5 ± 0.3 mm at 6 months. There was a significant effect of treatment, time point, and their interaction (p < 0.05). Specifically, the 6 weeks Injury and 6 months Injury groups were statistically different from all other groups. The tangent stiffness showed differences consistent with the results for peak elongation. For the Intact time points, the stiffness was similar at 6 weeks and 6 months, 370 ± 60 and 390 ± 80 N/mm, respectively. The Injury time points were much less stiff: 130 ± 30 N/mm at 6 weeks 220 ± 40 N/mm at 6 months. There was a significant effect of both treatment and time point on the tangent stiffness, and the 6 weeks Injury and 6 months Injury groups were statistically different from all other groups (p < 0.05).

FIGURE 5.

FIGURE 5

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Attachment tensile test data from the 10th cycle to 65 N tension. (A) Force versus displacement curves for each specimen and (B) 95% confidence intervals. Intact attachments had similar mechanics between 6 weeks and 6 months. Injury attachments had large displacements at 6 weeks and somewhat smaller displacements at 6 months, although still more than Intact attachments.

3.4 |. Correlations between measures

We observed some significant correlations between the meniscus position in the joint loading tests and attachment mechanics from the attachment tensile tests (Figure 6). The initial meniscus position in the medial direction was significantly correlated to both the attachment elongation in tensile testing (R2 = 0.34, p < 0.05, Figure 6) and the tangent stiffness (R2 = 0.16, p < 0.05). However, the displacement of the meniscus from Low Load to High Load was not correlated to either attachment tension measurement.

FIGURE 6.

FIGURE 6

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Initial position of the meniscus along the medial axis measured from MRI of Low Load joint compression is significantly correlated (p < 0.001) with peak attachment elongation measured from attachment tensile tests. Initial positions are plotted for (A) the whole meniscus (R2 = 0.34), (B) the anterior region (R2 = 0.28), and (C) the mid-body region (R2 = 0.49).

3.5 |. Cartilage wear

The percentage of tibia plateau cartilage staining positively with India ink was used to assess the amount of wear (Figure 7). The Intact joints had remarkably little wear with 0.18% area stained at 6 weeks and 0.84% area stained at 6 months. The Injury joints had much more staining with 1.77% area stained at 6 weeks and 2.13% area stained at 6 months (one outlier, 18%, was excluded from the intact 6 weeks group). By two-way ANOVA, there was a significant effect of Injury (vs. Intact) on the percentage of area stained (p < 0.05), but no significant effect of time.

FIGURE 7.

FIGURE 7

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India Ink staining of the tibial plateau indicated cartilage degeneration in the Injury group that was significantly different from Intact. (A) Representative samples of best, median, and worst cases at 6 months, (B) Mean and standard deviation of percent area stained at 6 weeks and 6 months. (* and ^ p < 0.05 for Injury compared to Intact at 6 weeks and 6 months, respectively).

3.6 |. Effect of repair

At 6 weeks post-surgery, we assessed the ability of the suture anchor repair to restore meniscus displacement under load to Intact levels (Figure 8). In our prior cadaver joint study, Acute repair partially restored meniscus displacement immediately after the repair was completed (Figure 8). Thus, there was the possibility to see further reduction of meniscus extrusion due to the combination of suture anchor repair and healing in the present animal study. However, Repair at 6 weeks showed meniscus displacement under load that was much closer to Injury at 6 weeks than to Intact at 6 weeks. Looking at the initial positions (Figure 8A–C), at 6 weeks the Injury and Repair menisci were further extruded from the joint space than menisci with the same treatments at the Acute time point in the cadaver study. With loading, there was a significant effect of treatment on the final position in the medial and anterior directions (p < 0.05) and a trending effect of time point on the anterior final position (p < 0.1). The final position for both Injury and Repair was significantly different from the Intact. The repair was, therefore, unable to restore joint function under in vivo loading conditions, even though there were initially promising results in the Acute condition. In contrast, despite the apparent inability of the repair to restore joint function, we observed some chondroprotection in terms of India Ink staining at 6 weeks (Repair, 1.45%, was not significantly different from Intact while Injury was, Figure 7B).

FIGURE 8.

FIGURE 8

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(A) Average meniscus initial position at Low Load (dots) and motion from Low Load to High Load (arrows) for Intact (gray), Injury (red), and Repair (blue) at the initial time point (Acute, light color) and at 6 weeks (dark color). There was no difference for the Intact group (gray) between Acute and 6-weeks. The Acute Repair (light blue) had a moderate improvement compared to the Acute Injury, with initial medial position of 0.54 mm and final medial displacement of 1.67 mm. However, at 6 weeks, the Repair (dark blue) initial position and final displacement (1.80 and 1.73 mm) were nearly the same as the Injury (dark red). Acutely, both Injury and Repair initial positions were significantly different from Intact in the cranial direction (*). At 6 weeks, both Injury and Repair initial positions were significantly different from Intact in the medial direction (*). Injury final position was significantly different from Intact in the medial direction acutely and at 6 weeks (^). Repair was significantly different from Intact in the medial direction acutely (^). These positions and motions were evaluated by their cranial and medial components in (B–G).

To help explain why the Acute repair partially restored meniscus displacements under joint loading, but the 6-week repair showed no benefit, we measured the tensile strength of the Acute Repair attachment using the same apparatus as was used to tensile test the Intact and Injury attachments at 6 weeks and 6 months. The Acute Repair attachments failed at 31–67 N, with 2–7 mm of elongation at failure (Figure 9). The Acute Repair, therefore, was much less stiff than the Intact attachment (cf. Figure 5), with strength almost always less than 65 N, the estimated tension in the attachment when the joint is compressed at 30° flexion with 1× body weight.

FIGURE 9.

FIGURE 9

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Individual force–elongation response for attachment tensile tests to failure of Acute Repair demonstrated that most repairs could not support the estimated 65 N of physiological tension, and that large elongations (2–7 mm) were required to reach the repair’s ultimate tensile strength. Note that in the 6 weeks and 6 months post-surgery groups (Figure 5), the attachment was strong enough to sustain 65 N cyclic tension.

4 |. DISCUSSION

In this study, we found that injury to the medial meniscal attachment (meniscotibial ligament) caused the meniscus to extrude out of the joint space after 6 weeks of normal activity, though the extrusion was reduced at 6 months, likely due to the formation of a fibrovascular scar. Tensile loading of the attachments (fibrovascular scar in injury group) proved that the fibrovascular scar was far less mechanically functional than the intact attachment; however, after 6 months, the fibrovascular scar was more mechanically functional than after 6 weeks. Despite the fibrovascular scar improving from 6 weeks to 6 months, the cartilage wear worsened over this time, indicating that this scar was insufficient to prevent cartilage degeneration. Thus, the rate of scar maturation was outpaced by the concomitant rate of chondral degeneration, as observed by India Ink staining and animal lameness.

4.1 |. Effect of injury

We observed that meniscus extrusion in the Injury group led to its inability to distribute loads across the joint and protect the underlying cartilage. The bulk of the medial extrusion in the Injury group was evident at Low Load, and the application of High Load did not meaningfully increase the amount of medial motion. This could indicate two possibilities: (1) that the meniscus was extruded outward in vivo and therefore was not carrying load in the joint, and/or (2) the meniscus was so weakly restrained that ex vivo the 50 N Low Load was enough to push the meniscus outward before applying the High Load. Either way, this amount of extrusion would indicate that the cartilage is likely experiencing high stress concentrations due to the meniscus’ failure to distribute load in the joint space, similar to other studies.7,8,26–28

Higher rates of OA are seen in patients when the meniscus fails to distribute load due to meniscal extrusion or meniscus tear.1,29–32 In our study, cartilage health was assessed via India Ink staining, which revealed worsening cartilage wear with time in the Injury group, likely due to this inability of the meniscus to distribute loads. This is particularly interesting considering that the 6-month Injury group saw some resolution of the medial meniscus extrusion and attachment mechanics compared to the 6-week Injury, yet it was not enough to prevent further cartilage wear. The fibrovascular scar that was occurring in the Injury group was likely not sufficiently mechanically robust as to prevent damaging stress concentrations on the underlying cartilage.

These findings are supported by our gait analyses at 6 months, which showed detectable abnormalities. Animals displayed a shortened stride in the affected limb, as well as a reduction in time spent in the stance phase, indicating that animals were increasing weightbearing on the unoperated limb. Further, the kinematic parameters assayed during the gait analysis showed a significant and consistent decrease in weight distribution on the Injured limb postoperative. Joint pain alters normal function, particularly locomotion, and is one of the clinical signs of osteoarthritis. The alterations in gait and loadbearing in pigs 6 months postoperatively may reflect painful ambulation secondary to OA changes because of the Injury.

One study limitation is that we measured meniscus motion and attachment tensile properties, but we did not directly measure the loading on the cartilage surface. The meniscus motion and tensile mechanics were correlated, where higher meniscus extrusion was associated with lower tensile properties. It is likely that these are also linked to higher stress concentrations and lower contact areas on the cartilage. Indeed, our measures match what we had previously seen in our animal model when we measured cartilage contact mechanics via a Tekscan pressure sensor.7,8 In addition, this study is somewhat limited by using only ink staining to evaluate cartilage wear; future work will add assessments of the micro-scale tissue outcomes.

4.2 |. Effect of repair

We found that an immediate suture anchor repair was inadequate to avoid the extrusion seen in the Injury group, so it was not continued for the 6-month group. After 6 weeks, the suture anchor repair was not outperforming the fibrovascular scar and had failed to re-establish joint loading seen in the Intact group. There are two potential reasons for why the repair failed, both compounded by the current iteration of our animal model’s inability to limit the range of motion of the operated joint and non-weightbearing and thus protect the repair: (1) the sutures pulled through the highly aligned anterior attachment at low loads, and (2) the sutures fatigued and stretched resulting in insufficient mechanical viability to preserve intact meniscus kinematics. Unfortunately, we do not know which was the true reason (or whether it was a combination of the two) due to the scarring around the sutures and attachment. To repeat this study, a better anchoring technique is needed.

Addressing limitations in the repair strategy and/or postsurgical joint unloading is warranted for this animal model, as the suture repair alone did not withstand the expected physiological loads (Figure 9). The meniscal root attachments in pigs are loose and more mobile than the body of the meniscus and arranged in small bundles resembling cruciate ligaments with fibers running horizontally from the body to their insertion sites on the tibial plateau. In human knees, the medial meniscus has broader insertion sites on the edge of the tibial plateau. The anterior medial meniscus insertional ligament in pigs is significantly longer and thinner compared to the human anterior and posterior meniscus insertional ligaments and offers a limited footprint for securely anchoring sutures during a repair. In the future, we could avoid this fixation failure by suturing closer to the horn where the fibers are less aligned and may provide better anchoring. Additionally, the pigs were allowed to walk the day after surgery; the repair was not protected by limiting the range of motion and degree of weight-bearing as a human would be following a meniscus repair. It is likely that the repair would have been more effective had some duration of healing been enabled through immobilization. This observation has important implications to study the impact of conservative and aggressive rehabilitation protocols following clinical repair surgery.

In addition to the repair limitations described above, some study limitations should be noted. First, the contralateral joint was intact, rather than a sham surgery. In our prior experience with this surgical model, we did use a sham control,7,8 with similar findings to the present work, so we do not think this is an important limitation. Whether intact or sham control, however, the contralateral control experienced altered weight bearing, which could have worsened the outcomes for the control but would not affect the overall findings comparing the treatment groups to each other and over time.

In conclusion, resection of the anterior attachment of the medial meniscus resulted in large amounts of meniscus extrusion at 6 weeks and 6 months post-injury, despite fibrovascular scar development. Anterior attachment tensile tests proved that the fibrovascular scar was not mechanically viable enough to mimic healthy, intact anterior attachment mechanics. As a result, higher levels of cartilage wear were seen in injured joints compared to intact joints, likely due to the stress concentrations on the cartilage resulting from loss of load distribution by the meniscus. Acutely repairing the meniscus tear showed no difference in joint kinematics compared to injury at 6 weeks, and alternative repair techniques and postsurgical immobilization are needed in this model. Ultimately, this study demonstrates that while meniscus injury is useful to study cartilage degeneration, a holistic consideration of the role of the meniscus itself, including its changing material properties and its impact on joint mechanics during injury, repair, and rehabilitation, are key factors contributing to overall joint health.

ACKNOWLEDGMENTS

The authors thank Edward D. Bonnevie for preparing Figure 1. This research was supported by the National Institute of Arthritis and Musculoskeletal and Skin Diseases R01 AR050052, R01 AR056624, and P30 AR069619, the National Institute of General Medical Sciences P20 GM139760, and the U.S. Department of Veterans Affairs I01 RX003375, I50 RX004845, and IK6 RX003416.

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