Abstract

Purpose: Post-traumatic osteoarthritis (PTOA) is a subset of osteoarthritis initiated by an injury such as a blunt impact. However, the local tissue mechanics associated with the earliest stages of PTOA remain undefined. Computational modeling techniques such as finite element analysis provide a means to evaluate mechanical environments that are challenging to measure experimentally. The objective of this work was to compare specimen-specific stresses and strains computed using finite element analysis to confocal microscopy images of chondrocyte viability. Methods: Four viable osteochondral explants (25 mm x 25 mm x 15 mm) were harvested from the tibial plateau of healthy bovine stifle joints (knee). Once removed, the explants were washed twice in HBSS, cultured in normal media (45% DMEM, 45% F12, 10% fetal bovine serum, 100 U/mL penicillin, 100 μg/mL streptomycin, and 2.5 μm/mL amphotericin B) and incubated at 37°C, 5% CO2, and 5% O2. The following day a surface scan of the articular surface of each explant (EinScan Pro 2X Pro; Shining 3D) and ultrasound measurements (Olympus 35DL) of cartilage thickness were acquired. Next, the explants were subject to a 2 J/cm2 impact using a 6 mm flat, beveled stainless-steel platen. After impact, the samples were placed in fresh porcine media and returned to the incubator for 24 h. Cartilage explants were cut with an Isomet 1000 (Buehler, Lake Bluff, IL) slightly off center of the impact and faced by trimming 0.5 mm of the cartilage with a scalpel. These provided a coronal plane cross-section of the explant through the center of the impact site. The explants were washed in phenol red free DMEM/F12 media and stained for 30 min in the incubator with 1 mg/mL Calcein Green AM and 200 nM MitoTracker Deep Red (Life Technologies, Waltham, MA). The samples were then washed and imaged on an Olympus FV1000 (Shinjuku, Japan) confocal microscope. Images were acquired on either edge of the impact, 1 mm inside each edge of the impact site, and in the center of the impact (Figure 1) using a 10X objective lens. An additional top-down view of the articular surface was acquired. Specimen-specific finite element models were generated using the surface scan, cartilage thickness measurements, and geometric measurements of each explant. Cartilage was modeled as hyperelastic with a first-order Ogden strain energy potential. Acceleration of the impact platen was measured (PCB Piezotronics accelerometer), and double integration was used to calculate platen displacement at max tissue compression and impact force. von Mises stress and normal z strain values were compared from corresponding nodes within confocal images (Figure 2). Cell death in the superficial zone and deeper regions of cartilage was assessed. Results: Cell death was present in the superficial zone in 4/4 peripheral edge locations, 3/4 locations interior to the impactor edge and 3/4 of the middle sites. There was no superficial cell death on the central edge of the impact zone. Locations experiencing > 40% strain were associated with high levels of cell death in the superficial zone. When strains exceeded 60%, cell death extended into the deeper zones of the cartilage. It was noteworthy that the highest strains were found at the 1-mm interior to the impact platen edge on all specimens. Von Mises stresses were consistently &gt 30 MPa on the edge of the impact site; however, no obvious relationships were identified between von Mises values and cell death in the superficial, middle, or deep zones. Conclusions: We found strong associations between cartilage strain and chondrocyte death. The highest normal z strain occurs just interior to the beveled edge of the impact platen, and not on the peripheral-most edge of the impact platen. While we did find associations between locations of elevated stress, strain, and cell death, the magnitudes of those values remain to be validated. While important, cell death is a relatively broad measure of injury and associations between cellular health and function and these mechanical measures should be explored.View Large Image Figure ViewerDownload Hi-res image Download (PPT)

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