We describe variations in the degree of mineralisation within the subchondral bone plate of the equine metacarpophalangeal
joint. A comparison of Optical Coherence Tomography, Micro CT, and SEM techniques was performed. These data are
compared between sites on a healthy sample and at points on an osteoarthritically degenerated sample. No significant
correlation was found between the optical scattering coefficient and the micro-CT derived BMD for comparisons between
different sites on the bone surface. Also OCT demonstrated a larger regional variation in scattering coefficient than did
micro CT for bone mineral density. This suggests that the optical scattering coefficient of bone is not related solely to the
volume-density of calcium-phosphate. Patches of lower optical scattering coefficient were found in the bone structure that
was related to the osteoarthritic lesion area on the overlying cartilage. Areas of microcracking, as revealed by both SEM and
micro CT produced distinctive granularity in the OCT images.
In further experiments, OCT was compared with micro CT and mechanical strength testing (3-point bending) in a
small animal model of cardiovascular disease (cholesterol overload in mice). In the cardiovascular diseased mice, micro-CT
of the trabecular bone did not demonstrate a significant change in trabecular bone mineral density before and after
administration of the high cholesterol diet. However mechanical testing demonstrated a decrease in mechanical strength and
OCT demonstrated a corresponding statistically significant decrease in optical scattering of the bone.
Polarization-sensitive optical coherence tomography has been used to solve fast-axis fibre orientation in three dimension
space. Previously we have demonstrated that the apparent variations in polar angle orientation of collagen fibers along
sagittal ridge of equine third metacarpophalangeal joint exist. A quantitative method based on multiple angles of
illumination has been proposed to determine the polar angle of the collagen fibers. This method however ignored the full 3D
structure by assuming that the collagen fibers long-axis lay within the plane of incidence.
A new quantitative method based on the theory of light propagation in uniaxial materials is described which avoids this
assumption. To test this method we have performed control experiments on a sample of equine tendon (this tissue has well
defined c-axis lying along the long-axis of the tendon). Several samples of tendon were cut to achieve a planar surface
inclined at -20° to the long axis. Additional 30° rotation provided non-zero azimuthal angle. The surface was then imaged
using incident beam angles -40°, -20°, 0, +20°, +40° in two orthogonal planes. Values for both the polar and azimuthal
angles were then derived using a numerical optimisation procedure. Results agreed qualitatively with the nominal values but
suggested that the accuracy was limited by our method of determining the apparent birefringence.
Polarization-sensitive optical coherence tomography has been used to solve fast-axis fibre orientation in three dimension
space. Previously we have demonstrated that the apparent variations in polar angle orientation of collagen fibers along
sagittal ridge of equine third metacarpophalangeal joint exist. A quantitative method based on multiple angles of
illumination has been proposed to determine the polar angle of the collagen fibers. This method however ignored the full
3-D structure by assuming that the collagen fibers long-axis lay within the plane of incidence.
A new quantitative method based on the theory of light propagation in uniaxial materials is described which avoids this
assumption. To test this method we have performed control experiments on a sample of equine tendon (this tissue has
well defined c-axis lying along the long-axis of the tendon). Several samples of tendon were cut to achieve a planar
surface inclined at -20° to the long axis. Additional 30° rotation provided non-zero azimuthal angle. The surface was
then imaged using incident beam angles -40°, -20°, 0, +20°, +40° in two orthogonal planes. Values for both the polar and
azimuthal angles were then derived using a numerical optimisation procedure. Results agreed qualitatively with the
nominal values but suggested that the accuracy was limited by our method of determining the apparent birefringence.
Equine articular cartilage has been imaged using both polarization-sensitive optical coherence tomography (PS-OCT)
and non-linear microscopy. PS-OCT has been used to spatially map the birefringence in the cartilage and we have found
that in the vicinity of the lesion the images display a characteristic disruption in the regular birefringence bands shown by
normal cartilage. We also note that significant (e.g. x2) variations in the apparent birefringence of samples taken from
young (18 month) animals that otherwise appear visually homogeneous are found over spatial scales of a few
millimeters. We have also imaged the cartilage using non-linear microscopy and compare the scans taken with second
harmonic generation (SHG) light and the two photon fluorescence (TPF) light. SHG images collected using 800 nm
excitation reveals the spatial distribution of collagen fibers, whilst TPF images clearly shows the distribution of
intracellular and pericellular fluorophores.
Osteoarthritis is a painful condition, causing restricted mobility in the articular joints. In this paper we present a review of
different optical techniques that might be used to clarify the etiology of degeneration of connective joint tissues, such as bone
and articular cartilage. Significant correlation (R2 = 0.8) between bone mineral density and scattering coefficient of cortical
bone tissue are found by using Integrating Sphere Technique. Optical Coherence Tomography and Polarization-Sensitive
Optical Coherence Tomography images of cartilage tissue are presented. They were performed as series of angle-dependant
measurements for different location along the surface. Method for spatial mapping the birefringence of equine articular
cartilage is proposed. Variations in band spacing of birefringence obtained from visually healthy and abnormal cartilage
samples are compared. Visible osteoarthritic lesions are characterized by a loss of the regular birefringence bands shown by
normal cartilage. We discuss the hypothesis that some of these variations may be due to changes in intrinsic structure of
tissue.
Polarization-sensitive optical coherence tomography has been used to spatially map the birefringence of equine articular cartilage. The polar orientation of the collagen fibers relative to the plane of the joint surface must be taken into account if a quantitative measurement of true birefringence is required. Using a series of images taken at different angles of illumination, we determine the fiber polar angle and true birefringence at one site on a sample of equine cartilage, on the assumption that the fibers lie within the plane of imaging. We propose a more general method based on the extended Jones matrix formalism to determine both the polar and azimuthal orientation of the collagen fibers as well as the true
birefringence as functions of depth.
Polarization-sensitive optical coherence tomography has been used to spatially map the birefringence of equine articular cartilage. Images obtained in the vicinity of visible osteoarthritic lesions display a characteristic disruption of the regular birefringence bands shown by normal cartilage. We also note that significant (e.g. ×2) variations in the apparent birefringence of samples taken from young (18 month) animals that otherwise appear visually homogeneous are found over spatial scales of a few millimeters. We suggest that whilst some of this variation may be due to changes in the intrinsic birefringence of the tissue, the 3-D orientation of the collagen fibers relative to the plane of the joint surface should also be taken into account. We propose a method based on multiple angles of illumination to determine the polar angle of the collagen fibers.
Previously we have demonstrated by an integrating-sphere technique a useful correlation between bone mineral density (BMD) and the optical scattering coefficient (μs). We have used OCT to study this relationship, with a view to assessing its viability as an in vivo measure of BMD. We used an OCT system operating at 1.3 microns to image various bone samples and to make images in different directions. To investigate the accuracy of OCT measurements we collected averaged images taken at different sites along the shaft of a bone sample. A small (about 3%) variation in measured coefficients, justifies the possibility of using the OCT technique for this kind of measurement. Measurements of bone both with and without periosteum, showed essentially the same optical properties. Applying the incident light beam perpendicular and parallel to the main direction of the collagen fibers gave differences in scattering coefficient of about 40% that confirms our previous suggestion about anisotropy of bone matter. Images of bone samples for different demineralisation time were collected. They allowed following changes in calcium distribution in cortical part of bone with the increasing time of acid activity.
The ability to measure changes in bone-mineral-density (BMD) in-vivo has potential applications in monitoring stress-induced
bone remodelling in, for example, competition race horses. In this study we have begun to investigate the potential of optical techniques to monitor such changes via changes in bone optical scattering. Using integrating spheres, we have investigated the optical properties of bone samples taken from the leg of the horse. Since our samples have stable characteristics over the time, we are able to use a single integrating-sphere technique. Diffuse reflection and transmission coefficients have been measured over the wavelength range 520 to 960 nm. Measurements were made on samples immersed
in formic acid solution for different lengths of time; this was to investigate the effect of reduction in BMD on the optical properties. The experimental results and a Monte-Carlo based inversion method were used to extract the absorption coefficient and unmodified scattering coefficient of the samples. After full demineralisation scattering coefficient fell by a factor 4. This shows that the calcium-content in bone influences its optical properties considerably. Our experiments confirm the possibility of using optical techniques to determine changes in the BMD of samples.
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