Bull Vet Inst Pulawy 51, 621-626, 2007
INTERRELATIONSHIPS BETWEEN DENSITOMETRIC,
MORPHOMETRIC, AND MECHANICAL PROPERTIES
OF THE TIBIA IN TURKEYS
WITOLD KRUPSKI, AND MARCIN R. TATARA1
II Department of Radiology, Medical University of Lublin, 20–081 Lublin, Poland
1
Department of Animal Physiology, Faculty of Veterinary Medicine,
Agricultural University of Lublin, 20–950 Lublin, Poland
[email protected]
Received for publication October 04, 2007
Abstract
The aim of the study was to evaluate the
interrelationships between parameters assessed with the use of
a computed tomography technique and mechanical properties
of the tibia in male turkeys at different stages of skeletal
development. Healthy male turkeys (N = 120) were kept under
standard rearing conditions and randomly selected from the
farm flock at the age of 4, 8, 12, and 20 weeks of life to obtain
a right tibia for the investigations. Using a computed
tomography technique, volumetric bone mineral density of the
trabecular and cortical bone, mean volumetric bone mineral
density, total bone volume, and cortical bone area of the tibia
were estimated. Geometrical properties, such as cross-sectional
area, second moment of inertia, mean relative wall thickness,
and cortical index, were derived from computed tomography
measurements of horizontal and vertical diameters of the tibia
in the midshaft. Using a three-point bending test, the
mechanical parameters were estimated in terms of maximum
elastic strength and ultimate strength of the tibia. A Pearson’s
correlation coefficient was determined for all the investigated
variables. The obtained results showed age-related changes of
the skeletal system in growing turkeys in terms of
morphological, densitometric, and mechanical properties.
Positive correlations among the investigated parameters were
found, except for mean relative wall thickness and cortical
index, which were negatively correlated with these parameters.
Morphometric and densitometric parameters assessed with the
use of computed tomography were significantly correlated
with maximum elastic strength and ultimate strength proving
the usefulness of this technique for predicting mechanical
properties of bones. In conclusion, this study showed
computed tomography as a precise and non-invasive technique
for the determination of skeletal system properties that may
serve to monitor in vivo the dynamics of the changes of bone
metabolism in studies on physiological, nutritional, and
pharmacological factors influencing the skeletal system.
Key words: turkey, tibia, bone mineral
density, mechanical endurance, quantitative computed
tomography.
Bone tissue plays supportive, locomotory,
protective, and metabolic functions in vertebrates. All
these functions are crucial for an effective production of
animals and optimal quality of life in humans, especially
in the elderly (8, 10, 17). Mechanical properties of bones
depend on many factors, among of which bone
geometry, bone quantity, and quality in terms of mineral
content, microarchitecture, microdamages, as well as
collagen structure and maturation belong to the most
important ones (11). During skeletal growth and during
aging of the skeleton, bone mechanical endurance is
related to constant changes of morphological properties
and bone mineral density. An increased bone size and
mineral density were reported as essential factors
improving the bone’s mechanical properties, whereas its
decreased values induced opposite effects (6, 12, 14, 15).
Bone mineral density determined with the use of dualenergy X-ray absorptiometry (DEXA) method was
reported as a good predictor for the vertebral fracture risk
(7, 18).
Computed tomography (CT), is a highly
advanced technique of diagnostic imaging invented by
Sir Godfrey Hounsfield in 1972 in Great Britain using
the theoretical basis provided by Allan Cormack. The
importance of the discovery of this diagnostic technique
was underlined by the Nobel Prize in Physiology or
Medicine for Hounsfield and Cormack in 1979. This
method provides highly precise imaging of anatomical
structures in both humans and animals with the
possibility for analysis of tissues and organs in series of
cross-sectional slices. As opposed to X-ray radiographs
showing two-dimensional anatomical structures,
advanced software of CT scanners enables the
conversion of obtained digital data into multiplanar
reconstructions (MPR) of the investigated structures and
its analysis at any trajectory. Moreover, three
dimensional (3D) reconstructions with the use of surface
shaded display (SSD) technique or volumetric analysis
using volume rendering technique (VRT) of organs may
be performed (17). Quantitative computed tomography
622
(QCT) is a highly respected and non-invasive method
for volumetric bone mineral density determination in
any part of the skeleton. In contrast to DEXA method,
which enables areal determination of the bone’s mineral
density (expressed in g/cm2), QCT allows volumetric
analysis of trabecular and cortical bone density
(expressed in g/cm3), independent of one another.
Another advantage of QCT method for the
determination of volumetric bone mineral density
(vBMD), in comparison with DEXA method is the fact
that with the use of this method, vBMD can be easy
measured without errors resulting from surrounding soft
tissues and possible osteoarthritic changes (6, 9).
The aim of this study was to evaluate the
interrelationships between parameters assessed with the
use of computed tomography technique and mechanical
properties of the tibia obtained from male turkeys at
different stages of skeletal development, namely 4, 8,
12, and 20 weeks of life. To achieve this goal, Pearson’s
correlation coefficient was estimated between such
variables as bone weight, bone length, vBMD of the
trabecular and cortical bone, mean vBMD, total bone
volume, cortical bone area, cross-sectional area, second
moment of inertia, mean relative wall thickness,
maximum elastic strength, and ultimate strength.
Material and Methods
Experimental
design
and
sampling
procedure. The study was performed on 120 healthy T5
male turkeys (Holly Berry Hatcheries Ltd., UK),
randomly selected from the farm flock at different stages
of their development, and divided into 4 agedifferentiated groups. The first group (N = 30) consisted
of turkeys slaughtered at the age of 4 weeks while to the
second group belonged 8-week-old animals (N = 30).
Turkeys slaughtered at the age of 12 and 20 weeks
belonged to the third (N = 30) and fourth (N = 30)
groups, respectively. During the course of the study, all
the animals were kept under standard rearing conditions
with constant access to fresh water and appropriate feed
supplied ad libitum in accordance with several stages of
the production cycle.
Morphological examination of the tibia.
Immediately after post-mortem isolation of right tibiae,
bone samples were cleaned from remaining soft tissues,
their weight and length were measured, and they were
frozen at –25ºC until further analyses.
Computed tomography evaluation of the
tibia. The bone samples were thawed for 2 h at room
temperature before being scanned with the use of
computed tomography technique in SOMATOM
EMOTION SIEMENS apparatus supplied with
Somaris/5 VB10B software (Version B10/2004A,
Siemens, Germany). All bone samples were scanned
using 2-mm thick, sequential, cross-sectional scans.
Using quantitative computed tomography method, the
volumetric bone mineral density (vBMD) of the
trabecular and cortical bone was determined. Trabecular
bone (Td) and cortical bone (Cd) mineral densities were
determined using 2-mm thick QCT scans placed in the
distal metaphysis and at 50% of the tibia length,
respectively. While Cd was determined in all groups of
turkeys, Td of the tibia was measured in 12 and 20week-old birds only. Furthermore, the cortical bone area
(ACb) on cross-section at 50% of tibia length was
measured in all the groups of turkeys. Using Volume
Evaluation application package (software Version
B10/2004), the total bone volume (Bvol) of each tibia
was determined automatically. For Bvol calculations, the
volume-of-interest (VOI) was defined by limiting the
minimum and maximum density for the investigated
bone at 60 and 3000 Hounsfield units (HU),
respectively. Volume Evaluation application package
was used also to determine the mean volumetric bone
mineral density (MvBMD) for each tibia.
Analysis of geometrical properties of the
tibia. On the basis of measurements of horizontal and
vertical diameters (both internal and external) performed
on CT mid-diaphyseal cross-section of the tibia, the
geometrical parameters such as cross-sectional area (A),
second moment of inertia (Ix), mean relative wall
thickness (MRWT), and cortical index were derived (2,
4, 5, 16).
Analysis of mechanical properties of the
tibia. Mechanical properties of tibia were estimated with
the use of the three-point bending test in INSTRON
3369 apparatus (Instron, Canton, USA) linked with a
computer registering the relationship between forces
perpendicular to the longitudinal axis of the bone and
the resulting displacement (4, 5). The distance between
bone supports was set at 40% of total tibia length and
the crosshead loaded bone samples at the midshaft with
constant speed of 20 mm/min. The values of maximum
elastic strength (Wy) and ultimate strength (Wf) were
obtained.
Statistical analysis. Statistical analysis was
performed using Statistica software (version 6.0). All the
data were presented as means ± SEM. The differences of
mean values among the age-differentiated groups were
tested for statistical significance with the use of one-way
ANOVA and post hoc Duncan’s test. Pearson’s
correlation coefficient was determined between all the
investigated variables. Differences showing P<0.05 were
considered statistically significant.
Results
Turkeys at the age of 4, 8, 12, and 20 weeks of
life reached body weight values of 1.17 ± 0.01 kg, 4.08 ±
0.11 kg, 8.53 ± 0.17 kg, and 14.96 ± 0.32 kg, respectively.
Statistically significant differences of body weight values
were found among all the investigated groups of turkeys
(P<0.001). The results of morphometric, densitometric,
and mechanical analyses of the tibia are shown in Table
1. The lowest values of bone length, bone weight,
trabecular bone mineral density, cortical bone mineral
density, mean volumetric bone mineral density, total
bone volume, cortical bone area, cross-sectional area,
second moment of inertia, maximum elastic strength,
and ultimate strength were in 4-week-old turkeys,
followed by the groups of 8 and 12-week-old birds.
623
Table 1
Morphometric, densitometric, and mechanical properties of the tibia of turkeys at different stages of their development
Parameter
Age of turkeys
Bone weight (g)
Bone length (mm)
Trabecular bone mineral density (g/cm3)
Cortical bone mineral density (g/cm3)
Mean volumetric bone mineral density (g/cm3)
Total bone volume (cm3)
Cortical bone area (mm2)
Cross-sectional area (mm2)
Second moment of inertia (mm4)
Mean relative wall thickness
Cortical index
Maximum elastic strength (N)
Ultimate strength (N)
4 weeks
10.6 a ± 0.2
104.8 a ± 0.5
–
1.939 a ± 0.025
1.333 a ± 0.005
6.3 a ± 0.1
14.3 a ± 0.3
16.7 a ± 0.3
41.3 a ± 1.2
0.613 a ± 0.020
37.6 a ± 0.7
106 a ± 6
193 a ± 5
8 weeks
48.8 b ± 0.8
176.5 b ± 0.9
–
2.190 b ± 0.026
1.390 b ± 0.006
27.9 b ± 0.5
39.5 b ± 1.5
44.5 b ± 2.5
408.9 b ± 43.0
0.394 b ± 0.018
27.6 b ± 1.2
370 b ± 21
542 b ± 28
12 weeks
86.1 c ± 1.1
221.7 c ± 1.2
1.194 a ± 0.008
2.221 b ± 0.024
1.436 c ± 0.011
48.2 c ± 0.7
62.1 c ± 2.5
67.6 c ± 2.2
1139.0 c ± 57.0
0.301 c ± 0.009
23.0 c ± 0.5
547 c ± 32
913 c ± 48
20 weeks
116.3 d ± 2.0
251.6 d ± 1.2
1.243 b ± 0.005
2.300 c ± 0.023
1.521 d ± 0.008
60.0 d ± 1.1
74.4 d ± 1.3
81.7 d ± 1.3
2004.2 d ± 67.3
0.251 d ± 0.006
19.9 d ± 0.4
834 d ± 24
1212 d ± 28
abcd
Values within a row that do not share common superscript letter differ significantly for P<0.05.
The highest values of these parameters were
obtained in 20-week-old turkeys. The lowest values of
mean relative wall thickness and cortical index were
found in 20-week-old turkeys, followed by the groups of
12- and 8-week-old turkeys. The highest values of
MRWT and CI were found in turkeys at the age of 4
weeks (Table 1). The values of Pearson’s correlation
coefficient between all the investigated parameters are
shown in Table 2. Significantly positive correlations
were found between such parameters as bone weight,
bone length, Td, Cd, MvBMD, Bvol, ACb, A, Ix, Wy, and
Wf. A positive correlation was also found between
MRWT and CI (r = 0.982); however, both these
parameters were negatively correlated with all the other
investigated parameters (Table 2).
Discussion
The growth and development of the skeletal
system in mammals and birds until the achievement of
peak bone mass (PBM) is characterised by changes in
bone length, bone geometry, and bone mineral density.
The Bone’s length, geometrical parameters, and bone
mineral density, increase with the age until the end of
the skeletal maturation process when the maximum
amount of bone tissue is reached. After this event, the
bone mass of the skeleton results from the amount of
bone acquired at skeletal maturity and age-related bone
loss (12, 13). In the current study, significant changes of
the investigated parameters of the tibia among the agedifferentiated groups of turkeys were demonstrated in
accordance with body weight gain. Bone weight, bone
length, mean volumetric bone mineral density, total
bone volume, cortical bone area, cross-sectional area,
and second moment of inertia were significantly
different at each of the investigated stages of skeletal
development of the turkeys and a gradual increase in
these values with the age was observed. Similar changes
were observed when volumetric bone mineral density of
the trabecular and cortical bone was analysed. As the
consequence of age-related increase in bone size, mainly
in external dimensions of the tibia, the values of mean
relative wall thickness and cortical index showed regular
decrease in accordance with the increasing age of the
turkeys. All these age-related changes of the
morphometric and densitometric properties of the tibia
resulted in gradually increasing values of the mechanical
parameters, such as maximum elastic strength and
ultimate strength.
The
obtained
results
also
showed
interrelationships between morphometric, densitometric,
and mechanical properties of the tibia in the turkeys.
Positive correlations were found among bone weight,
bone length, total bone volume, cortical bone area,
cross-sectional area, second moment of inertia and
mechanical parameters of the tibia, such as maximum
elastic strength and ultimate strength, clearly indicate
that developmental changes of the skeletal system,
which increase in all these morphometrical parameters,
improve mechanical endurance of bones to withstand
higher skeletal loading related to body weight gain.
Positive correlations of trabecular bone mineral density,
cortical bone mineral density, and mean volumetric bone
mineral density with maximum elastic strength and
ultimate strength of the tibia show significant
contribution of bone mineralisation to mechanical
properties of bones. The fact that MvBMD and Cd seem
to be better predictors of bone mechanical properties
than Td is noteworthy. This fact may be explained by
different metabolic activity and functions of both these
bone compartments in the skeleton (3). Metabolic
turnover of the trabecular bone is very high when
compared to the cortical bone. While trabecular bone
compartment ensures resistance to compressive stress,
cortical bone, characterised by higher bone mineral
density, is mainly responsible for withstanding bending
forces (1).
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Table 2
The values of Pearson’s correlation coefficient among all the investigated parameters of turkey tibia
Bone weight
Bone length
Td*
Cd
MvBMD
Bvol
ACb
A
Ix
MRWT
CI
Wy
Wf
Parameter
Bone weight
x
0.981
0.522
0.677
0.830
0.994
0.944
0.939
0.945
– 0.829
– 0.814
0.897
0.923
Bone length
0.981
x
0.499
0.701
0.808
0.986
0.934
0.930
0.885
– 0.862
– 0.840
0.882
0.901
Td*
0.522
0.499
x
0.334
0.571
0.521
0.316
0.398
0.506
– 0.286
– 0.293
0.505
0.413
Cd
0.677
0.701
0.334
x
0.827
0.695
0.704
0.730
0.628
– 0.557
– 0.512
0.727
0.744
MvBMD
0.830
0.808
0.571
0.827
x
0.818
0.874
0.859
0.854
– 0.611
– 0.593
0.893
0.887
Bvol
0.994
0.986
0.521
0.695
0.818
x
0.945
0.939
0.926
– 0.838
– 0.820
0.890
0.919
ACb
0.944
0.934
0.316
0.704
0.874
0.945
x
0.960
0.911
– 0.740
– 0.715
0.909
0.937
A
0.939
0.930
0.398
0.730
0.859
0.939
0.960
x
0.935
– 0.695
– 0.652
0.911
0.940
Ix
0.945
0.885
0.506
0.628
0.854
0.926
0.911
0.935
x
– 0.690
– 0.675
0.901
0.915
MRWT
– 0.829
– 0.862
– 0.286
– 0.557
– 0.611
– 0.838
– 0.740
– 0.695
– 0.690
x
0.982
– 0.683
– 0.704
CI
– 0.814
– 0.840
– 0.293
– 0.512
– 0.593
– 0.820
– 0.715
– 0.652
– 0.675
0.982
x
– 0.665
– 0.681
Wy
0.897
0.882
0.505
0.727
0.893
0.890
0.909
0.911
0.901
– 0.683
– 0.665
x
0.965
Wf
0.923
0.901
0.413
0.744
0.887
0.919
0.937
0.940
0.915
– 0.704
– 0.681
0.965
x
*Pearson’s correlation coefficient between trabecular bone mineral density and all the investigated parameters of the tibia was estimated for the values obtained from 12- and 20week-old turkeys.
625
However, MvBMD value representing mineral
density for both the trabecular and cortical bone
compartments measured within whole bone sample,
reached the highest correlation with maximum elastic
strength and ultimate strength of the tibia. The results
obtained in the current study are in accordance with data
presented by Ferretti et al. (5), which have shown
positive correlations between cross-sectional area and
moment of inertia with stiffness and strength of the
femur in rats. In contrast to our results showing negative
correlations between MRWT and all other
morphometric,
densitometric,
and
mechanical
parameters (except for cortical index) of the tibia, the
mentioned above authors have shown positive
correlation of this parameter with bone stiffness and
strength of the femur. However, their study was
performed on rats of both sexes at the age between 18
and 44 weeks. Clearing, morphological differences of
the skeleton conditioned by bone modelling and
remodelling related to the periods of skeletal
development and aging may explain this discrepancy.
Negative correlations of MRWT and CI with all other
investigated parameters of the tibia point to an inverse
relationship of both these parameters with other
morphometrical, densitometric, and mechanical
properties in the growing turkeys.
Considering significant positive correlations
among parameters such as Td, Cd, MvBMD, Bvol, ACb,
A, Ix, obtained using computed tomography technique,
with the values of Wy and Wf, computed tomography
may be postulated as a useful tool not only for the
determination of volumetric bone mineral density but
also for predicting bone mechanical properties.
Computed tomography technique also makes possible a
precise determination of bone geometrical properties
and total bone volume – the parameters strictly
reflecting bone morphological properties. The data
obtained from densitometric and morphometric analyses
may
be
completed
with
three-dimensional
reconstructions of the investigated bone. Furthermore,
non-invasive imaging of the internal structure of a bone
at any projection may be easily performed. All these
data indicate that computed tomography technique has
great value for not only investigations of
morphometrical and densitometric properties of bones
isolated ex vivo or in vivo, but it may also be used to
predict bone mechanical endurance. Advantages of this
technique give the possibility for precise and noninvasive monitoring of skeletal system properties in vivo
during different stages of the development of animals,
which is important for optimal breeding selection. Due
to repeated assessment of skeletal properties in
experimental animals, this technique may serve to
monitor dynamics of the changes of bone metabolism in
studies aimed at physiological, nutritional, and
pharmacological factors influencing the skeletal system.
Acknowledgments: Dr Marcin Tatara was
awarded a scholarship for young scientists from the
Foundation for Polish Science in 2006-2007.
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