A Large Animal Model for Orthopedic Foot and Ankle Research

Abstract

Trauma to the soft tissues of the ankle joint distal syndesmosis often leads to syndesmotic instability, resulting in undesired movement of the talus, abnormal pressure distributions, and ultimately arthritis if deterioration progresses without treatment. Historically, syndesmotic injuries have been repaired by placing a screw across the distal syndesmosis to provide rigid fixation to facilitate ligament repair. While rigid syndesmotic screw fixation immobilizes the ligamentous injury between the tibia and fibula to promote healing, the same screws inhibit normal physiologic movement and dorsiflexion. It has been shown that intact screw removal can be beneficial for long-term patient success; however, the exact timing remains an unanswered question that necessitates further investigation, perhaps using animal models. Because of the sparsity of relevant preclinical models, the purpose of this study was to develop a new, more translatable, large animal model that can be used for the investigation of clinical foot and ankle implants. Eight (8) skeletally mature sheep underwent stabilization of the left and right distal carpal bones following transection of the dorsal and interosseous ligaments while the remaining two animals served as un-instrumented controls. Four of the surgically stabilized animals were sacrificed 6 weeks after surgery while the remaining four animals were sacrificed 10 weeks after surgery. Ligamentous healing was evaluated using radiography, histology, histomorphometry, and histopathology. Overall, animals demonstrated a high tolerance to the surgical procedure with minimal complications. Animals sacrificed at 10 weeks post-surgery had a slight trend toward mildly decreased inflammation, decreased necrotic debris, and a slight increase in the healing of the transected ligaments. The overall degree of soft tissue fibrosis/fibrous expansion, including along the dorsal periosteal surfaces/joint capsule of the carpal bones was very similar between both timepoints and often exhibited signs of healing. The findings of this study indicate that the carpometacarpal joint may serve as a viable location for the investigation of human foot and ankle orthopedic devices. Future work may include the investigation of orthopedic foot and ankle medical devices, biologic treatments, and repair techniques in a large animal model capable of providing translational results for human treatment.

Keywords: carpus; fixation; foot and ankle; sheep; syndesmosis.

Figure 1

Comparison of the (left) human syndesmotic structure and the (right) sheep distal carpal bones. (Left) The human structure contains the fibula and tibia joined anteriorly by the anterior tibiofibular ligament (ATIFL), posteriorly by the posterior tibiofibular ligament (PTIFL), and centrally by an interosseous ligament (IL) located between a layer of cartilage (C) on the two bones. (Right) The distal carpal bones of the sheep (MDCB, medial distal carpal bone; LDCB, lateral distal carpal bone) share similar cross-sectional morphometry as the human fibula and tibia, ligamentous attachments (DL, dorsal ligament; VL, ventral ligament; IL, interosseous ligament) and cartilage (C) zone as the human syndesmotic complex. Illustration courtesy of Kelsea Erickson, DVM.

Figure 2

Step (1) An Anterior-posterior fluoroscopic image of the carpus was taken. Step (2) 22-gauge needles were placed within the carpometacarpal joint and the middle carpal joint as well as the intercarpal joint between the 2nd/3rd and 4th Carpal bones to aid in alignment. Step (3) A beaver blade was placed within the 2nd/3rd and 4th inter carpal bone to transect the anterior inter carpal ligament. Step (4) A 1.2mm K-wire was placed midway between the carpometacarpal joint and middle carpal joint. Step (5) The 1.2mm K-wire was inserted from the medial aspect of the fused 2nd/3rd carpal bone across the intercarpal joint and through the 4th carpal bone. The K-wire angled in ~15 degrees anteromedial to posterolateral direction in order to stay central within the 2nd/3rd and 4th carpal bones in both the dorsal and transverse planes. Step (6) A cannulated countersink tool was placed over the K-wire to countersink the medial edge of 2nd/3rd carpal bone. Step (7) The K-wire distance was measured and a 2.6mm cannulated drill bit was used to drill a hole of matching length. Step (8) A 4.0mm tap is placed over the K-wire to thread the drill hole. Step (9) A 4.0 mm cannulated screw was placed over the K-wire and driven across the fused 2nd/3rd and 4th carpal bones in a neutral position to stabilize the intercarpal joint. Step (10) The K-wire was removed from the cannulated screw.

Figure 3

The dorsal ligament area was histomorphometrically analyzed. (A) Original histology section. (B) The dorsal ligament area was quantified around the dorsal peripheral surface of the carpal bones.

Figure 4

Mean dorsal ligament area around the periphery of the carpal bones was similar between 6-week and 10-week specimens and increased as compared to the Control group; however, those differences were not statistically significant between groups.

Figure 5

Representative photomicrographs of histological scoring parameters. (A–C) Collagen organization and neo-vascularization scoring. (A) Control animal 1 - left. Photomicrograph demonstrating a collagen fiber organization score of 3 (dense/mature) and a neovascularization score of 0 (none). Interosseous ligament collagen is comprised entirely of bundles of densely packed collagen fibers with a crimped appearance and which contain few flattened, hyperchromatic nuclei of ligament fibroblasts. No blood vessels are observed within this ligament tissue. (B) Animal PG04 - right, 6-week timepoint. Photomicrograph demonstrating a collagen fiber organization score of 0 (none/no organization) and a neovascularization score of 2 (mild). Newly produced fibrous tissue surrounding carpal bones is characterized by reactive and disorganized fibroplasia with haphazardly intersecting collagen bundles, and numerous plump reactive fibroblasts. 3–5 capillary-sized neo-vessels per high powered (400x magnification) field are present throughout this fibrous tissue (arrows). (C) Animal PG07 - right, 12-week timepoint. Photomicrograph demonstrating a collagen fiber organization score of 2 (moderate) and a neovascularization score of 2 (mild). Newly produced fibrous tissue is being progressively re-organized into more parallel and tightly packed bundles of collagen fibers, similar to native interosseous ligament observed in healthy control animals. Similar to the 6-week animal shown in (B), 3–5 capillary-sized neo-vessels per high powered (400x magnification) field remain present throughout this fibrous tissue (arrows). (D–F) Inflammation scoring. (D) Control animal 4 - left. Photomicrograph demonstrating a cumulative inflammation score 0. No inflammatory cells were observed in the carpal ligament tissues of any healthy control animals. (E) Animal PG06 – right, 6-week timepoint. Photomicrograph demonstrating a cumulative inflammation score of 6/20. Multifocal nodular clusters of predominately lymphocytes and fewer plasma cells (arrow), as well as scattered individual infiltrating macrophages, were observed throughout the newly produced fibrous tissue. (F) Animal PG05 – left, 12-week timepoint. Photomicrograph demonstrating a cumulative inflammation score of 8/20. Similar to animal PG06-right, clusters of infiltrating lymphocytes, plasma cells and macrophages (arrows) are multifocally present throughout the newly produced fibrous tissue. All images are 20x magnification.