Relationship between cartilage and subchondral bone lesions in repetitive impact trauma-induced equine osteoarthritis

ArticleinOsteoarthritis and Cartilage 20(6):572-83 · February 2012with51 Reads
Impact Factor: 4.17 · DOI: 10.1016/j.joca.2012.02.004 · Source: PubMed
Abstract

To correlate degenerative changes in cartilage and subchondral bone in the third carpal bone (C3) of Standardbred racehorses with naturally occurring repetitive trauma-induced osteoarthritis. Fifteen C3, collected from Standardbred horses postmortem, were assessed for cartilage lesions by visual inspection and divided into Control (CO), Early Osteoarthritis (EOA) and Advanced Osteoarthritis (AOA) groups. Two osteochondral cores were harvested from corresponding dorsal sites on each bone and scanned with a micro-computed tomography (CT) instrument. 2D images were assembled into 3D reconstructions that were used to quantify architectural parameters from selected regions of interest, including bone mineral density and bone volume fraction. 2D images, illustrating the most severe lesion per core, were scored for architectural appearance by blinded observers. Thin sections of paraffin-embedded decalcified cores stained with Safranin O-Fast Green, matched to the micro-CT images, were scored using a modified Mankin scoring system. Subchondral bone pits with deep focal areas of porosity were seen more frequently in AOA than EOA but never in CO. Articular cartilage damage was seen in association with a reduction in bone mineral and loss of bone tissue. Histological analyses revealed significant numbers of microcracks in the calcified cartilage of EOA and AOA groups and a progressive increase in the score compared with CO bones. The data reveal corresponding, progressive degenerative changes in articular cartilage and subchondral bone, including striking focal resorptive lesions, in the third carpal bone of racehorses subjected to repetitive, high impact trauma.

Full-text

Available from: Guy Beauchamp, Oct 14, 2015
Relationship between cartilage and subchondral bone lesions in repetitive
impact trauma-induced equine osteoarthritis
M. Lacourt y, C. Gao z,A.Liz, C. Girard x, G. Beauchamp x, J.E. Henderson z, S. Laverty y
*
y Comparative Orthopaedic Research Laboratory, Département de Sciences Cliniques, Faculté de Médecine Vétérinaire, Université de Montréal, C.P. 5000,
Saint-Hyacinthe (QC), J2S 7C6, Canada
z Division of Experimental Medicine, McGill University, JTN Wong Lab for Bone Engineering, McGill University Health Centre, 740 Ave Dr Peneld, Montreal, Quebec H3A 1A4, Canada
x Département de pathologie et microbiologie vétérinaires, Faculté de Médecine Vétérinaire, Université de Montréal, C.P. 5000, Saint-Hyacinthe (QC), J2S 7C6, Canada
article info
Article history:
Received 4 June 2011
Accepted 9 February 2012
Keywords:
Osteoarthritis
Horse
Subchondral bone
Cartilage
Microcracks
summary
Objective: To correlate degenerative changes in cartilage and subchondral bone in the third carpal bone
(C3) of Standardbred racehorses with naturally occurring repetitive trauma-induced osteoarthritis.
Design: Fifteen C3, collected from Standardbred horses postmortem, were assessed for cartilage lesions
by visual inspection and divided into Control (CO), Early Osteoarthritis (EOA) and Advanced Osteoar-
thritis (AOA) groups. Two osteochondral cores were harvested from corresponding dorsal sites on each
bone and scanned with a micro-computed tomography (CT) instrument. 2D images were assembled into
3D reconstructions that were used to quantify architectural parameters from selected regions of interest,
including bone mineral density and bone volume fraction. 2D images, illustrating the most severe lesion
per core, were scored for architectural appearance by blinded observers. Thin sections of parafne
embedded decalcied cores stained with Safranin O-Fast Green, matched to the micro-CT images,
were scored using a modied Mankin scoring system.
Results: Subchondral bone pits with deep focal areas of porosity were seen more frequently in AOA than
EOA but never in CO. Articular cartilage damage was seen in association with a reduction in bone mineral
and loss of bone tissue. Histological analyses revealed signicant numbers of microcracks in the calcied
cartilage of EOA and AOA groups and a progressive increase in the score compared with CO bones.
Conclusion: The data reveal corresponding, progressive degenerative changes in articular cartilage and
subchondral bone, including striking focal resorptive lesions, in the third carpal bone of racehorses
subjected to repetitive, high impact trauma.
Ó 2012 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.
Introduction
Osteoarthritis (OA) is a characterized by ssures and erosion of
articular cartilage combined with osteophytosis, sclerosis, and cysts
in the subchondral bone. Microcracks in the calcied tissues have
also been observed in naturally occurring OA in humans
1
, following
joint impact trauma in animal models
2,3
and in equine joint dis-
ease
4e8
. This structural change may enhance cellular activity
leading to increased bone remodeling
9,10
. Although the interplay
between altered biomechanical and cellular signaling in articular
cartilage and subchondral bone promotes joint tissue degeneration,
there is no consensus regarding the initiating events
11
.
An early decrease in subchondral bone mineral density occurs in
acutely induced animal models of OA
12e15
, with a rebound increase
observed in longer duration studies
16e19
. These observations
suggest an important role for early subchondral bone changes in
rapidly progressive experimental OA but may not reect the clinical
situation where OA progresses slowly over many years. In a guinea
pig model of naturally occurring OA, subchondral bone changes
occurred before alterations in articular cartilage
20
.
The racehorse is an athlete that serves as a model for naturally
occurring, repetitive impact trauma-induced OA. Human athletes
that engage in strenuous sports also suffer from occupational-
induced repetitive compression trauma OA. Examples include OA in
soccer players knees and ballet dancers ankles
21,22
.
Osteochondral lesions, both OA and fractures, occur frequently on
the dorsal aspect of the third carpal cuboidal bone (C3) in race-
horses
23e27
. C3 is subject to repetitive compression forces during
strenuous exercise that induce changes in both the articular cartilage
and subchondral bone. Fibrillation and loss of cartilage proteoglycan
*
Address correspondence and reprint requests to: S. Laverty, Département de
sciences cliniques, Faculté de Médecine, Vétérinaire, Université de Montréal, C.P.
5000, Saint-Hyacinthe, Quebec J2S 7C6, Canada. Tel: þ1-4507788100.
E-mail addresses: MatLacourt@aol.com (M. Lacourt), sheila.laverty@umontreal.ca
(S. Laverty).
1063-4584/$ e see front matter Ó 2012 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.
doi:10.1016/j.joca.2012.02.004
Osteoarthritis and Cartilage 20 (2012) 572e583
Page 1
been identied in C3 following strenuous athletic activity
28
. Radio-
graphic sclerosis
29
, increased bone mineral density
30,31
and
increased stiffness also arise in the subchondral bone
32
.
Recent advances in micro-computed tomography (CT) tech-
nology have enabled quantication of bone architectural parame-
ters from 3D reconstructions of high resolution non-destructive
imaging of bone
10,33,34
.In the current study we correlated micro-CT
assessment of subchondral bone architecture and composition with
histological analysis of overlying cartilage in the C3 bone of equine
athletes at varying stages of OA.
Method
Tissue procurement
Tissue was harvested from racing Standardbred horses at a local
abattoir. The history of a prior racing career was determined by the
presence of a racing shoe and/or a tattoo. Palpation of the carpal
joints was performed to detect effusion and age was estimated by
teeth examination
35
. Limbs were refrigerated at þ4
C until harvest
of C3, within 10 h of death.
Macroscopic assessment of C3 articular surface
The greatest length and width of the macroscopic cartilage lesion
was measured and the product was employed to estimate lesion
area. The lesions were scored as: Control (CO), no visible lesions on
the articular surface; Early Osteoarthritis (EOA), ssures and/or
partial thickness erosions in an area 100 mm
2
: or Advanced
Osteoarthritis (AOA), with partial to full thickness erosions or
ulcerations in an area 100 mm
2
(Fig. 1). This terminology was
selected based on a recent report conrming that cartilage lesions
increase in size with progression of OA
36
. Limb harvest was
continued until ve specimens were identied in each group. Mean
(SD) age of animals was: 6.6 1.6 years (CO), 5.6 2.0 years (EOA)
and 5.0 2.1 (AOA). They included three females and two geldings
in the CO Group, two females, one male and two geldings in the EOA
group, and three females and two geldings in the AOA group.
Two 1 cm diameter osteochondral cores were harvested (Fig. 2).
Core 1 was removed from the classical location for OA in this bone
and Core 2 from a site that is rarely affected. The dorsal aspect of
each core was notched to facilitate orientation (Fig. 2) and xed in
4% paraformaldehyde for 24 h at þ4
C, rinsed and stored in PBS
at þ4
C until micro-CT imaging.
Micro-CT analysis
The specimens were scanned using a Skyscan 1172 (Skyscan,
Antwerp, Belgium). (Supplementary data online)
Qualitative micro-CT analysis
Micro-CT visual assessment score
2D reconstructions, based on the most severe bone lesion per
core, were examined (two readers) to identify a pattern of bone
degenerative changes for a consensual structural score: Score 1:
absence of surface irregularities; Score 2: surface irregularities and
pits, sometimes associated with focal porosity in subchondral
bone; Score 3: surface irregularities and pits with marked focal
porosity throughout the full depth of subchondral bone (Fig. 2).
Fig. 1. Illustration of macroscopic appearance of the proximal articular surface of the third carpal bone (C3), micro-CT image from the most severe lesion in core 1 from the same
bone and the corresponding histological slide (Safranin O fast green stain-SOFG) from the CO group, EOA group and AOA group. Black arrows indicate sites of focal articular cartilage
lesions; the subchondral bone pits (white arrow heads) are shown on the micro-CT images. Note focal patches of remodeling porosity under and peripheral to the subchondral pits.
In the early OA histological section, although the cartilage appears normal, some cracks are present in the mineralized cartilage. A focal loss of subchondral bone with presence of
cartilage is observed. Underlying bone pathology was not suspected from surface inspection. In the AOA section the articular cartilage surface is minimally eroded but there is loss of
staining and a ssure is present in the cartilage as well as under it. Extensive underlying bone pathology and remodeling are visible. An island of cartilage (grey arrow head) is
present deep in the bone. Large trabecular spaces, probably due to remodeling, are also present deeper in the bone.
M. Lacourt et al. / Osteoarthritis and Cartilage 20 (2012) 572e583 573
Page 2
Fig. 2. Micro CT assessment of harvested specimens. Core harvest: (A) Proximal articular surface of C3 illustrating the location of cores 1 and 2 in the radial fossa from a specimen
with an advanced OA lesion. (B) Latero-medial view of a core with dorsal notch (white arrow) for orientation purposes. Structural score: This was an overall subjective score for
structural changes in the most severe section on micro-CT. (A) Score 1:absence of articular surface irregularity; (B) Score 2: absence of irregularities and pits at the articular surface
of the subchondral bone with or without moderate focal porosity of the underlying subchondral bone and (C) Score 3: irregularities and pits at the articular surface of the sub-
chondral bone with marked concomitant focal porosity throughout the full depth of the core. ROIs: Quantication: Illustration of ROI where bone morphometric parameters were
assessed on micro CT 2D reconstruction of samples with (A) and without (B) pit. In the absence of a pit 3 ROI were dened, a dorsal ROI (D) between the dorsal edge and the middle
of the core, a central ROI (C) in the middle of the core and a palmar ROI (P) between the middle and the palmar edge. In presence of a SubCondral Bone (SCB) pit 5 ROI were selected.
The ROI L is centered in the SCB immediately below the pit, two adjacent perilesional ROI in the dorsal (DPL) and palmar aspect (PPL) of the lesion were evaluated. Two remote
perilesional ROI one dorsal (DR) and palmar (PR), 1 mm from ROI L were also studied.
M. Lacourt et al. / Osteoarthritis and Cartilage 20 (2012) 572e58357 4
Page 3
Four other observers then independently scored all the images
employing the atlas with designated consensual score.
Quantitative micro-CT analyses
Regions of interest (ROI) were identied for quantitative analyses
and included: D, dorsal; C, central; P, palmar; L, lesion; DPL, dorsal
perilesional; PPL, palmar perilesional sites. Two remote perilesional
ROIs, one dorsal (DR) and palmar (PR), 1 mm from ROI L were also
studied (Fig. 2). Parameters measured included bone mineral density
(BMD), bone volume fraction (BV/TV), trabecular thickness (Tb.Th),
spacing (Tb.Sp) and number (TbN), fragmentation index (Tb.Pf),
structure model index (SMI) and degree of anisotropy (Da).
Histological analyses of articular cartilage and subchondral bone
Specimens were decalcied in 10% EDTA, embedded in parafn
and sectioned at 4
m
m, to correspond to the 2D micro-CT recon-
structions. Safranin O stained sections were assessed by two blin-
ded observers for degenerative changes using a modied Mankin
grading system
37
(Supplementary data online. (Table. 1)). The
number of cracks in the calcied cartilage and pits in subchondral
bone were also counted (over 1 cm length).
Statistical analysis
Micro-CT visual assessment score
Inter-observer agreement was assessed employing a Kappa
coefcient. A Kruskal-Wallis test examined differences in scores
between the groups (CO, EOA, AOA).
Quantitative micro CT analyses
Mean Micro-CT ROI values from the three sites (C, D and P) in
the CO group were compared using a repeated-measures linear
model, with ROI (three levels: C, D and P) and core sample (dorsal
and palmar cores) as within-subject factors.
The EOA and AOA groups were also compared using a repeated-
measures linear model, with ROI (four levels: L, DPL, PPL and R) as
within subject factors and disease status (EOA and AOA) as
between-subject factors.
ROIs C and L were compared in the three groups using a linear
model with disease status (three levels) as between-subject factors.
A priori contrasts between pairs of means were conducted with
a Bonferroni sequential adjustment procedure to ensure a family
wise level of statistical signicance at the set alpha level. Tukeys
post hoc tests were also used for unplanned comparisons.
Comparison Core 1 and core 2
A priori contrasts with the Bonferroni adjustment procedure
were also used to compare the mean Micro CT quantitative
parameters of ROIs C or L of core 1 and core 2 in each group.
Histological analyses of cartilage and subchondral bone
Intra-class correlation coefcients for histological scores were
also assessed. Histological score difference between groups was
analyzed using a KruskaleWallis test. The median number of cracks
in the calcied cartilage was compared between the three groups
and also the combined OA groups (EOA and AOA) and the CO group
with a Mann-Whitney test. The prevalence of pits in the SCB was
compared between CO group and the combined OA groups with an
exact chi-square test.
Correlations between cartilage and bone changes
Associations between the cartilage macroscopic and histological
scores and Micro-CT visual assessment scores, quantitative data
and calcied cartilage crack numbers were investigated employing
Spearman non-parametric correlations.
A level of P < 0.05 was considered statistically signicant. SAS v.
9.2 (Cary, N.C.) was employed to perform the statistical analyses.
Results
Synovial distension was noted in four of ve of the joints that fell
into the AOA group but not in any of the other joints.
Macroscopic assessment of C3 articular surface
The C3 articular surface in the CO group was smooth, whereas the
EOA group exhibited partial thickness erosions of 44 42.8 mm
2
(mean SD) and the AOA group had partial and/or full thickness
erosions or ulcerations of 178.6 40.1 mm
2
(Fig. 1).
Micro-CT analysis
Qualitative micro-CT analysis
There was a high level of inter-observer agreement for scores of
visual assessment of the 2D micro-CT reconstructions (kappa coef-
cient ranging between 0.85 and 0.92). The mean scores of the core
SCB architecture in 9/10 specimens (cores 1 in CO and cores 2 in CO,
EOA and AOA) were considered to be of homogeneous density
without visible structural change (Supplementary data online:
Table 2). No signicant differences were observed between Cores
1 and 2 in the CO group or between Cores 2 in all three groups.
A signicant heterogeneity (P ¼ 0.006) was identied in scores
from cores 1 among groups and postehoc tests revealed a statisti-
cally signicant difference between scores in the CO and AOA
groups. Irregularities of the articular surface, subchondral pits and
porosity were a feature of Core 1 in AOA specimens and although
not statistically signicant, were also observed in the EOA group. An
area of focal porosity in the underlying bone was typically associ-
ated with subchondral bone pits (Figs. 1 and 2).
Bone loss, with increasing severity of OA, was also observed on
volume rendered models of bone structure (Fig. 3).
Quantitative micro-CT analysis (Core 1)
Results of the Micro-CT quantitative analyses are illustrated in
Fig. 4 (Supplementary data Online. Table 3).
Within group comparisons
There were no statistically signicant differences in any of the
bone parameters assessed between the three ROI sites within the
CO or EOA groups.
Statistically signicant differences in bone parameters between
ROI sites were noted within the AOA group alone. Furthermore,
they occurred uniquely between the ROI site below the lesion (L)
and the remote sites (R) and included: a lower BMD (P ¼ 0.001)
below the lesion [Fig. 4(A)], a lower BV/TV (P ¼ 0.003) [Fig. 4(B)],
a higher TbSp (P ¼ 0.002) [Fig. 4(E)] and a lower TbTh (P ¼ 0.001)
[Fig. 4 (F)] when compared to the remote sites in the AOA group.
Trabecular number (TbN)(mm
-1
), SMI, and Fragmentation Index
(Tb.Pf)(mm
-1
), Degree of anisotropy (Da)
No statistically signicant differences were detected for these
parameters within groups (Fig. 4C, D, E and H).
Intergroup comparisons
As there were no statistically signicant differences in any of the
bone parameters assessed between the three ROI sites of the CO
group, the mean value was chosen for the subsequent intergroup
M. Lacourt et al. / Osteoarthritis and Cartilage 20 (2012) 572e583 575
Page 4
comparisons. No statistically signicant differences were detected
on intergroup comparisons (CO, EOA and AOA).
Quantitative micro-CT analysis: Core 1 and Core 2 comparisons
No statistically signicant differences were detected in the CO
group or the EOA group between cores 1 and 2. In the AOA group
the BMD (P ¼ 0.02), BV/TV (P ¼ 0.016) and Tb.Th (P ¼ 0.02) were
signicantly lower in core 1 when compared to core 2 but the Tb.Sp
was signicantly higher (P ¼ 0.00 06) (Supplementary data online
Table 3).
Histological analysis of articular cartilage
There was a good intra-class coefcient of variation (71%) for the
mean histological scores between observers.
Histological score
Representative histological images of specimens in different
groups with a description of typical lesions are provided in Fig. 5
(AeG). The total histological score was signicantly increased
(P ¼ 0.02) in the AOA group when compared to the CO group in Core
1, but not when compared to the EOA group (Fig. 6).
Correlations between cartilage and bone changes
There was a signicant positive correlation between the artic-
ular cartilage macroscopic and histologic scores and the micro-CT
visual assessment score (bone structure) in Core 1; r ¼ 0.84;
P < 0.001 and r ¼ 0.76, P < 0.001 respectively.
Signicant negative correlations were identied between the
cartilage macroscopic score and BMD (r ¼0.55; P ¼ 0.03) and
BVTV (r ¼0.61; P ¼ 0.02) and a positive correlation with TbSp
(r ¼ 0.76; P ¼ 0.002).
Associations between cartilage histological scores and micro-CT
quantitative bone parameters are illustrated in Fig. 7 (AeH). There
was a signicant
negative correlation between histological scores
and BV/TV (r ¼0.69, P ¼ 0.005) [Fig. 7(B)] but a positive correla-
tion with Tb.Sp (r ¼ 0.82, P < 0.001)[Fig. 7 (E)].
Calcied cartilage microcracks
One core from the AOA Group was excluded from the calcied
cartilage crack count due to a large zone of ulceration of the
cartilage on the section. The cracks were oblique and oriented at
30
and often localized close to the tidemark [Fig. 5(G)]. They were
less frequent in the CO group in comparison to EOA and AOA groups
(Supplementary data online: Table. 4). In EOA and AOA groups
cracks occasionally entered the SCB [Fig. 5(E)] and a coalescence of
the cracks was observed. In some specimens the articular cartilage
appeared to be collapsed, probably due to the coalescence of many
cracks (Fig. 5D, F and G).
There was signicantly greater (P ¼ 0.04) number of cracks in
the combined OA (EOA þ AOA) groups in comparison to the CO
group.
Correlations
There was a signicant positive correlation (r ¼ 0.52; P ¼ 0.03)
between the cartilage macroscopic score and the number of calci-
ed cartilage cracks. There was also a trend (r ¼ 0.50; P ¼ 0.08) for
a correlation between the cartilage histological score and the
number of microcracks.
Histological analysis of subchondral bone
SCB pits were observed in 10 of 15 Core 1 specimens. The
number of pits was signicantly greater (P ¼ 0.04) in the combined
OA groups compared with the CO group (Supplementary data
online: Table. 4).
Subchondral bone resorption was observed below some of the
pits and appeared to be organized in the vertical axis of the core
[Fig. 5 (D, E and G)]. The lacunae appeared to be oriented towards
the tidemark so that there was a progressive loss of continuity and
Fig. 3. Volume-rendering (upper panel) and 3D (lower panel) models of bone structure located in the central ROI (C) of CO group and ROIs L of EOA and AOA groups. Volume model
and 3D model were generated when loading dataset images inside of each ROI in CTVox and CTVol respectively. The boundaries of ROI were shown as the purple clip boxes in
volume models whereas bone is the white objects within. Different colors (brown, blue and green) are applied to the 3D models of bone structure within aforementioned ROIs.
There is an obvious trend of increased bone loss and lack of connectivity from CO to AOA (left to right).
M. Lacourt et al. / Osteoarthritis and Cartilage 20 (2012) 572e583576
Page 5
formation of pits and crevasses. Although not quantied, this
appeared to occur more frequently in the EOA and AOA groups.
A loss of the tidemark continuity was observed and subchondral
bone was replaced by cartilage in some cores of both EOA and AOA
groups [Fig. 5(G)]. In some specimens an intact articular cartilage
surface on histological slides masked the presence of obvious pits
(visible on matched Micro-CT sections). Pits contained cartilage
with chondrocytes and proteoglycans stained with SOFG replacing
large areas of bone resorption. Fibrous tissue and chondrocytes
were also observed in these defects. Areas of bone resorption and
the presence of osteoclasts in the periphery of resorptive lacunae
were also detected [Fig. 8(A and B)].
Fig. 4. Results of micro-CT quantitative analyses of bone morphometric parameters from ROIs from core I in CO (C), early OA (EOA) and advanced OA (AOA) groups. Bone parameters
assessed included BMD, BV/TV, SMI, TbSp, TbTh, TbN, TbPf and Da. D ¼ Dorsal ROI, C ¼ Central ROI, P ¼ Palmar ROI, DPL ¼ Dorsal perilesional ROI, L ¼ Lesional ROI, PPL ¼ Palmar
perilesional ROI, R ¼ dorsal or palmar ROI (see Fig. 2). The boxes dene the twenty-fth and seventy-fth percentile with a line at the median. Whiskers dene the maximum and
minimum values. Statistically signicant differences are illustrated. (see also Supplementary data online e Table 3 ).
M. Lacourt et al. / Osteoarthritis and Cartilage 20 (2012) 572e583 577
Page 6
Discussion
Using a combination of micro-CT and histochemical assessment,
focal microstructural changes were identied in the calcied carti-
lage and underlying subchondral bone in equine naturally occurring
repetitive trauma OA. The defects include calcied cartilage cracks,
focal pits extending through the calcied cartilage and underlying
subchondral bone in addition to striking areas of vertically oriented
resorptive remodeling in the subchondral cancellous bone, deep to
the pits. Microstructural changes deeper in the bone, below the pits,
were conrmed on micro-CT morphometric measurements that
revealed a lower bone mineral density, bone volume, and trabecular
thickness but a greater trabecular spacing when compared to
remote sites in the advanced OA specimens. Importantly, the sub-
chondral bone changes correlated with the cartilage lesions,
providing additional evidence that the subchondral bone is involved
and plays a role in the pathogenesis of naturally occurring repetitive
impact OA.
A lower bone mineral density due to hypomineralisation of the
subchondral bone and an increase in the spacing between trabec-
ulae, indicating trabecular erosion, has also been previously iden-
tied by other investigators in human OA tissues
10,38
. Conversely,
however, the bone volume and trabecular thickness increased in
most studies of human OA specimens
10,33,38
and may reect the
longer trajectory or different pathogenesis of primary human OA.
Bone mineral density and structure have previously been
investigated in osteoarthritic equine metacarpi employing clinical
CT
39,40
. Focal resorptive lesions and a heterogeneous subchondral
bone were also observed
39
. The study herein, using much higher
resolution micro e CT to study C3 microarchitecture, conrms and
extends these previous observations, but in a different joint. Stover
has previously illustrated similar, but milder, pathology in a C3
bone on microradiographs
41
.
It has been known for some time that adaptive changes occur in
the subchondral bone of equine C3 due to the high cyclic strains of
Fig. 6. The total histological scores from cores 1 of the different groups. There is a
statistically signicant difference in the histological scores between the CO groups and
advanced OA groups. The boxes dene the twenty-fth and seventy-fth percentile
with a line at the median. Whiskers dene the maximum and minimum values.
Fig. 5. Histological sections of specimens with different lesion scores stained with Safranin O fast green (AeI) to illustrate pathological ndings. A e CO sample, the red coloration of
the hyaline cartilage, reecting proteoglycan content, is homogenous and the surface of the cartilage is smooth. The SCB bone structure appears regular. B e EOA. A small vertical
crack is present in the calcied cartilage just above the tidemark. C e EOA. Multiple vertical cracks at 30
obliquity in the calcied cartilage above the tidemark. D e EOA. This
section illustrates an example of coalescence of the cracks in the calcied cartilage. The calcied cartilage is lost, the overlying hyaline cartilage has reduced proteoglycans (loss of
staining), a reduced thickness (erosion) is diffusely hypocellular and clones are present at the periphery of the changes. There appears to be an increase in the number and size of
bone lacunae under the pits. E e A large vertical crack running through hyaline cartilage, calcied cartilage and subchondral bone is present (right side of the picture) and a focal
total thickness ulceration is present (left side of the picture). F e Discontinuity in the calcied cartilage zone and the subchondral bone. The overlying hyaline cartilage has reduced
proteoglycans compared to adjacent normal cartilage. The pit contains cartilage, either an attempt at healing or a result of collapse into the pit. The articular cartilage surface is
irregular and a focal subchondral bone porosity is present. G e This core has a large area of focal porosity underlying an area with islands of cartilage inside in the subchondral bone
defect. (G2) The overlying articular cartilage has reduced staining but only a single crack or ssure without loss of articular cartilage (no erosion). A loss of calcied cartilage is
centered over the bone lesion. Cracks in the calcied cartilage, with approximately 30
oblique angles are present dorsal and palmar aspect to the lesion (G 1 and 3).
M. Lacourt et al. / Osteoarthritis and Cartilage 20 (2012) 572e583578
Page 7
Fig. 7. Correlations between the histological score and the micro-CT quantitative bone parameters and the number of microcracks in core 1. (A) BMD, (B) BV/TV, (C) TbN, (D) SMI,
(E) TbSp, (F) TbTh, (G) TbPf, (H) Da, (I) number of microcracks counted on histological sections. P values are indicated on gure when statistically signicant or illustrating a trend.
The diamonds represent the different specimens (White ¼ CO, Grey ¼ EOA and Black ¼ AOA). .
M. Lacourt et al. / Osteoarthritis and Cartilage 20 (2012) 572e583 579
Page 8
strenuous exercise
23
, leading to regional radiographic sclerosis
29
,
increased bone stiffness
5,32
and mineral density
5,30,32,42
. Yet, it is
unclear how these adaptive changes are linked to the chon-
droosseus changes described here. Radiolucencies within dorsal
sclerotic areas in C3 are associated with lameness in racehorses
43
and probably correspond to the calcied tissue remodelling
events illustrated here with micro-CT.
In the current study, osteoclasts were observed in resorptive
lacunae in some of the specimens. There is limited knowledge on
the role of osteoclasts in OA. Subchondral bone resorption pits, with
enhanced metalloproteinase expression, have been observed to cut
through calcied cartilage and postulated to contibute to hyper-
remodelling in OA patients
44
. Also osteoclastic activity increases in
the subchondral bone of OA patients and correlates to cartilage
damage
38
.
The calcied cartilage layer of articular cartilage transfers joint
forces at the cartilage-bone interface. Structural or molecular
alterations in this transitional zone induced by repetitive trauma,
could result in fragility that upsets the normal biomechanical
equilibrium that exists between cartilage and bone in healthy joints.
Consistent with previous investigations of both naturally occurring
and experimental OA
2,3,11,45e47
cracks were observed in the calcied
cartilage. Calcied cartilage cracks have also been previously iden-
tied in equine C3 on microradiographs
5
and SEM
4,5
but have been
more extensively investigated in the palmar condyles of the meta-
carpal bones
4,6,7
. However, calcied cartilage cracks have not, to our
knowledge, previously been related to OA cartilage disease severity
in naturally occurring repetitive impact OA.
It is very important to emphasize here that there is a debate
whether microcracks observed in the calcied cartilage of decal-
cied specimens are processing artifacts or splits
4,48,49
. Bulk
staining of specimens with basic fuchsin
50,51
or SEM
4
has been
recommended to infer their presence in vivo. Yet, both here and in
a previous study investigating human OA decalcied tissues
45
, few
cracks were located in normal appearing cartilage, whereas the
numbers increased signicantly with OA and were of a similar
magnitude in both studies. Furthermore, the numbers correlated
with the macroscopic extent of the disease. It is interesting that
Muir et al. compared the number of microcracks in the calcied
cartilage, identied with bulk basic fuchsin staining, between
racing and non racing horses and revealed that they increased with
racing
51
. Scanning electron microscopic studies of equine calcied
cartilage have also revealed that healing of microcracks occurs
through mineralization of the cracks and this provides denitive
evidence of their existence in vivo
4,49,52
. The cracks or splits we
observed may, at the very least, reect fragility of this interface
tissue associated with the pathophysiology of disease and could be
a step in the pathway to post-traumatic OA resulting from repeti-
tive loading, but this remains to be elucidated. Future studies to
quantify cracks in calcied cartilage in demineralised parafn
sections of OA tissues with site-matched SEM studies would shed
further light on these issues.
We postulate that the focal microstructural bone resorptive
changes identied in the subchondral bone in the OA groups with
micro-CT are induced by repeated high compressive forces acting
on this dorsal site during athletic activity
53
leading to accumulation
of calci ed tissue microdamage at this site. Although the study of
bone microcracks was not an objective of this investigation, some
calcied cartilage cracks penetrated the subchondral bone.
A topographic relationship has previously been identied between
microcracks in the calcied cartilage and sites of active remodeling
in human
48
and equine
7,8,52,54
OA bone. In experimental models of
impact trauma to the joint these changes have also been associated
with deterioration of the overlying hyaline articular cartilage
2,55
.
Combined these studies suggest a link between calcied tissue
structural damage, bone remodeling and cartilage degradation.
It is well known that cyclic loading of bone causes linear
microscopic cracks in the matrix that may occur faster than the
bones capacity to repair them
48
and induce focal targeted
remodeling
47
. Osteocytes sense these mechanical strains (reviewed
by Bonewald (2011))
56
but also undergo apoptosis beside the linear
microcracks
57e59
and may orchestrate remodelling. However Muir
reported no signicant reduction of osteocyte density in meta-
carpal bones of raced horses, despite a signicant increase in
microcracks, when compared to unraced animals
51
. It is likely that
the events we observe at the surface of equine joints share similar
features with microdamage stress remodeling that has been
studied for years in bone but is necessarily more complex due to the
interplay with overlying hyaline and calcied cartilage in the
injury
11, 41, 47
.
Coalescence of the cracks and collapse of the calcied cartilage
was also observed in more advanced OA in the present study.
Safranin O stained cartilage, was visible in the larger subchondral
bone pits and has also been observed previously in human
45
and
equine
23
tissues. These lesions are very similar to those shown by
Thompson et al., following single impact trauma in a canine
model
2
and include collapse of the subchondral plate, the occa-
sional presence of cartilage in the defect and increased porosity of
the underlying bone. The observation of these focal islands of
cartilage in the bone in both human and animal OA tissues lends
Fig. 8. Histological section of a core from an EOA sample stained with HES (A). Some lacunae contain brous tissue and osteoclasts (arrow) in (B).
M. Lacourt et al. / Osteoarthritis and Cartilage 20 (2012) 572e583580
Page 9
credence to Sokoloffs theory that calcied cartilage microcracks
might induce an ineffective repair involving endochondral ossi-
cation in these tissues and could potentially contribute to disease
progression
45
.
C3 also frequently fractures at the site we investigated. The site
specic remodeling and porosity we observed may lead to a focal
weakness in the bone matrix that could facilitate fracture propa-
gation. A relationship between joint surface microdamage, stress
fractures and subsequent clinical catastrophic fracture through the
joint surface is postulated to exist in equine distal metacarpal
bones
4,8,60
. Muir (2008) has proposed that remodeling targeted at
microcracks may facilitate crack propagation by increasing porosity
of the subchondral plate and could also potentially be responsible
for the lesions we observed
51
. The bone pathology described here
supports the theory that C3 fractures may also be pathological
stress fractures
41
. Combined, the ndings of these investigations of
different equine joints, suggest that racing and training of equine
athletes induces site specic accumulation of microdamage in the
calcied tissues of the joints due to repeated high compressive
stresses
7,8,51
. Together, these events may lead to mechanical failure
of the joint surface. In addition to the structural injury, a parallel
cascade of cellular metabolic events likely occurs in the hyaline
cartilage and underlying bone similar to post-traumatic OA
(recently reviewed by Lotz 2010)
61
.
There are several limitations to this study. An important limi-
tation is that we studied a small number of bones. In addition,
a precise exercise or medication history was not available for the
animals. Consequently, as bone adapts to exercise, varying exercise
regimens could introduce variability in the bone parameters and
the changes observed could, in part, be due to exercise effects and
not exclusively to the OA process. Equally, if intraarticular medi-
cation such as corticosteroids had been administered, osteochon-
dral effects could arise either through pain alleviation and
increased usage or direct effects on bone
62
.
It is important to note that the specimens investigated here
were from young adult equine athletes alone and that the
pathology observed in both cartilage and bone are likely due to
loading history, and not related to age. Consequently, this is not
a model for primary human OA occurring in older human adults
where severe changes in the subchondral bone develop over many
years.
In conclusion, the current data reveals degeneration in hyaline
cartilage and calcied cartilage associated with striking focal
microstructural changes deep in the underlying trabecular bone of
C3 in equine athletes undergoing repetitive compressive loads.
Bone porosity extended deeper into the bone with advancing
cartilage degeneration of the surface, probably due to continued
loading of the microdamaged tissue.
Contributions
Mathieu Lacourt (MatLacourt@aol.com): conception and design,
analysis and interpretation of the data, drafting of the article, nal
approval of the article, collection and assembly of data, provision of
study materials or patients.
Chan Gao (chan.gao@mail.mcgill.ca): Conception and design,
collection and assembly of data, analysis and interpretation of the
data, drafting of the article, nal approval of the article.
Ailian Li (ailian.li@mcgill.ca): Technical support.
Christiane Girard (christiane.girard@umontreal.ca): Analysis
and interpretation of the data drafting of the article, critical revision
of the article for important intellectual content, nal approval of
the article.
Guy Beauchamp (guy.beauchamp@umontreal.ca): Statistical
expertise, Drafting of the article, nal approval of the article.
Janet E Henderson (janet.henderson@mcgill.ca): Drafting of the
article, critical revision of the article for important intellectual
content, nal approval of the article.
Sheila Laverty (sheila.laverty@umontreal.ca): Conception and
design, analysis and interpretation of the data, drafting of the
article, critical revision of the article for important intellectual
content, nal approval of the article.
Competing interest
None of the authors has received any nancial contribution from
commercial sources for this work, nor do we have other nancial
interests that would create a potential conict of interest with
regards to the work.
Acknowledgements
Sheila Laverty is funded by the Canadian Arthritis Network.
Chan Gao is the recipient of awards from the CIHR-MENTOR and RI-
MUHC training programs and Janet E Henderson is the recipient of
funding for trans-disciplinary research from the FRSQ-RSBO. We
acknowledge the assistance of Helene Richard, Dr Judith Farley & Dr
Erin Gillam for assistance with technical aspects of this project.
Supplementary material
Supplementary data related to this article can be found online at
doi:10.1016/j.joca.2012.02.004
References
1. Radin EL, Rose RM. Role of subchondral bone in the initiation
and progression of cartilage damage. Clin Orthop Relat Res
1986;213:34e40.
2. Thompson Jr RC, Oegema Jr TR, Lewis JL, Wallace L. Osteo-
arthrotic changes after acute transarticular load. An animal
model. J Bone Joint Surg Am 1991;73(7):990e1001.
3. Isaac DI, Meyer EG, Kopke KS, Haut RC. Chronic changes in the
rabbit tibial plateau following blunt trauma to the tibiofemoral
joint. J Biomech 2010;43(9):1682e8.
4. Boyde A. The real response of bone to exercise. J Anat 2003;
203(2):173e89.
5.
Young A. Microradiographic and histologic changes in the
third carpal bone of the racing thoroughbred. Masters thesis,
University of California-Davis, 1987.
6. Riggs CM, Boyde A. Effect of exercise on bone density in distal
regions of the equine third metacarpal bone in 2-year-old
thoroughbreds. Equine Vet J Suppl 1999;30:555e60.
7. Norrdin RW, Stover SM. Subchondral bone failure in overload
arthrosis: a scanning electron microscopic study in horses.
J Musculoskelet Neuronal Interact 2006;6(3):251e7.
8. Muir P, McCarthy J, Radtke CL, Markel MD, Santschi EM,
Scollay MC, et al. Role of endochondral ossication of articular
cartilage and functional adaptation of the subchondral plate in
the development of fatigue microcracking of joints. Bone
2006;38(3):342e9.
9. Matsui H, Shimizu M, Tsuji H. Cartilage and subchondral bone
interaction in osteoarthrosis of human knee joint: a histolog-
ical and histomorphometric study. Microsc Res Tech 1997;
37(4):333e42.
10. Bobinac D, Spanjol J, Zoricic S, Maric I. Changes in articular
cartilage and subchondral bone histomorphometry in osteo-
arthritic knee joints in humans. Bone 2003;32(3):284e90.
11. Burr DB, Schafer MB. The involvement of subchondral
mineralized tissues in osteoarthrosis: quantitative microscopic
evidence. Microsc Res Tech 1997;37(4):343e57.
M. Lacourt et al. / Osteoarthritis and Cartilage 20 (2012) 572e583 581
Page 10
12. Boyd SK, Matyas JR, Wohl GR, Kantzas A, Zernicke RF. Early
regional adaptation of periarticular bone mineral density after
anterior cruciate ligament injury. J Appl Physiol 2000;89(6):
2359e64.
13. Batiste DL, Kirkley A, Laverty S, Thain LM, Spouge AR,
Holdsworth DW. Ex vivo characterization of articular cartilage
and bone lesions in a rabbit ACL transection model of osteo-
arthritis using MRI and micro-CT. Osteoarthritis Cartilage
2004;12(12):986e96.
14. Cake MA, Read RA, Appleyard RC, Hwa SY, Ghosh P. The nitric
oxide donor glyceryl trinitrate increases subchondral bone
sclerosis and cartilage degeneration following ovine menis-
cectomy. Osteoarthritis Cartilage 2004;12(12):974e81.
15. Wang SX, Laverty S, Dumitriu M, Plaas A, Grynpas MD. The
effects of glucosamine hydrochloride on subchondral bone
changes in an animal model of osteoarthritis. Arthritis Rheum
2007;56(5):1537e48.
16. Pastoureau PC, Chomel AC, Bonnet J. Evidence of early sub-
chondral bone changes in the meniscectomized guinea pig. A
densitometric study using dual-energy X-ray absorptiometry
subregional analysis. Osteoarthritis Cartilage 1999;7(5):
466e73.
17. Chalmers HJ, Dykes NL, Lust G, Farese JP, Burton-Wurster NI,
Williams AJ, et al. Assessment of bone mineral density of the
femoral head in dogs with early osteoarthritis. Am J Vet Res
2006;67(5):796e800.
18. McErlain DD, Appleton CT, Litcheld RB, Pitelka V, Henry JL,
Bernier SM, et al. Study of subchondral bone adaptations in
a rodent surgical model of OA using in vivo micro-computed
tomography. Osteoarthritis Cartilage 2008;16(4):458e69.
19. Bouchgua M, Alexander K, Carmel EN, dAnjou MA,
Beauchamp G, Richard H, et al. Use of routine clinical multi-
modality imaging in a rabbit model of osteoarthritisepart II:
bone mineral density assessment. Osteoarthritis Cartilage
2009;17(2):197e204.
20. Muraoka T, Hagino H, Okano T, Enokida M, Teshima R. Role of
subchondral bone in osteoarthritis development: a compara-
tive study of two strains of guinea pigs with and without
spontaneously occurring osteoarthritis. Arthritis Rheum
2007;56(10):3366e74.
21. van Dijk CN, Lim LS, Poortman A, Strubbe EH, Marti RK.
Degenerative joint disease in female ballet dancers. Am J
Sports Med 1995;23(3):295e 300.
22. Molloy MG, Molloy CB. Contact sport and osteoarthritis. Br J
Sports Med 2011;45(4):275e 7.
23. Pool RR, Meagher DM. Pathologic ndings and pathogenesis of
racetrack injuries. Vet Clin North Am Equine Pract 1990;6(1):
1e30.
24. Bramlage LR, Schneider RK, Gabel AA. A clinical perspective on
lameness originating in the carpus. Equine Vet J Suppl
1988;6:12e8.
25. Park RD, Morgan JP, OBrien T. Chip fractures in the carpus of
the horse: a radiographic study of their incidence and location.
J Am Vet Med Assoc 1970;157(10):1305e12.
26. Palmer SE. Prevalence of carpal fractures in thoroughbred and
standardbred racehorses. J Am Vet Med Assoc 1986;188(10):
1171e3.
27. McIlwraith CW, Yovich JV, Martin GS. Arthroscopic surgery for
the
treatment of osteochondral chip fractures in the equine
carpus. J Am Vet Med Assoc 1987;191(5):531e40.
28. Murray RC, Henson FMD, Zhu CF, Goodship AE, Agrawal CM,
Athanasiou KA. The effects of strenuous training on equine
carpalarticular cartilage mechanical behaviour and morphology.
Trans Orthop Res Soc 1998;23:200.
29. OBrien T. Third carpal bone lesions of the racing thoroughbred.
Proc Am Ass of Equine Practnrs 1985;31:515e24.
30. Firth EC, van Weeren PR, Pfeiffer DU, Delahunt J, Barneveld A.
Effect of age, exercise and growth rate on bone mineral density
(BMD) in third carpal bone and distal radius of Dutch Warm-
blood foals with osteochondrosis. Equine Vet J Suppl 1999;31:
74e8.
31. Firth EC, Rogers CW. Musculoskeletal responses of 2-year-old
Thoroughbred horses to early training. Conclusions. N Z Vet J
2005;53(6):377e83.
32. Young DR, Richardson DW, Markel MD, Nunamaker DM.
Mechanical and morphometric analysis of the third carpal
bone of Thoroughbreds. Am J Vet Res 1991;52(3):402e9.
33. Ding M, Odgaard A, Hvid I. Changes in the three-dimensional
microstructure of human tibial cancellous bone in early oste-
oarthritis. J Bone Joint Surg Br 2003;85(6):906e12.
34. Day JS, Ding M, van der Linden JC, Hvid I, Sumner DR,
Weinans H. A decreased subchondral trabecular bone tissue
elastic modulus is associated with pre-arthritic cartilage
damage. J Orthop Res 2001;19(5):914e8.
35. Martin M. Guide for determining the age of the horse: American
Association of Equine Practitioners, AAEP; 2002.
36. Stahl R, Jain SK, Lutz J, Wyman BT, Le Graverand-
Gastineau MP, Vignon E, et al. Osteoarthritis of the knee at 3.0
T: comparison of a quantitative and a semi-quantitative score
for the assessment of the extent of cartilage lesion and bone
marrow edema pattern in a 24-month longitudinal study.
Skeletal Radiol 2011;40(10):1315e27.
37. Mankin HJ, Dorfman H, Lippiello L, Zarins A. Biochemical
and metabolic abnormalities in articular cartilage from
osteo-arthritic human hips. II. Correlation of morphology with
biochemical and metabolic data. J Bone Joint Surg Am 1971;
53(3):523e37.
38. Grynpas MD, Alpert B, Katz I, Lieberman I, Pritzker KP. Sub-
chondral bone in osteoarthritis. Calcif Tissue Int 1991;49(1):
20e6.
39. Young BD, Samii VF, Mattoon JS, Weisbrode SE, Bertone AL.
Subchondral bone density and cartilage degeneration patterns
in osteoarthritic metacarpal condyles of horses. Am J Vet Res
2007;68(8):841e9.
40. Olive J, DAnjou MA, Girard C, Laverty S, Theoret CL. Imaging
and histological features of central subchondral osteophytes in
racehorses with metacarpophalangeal joint osteoarthritis.
Equine Vet J 2009;41(9):859e64.
41. Stover SM, Murray A. The California Postmortem Program:
leading the way. Vet Clin North Am Equine Pract 2008;24(1):
21e36.
42. Tidswell HK, Innes JF, Avery NC, Clegg PD, Barr AR,
Vaughan-Thomas A, et al. High-intensity exercise induces
structural, compositional and metabolic changes in cuboidal
bonesendings from an equine athlete model. Bone 2008;
43(4):724e33.
43. Uhlhorn H, Carlsten J. Retrospective study of subchondral
sclerosis and lucency in the third carpal bone of Standardbred
trotters. Equine Vet J 1999;31(6):500e5.
44. Shibakawa A, Yudoh K, Masuko-Hongo K, Kato T, Nishioka K,
Nakamura
H. The role of subchondral bone resorption pits in
osteoarthritis: MMP production by cells derived from bone
marrow. Osteoarthritis Cartilage 2005;13(8):679e87.
45. Sokoloff L. Microcracks in the calcied layer of articular
cartilage. Arch Pathol Lab Med 1993;117(2):191e5.
46. Oegema Jr TR, Carpenter RJ, Hofmeister F, Thompson Jr RC. The
interaction of the zone of calcied cartilage and subchondral
bone in osteoarthritis. Microsc Res Tech 1997;37(4):324e32.
M. Lacourt et al. / Osteoarthritis and Cartilage 20 (2012) 572e583582
Page 11
47. Burr DB, Radin EL. Microfractures and microcracks in sub-
chondral bone: are they relevant to osteoarthrosis? Rheum Dis
Clin North Am 2003;29(4):675e85.
48. Mori S, Harruff R, Burr DB. Microcracks in articular calcied
cartilage of human femoral heads. Arch Pathol Lab Med
1993;117(2):196e8.
49. Boyde A, Riggs CM, Bushby AJ, McDermott B, Pinchbeck GL,
Clegg PD. Cartilage damage involving extrusion of miner-
alisable matrix from the articular calcied cartilage
and subchondral bone. Eur Cell Mater 2011;21:470e8.
Discussion 8.
50. Frost H. Presence of microscopic cracks in vivo in bone. Henry
Ford Hosp Med Bull 1960;8:25e35.
51. Muir P, Peterson AL, Sample SJ, Scollay MC, Markel MD,
Kalscheur VL. Exercise-induced metacarpophalangeal joint
adaptation in the Thoroughbred racehorse. J Anat 2008;
213(6):706e17.
52. Boyde A, Firth EC. High resolution microscopic survey of third
metacarpal articular calcied cartilage and subchondral bone
in the juvenile horse: possible implications in chondro-
osseous disease. Microsc Res Tech 2008;71(6):477e88.
53. Palmer JL, Bertone AL, Litsky AS. Contact area and pressure
distribution changes of the equine third carpal bone during
loading. Equine Vet J 1994;26(3):197e202.
54. Boyde A, Firth EC. Musculoskeletal responses of 2-year-old
Thoroughbred horses to early training. 8. Quantitative back-
scattered electron scanning electron microscopy and confocal
uorescence microscopy of the epiphysis of the third meta-
carpal bone. N Z Vet J 2005;53(2):123e32.
55. Vener MJ, Thompson Jr RC, Lewis JL, Oegema Jr TR. Sub-
chondral damage after acute transarticular loading: an in vitro
model of joint injury. J Orthop Res 1992;10(6):759e65.
56. Bonewald LF. The amazing osteocyte. J Bone Miner Res
2011;26(2):229e38.
57. Noble BS, Peet N, Stevens HY, Brabbs A, Mosley JR, Reilly GC,
et al. Mechanical loading: biphasic osteocyte survival and
targeting of osteoclasts for bone destruction in rat cortical
bone. Am J Physiol Cell Physiol 2003;284(4):C934e43.
58. Verborgt O, Gibson GJ, Schafer MB. Loss of osteocyte integrity
in association with microdamage and bone remodeling after
fatigue in vivo. J Bone Miner Res 2000;15(1):60e7.
59. Verborgt O, Tatton NA, Majeska RJ, Schafer MB. Spatial
distribution of Bax and Bcl-2 in osteocytes after bone fatigue:
complementary roles in bone remodeling regulation? J Bone
Miner Res 2002;17(5):907e14.
60. Radtke CL, Danova NA, Scollay MC, Santschi EM, Markel MD,
Da Costa Gomez T, et al. Macroscopic changes in the distal
ends of the third metacarpal and metatarsal bones of Thor-
oughbred racehorses with condylar fractures. Am J Vet Res
2003;64(9):1110e6.
61. Lotz MK, Otsuki S, Grogan SP, Sah R, Terkeltaub R, DLima D.
Cartilage cell clusters. Arthritis Rheum 2010;62(8):2206e18.
62. Weinstein RS. Clinical practice. Glucocorticoid-induced bone
disease. N Engl J Med 2011;365(1):62e70.
M. Lacourt et al. / Osteoarthritis and Cartilage 20 (2012) 572e583 583
Page 12
    • "Future studies evaluating MMP and VDR, in this same animal model, will be necessary to better understand the role of vitamin D on articular cartilage. Different studies have shown subchondral bone changes in osteoarthritis [12, 19]. In our study, we did not observe significant differences in the subchondral bone between the groups. "
    [Show abstract] [Hide abstract] ABSTRACT: Background Vitamin D appears to play an important role in bone and cartilage metabolism since its receptors are widely found in human articular chondrocytes. Thus, effects of variation of vitamin D may directly impact cartilage and bone biology. Questions/Purposes The aims of this study are to compare (1) articular cartilage structure and composition and (2) trabecular and cortical bone microstructure in rats with normal versus insufficient vitamin D levels. Methods Twenty-five mature, male Sprague-Dawley rats were allocated to two groups: (1) control arm (vitamin D replete—12 rats) and (2) an experimental arm (vitamin D deficient—13 rats). Vitamin D deficiency was induced using a vitamin D-deficient diet and UV light restriction. Rats were sacrificed after 4 weeks vitamin D deficiency was confirmed. The right knee was harvested for analysis of both the medial (MFC) and lateral femoral condyles (LFC). A region of interest was established on both condyles to correlate subchondral bone architecture and the overlying cartilage. Histological analysis was performed and graded using the modified Mankin score. Subchondral and cortical bony architecture was evaluated with micro-CT. Results After 4 weeks, the vitamin D-deficient group had statistically significant changes in cartilage structure in both the MFC and LFC [1.55 ± 0.6 vs. 4.23 ± 4.1 (p = 0.035) and 1.55 ± 0.6 vs. 3.53 ± 2.4 (p = 0.009), respectively]. Micro-CT analysis revealed no correlation between subchondral bone values and the overlying cartilage Mankin score (p = 0.460). No significant difference was evident between the subchondral bone of the control and study group. Conclusions Low levels of vitamin D have a deleterious effect on the cartilage. Given the high prevalence of vitamin D deficiency in the general population, these findings raise important questions about the potential role of vitamin D in articular cartilage health.
    Full-text · Article · Mar 2016 · HSS Journal
    • "Tissue was harvested at a local abattoir from Standardbred racehorses with a previous racing career, as previously described [4]. A scoring system of the macroscopic cartilage degeneration in the third carpal bones was used to create control (C), early osteoarthritis (EOA) and advanced OA (AOA) groups. "
    [Show abstract] [Hide abstract] ABSTRACT: We studied changes in articular calcified cartilage (ACC) and subchondral bone (SCB) in the third carpal bones (C3) of Standardbred racehorses with naturally-occurring repetitive loading-induced osteoarthritis (OA). Two osteochondral cores were harvested from dorsal sites from each of 15 post-mortem C3 and classified as control or as showing early or advanced OA changes from visual inspection. We re-examined X-ray micro-computed tomography (µCT) image sets for the presence of high-density mineral infill (HDMI) in ACC cracks and possible high-density mineralized protrusions (HDMP) from the ACC mineralizing (tidemark) front (MF) into hyaline articular cartilage (HAC). We hypothesized and we show that 20-µm µCT resolution in 10-mm diameter samples is sufficient to detect HDMI and HDMP: these are lost upon tissue decalcification for routine paraffin wax histology owing to their predominant mineral content. The findings show that µCT is sufficient to discover HDMI and HDMP, which were seen in 2/10 controls, 6/9 early OA and 8/10 advanced OA cases. This is the first report of HDMI and HDMP in the equine carpus and in the Standardbred breed and the first to rely solely on µCT. HDMP are a candidate cause for mechanical tissue destruction in OA.
    Full-text · Article · May 2015 · International Journal of Molecular Sciences
    • "ndrocyte death associated with mechanically injured regions (Lewis et al. 2003; K€ uhn et al. 2004; Novakofski et al. 2014). In a series of in vitro studies of bovine cartilage, Broom and his colleagues have shown that the mode of fracture of HAC and ACC under high loading rates is influenced by previous static creep loading (Thambyah et al. 2012). Lacourt et al. (2012) used both microtomography and decalcified section to quantify cracks in the equine third carpal bone as a natural model of repetitive injuryinduced arthritis. X-ray microtomography was introduced to the bone field by Elliott & Dover (1982, 1984). Although there have been several relevant XMT studies of equine and human joints to date, n"
    [Show abstract] [Hide abstract] ABSTRACT: High density mineralised protrusions (HDMP) from the tidemark mineralising front into hyaline articular cartilage (HAC) were first described in Thoroughbred racehorse fetlock joints and later in Icelandic horse hock joints. We now report them in human material. Whole femoral heads removed at operation for joint replacement or from dissection room cadavers were imaged using magnetic resonance imaging (MRI) dual echo steady state at 0.23 mm resolution, then 26-μm resolution high contrast X-ray microtomography, sectioned and embedded in polymethylmethacrylate, blocks cut and polished and re-imaged with 6-μm resolution X-ray microtomography. Tissue mineralisation density was imaged using backscattered electron SEM (BSE SEM) at 20 kV with uncoated samples. HAC histology was studied by BSE SEM after staining block faces with ammonium triiodide solution. HDMP arise via the extrusion of an unknown mineralisable matrix into clefts in HAC, a process of acellular dystrophic calcification. Their formation may be an extension of a crack self-healing mechanism found in bone and articular calcified cartilage. Mineral concentration exceeds that of articular calcified cartilage and is not uniform. It is probable that they have not been reported previously because they are removed by decalcification with standard protocols. Mineral phase morphology frequently shows the agglomeration of many fine particles into larger concretions. HDMP are surrounded by HAC, are brittle, and show fault lines within them. Dense fragments found within damaged HAC could make a significant contribution to joint destruction. At least larger HDMP can be detected with the best MRI imaging ex vivo.
    Full-text · Article · Oct 2014 · Journal of Anatomy
    A Boyde A Boyde G.R. Davis G.R. Davis D. Mills D. Mills +9 more authors... T. Zikmund T. Zikmund
Show more

Similar publications

Discover more