In vitro method for 3D morphometry of human articular cartilage chondrons based on micro-computed tomography

I Kestilä,J Thevenot,M.A Finnilä,S.S Karhula,I Hadjab,S Kauppinen,M Garon,E Quenneville,M Haapea,L Rieppo,K.P Pritzker,M.D Buschmann,H.J Nieminen,S Saarakkala

doi:10.1016/j.joca.2018.05.012

I Kestilä, J Thevenot + Show 12 more

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https://doi.org/10.1016/j.joca.2018.05.012

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SummaryObjectiveThe aims of this study were: to 1) develop a novel sample processing protocol to visualize human articular cartilage (AC) chondrons using micro-computed tomography (μCT), 2) develop and validate an algorithm to quantify the chondron morphology in 3D, and 3) compare the differences in chondron morphology between intact and osteoarthritic AC.MethodThe developed protocol is based on the dehydration of samples with hexamethyldisilazane (HMDS), followed by imaging with a desktop μCT. Chondron density and depth, as well as volume and sphericity, were calculated in 3D with a custom-made and validated algorithm employing semi-automatic chondron selection and segmentation. The quantitative parameters were analyzed at three AC depth zones (zone 1: 0–10%; zone 2: 10–40%; zone 3: 40–100%) and grouped by the OARSI histological grades (OARSI grades 0–1.0, n = 6; OARSI grades 3.0–3.5, n = 6).ResultsAfter semi-automatic chondron selection and segmentation, 1510 chondrons were approved for 3D morphometric analyses. The chondrons especially in the deeper tissue (zones 2 and 3) were significantly larger (P < 0.001) and less spherical (P < 0.001), respectively, in the OARSI grade 3–3.5 group compared to the OARSI grade 0–1.0 group. No statistically significant difference in chondron density between the OARSI grade groups was observed at different depths.ConclusionWe have developed a novel sample processing protocol for chondron imaging in 3D, as well as a high-throughput algorithm to semi-automatically quantify chondron/chondrocyte 3D morphology in AC. Our results also suggest that 3D chondron morphology is affected by the progression of osteoarthritis (OA).

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With osteoarthritis (OA), chondrocytes are known to become hypertrophic and to form clusters, indicating that they may play a critical role in the increased metabolism observed in this condition[1,2]
One explanation is that the chondrocytes produce more extracellular matrix (ECM) macromolecules trying to compensate the increased cartilage degeneration, which leads to increased cellular clustering and hypertrophy[1]
Volumetric visualizations of HMDS-dried osteochondral samples from Osteoarthritis Research Society International (OARSI) grades 0e4 imaged with desktop micro-computed tomography (mCT) are shown in Automatic selection and segmentation performance

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Introduction

With osteoarthritis (OA), chondrocytes are known to become hypertrophic and to form clusters, indicating that they may play a critical role in the increased metabolism observed in this condition[1,2]. One explanation is that the chondrocytes produce more extracellular matrix (ECM) macromolecules trying to compensate the increased cartilage degeneration, which leads to increased cellular clustering and hypertrophy[1]. At the level of the chondron e the chondrocyte and its complex pericellular microenvironment5e7 e previous studies have suggested that the cleavage of fibrillary collagens initiates chondron enlargement, which continues due to pericellular matrix deposition[8,9]. Both the chondrocyte and its pericellular matrix (PCM) appear to be modulated by OA8,10

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