Analysis of cartilage matrix fixed charge density and three-dimensional morphology via contrast-enhanced microcomputed tomography

Abstract

Small animal models of osteoarthritis are often used for evaluating the efficacy of pharmacologic treatments and cartilage repair strategies, but noninvasive techniques capable of monitoring matrix-level changes are limited by the joint size and the low radiopacity of soft tissues. Here we present a technique for the noninvasive imaging of cartilage at micrometer-level resolution based on detecting the equilibrium partitioning of an ionic contrast agent via microcomputed tomography. The approach exploits electrochemical interactions between the molecular charges present in the cartilage matrix and an ionic contrast agent, resulting in a nonuniform equilibrium partitioning of the ionic contrast agent reflecting the proteoglycan distribution. In an in vitro model of cartilage degeneration we observed changes in x-ray attenuation magnitude and distribution consistent with biochemical and histological analyses of sulfated glycosaminoglycans, and x-ray attenuation was found to be a strong predictor of sulfated glycosaminoglycan density. Equilibration with the contrast agent also permits direct in situ visualization and quantification of cartilage surface morphology. Equilibrium partitioning of an ionic contrast agent via microcomputed tomography thus provides a powerful approach to quantitatively assess 3D cartilage composition and morphology for studies of cartilage degradation and repair.noninvasive imaging ͉ proteoglycans ͉ cartilage degeneration ͉ osteoarthritis A nalysis of small-animal models is limited by the availability of quantitative evaluation techniques for studying the extracellular matrix (ECM) changes associated with osteoarthritis (OA) and cartilage repair. Histology is traditionally used to monitor the spatial distribution of matrix macromolecules but is time-consuming and subject to distortion artifacts and tissue damage, and it produces only semiquantitative analysis of 2D sections that may provide inaccurate 3D representations. Biochemical assays are available to quantify the amount and type of matrix macromolecules in cartilage, but these assays fail to provide their spatial distributions, particularly in small animals where the limited thickness and volume of cartilage make it difficult or impossible to extract samples from multiple regions. Additionally, longitudinal monitoring of changes with time are impossible because of the destructive nature of these histological and biochemical techniques.Proteoglycans (PGs) are a particularly appropriate target for studying OA and for evaluating the efficacy of cartilage defect repair. PGs comprise 5-10% of articular cartilage by wet mass (1) and are key regulators of its equilibrium and dynamic mechanical properties. This regulation is the result of interactions between ionic interstitial fluid and negatively charged sulfated glycosaminoglycans (sGAGs) attached to the PG backbone (2). The amount and distribution of PGs changes substantially during development (3), during degeneration and repair (4, 5), and in response to blunt trauma (6). Of particular...

Topographical and depth-dependent glycosaminoglycan concentration in canine medial tibial cartilage 3 weeks after anterior cruciate ligament transection surgery—a microscopic imaging study

“…In the µCT experiments, the baseline X-ray attenuation (without the use of a contrast agent) was found to be indifferent for all topographical locations in the medial tibia and between the ACLT and contralateral cartilage. This observation confirmed the difficulty in the detection of a cartilage lesion without the contrast agent, because of the low attenuation of X-rays in soft tissues (34,41). It should be noted that the baseline attenuation in CT could measure some other properties of cartilage (e.g., cartilage thickness) and the bone structure (e.g., porosity, bone mineral density) (31,42).…”

“…The reduction of T1 can be correlated with the Gd concentration in cartilage, which is inversely proportional to the GAG concentration in cartilage (26)(27)(28)(29)(30) with human CT scanners, can measure a number of changes in cartilage and bone, including the tissue volume, X-ray attenuation, and bone mineral density (31)(32)(33). The use of a negatively charged contrast agent ioxaglate enables µCT to measure the GAG concentration in cartilage (34)(35)(36). Similar to the use of Gd in MRI, the negatively charged ioxaglate ions distribute inversely proportional to the GAG molecules in cartilage, which can be used to calculate the GAG concentration.…”

Effects of diacerein on cartilage and subchondral bone in early stages of osteoarthritis in a rabbit model

“…Although normally when the cartilage structure was quantified using micro-CT, the process involved complex staining techniques with radiopaque agents [38][39][40], in the present study the scan conditions have been adjusted to achieve a correct cartilage visualisation without contrast agents and it was enough to be able to quantify its morphology.…”

No Effect of Hole Geometry in Microfracture for Talar Osteochondral Defects

“…To compare the quality of the repair tissue between the 2-mm and 4-mm deep groups, a noncontrast-enhanced micro-CT scan (lCT) and an equilibrium partitioning of an ionic contrast agent (EPIC) through lCT (EPIC-lCT) scan were acquired of each entire talus using a lCT scanner (Skyscan1076; Skyscan, Kontich, Belgium) [30,49] before histologic analysis. The EPIC-lCT was acquired after the tali had been saturated for 24 hours in a 40% Hexabrix 360 dilution (Guebet BV, Gorinchem, The Netherlands) [49].…”