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Biological Journal of the Linnean Society, 2014, 112, 765–781. With 16 figures
The three-front model: a developmental explanation of
long bone diaphyseal histology of Sauropoda
JESSICA MITCHELL* and P. MARTIN SANDER
Steinmann-Institut für Geologie, Mineralogie und Paläontologie, Universität Bonn, Nussallee 8,
53115 Bonn, Germany
Received 16 December 2013; revised 24 March 2014; accepted for publication 31 March 2014
The bone histology of non-avian dinosaurs is enlightening for understanding many aspects of the growth and
development of these long-extinct animals. The rate of bone apposition and remodelling in the shaft of long bones
appears to be accelerated in some groups and decelerated in others. We propose a developmental model to illustrate
these fundamental aspects of long bone diaphyseal histology at different growth stages. We developed the model
based on an ontogenetic series of long bones of the sauropod dinosaur Apatosaurus. The model describes the
histology and microanatomy based on three fronts that move radially: the apposition front, the Haversian
substitution front, and the resorption front. When applied to additional sauropod dinosaurs, differences and
similarities observed in the microstructure of the different taxa could be explained with the model. The benefit of
this model is that it is not limited to Sauropoda but appears to be applicable to a broad range of terrestrial amniote
long bones and thus could provide unique insights into life history and evolutionary patterns of bone development. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781.
ADDITIONAL KEYWORDS: apposition front – Haversian substitution front – palaeohistology – resorption
front.
INTRODUCTION
Bone histology has been a major asset to understanding growth and development of dinosaurs (de Ricqlès,
1968; Reid, 1987; Curry, 1999; Starck & Chinsamy,
2002; Padian, Horner & de Ricqlès, 2004; Erickson,
2005; Klein & Sander, 2008; Lehman & Woodward,
2008; Hayashi, Carpenter & Suzuki, 2009; Stein
et al., 2010; Klein et al., 2012; Griebeler, Klein &
Sander, 2013). Over the past four decades, research
has shown that dinosaurs have a unique growth
pattern compared to living organisms but are most
similar to mammals in histology, as opposed to
modern reptiles. We have come to understand that
dinosaurs are most likely endothermic, having fast
growth rates as juveniles and a determinate growth
pattern, reaching reproductive maturity before skeletal maturity (Sander, 2000; Erickson, 2005; Gillooly,
Allen & Charnov, 2006; Lee et al., 2013).
*Corresponding author. E-mail: [email protected]
PRINCIPLES
OF BONE SHAFT GROWTH: THREE FRONTS
When studying ontogenetic series, it becomes clear
that periosteal bone development (i.e. growth in diameter) begins at the innermost cortex and moves
outward. This is obtained through apposition of new
bone on the periosteal surface, forming what we will
refer to as the apposition front (AF). As new bone is
being deposited, the cortex is being resorbed along the
endosteal surface, maintaining a biomechanically
ideal cortical thickness through development (Currey,
2002). This is what we term the resorption front (RF).
Haversian substitution also begins in the inner cortex
and moves outwards in a more or less well defined
Haversian substitution front (HSF). Together, these
three fronts can describe the progression of bone
formation, Haversian substitution, and resorption
over time. The apposition and remodelling fronts
occur in all long bones of all tetrapods; however, the
Haversian substitution front, is not always present
(e.g. in amphibians and crocodiles). Each front progresses at a different rate and influences the histology
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
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J. MITCHELL and P. M. SANDER
and microanatomy of the specimen. The present study
develops a model, using sauropod dinosaurs as model
organisms, that describes the interplay of the AF,
HSF, and RF, which are responsible for the histology
and microanatomy observed in the midshaft crosssection of the long bones of dinosaurs and large
mammals.
Three-front models are thus used to visualize these
fundamental aspects of bone development in the middiaphysis of sauropod long bones and to visualize the
similarities and differences between taxa. Put into a
phylogenetic framework, the comparison of threefront models informs us about evolutionary questions,
such as changes in body size and life history.
Specifically, we studied a growth series of the
diplodocoid Apatosaurus and compared it with
the growth series of an indeterminate Tendaguru
diplodocid and the basal macronarians Giraffatitan
and Camarasaurus. All share very similar histologies,
as already noted by Klein & Sander (2008) and
Sander et al. (2011), although the three-front models
greatly aid in formalizing and visualizing this similarity. Most importantly, three-front models provide
the developmental explanation for this similarity. The
comparison is then extended to individuals of the
dwarf sauropods Europasaurus and Magyarosaurus
and of the slow-growing titanosaur Ampelosaurus.
Finally, we ‘spot-check’ the usefulness of three-front
models for describing mammalian bone histology
based on a Cervus sample. This work suggests that
three-front models will be applicable to dinosaurs and
mammals in general.
DINOSAUR
HISTOLOGY
Primary cortical bone histology
To understand three-front models, background information on dinosaur bone histology and principles of
bone tissue formation and remodelling are necessary,
particularly to estimate the speed of the fronts.
The discovery that bone tissue type is linked to
local apposition rate was made by Amprino (1947)
and is known as Amprino’s rule (Cubo et al., 2012).
Although it has been difficult to quantify, it is clear
that the two major bone tissue types, lamellar zonal
bone and fibrolamellar bone, differ by an order of
magnitude in their apposition rates. Observed apposition rates for lamellar zonal bone vary from 1 to
10 μm day−1, whereas those of fibrolamellar bone
exceed 10 μm day−1 and may be as high as
171 μm day−1 (Castanet et al., 2000; de Margerie,
Cubo & Castanet, 2002; de Margerie et al., 2004;
Cubo et al., 2012).
Dinosaur bone histology is dominated by highly
vascularized fibrolamellar bone. Fibrolamellar bone
consists of a framework of woven bone, in which
vascular canals are centripetally filled in with either
lamellar or parallel-fibred bone (i.e. primary osteons)
(Francillon-Vieillot et al., 1990). In some cases, such
as in Ampelosaurus, this woven framework is
replaced with parallel-fibred bone (Klein et al., 2012;
Stein & Prondvai, 2013). The vascular organization varies as well. Laminar fibrolamellar bone
is dominated by circumferential vascular canals;
plexiform fibrolamellar bone has radial and circumferential canals; and reticular fibrolamellar bone
has an irregular arrangement of canals (de Ricqlès,
1975; Francillon-Vieillot et al., 1990; Huttenlocker,
Woodward & Hall, 2013). Typically, as the individual
ages, the bone becomes less vascularized and a relative increase occurs in parallel-fibred and lamellar
bone. Lines of arrested growth (LAGs), useful for
determining age of individuals and growth rates, are
a common observation in the cortices of ectotherms
(Castanet et al., 1993), some mammals (Horner, de
Ricqlès & Padian, 1999; Sander & Andrassy, 2006;
Köhler et al., 2012), and many dinosaurs (Erickson,
2005), with sauropods being the exception (Sander
et al., 2011). When full size is attained, closely spaced
LAGs can usually be observed in lamellar bone in the
outermost cortex known as the external fundamental
system (EFS; Ham, 1953).
Because sauropod dinosaurs lack LAGs, age and
growth rates are difficult or impossible to assess and
compare between taxa. This is why Klein & Sander
(2008) developed histological ontogenetic stages
(HOS) for Neosauropoda to describe the change in
tissue type and vascular organization from small to
large specimens. Sexual maturity was determined to
be around HOS 8 and full size was reached at HOS 12
(indicated by an EFS). The final stage, HOS 13, is
described as complete Haversian bone. Stein et al.
(2010) added an additional stage, HOS 14, to distinguish the highly remodelled bone of Magyarosaurus.
Griebeler et al. (2013) were able to quantify the sauropod growth record, and determined that sauropods
had fast growth rates similar to megaherbivores and
ratites.
Remodelling
Bone differs from other hard tissues, such as enamel
and mollusc shell, in that it is a living tissue and can
be modified after deposition by a process known as
remodelling. Remodelling consists of the resorption
of bone tissue followed by redeposition; within the
cortex, secondary osteons form, which are also known
as Haversian systems.
Secondary osteons result from two cellular components, the osteoclasts and osteoblasts, known collectively as a basic multicellular unit (BMU; Frost,
1969). Osteoclasts form a resorption cavity, forming a
tunnel in cortical bone. Osteoblasts then centripetally
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
THREE-FRONT MODEL OF LONG BONE HISTOLOGY
secrete lamellar bone layers around a vascular canal
(Haversian canal). Typically in sauropods, secondary
osteons do not exceed 400 μm in diameter (Mitchell,
2012). The secondary osteon forms along the path of
least strain, which is the longitudinal direction in
long bones (van Oers et al., 2008).
A prominent theory suggests that the activation of
the BMU is triggered via osteocyte apoptosis (programmed cell death), which in turn is caused primarily by fatigue damage to cortical bone tissue (Burr
et al., 1985; Burr & Martin, 1989; Mori & Burr, 1993;
Parfitt, 2002; Martin, 2003; Aguirre et al., 2006;
Hedgecock et al., 2007; van Oers et al., 2008; Cardoso
et al., 2009). Thus, secondary osteons form primarily
to repair areas that are damaged. Although Haversian
bone is mechanically weaker than primary bone
(Currey, 1959, 1962), secondary osteons are known to
inhibit crack propagation and increase the bone’s
fracture toughness (Currey, 1962; Ascenzi & Bonucci,
1967; Burr et al., 1985). As secondary functions,
Haversian
substitution
aids
with
calcium
homoeostasis, supplying the body with additional
calcium and phospate, and also with haematopoiesis
(formation of blood cellular components). However,
these activities mainly involve cancellous bone, and
not cortical bone (Parfitt, 1981; Parfitt, 2002).
There are many terms used for secondary bone
structure. Remodelling is used frequently to describe
the formation of secondary osteons, although the term
itself can have a broader meaning, encompassing all
changes to bone after initial deposition, including
expansion of the medullary region. For clarity, we use
the following terms: Haversian substitution for the
process of primary bone replacement in the cortex;
secondary osteons for the individual units, which
replace primary bone; and Haversian bone to describe
bone with multiple generations of secondary osteons.
Interestingly, cortical Haversian substitution
usually follows a specific pattern in dinosaurs: secondary osteons always begin forming in the innermost cortex and extend out to the outermost cortex; a
process that continues until the animal dies (Sander,
2000; Klein & Sander, 2008).
MATERIAL AND METHODS
MATERIAL
Growth series of several different sauropod dinosaur
taxa used in previous histological studies (Sander,
2000; Sander et al., 2006; Klein & Sander, 2008;
Stein et al., 2010; Klein et al., 2012) were examined
along with a single mammalian specimen (Table 1).
Giraffatitan brancai Janensch, 1914 and an indeterminate diplodocid are from the Late Jurassic
Tendaguru Formation, Tanzania (Sander, 2000).
767
Europasaurus holgeri Mateus, Laven, & Knötschke in
Sander et al. (2006) is a dwarf sauropod from the Late
Jurassic of Germany (Sander et al., 2006). Apatosaurus sp. Marsh, 1877 and Camarasaurus sp. Cope,
1877 are from the Late Jurassic Morrison Formation
of the USA (Klein & Sander, 2008). Magyarosaurus
dacus Nopcsa, 1914 is a dwarf titanosaur from
the early Maastrichtian of Romania (Stein et al.,
2010). Ampelosaurus atacis le Loeuff, 1995 is
another titanosaur from the late Campanian to early
Maastrichtian from southern France (Klein et al.,
2012). Cervus elaphus Linnaeus 1758 is a mammalian
Pleistocene specimen. All sections are housed
in the Steinmann-Institut Bereich Paläontologie,
Universität Bonn.
HISTOLOGICAL
METHODS
We focus on the histology of the mid-diaphysis of the
large stylopodial long bones: the humerus and femur.
In an earlier study (Klein & Sander, 2008), it was
shown that, at least in sauropods, humeri and femora
do not differ from each other histologically, which is
why, in the present study, we use samples from both
bones interchangeably. We are well aware that this
may not be possible in other tetrapods where the
forelimb and hindlimbs are less columnar and differ
more in function and loading regime.
Thin sections were obtained from the midshaft of
the humerus and femur because the midshaft region
contains the most complete growth record (Sander,
2000; Stein & Sander, 2009). Histology varies within
a single bone; thus, it is crucial that a homologous
region is used when comparing the histological structure of bone. This is also true if comparing different
elements of the skeleton (Werning, 2012) because
different amounts of periosteal tissue need to be laid
down at the same time in different elements, such as
a small bone like a rib compared to a large bone, such
as a femur.
For previous studies, core drilling was used for
most samples (for a description of the process, see
Stein & Sander, 2009). This included all sauropods
with the exception of the Europasaurus sections
DFMMh/FV 495.9 and 372, the Magyarosaurus
section FGGUB R1992, all Ampelosaurus sections
examined (MDE C3 238 and C3 270, LCE Cruzy I),
and the Cervus section IPB M6007 (Table 1). Thinsections had been prepared in accordance with standard methods (Enlow & Brown, 1956; Chinsamy &
Raath, 1992; Wilson, Leiggi & May, 1994). They
were examined in transmitted and polarized transmitted light using a Leica DMLP Polarizing Microscope configured with a 360° rotating stage. Images
were acquired with a Leica DFC420 colour camera
attachment and IMAGE ACCESS EASYLAB, version
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
OMNH 1278
BYU 681-17014
BYU 681-11940
BYU 601-17328
OMNH 4020
MfN
MfN
MfN
MfN
MfN
MfN
MfN
MfN
MfN
OMNH 1794
BYU 725-12173
OMNH 2115
BYU 681-4742
SMA 0002
DFMMh/FV
DFMMh/FV
DFMMh/FV
DFMMh/FV
MAFI Ob 3092
FGGUB R1047
FGGUB R1046
FGGUB R1992
MDE C3 270
MDE C3 238
LCE Cruzy I
IPB M6007
Apatosaurus sp.
Giraffatitan brancai
Diplodocid indet.
Camarasaurus sp.
Europasaurus holgeri
Magyarosaurus dacus
Ampelosaurus atacis
Cervus elaphus
H
H
H
H
H
H
F
F
F
F
F
H
F
F
H
H
H
H
H
H
H
F
F
F
H
H
F
F
F
F
F
Element
–
7
13
13
> 180
> 340
> 600
–
12
13
14
14
9
10
11
6
8
4
7
13
5
9
9
10.5
8
10
12
6
11
4
8
9
12
13
HOS
222
403
525
540
400
475
316 (est)
450
600
1330
227
615
705
435
990
730
805
880
1350
2190
690
1760
258
970
1330
1580
> 1800
Length (mm)
3.9
8
14
14
10
20
14
16
6.9
7
8
6.5
14
32
9
20
27
18
29
19
20
18
33
40
13
39
25
35
37
70
CT (mm)
75
170
270
250
115
183
193
195
181
185
175
174
290
570
120
285
–
220
480
330
370
340
620
820
235
620
–
370
535
680
910
CaM (mm)
0.052
0.047
0.052
0.056
0.087
0.109
0.073
0.082
0.038
0.038
0.046
0.037
0.048
0.056
0.075
0.070
–
0.082
0.060
0.058
0.054
0.053
0.053
0.049
0.055
0.063
–
0.068
0.065
0.054
0.077
CT/CaM
–
Klein et al. (2012)
Stein et al. (2010)
Sander et al. (2006)
Klein & Sander (2008)
Sander (2000)
Sander (2000)
Klein & Sander (2008)
Reference
Most specimens were sampled using a core drill, whereas some are entire or half cross-sections. The cortical thickness was measured as the length of the core
not including the medullary region, if present. The sampling location of the femora during coring is the anterior side and, for humeri, the posterior side (Sander,
2000; Stein & Sander, 2009). The same relative locations were measured for the cross-sections. The ratio is the cortical thickness over circumference. Most
circumference data were collected at the time of sampling. A few cross-section circumferences were measured using IMAGEJ. CaM, circumference at midshaft;
CT, cortical thickness; F, femur; H, humerus.
495.9
403.3
555.2
372
G91
A1
XVI 64I
XI a7
IX 1
dd452
XV
XX19
II28 e
Specimen ID
Species
Table 1. List of specimens studied, including element type, length, and histological ontogenetic stage (HOS)
768
J. MITCHELL and P. M. SANDER
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
THREE-FRONT MODEL OF LONG BONE HISTOLOGY
769
7 (Leica). Cross-sections were scanned with an
Epson V740 PRO high resolution scanner (800 to
1200 d.p.i.). High-resolution thin section images of
specimens used in this study are available in a digital
repository at MorphoBank (www.morphobank.org;
project number: P1168).
RELATIVE
CORTICAL THICKNESS MEASUREMENTS
We used the ratio of cortical thickness to circumference at midshaft to obtain relative thickness data of
the specimens (Table 1). We measured the thickness
of the cortex along the anterior side of femora and
the posterior side of the humeri. The circumference
at midshaft was obtained during sampling for most
specimens. The Cervus sample and Europasaurus
DFMMh/FV 372 were measured using IMAGEJ
(NIH).
Institutional abbreviations
BYU, Museum of Earth Sciences, Brigham Young
University,
Provo,
Utah,
USA;
DFMMh/FV,
Dinosaurier-Freilichtmuseum Münchehagen/Verein
zur Förderung der Niedersächsischen Paläontologie
(e.V.), Germany; FGGUB, Faculty of Geology and
Geophysics of the University of Bucharest, Romania;
IPB, Bereich Paläontologie, Steinmann-Institut,
Universität Bonn, Germany; LCE, Musée de Cruzy,
Cruzy, France; MAFI, Geological Survey of Hungary,
Budapest, Hungary; MDE, Musée des Dinosaures,
Esperaza, France; MfN, Museum für Naturkunde,
Berlin, Germany; OMNH, Oklahoma Museum of
Natural History, Norman, Oklahoma, USA; SMA,
Sauriermuseum Aathal, Canton Zürich, Switzerland.
RESULTS
APPOSITION FRONT
The AF represents the deposition of new bone on the
periosteal surface. It is always distinct as a result
of its very nature of representing the outer edge of
bone formation. The AF is based on the primary bone
histology, with each tissue type being assigned a
relative speed of apposition (Fig. 1). Thus, woven bone
is deposited faster than fibrolamellar bone, which
in turn is deposited faster than modified laminar
bone (i.e. parallel-fibered framework instead of woven
bone; sensu Klein et al., 2012), and so on (Fig. 1). The
slowest apposition rate was assigned to tissue in the
EFS, which records an extremely slow apposition
(Fig. 1). Note that we average the speed of the AF
over an entire year. Thus, the presence of LAGs
interrupting the apposition of fibrolamellar bone indicates that AF speed was slower than recorded by
fibrolamellar bone lacking LAGs. At this stage in the
Figure 1. Determination of the position of the arrow on
the apposition front (AF) part of the three-front model.
This is based on the diagram to the left, which shows
different coloured regions indicating different tissue and
matrix types. Woven bone is considered the fastest, and
the external fundamental system is considered the slowest
because it indicates that the animal has attained full size.
Note that the placement of the AF arrow can vary within
a tissue type as a result of lines of arrested growth, which
indicates a decrease in the overall growth rate, and the
amount of infilling of lamellar bone, which increases over
time, suggesting a relatively slower growth. EFS, external
fundamental system.
development of the model, we ignore differences in
apposition rate indicated by vascular organization of
fibrolamellar bone.
HAVERSIAN SUBSTITUTION FRONT
The redeposition of bone in our samples occurs in the
form of secondary osteons, as noted above. They
advance from the inner cortex to the outer cortex over
time. In some cases, the HSF can be distinct, with a
separation of primary and Haversian bone, or it can
be more diffuse with scattered secondary osteons, as
already noted by Sander (2000) for the sauropods
from the Tendaguru Beds. Over time, the entire
cortex can be converted to Haversian bone.
The absolute speed of HSF currently cannot be
determined from the fossil sauropod bone. However,
the advancement of the HSF observed during ontogeny suggest, at least qualitatively, that HSF speed is
essentially constant throughout the ontogeny of a
sauropod taxon. Thus, in our model, we use a constant rate of advancement of the HSF, but HSF
speeds may vary between different sauropod taxa.
RESORPTION FRONT
The RF marks the expansion of the medullary cavity
and the removal of bone in the deep cortex. This
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
770
J. MITCHELL and P. M. SANDER
Figure 2. Three-front model and cortical midshaft histology of juvenile Apatosaurus OMNH 1278 (HOS 4), femur,
length = 258 mm. Inset shows highly vascularized fibrolamellar bone and large resorption spaces. Photograph taken in
normal transmitted light. The blue dotted line indicates the position of the resorption front. The apposition front is not
indicated because it is always the outermost surface. The three-front model shows the relative positions of the apposition
front (AF), which is very rapid, the Haversian substitution front (HSF), which is constant, and the resorption front (RF),
which is more rapid than the HSF.
region is typically marked by large resorption spaces
and a region of cancellous bone that grades into
compact bone. It can be more abrupt with little to no
cancellous bone. When it is more gradual, the RF
can overlap the HSF. For estimating RF speed, we
used relative cortical thickness, with a relatively
thinner cortex suggesting a faster RF speed than a
thicker one. Cortical thickness also depends on AF;
thus, if two specimens have a similar RF but different AF, then the one with the faster AF will have a
thicker cortex. Our proxy for relative cortical thickness is the ratio of absolute cortical thickness (CT)
at the histological sample site to circumference at
midshaft (CaM), yielding the thickness index
CT/CaM. Cortical thickness at the sample site is
considered as a valid proxy for overall relative cortical thickness as a result of the homologous location
of all sample sites. We distinguish between a thin
cortex (CT/CaM < 0.05), a cortex of average thickness
(0.05 < CT/CaM < 0.08), and a thick cortex (CT/
CaM > 0.08) (Table 1). These distinctions are somewhat arbitrary but further quantification is beyond
the scope of the present study because we only look
at the relative speed of the three fronts, and not
their absolute speed.
APPLICATION
OF MODEL TO
APATOSAURUS
We recorded the histological characters of an
ontogenetic series of Apatosaurus femora (Klein &
Sander, 2008) and described the histology and
microanatomy of four representative specimens from
a much more extensive growth series in the framework of a three-front model.
In the smallest individual, represented by a femur
(OMNH 1278, HOS 4, 258 mm long), the primary
bone is highly vascularized (Fig. 2). This juvenile
specimen has neither LAGs, nor other types of rest
lines. The AF was thus advancing rapidly, and the
large resorption spaces in the inner cortex suggest
that the RF was also fast. However, the RF lagged
somewhat behind the AF to increase cortical thickness with the growth of the individual. No Haversian
substitution is observed; thus, the RF had progressed
more rapidly than the HSF, preventing the formation
of secondary osteons.
In BYU 681-17014 (femur, HOS 8, 970 mm long),
the resorption cavities increase in size towards the
medullary cavity, indicating that the RF was still
quite active. The primary bone is still fibrolamellar
bone, although the vascular spaces are narrower.
Growth was still rapid but not as fast as in the
juvenile. The AF, accordingly, was slower. Haversian
substitution had only begun, with the formation of a
few immature secondary osteons (Fig. 3). The RF still
dominated over the HSF but had begun to slow down,
as demonstrated by the relative decrease of resorption
cavities.
The second largest specimen is a femur of 1580 mm
in length (BYU 601-17328, HOS 12). An EFS is
present indicating that the animal attained full size;
thus, the AF had slowed down significantly. The RF
lagged behind the AF, resulting in a much thicker
cortex compared to the previous specimen. Secondary
osteons cover almost two-thirds of the thickness of the
cortex, indicating that the HSF had not stopped but
had advanced steadily and faster than the AF and RF
(Fig. 4).
Finally, OMNH 4020 (HOS 13) is an incomplete
femur that must have exceeded 1800 mm in length. It
is from a fully grown, senescent individual. The thick
cortex is made of dense Haversian bone with multiple
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
THREE-FRONT MODEL OF LONG BONE HISTOLOGY
771
Figure 3. Three-front model and cortical midshaft histology of Apatosaurus BYU 681-17014 (HOS 8), femur,
length = 970 mm. The three-front model indicates that the apposition and resorption rates have started to decrease. The
yellow dotted line indicates the position of the Haversian substitution front. Inset shows resorption spaces, immature
secondary osteons and fibrolamellar bone. Photograph taken in normal transmitted light. AF, apposition front; HSF,
Haversian substitution front; RF, resorption front.
Figure 4. Three-front model and cortical midshaft histology of fully grown Apatosaurus BYU 601-17328 (HOS 12),
femur, length = 1580 mm. The three-front model indicates
growth and resorption have almost stopped, and
Haversian substitution remains the same. Inset shows
external fundamental system. Photograph taken in
normal transmitted light. AF, apposition front; HSF,
Haversian substitution front; RF, resorption front.
Figure 5. Three-front model and cortical midshaft histology of fully grown Apatosaurus OMNH 4020 (HOS 13),
femur, length > 1800 mm. The three-front model indicates
that apposition and resorption have completely stopped
but Haversian substitution continues. Inset shows complete Haversian bone. Photograph taken in polarized light
with lambda filter. AF, apposition front; HSF, Haversian
substitution front; RF, resorption front.
generations of secondary osteons (Fig. 5). There is
very little primary bone visible. The AF and RF would
have completely halted at this point, whereas the
HSF was still active because secondary osteons form
continuously until death.
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
772
J. MITCHELL and P. M. SANDER
-
-
Figure 6. The three-front model: comparison of juvenile
to senescent histologies in Apatosaurus. Note how the
gradual slow-down of the apposition and resorption fronts
combined with a constant rate of the Haversian substitution front explain the histology and microanatomy of
the samples. Histology concerns the relative thickness
and distribution of the different bone tissue types;
microanatomy concerns the thickness of the cortex. The
apposition front moves faster than the resorption front,
resulting in a gradual thickening of the bone cortex. As
apposition comes to a stop, so does resorption to maintain
the biomechanically necessary thickness of the cortex.
Haversian substitution remains the same in all stages.
AF, apposition front; HSF, Haversian substitution front;
RF, resorption front.
Figure 6 compares the three-front models for all
four Apatosaurus specimens. The comparison clearly
shows the ontogenetic decrease in apposition and
resorption rates. Because the cortex thickens when
the animal is growing, the RF procedes slower than
the AF. The HSF, on the other hand, maintains the
same speed throughout ontogeny but only becomes
visible in histology when the RF is slower than the
HSF. The comparison of the three-front models for the
ontogenetic stages of Apatosaurus thus indicates that
this taxon was characterized by fast growth until it
reached full size, with the RF lagging behind the AF,
resulting in a constant thickening of the cortex. Both
fronts were faster than the HSF until full size was
reached, meaning that complete Haversian substitution of the cortex occurred only after growth had
effectively ceased.
APPLICATION
OF MODEL TO INDIVIDUALS OF
OTHER TAXA
The three-front model was applied to the Tendaguru
diplodocids as well. In general, they all have an
average cortical thickness. The juvenile specimen
(humerus MfN G91, HOS 5, 435 mm long) has a
similar histology to the juvenile Apatosaurus: highly
vascularized fibrolamellar bone, Haversian substitu-
tion (Sander, 2000). The model of OMNH 1278 (HOS
4) can be applied to this juvenile as well (Fig. 7), with
rapid AF and RF. The larger specimen of the
Tendaguru diplodocid (humerus MfN A1, HOS 9,
990 mm long) has laminar fibrolamellar bone (Fig. 8)
and a few secondary osteons in the inner cortex. The
histology is similar to Apatosaurus BYU 681-17014
(HOS 8). The AF was not as fast as in the juvenile. In
this
specimen,
the
RF
was
still
active
but slowing down, and the HSF was close behind it
because there are scattered secondary osteons in the
inner cortex of this specimen.
The Camarasaurus specimen (SMA 0002, HOS 13,
humerus, 705 mm long) is a fully grown individual
with the cortex completely consisting of Haversian
bone (Klein & Sander, 2008). This core section (Fig. 9)
preserves the entire anterior side of the cortex and
medullary cavity region along with some of the cortex
from the posterior side. The AF and RF had stopped
advancing, yet the HSF continued, as seen in the
largest Apatosaurus. Camarasaurus also shows an
average cortical thickness (Table 1).
The Giraffatitan specimen MfN dd452 (HOS 10,
femur, 1350 mm long) has typical fibrolamellar bone
(Sander, 2000). The RF is slowing down, as is the AF,
although growth is still occurring (Fig. 10). Scattered
secondary osteons extend into the mid cortex. Again,
similar to the previous taxa, Giraffatitan has an
average cortical thickness.
The specimen of the dwarf Europasaurus
(DFMMh/FV 495.9, HOS 9, femur approximately
400 mm long) has primary tissue consisting of
fibrolamellar bone (Sander et al., 2006). No Haversian
substitution is present, and the cortex is much
thinner than in other sauropods (Table 1). The AF
was not as fast as in Apatosaurus BYU 681-17014
because of the LAGs interrupting fibrolamellar bone
apposition. In the femora and humeri of large-bodied
sauropods, LAGs only appear very late in ontogeny,
shortly before the EFS, and signal a major slowdown
in growth (Klein & Sander, 2008; Sander et al., 2011).
The RF was quite fast, as indicated by the large
medullary cavity, and faster than the HSF, as indicated by the lack of secondary osteons (Fig. 11).
The dwarf sauropod Magyarosaurus shows a
strikingly different histology from Europasaurus,
despite both of them being dwarfs. Similar to other
Magyarosaurus samples (Stein et al., 2010), the
sample figured in the present study (FGGUB R1992,
HOS 14, femur approximately 540 mm long) has a
thick cortex (Table 1) with extremely dense Haversian
bone, which indicates that growth had likely ceased;
thus, AF and RF had both halted and the HSF continued at a constant rate (Fig. 12). However, during
the growth phase, the RF was considerably slower
than the AF, resulting in a thick cortex. Assuming a
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
THREE-FRONT MODEL OF LONG BONE HISTOLOGY
773
Figure 7. Three-front model and cortical midshaft histology of juvenile Tendaguru diplodocid indet. MfN G91 (HOS 5),
humerus, length = 435 mm. The three-front model indicates rapid advancement of the apposition and resorption fronts,
similar to Fig. 2. Inset shows fibrolamellar bone. Photograph taken in normal transmitted light. AF, apposition front;
HSF, Haversian substitution front; RF, resorption front.
Figure 8. Three-front model and cortical midshaft histology of Tendaguru diplodocid indet. MfN A1 (HOS 9), humerus,
length = 990 mm. The three-front model indicates resorption and apposition have slowed, similar to Figure 3. Inset shows
laminar fibrolamellar bone with few secondary osteons and resorption spaces. Photograph taken in normal transmitted
light. AF, apposition front; HSF, Haversian substitution front; RF, resorption front.
constant HSF speed in sauropods, the histology of
Magyarosaurus would suggest that both the AF and
the RF were much slower in this taxon than in other
sauropods (Stein et al., 2010).
The Ampelosaurus juvenile (MDE C3 270, HOS 7,
humerus > 180 mm long) has a thin cortex, no secondary osteons, and a type of presumably slowgrowing fibrolamellar bone called modified laminar
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
774
J. MITCHELL and P. M. SANDER
Figure 9. Three-front model and cortical midshaft histology of Camarasaurus SMA 0002 (HOS 13), humerus,
length = 705 mm. The three-front model indicates that the
apposition and resorption fronts have completely stopped.
Inset shows dense Haversian bone, similar to Fig. 5. Photograph taken in normal transmitted light. AF, apposition
front; HSF, Haversian substitution front; RF, resorption
front.
bone (Klein et al., 2012). This suggests that AF was
relatively rapid but not as rapid as in the large
Jurassic sauropods, and that the RF was more active
compared to the HSF (Fig. 13), very similar to
the Europasaurus specimen. The adult individual
(MDE C3 238, HOS 13, humerus > 340 mm long)
has, in comparison, a thicker cortex and dense
Haversian bone. The AF and RF had ceased advancing (Fig. 14).
All sauropod dinosaur three-front models are summarized in Figure 15. Additional specimens are used
and their histological descriptions can be obtained
from the references listed in Table 1. Figure 15 shows
a direct comparison of the different sauropod
three-front models and that most sauropods fit into a
similar pattern with Apatosaurus. However, the two
dwarfs, Europasaurus and Magyarosaurus, show a
unique growth pattern, as does Ampelosaurus.
During growth, Europasaurus maintains a relatively
fast RF, whereas Magyarosaurus has a much slower
RF. Ampelosaurus has a slower AF than the
diplodocids and basal macronarians but develops
dense Haversian tissue later in ontogeny.
The primary bone histology of Cervus elaphus, IPB
M6007, humerus (element length unknown) consists
of plexiform fibrolamellar bone. The outer cortex has
less vascularization in some areas compared to the
mid cortex. A thin lamellar layer partially rims the
endosteal surface of the cortex. No resorption pits are
observed. Haversian substitution is only observed on
one side of the cortex and extends into the outer
cortex, which likely is a result of drift (Fig. 16). Vascular canals can be observed at the outer surface,
Figure 10. Three-front model and cortical midshaft histology of Giraffatitan MfN dd452 (HOS 10), femur,
length = 1350 mm. The three-front model indicates slowing down of resorption and apposition relative to Haversian
substitution. Inset shows highly vascularized fibrolamellar bone with scattered secondary osteons. Photograph taken in
polarized light with lambda filter. AF, apposition front; HSF, Haversian substitution front; RF, resorption front.
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
THREE-FRONT MODEL OF LONG BONE HISTOLOGY
which suggests that appositional growth was not completed. However, it has likely slowed based on the
decreased vascularization. The RF, on the other hand,
has likely stopped; the endosteal surface shows
no signs of resorption. The overall appearance and
location of primary and Haversian bone suggests
that during growth the bone had drifted (i.e. bone
began to be resorbed on one side and deposited on the
other).
DISCUSSION
The three-front model based on an ontogenetic series
of Apatosaurus specimens provides a descriptive comparison of microstructure and histology of diaphyseal
long bone development. This model can be applied to
other sauropods as well and, in comparison with
Apatosaurus, reveals the AF or RF moving at similar
or different rates. The three-front model assumes a
constant Haversian substitution rate, whereas apposition and resorption decrease over time. Under these
circumstances, all changes to microstructure and histology result from relative changes in the rates of
apposition and resorption.
DIFFERENCE BETWEEN HOS
THREE-FRONT MODELS
Figure 11. Three-front model and cortical midshaft histology of dwarf sauropod Europasaurus DFMMh/FV 495.9
(HOS 9), femur, length ∼ 400 mm. The three-front model
indicates that apposition and resorption are faster than
Haversian substitution. Inset shows fibrolamellar bone.
Photograph taken in normal transmitted light. Note the
lack of Haversian substitution and thin cortex. Lines of
arrested growth are present but difficult to see in the
section images. AF, apposition front; HSF, Haversian substitution front; RF, resorption front.
775
AND
Although the HOS approach and three-front models
may appear to overlap to a certain extent because
they use similar features of bone histology, this is not
the case. HOS detect the relative ontogenetic age of
individuals, providing answers to questions such as,
‘Is the specimen in question an adult from a smallbodied species or a juvenile of a large-bodied species?’.
Three-front models, on the other hand, provide an
ontogenetic explanatory framework for observed
histologies. Also, unlike the three-front model, HOS
are specific to variably inclusive clades such as
Diplodocidea or Sauropoda and allow comparisons of
individuals from such a clade. Three-front models are
in principle applicable to all amniotes.
APPLICATION
TO OTHER SAUROPODS
Although slight variations in the amount of
Haversian bone in the Late Jurassic large-bodied
sauropods indicate that the RF may be more active in
some specimens and less active in others, the progression of all three fronts is comparable to Apatosaurus
Figure 12. Three-front model and cortical midshaft histology of dwarf titanosaur Magyarosaurus FGGUB R1992 (HOS
14), femur, length ∼ 540 mm. The three-front model indicates that growth has completely stopped. Inset shows extensive
Haversian bone. Photograph taken in cross-polarized light. Note the thick cortex. AF, apposition front; HSF, Haversian
substitution front; RF, resorption front.
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
776
J. MITCHELL and P. M. SANDER
Figure 13. Three-front model and cortical midshaft histology of Ampelosaurus MDE C3 270 (HOS 7), humerus,
length > 180 mm. Note the lack of Haversian substitution and thin cortex. The three-front model indicates rapid
apposition and resorption fronts. Inset shows laminar fibrolamellar bone that occurs throughout the cortex.
Photograph taken in polarized light with lambda filter. AF, apposition front; HSF, Haversian substitution front; RF,
resorption front.
Figure 14. Three-front model and cortical midshaft histology of fully grown Ampelosaurus MDE C3 238 (HOS 13),
humerus, length > 340 mm. Note the thick cortex. The three-front model indicates complete cessation of apposition and
resorption. The inset shows contact of primary bone (PB) and Haversian bone (HB) in the anterior cortex. Most of the
cortex is replaced by Haversian bone. Photograph taken in normal transmitted light. AF, apposition front; HSF,
Haversian substitution front; RF, resorption front.
and, thus, the three-front model of Apatosaurus can
be applied to these other sauropods (Fig. 15).
The extreme amount of Haversian bone observed in
the titanosaurs Magyarosaurus and Ampelosaurus
may be the result of a slow AF demonstrated by
parallel-fibred bone substituting for woven bone in
the fibrolamellar bone, a specific tissue called modified laminar bone (Klein et al., 2012). Klein et al.
(2012) suggested that the Haversian formation rate
(i.e. the HSF) stayed relatively the same but the
slower AF in Ampelosaurus allowed for more secondary reconstruction in the wake of the HSF.
A slow AF being closely followed by the HSF apparently does not explain the cortical histology of
Europasaurus because, even though it also shows
phylogenetically decreased appositional growth, it has
considerably less Haversian substitution compared
to the titanosaurs reviewed here and less than the
other sauropods as well. However, if we also consider
the RF and cortical thickness, the differences can be
explained. Europasaurus has a comparatively thin
cortex compared to Magyarosaurus (Figs 11, 12).
Indeed, its histology is similar to the Triassic
sauropodomorph Plateosaurus with respect to not
only having thin cortices, but also LAGs and little
Haversian bone (Klein & Sander, 2007). The RF is
interpreted to have been faster in Europasaurus than
Magyarosaurus, which may have prevented secondary osteons from forming because the bone was
removed faster than osteons could form.
Although their three-front models are very different, both Europasaurus and Magyarosaurus are
paedomorphic dwarfs (Sander et al., 2006; Stein et al.,
2010; Carballido & Sander, 2013). The difference
between the histology of the two taxa may be
explained by a much longer history of island evolution
in Magyarosaurus compared to Europasaurus.
Although Europasaurus may represent an initial
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
THREE-FRONT MODEL OF LONG BONE HISTOLOGY
777
Figure 15. Comparison of three-front models for the different sauropod taxa. The sauropods Apatosaurus,
Camarasaurus, Giraffatitan, and diplodocid indet. fit into a single model for an ontogenetic series of humeri and femora,
based on their histology and microstructure. The three-front model for Europasaurus, Magyarosaurus, and Ampelosaurus
is different, each showing a difference in mid-diaphyseal bone growth. Note that models in grey do not have specimens
of that ‘age’ group. AF, apposition front; HSF, Haversian substitution front; RF, resorption front.
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
778
J. MITCHELL and P. M. SANDER
Figure 16. Mammal specimen, Cervus elaphus IPB M6007, humerus, length unknown. Main primary bone tissue is
fibrolamellar bone. Haversian bone only occurs along one side of the element. A thin, incomplete layer of lamellar bone
encircles the endosteal region. The three-front model indicates apposition still continuing but resorption greatly slowed
down. Inset shows the abrupt transition from fibrolamellar bone in the outer half and Haversian bone in the inner half.
Photograph taken in polarized light with lambda filter. AF, apposition front; HSF, Haversian substitution front; RF,
resorption front.
stage of dwarfing in which the animal is simply a
scaled-down version of a large ancestor, Magyarosaurus experienced shape change in its skeleton after
size reduction. Dwarfing in Europasaurus evolved by
a decrease in growth rate, expressed in the mid-shaft
region as a slow-down of the AF. The fast RF of
Europasaurus may have resulted from the decrease
in axial loading resulting from the scaling relationships of body mass and bone shaft cross-section.
Magyarosaurus may have gone through the same
stage as Europasaurus but, upon further evolution
in isolation on an island, may have adapted its
long bones to the axial loading regime typical for
sauropods with their thick cortex and small medullary region by reducing the speed of the RF. Assuming
a constant speed of the HSF across taxa, the slowdown in the RF would at least partially be responsible
for the heavy remodelling seen in Magyarosaurus. An
example of such initial size change followed by shape
change in the evolution of an island dwarf is the
goat-like bovid mammal Myotragus. Over five million
years of isolation, this lineage from the Spanish
island of Mallorca consists of consecutive species in
which size reduction preceded morphological and histological change (Köhler & Moyà-Solà, 2004, 2009).
The two Ampelosaurus specimens show similar histologies to the above-mentioned dwarfs. The specimen
with a HOS 7 has a thin cortex and no secondary
osteons, whereas the adult, with an average cortical
thickness, has dense Haversian bone. This may
suggest that Ampelosaurus resorption rates start out
quite fast but slow down considerably to allow a
thicker cortex to form, which in turn, allows
Haversian substitution to occur. If the RF were to
remain more active, fewer secondary osteons could
form.
APPLICATION
TO OTHER AMNIOTES
Based on our initial trial of the humerus of the Cervus
specimen, we propose that the three-front model
could very easily work on mammals. We can illustrate
the microstructure with the three-front model and,
given an ontogenetic series, we should be able to
formulate how the different fronts changed during
growth. Ideally, an ontogenetic series from a single
element of several types of mammals should be tested
for a more thorough result. With that information, we
could compare variation in diaphyseal bone development between these groups.
Conceptually, we could extend this model to any
amniote. For example, based on testable hypotheses
and a three-front model, we could explain why crocodiles lack Haversian remodelling: assuming that HSF
speed is very low in crocodiles compared to their AF,
the HSF would always be overtaken by the RF and
never become visible. Further study is needed to
confirm whether this model would work in all
amniotes.
CONCLUSIONS
The three-front model illustrates fundamental
aspects of sauropod long bone diaphyseal histology at
different growth stages and provides a developmental
explanation for the similarities and differences of
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
THREE-FRONT MODEL OF LONG BONE HISTOLOGY
cortical midshaft bone histology and microanatomy
between different taxa. We can show that Apatosaurus, Camarasaurus, Giraffatitan, and a diplodocid
indet. share similar three-front models. The distinct
histologies of two dwarf sauropods, Europasaurus and
Magyarosaurus, and Ampelosaurus are explained in
the three-front model. The most intriguing aspect of
this model is that it has far-reaching, practical uses
because it appears to be useful for all amniotes. Thus,
a deeper and more comprehensive understanding of
mid-diaphyseal bone development in relation to life
history can be determined with the application of this
model to other taxonomic groups.
ACKNOWLEDGEMENTS
We would like to thank O. Dülfer, R. Schwarz, G.
Oleschinski, D. Kranz, K. Wiersma, and R. Hofmann
for preparing, photographing, and scanning thinsections (Steinmann-Institut Bereich Paläontologie,
Universität Bonn). For generously allowing access to
materials for coring or sectioning, we thank R. Cifelli
and K. Davies (OMNH), W.-D. Heinrich and H.-P.
Schultze (MfN), L. Kordos (FGGUB), N. Knötschke
(DFMMh/FV), J. Le Loeuff (MDE), K. Stadtman
(BYU), and H. J. Siber (SMA). We thank the reviewers S. Werning and H. N. Woodward for their helpful
and insightful comments that improved this manuscript. This is contribution number 163 of the DFG
Research Unit 533 ‘Biology of the Sauropod Dinosaurs: The Evolution of Gigantism’.
REFERENCES
Aguirre JI, Plotkin LI, Stewart SA, Weinstein RS,
Parfitt AM, Manolagas SC, Bellido T. 2006. Osteocyte
apoptosis is induced by weightlessness in mice and precedes
osteoclast recruitment and bone loss. Journal of Bone and
Mineral Research 21: 605–615.
Amprino R. 1947. La structure du tissu osseux envisagée
comme expression de différences dans la vitesse de
l’accroissement. Archives de Biologie 58: 317–330.
Ascenzi A, Bonucci E. 1967. The tensile properties of single
osteons. The Anatomical Record 158: 375–386.
Burr DB, Martin RB. 1989. Errors in bone remodeling:
toward a unified theory of metabolic bone disease. American
Journal of Anatomy 186: 186–216.
Burr DB, Martin RB, Schaffler MB, Radin EL. 1985. Bone
remodeling in response to in vivo fatigue microdamage.
Journal of Biomechanics 18: 189–200.
Carballido JL, Sander PM. 2013. Postcranial axial
skeleton of Europasaurus holgeri (Dinosauria, Sauropoda)
from the Upper Jurassic of Germany: implications for sauropod ontogeny and phylogenetic relationships of basal
Macronaria. Journal of Systematic Palaeontology 12: 335–
387.
779
Cardoso L, Herman BC, Verborgt O, Laudier D, Majeska
RJ, Schaffler MB. 2009. Osteocyte apoptosis controls
activation of intracortical resorption in response to bone
fatigue. Journal of Bone and Mineral Research 24: 597–605.
Castanet J, Curry Rogers K, Cubo J, Boisard J. 2000.
Periosteal bone growth rates in extant ratites (ostriche and
emu). Implications for assessing growth in dinosaurs.
Comptes Rendus de l’Académie des Sciences – Séries
III – Sciences de la Vie 323: 543–550.
Castanet J, Francillon-Vieillot H, Meunier FJ, de
Ricqlès A. 1993. Bone and individual aging. In: Hall BK,
ed. Bone, Vol. 7: bone growth. London: CRC Press, 245–283.
Chinsamy A, Raath MA. 1992. Preparation of fossil bone for
histological examination. Palaeontologia africana 29: 39–44.
Cope ED. 1877. On a gigantic saurian from the Dakota Epoch
of Colorado. Palaeontological Bulletin 25: 5–10.
Cubo J, Le Roy N, Martinez-Maza C, Montes L. 2012.
Paleohistological estimation of bone growth rate in extinct
archosaurs. Paleobiology 38: 335–349.
Currey JD. 1959. Differences in the tensile strength of bone
of different histological types. Journal of Anatomy 93:
87–95.
Currey JD. 1962. Stress concentrations in bone. Quarterly
Journal of Microscopical Science 3: 111–133.
Currey JD. 2002. Bones: structure and mechanics. Princeton,
NJ: Princeton University Press.
Curry K. 1999. Ontogenetic histology of Apatosaurus
(Dinosauria: Sauropoda): new insights on growth rates and
longevity. Journal of Vertebrate Paleontology 19: 654–
665.
Enlow DH, Brown SO. 1956. A comparative histological
study of fossil and recent bone tissues. Part I. Texas.
Journal of Science 8: 405–443.
Erickson GM. 2005. Assessing dinosaur growth patterns: a
microscopic revolution. Trends in Ecology & Evolution 20:
677–684.
Francillon-Vieillot H, Buffrénil V de, Castanet J,
Géraudie J, Meunier FJ, Sire JY, Zylberberg L,
Ricqlès A de. 1990. Microstructure and mineralization of
vertebrate skeletal tissues. In: Carter JG, ed. Skeletal
biomineralization: patterns, processes and evolutionary
trends. New York, NY: Van Nostrand Reinhold, 471–529.
Frost HM. 1969. Tetracycline-based histological analysis
of bone remodeling. Calcified Tissue International 3: 211–
237.
Gillooly JF, Allen AP, Charnov EL. 2006. Dinosaur fossils
predict body temperatures. PLoS Biology 4: 1467–1469.
Griebeler EM, Klein N, Sander PM. 2013. Aging, maturation and growth of sauropodomorph dinosaurs as deduced
from growth curves using long bone histological data: an
assessment of methodological constraints and solutions.
PLoS ONE 8: e67012.
Ham AW. 1953. Histology. Philadelphia, PA: Lippincott
Company.
Hayashi S, Carpenter K, Suzuki D. 2009. Different growth
patterns between the skeleton and osteoderms of Stegosaurus (Ornithischia: Thyreophora). Journal of Vertebrate
Paleontology 29: 123–131.
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
780
J. MITCHELL and P. M. SANDER
Hedgecock NL, Hadi T, Chen AA, Curtiss SB, Martin
RB, Hazelwood SJ. 2007. Quantitative regional associations between remodeling, modeling, and osteocyte
apoptosis and density in rabbit tibial midshafts. Bone 40:
627–637.
Horner JR, de Ricqlès A, Padian K. 1999. Variation in
dinosaur skeletochronology indicators: implications for age
assessment and physiology. Paleobiology 25: 295–304.
Huttenlocker AK, Woodward HN, Hall BK. 2013. The
biology of bone. In: Padian K, Lamm ET, eds. Bone histology
of fossil tetrapods. Advancing methods, analysis, and interpretation. Berkeley, CA: University of California Press,
13–34.
Janensch W. 1914. Übersicht über die Wirbeltierfauna der
Tendaguruschichten, nebst einer kurzer Charakterisierung
der neu aufgeführten Arten von Sauropoden. Archiv für
Biontologie 3: 81–110.
Klein N, Sander PM. 2007. Bone histology and growth of the
prosauropod dinosaur Plateosaurus engelhardti von Meyer,
1837 from the Norian bonebeds of Trossingen (Germany)
and Frick (Switzerland). Special Papers in Palaeontology
77: 169–206.
Klein N, Sander PM. 2008. Ontogenetic stages in the long
bone histology of sauropod dinosaurs. Paleobiology 34: 247–
263.
Klein N, Sander PM, Stein K, Le Loeuff J, Carballido JL,
Buffetaut E. 2012. Modified laminar bone in Ampelosaurus
atacis and other titanosaurs (Sauropoda): implications for
life history and physiology. PLoS ONE 7: e36907.
Köhler M, Marín-Moratalla N, Jordana X, Aanes R. 2012.
Seasonal bone growth and physiology in endotherms shed
light on dinosaur physiology. Nature 487: 358–361.
Köhler M, Moyà-Solà S. 2004. Reduction of brain and
sense organs in the fossil insular bovid Myotragus. Brain,
Behavior and Evolution 63: 125–140.
Köhler M, Moyà-Solà S. 2009. Physiological and life history
strategies of a fossil large mammal in a resource-limited
environment. Proceedings of the National Academy of
Sciences of the United States of America 106: 20354–20358.
Lee AH, Huttenlocker AK, Padian K, Woodward HN.
2013. Analysis of growth rates. In: Padian K, Lamm ET,
eds. Bone histology of fossil tetrapods. Advancing methods,
analysis, and interpretation. Berkeley, CA: University of
California Press, 217–251.
Lehman TM, Woodward HN. 2008. Modeling growth rates
for sauropod dinosaurs. Paleobiology 34: 264–281.
Linnaeus C. 1758. Systema naturæ per regna tria naturæ,
secundum classes, ordines, genera, species, cum characteribus, differentiis, synonymis, locis. Stockholm: Salvius.
le Loeuff J. 1995. Ampelosaurus atacis (nov. gen., nov. sp.), a
new titanosaurid (Dinosauria, Sauropoda) from the Late
Cretaceous of the Upper Aude Valley (France). Comptes
Rendus de l’Académie des Sciences - Série IIa - Sciences de
la Terre et des planètes 321: 693–699.
de Margerie E, Cubo J, Castanet J. 2002. Bone typology
and growth rate: testing and quantifying ‘Amprino’s rule’ in
the mallard (Anas platyrhynchos). Comptes Rendus Biologies 325: 221–230.
de Margerie E, Robin JP, Verrier D, Cubo J, Groscolas
R, Castanet J. 2004. Assessing a relationship between
bone microstructure and growth rate: a fluorescent labelling
study in the king penguin chick (Aptenodytes patagonicus).
Journal of Experimental Biology 207: 869–879.
Marsh OC. 1877. Notice of new dinosaurian reptiles from the
Jurassic Formation. American Journal of Science XIV: 514–
516.
Martin RB. 2003. Fatigue microdamage as an essential
element of bone mechanics and biology. Calcified Tissue
International 73: 101–107.
Mitchell J. 2012. Bone remodeling in sauropod dinosaurs:
using secondary osteons as ontogenetic indicators. Master’s
Thesis, University of Bonn.
Mori S, Burr D. 1993. Increased intracortical remodeling
following fatigue damage. Bone 14: 103–109.
Nopcsa F. 1914. Über das Vorkommen der Dinosaurier in
Siebenbürgen. Verhandlungen der Zoologisch-Botanischen
Gesellschaft Wien 54: 12–14.
van Oers RFM, Ruimerman R, Tanck E, Hilbers PAJ,
Huiskes R. 2008. A unified theory for osteonal and hemiosteonal remodeling. Bone 42: 250–259.
Padian K, Horner JR, de Ricqlès A. 2004. Growth in small
dinosaurs and pterosaurs: the evolution of archosaurian
growth strategies. Journal of Vertebrate Paleontology 24:
555–571.
Parfitt AM. 1981. Integration of skeletal and mineral
homeostasis. In: Deluca H, Frost HM, Jee W, Johnston C,
Parfitt A, eds. Osteoporosis. Recent advances in pathogenesis
and treatment. Baltimore, MD: University Park Press, 115.
Parfitt AM. 2002. Targeted and nontargeted bone
remodeling: relationship to basic multicellular unit origination and progression. Bone 30: 5–7.
Reid REH. 1987. Bone and dinosaurian ‘endothermy’.
Modern Geology 11: 133–154.
de Ricqlès A. 1968. Recherches paléohistologiques sur les os
longs des tétrapodes: origine du tissu osseux plexiforme des
dinosauriens sauropodes. Annales de Paléontologie 54: 133–
145.
de Ricqlès A. 1975. Recherches paléohistologiques sur les os
longs des tétrapodes VII. Sur la classification, la signification fonctionnelle et l’histoire des tissus osseux des
tétrapodes. Annales de Paléontologie 61: 51–129.
Sander PM. 2000. Longbone histology of the Tendaguru
sauropods: implications for growth and biology. Paleobiology
26: 466–488.
Sander PM, Andrassy P. 2006. Lines of arrested growth and
long bone histology in Pleistocene large mammals from
Germany: what do they tell us about dinosaur physiology?
Palaeontographica Abteilung A 277: 143–159.
Sander PM, Klein N, Stein K, Wings O. 2011. Sauropod
bone histology and implications for sauropod biology.
In: Klein N, Remes K, Gee CT, Sander PM, eds. Biology of
the sauropod dinosaurs. Understanding the life of giants.
Bloomington, IN: Indiana University Press, 276–302.
Sander PM, Mateus O, Laven T, Knötschke N. 2006. Bone
histology indicates insular dwarfism in a new Late Jurassic
sauropod dinosaur. Nature 441: 739–741.
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781
THREE-FRONT MODEL OF LONG BONE HISTOLOGY
Starck JM, Chinsamy A. 2002. Bone microstructure and
developmental plasticity in birds and other dinosaurs.
Journal of Morphology 254: 232–246.
Stein K, Csiki Z, Rogers KC, Weishampel DB,
Redelstorff R, Carballido JL, Sander PM. 2010. Small
body size and extreme cortical bone remodeling indicate
phyletic dwarfism in Magyarosaurus dacus (Sauropoda:
Titanosauria). Proceedings of the National Academy of Sciences of the United States of America 107: 9258–9263.
Stein K, Prondvai E. 2013. Rethinking the nature of
fibrolamellar bone: an integrative biological revision of
781
sauropod plexiform bone formation. Biological Reviews 89:
24–47.
Stein K, Sander PM. 2009. Histological core drilling: a less
destructive method for studying bone histology. In: Brown
MA, Kane JF, Parker WG, eds. Methods in fossil
prepraration: proceedings of the first annual fossil preparation and collections symposium, 69–80.
Werning S. 2012. The ontogenetic osteohistology of
Tenontosaurus tilletti. PLoS ONE 7: e33539.
Wilson JW, Leiggi P, May P. 1994. Histological techniques.
Vertebrate Paleontological Techniques 1: 205–234.
SHARED DATA
High-resolution thin section images of specimens used in this study are available at MorphoBank
(www.morphobank.org; project number: P1168).
© 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112, 765–781