J. Anat. (1998) 192, pp. 99 106, with 7 figures Printed in the United Kingdom 99
Quantification of the collagen fibre architecture of human
cranial dura mater
MARIA C. JIMENEZ HAMANN1, MICHAEL S. SACKS1 AND THEODORE I. MALININ2
Tissue Mechanics Laboratory, Department of Biomedical Engineering, University of Miami, and Tissue Bank,
Department of Orthopaedics and Rehabilitation, School of Medicine, University of Miami, FL, USA
(Accepted 28 October 1997)
abstract
The purpose of this study was to quantify and map the gross fibre architecture of the cranial dura mater
(CDM) using small angle light scattering (SALS). In SALS, HeNe laser light is passed through the tissue,
and the resultant scattering pattern is analysed to determine the preferred fibre direction and degree of
orientation. The dura mater was found to be a complex structure with fibre orientations ranging from highly
aligned to nearly random. In the temporal region, 80% of the samples (n 20) were found to have regions
composed of highly oriented fibres with a mean fibre direction of 6 3 0 8 with respect to the sagittal
plane (i.e. almost parallel to the superior sagittal sinus). These highly aligned regions were found in
symmetric anatomical locations about the median sagittal sinus and had similar fibre orientations over both
hemispheres. Although our sample size was small, we found that the size of the symmetric regions, which
covered 14 4 1 6% of the total CDM area, was not influenced by subject s age or sex. The presence of
these highly oriented fibre regions in CDM may be due to mechanical forces exerted on dura mater during
its development. These forces may have induced realignment of the collagen fibres in the direction of tensile
pull, although the exact basis for the unique gross fibre architecture of CDM remains unknown.
Key words: Collagen fibre orientation; small angle light scattering; cranial sutures; structural anisotropy.
Knowledge of the fibre architecture of the CDM is
introduction
important in defining the anatomy of this structure.
The brain and spinal cord are contained by the Since 1955, cranial dura mater allografts (CDMA)
cranium, vertebral column, meninges, and cerebro- have been used for the closure of cranial defects due to
spinal fluid (CSF). The meninges are unique mem- their biocompatibility with host tissue (Campbell et
branous structures with 3 distinct constituents; the pia al. 1958). CDM has also been successfully used in the
mater, the arachnoid, and the dura mater. The cranial repair of pelvic, abdominal, and periodontal defects.
dura mater (CDM), or pachymeninx, is the outermost However, despite the importance of dura mater, its
and most substantial of these layers. It consists of an structural details have not been fully quantified.
outer periosteal layer rich in blood vessels, nerves, and Recently, we have quantified the collagen fibre
large collagenous bundles, and an inner meningeal architecture of CDMA using small angle light
layer composed of flattened mesothelial cells with scattering (SALS) (Sacks et al. unpublished results).
dense cytoplasm (Carpenter, 1976; Nolte, 1988; Gray, SALS is a nondestructive optical technique which
1993). CDM mechanically protects and supports the permits structural characterisation of large fibrous
brain, functions in the collection of venous blood, tissues on a size scale intermediate between optical
CSF drainage, and possibly influences cranial suture microscopy and gross visual analysis. The fibre
formation (Moss, 1960; Smith & Tondury, 1978; architecture of several tissues, including the central

Kokich, 1986; Drake et al. 1993; Hobar et al. 1993; tendon of the diaphragm, pericardium and aortic
Opperman et al. 1993, 1995; Roth et al. 1996. valve leaflets, have been determined using SALS
Correspondence to Dr Michael S. Sacks, Department of Bio-engineering, University of Pittsburgh, Benedum Hall, 7th Floor, 3700 O Hara
St., Pittsburgh, PA 15261, USA.
100 M. C. Jimenez Hamann, M. S. Sacks and T. I. Malinin
(Chuong et al. 1991; Sacks & Chuong, 1992; Hiester with the posterior portion of the superior sagittal
& Sacks, 1997a, b; Sacks et al. unpublished results). sinus towards the examiner. The longest and widest
SALS analysis of CDMA demonstrated that regions dimensions of each sample were recorded, and the
composed of highly aligned collagen fibres were tissue was sectioned in half by cutting along both sides
located in corresponding anatomical positions on the of the superior sagittal sinus. The tissue was then
right and left sides of the superior sagittal sinus, taken through graded aqueous solutions of 50, 75, 85,
indicating that this tissue had symmetric regions along and 90% glycerol in order to make the tissue
the median sagittal plane. Uniaxial mechanical testing translucent for SALS testing.
of tissue samples obtained from symmetric regions
with fibres aligned parallel and perpendicular to the
SALS measurements
uniaxial stress showed that the tissues with parallel
fibre direction were stronger than those with per- A detailed description of the SALS device has been
pendicular fibre direction (Sacks et al. unpublished provided elsewhere (Sacks et al. 1997c). Briefly, a
results). These studies suggested that CDMA is 4 mW HeNe ( 632 8 nm) laser beam is passed
structurally and mechanically an anisotropic material, through the tissue specimen, which scatters light
i.e. its mechanical behaviour is dependent on the according to the internal fibre structure within the
orientation of its local fibre network. light beam envelope. The resulting angular distri-
These preliminary results led us to conduct a bution of scattered light intensity, ™(Åš) vs Åš, about the
detailed study on the fibre architecture of CDM. laser axis represents the distribution of fibre angles
Previous investigators have studied dura mater by within the light beam envelope at the current tissue
conventional methods, such as histology and electron location. The information obtainable from the ™(Åš)
microscopy (EM) (van Noort et al. 1981; McGarvey distribution includes (1) the preferred fibre direction
et al. 1984; Wolfinbarger et al. 1994). However, these computed from the centroid of the fibre orientation
do not allow for quantification of the gross fibre distribution (Åšc), and (2) the orientation index, OI,
architecture of this tissue. Detailed findings of the defined as the angle that contains one half of the total
fibre architecture of CDM may contribute to the area under the ™(Åš) distribution, representing 50% of
understanding of its mechanical behaviour and may the total number of fibres (Sacks et al. 1997c). Thus,
shed some light on the developmental processes that highly oriented fibre networks will have low OI values,
influence its specialised structure. Thus the objective while more randomly oriented networks will have
of this study was the detailed characterisation of the greater OI values. Using the intensity distributions at
collagen fibre architecture of human CDM using each test location, vector maps indicating Åšc and
SALS. colour fringe plots of the OI were generated for the
tissue samples. The fibre orientation distribution of
whole CDM halves were obtained using a 2 54 mm
methods rectilinear grid.
Tissue excision and preparation
Fibre architecture analysis
The clinical history, autopsy findings, and results of
serological laboratory studies of the subjects were Based on our preliminary results (Sacks et al.
used in determining their suitability for this study. unpublished results) we found areas of highly aligned
Dura mater samples were obtained from bodies of collagen fibres with similar fibre orientations in
individuals free of chronic debilitating diseases, symmetric anatomical locations on the right and left
malignancy, or the presence of potentially trans- halves of CDM. In order to quantify the extent and
missible infectious diseases. Tissue samples were size of these highly aligned regions in CDM, we
removed from 20 cadavers (44 5 y) at autopsy developed software which was used to superimpose
performed less than 24 h postmortem, placed in the acquired SALS data of the sample s right half
containers with saline solution, and rapidly frozen in onto its corresponding left half. The difference in Åšc
liquid nitrogen vapour. Each CDM sample was was calculated and used to determine the tissue
composed of the calvarial section of the tissue and the regions where the dural halves had similar fibre
superior sagittal sinus. orientations. Symmetry was defined as a difference in
CDM was thawed overnight at 4 C and then rinsed the fibre s preferred direction between the right and
under running tap water for 5 min. The specimen was left tissue halves of less than 20 , i.e. "Åšc 20 . Grey
laid flat on a Teflon cutting board on its parietal side scale plots of the results were constructed to visually
Fibre architecture of dura mater 101
Fig. 1. SALS scattering patterns illustrating (a) single and (b) double fibre populations with their corresponding (c, d) normalised intensity
plots.
assess regions of symmetry between the dural halves, solution mixture. Paraffin sections were cut trans-
where black regions represented areas with high versely and longitudinally and stained with haema-
degrees of symmetry ("Åšc 20 ) and grey regions toxylin and eosin-, Romanovsky-Giemsa, Masson s
illustrated nonsymmetric areas. trichrome, and Verhoeff s elastic tissue stains. Polar-
In order to determine the anatomical locations of ised light images of the samples were obtained in order
the symmetric regions, a template of CDM was to determine the orientation of multiple fibre popu-
constructed. The CDM template was produced by lations.
using the longest and widest dimensions of each dural
half and visually dividing the tissue samples into 3
coronal and 3 sagittal sections. Partitioning of the Vasculature study
tissue aided in the quick determination of the
We had originally hypothesised that the orientation of
anatomical locations having symmetric regions.
collagen fibres around the meningeal vasculature
would be influenced by the orientation of the vessels.
Therefore, we performed SALS analysis at a high
Histological evaluation
scanning density (using 0 254 mm intervals) around
CDM specimens were examined by light microscopy the vasculature of CDM. The location of the arteries
in order to further evaluate tissue structure. For in dura mater was determined by Micropaque solution
histological studies, representative pieces of fresh dura injection and analysis of radiographs without mag-
matter were fixed in 10% formalin-balanced salt nification. This was performed on 3 CDM samples
102 M. C. Jimenez Hamann, M. S. Sacks and T. I. Malinin
and 2 whole hemisphere dura maters. Markers were samples had certain common structural character-
placed on the injected vessels for recognition. istics. At each test location, SALS results demon-
strated that the local fibre orientation of the CDM
results was mainly composed of single fibre populations (Fig.
1a), which were easily determined by plotting the
Fibre architecture
intensity distribution at each test location (Fig. 1c). A
Even though fibre structural analysis of the dural few double fibre populations were also present in the
halves revealed interspecimen variability, all CDM fibre distribution (Fig. 1b, d). On a regional scale,
Fig. 2. (a) SALS CDM vector plot representing the local preferred fibre directions obtained from the right half of a CDM sample. An area
of structural anisotropy has been outlined. Note how the area of structural anisotropy is located in the temporal region of CDM. (b) Vector
plot of the left half of the same sample. (c) Superposition of the mirror image of the right half onto the left half. Software was developed
to superimpose corresponding vector plots of CDM halves in order to determine symmetric regions easily, as outlined with solid lines.
Fig. 3. (a) Illustration of the superposition of the mirror image of a donor right half onto its corresponding left half. The symmetric area
is outlined for easy visualisation. (b) Grey scale error plot of the same CDM sample illustrating the symmetric area which is represented by
the dark region. The difference in the preferred fibre direction ("Åšc) in this region is 20 .
Fibre architecture of dura mater 103
Table. Summary of all CDM donor information used in this
study*
Specimen Area 1 Area 2 Area 3
no. Age Sex (%) (%) (%)
1 0 M 25  
20 M 12 8 
317 M 27 8 
4 23 M 13 11 
524 M 9 6 
6 31 M 19 12 5
7 36 M 17 12 
8 39 M 995
9 41 M 23  
10 47 M 17 9 
11 49 M 14  
12 52 M 11 9 
13 53 M 3 3 
14 55 M 15 6 
15 59 F 3 3 
16 59 F 24  
17 67 F 10  
18 67 M 21 14 14
19 69 M 9  
20 92 F 7  
* The sizes of the symmetric areas were calculated by dividing the
area of the symmetric region by the total sample area.
the CDM specimens also corresponded to areas
having highly aligned fibres, i.e. tissue regions where
the fibres were oriented in the same direction. The
Fig. 4. (a) Template used to partition CDM samples visually into
grey scale plots (Fig. 3) and CDM templates, which
7 anatomical areas. The longest and widest dimensions of each
sample were used to divide the tissue into 3 coronal and 3 sagittal
divided the CDM samples into 7 anatomical regions,
sections. Region 3 indicates the temporal region of CDM. (b)
were used to determine the tissue locations which were
Graphical illustration depicting the percentage of samples having
symmetric (Fig. 4).
symmetric areas in the specified numbered anatomical areas
from a. The majority (80%) of the samples (n 20) showed
consistent symmetric areas in the temporal region and
the area extending laterally to the superior sagittal
vector maps of CDM halves showed fibres with sinus (Fig. 4a, b) while the frontal and dorsal regions
multidirectional orientations, which did not follow a did not appear to have as many symmetric regions.
consistent pattern in the majority of the tissue (Fig. Even though the largest symmetric regions were
2a, b). These results were similar to previous studies usually found in the temporal region, more than half
reporting structural isotropy in CDM (van Noort et (65%) of the CDM specimens also had 2 or 3 smaller
al. 1981; McGarvey et al. 1984; Wolfinbarger et al. symmetric regions in other anatomical locations
1994). However, on closer examination, localised (Table). Of the total sample population, 20% did not
regions with highly aligned collagen fibres became have large symmetric regions (area of symmetric
evident. The mean preferred fibre direction of all test regions 10% of the total area) indicating that
points within these highly aligned regions was meas- symmetry was not present in all dural specimens.
ured to be 6 3 0 8 with respect to the sagittal plane The areas of the symmetric regions with respect to
(i.e. almost parallel to the superior sagittal sinus) (Fig. the total area of its corresponding sample were
2a, b). calculated from digitised images of each CDM
Superposition of the mirror image of the left half specimen (Table). Although our sample size was
onto its corresponding right half revealed tissue small, linear regression results (r 0 15 and 0 06,
regions with similar fibre alignment in both halves or P 0 05) demonstrated that an evident relationship
symmetric tissue regions along the superior sagittal did not exist between the areas of the symmetric
sinus (Fig. 2c). The symmetric regions found within regions and the donor s age or sex, respectively. Since
104 M. C. Jimenez Hamann, M. S. Sacks and T. I. Malinin
Fig. 6. (a) Polarised light photograph of CDM injected with
Fig. 5. (a) Longitudinal section of human CDM. The photograph
radiopaque dye illustrating how the collagen fibres are distributed
shows the distribution and orientation of elastic fibres. Some of the
around the vasculature. (b) OI plot of the dural section outlined in
fibres are arranged in parallel arrays while others cross each other
a demonstrating that the collagen fibres are not preferentially
at angles. Verhoeff s elastic tissue stain, 250. (b) Same field used
aligned along the meningeal vasculature. (c) A vector plot of the
as in a photographed in polarised light with xenon light as an
same section portraying the preferred fibre direction at 0 254 mm
illumination source. Different orientation of doubly refractile
intervals.
collagen fibres is evident. H & E, 100.
Radiographic study
these results were based on a small and unevenly
distributed sampling pool, they may not hold true for
SALS analysis of radiopaque dye injected CDM
a larger sample size. However, our findings suggest
halves demonstrated that there was no relationship
that each donor s CDM fibre architecture is unique.
between the orientation of the meningeal vasculature
and the collagen fibre architecture (Fig. 6). The
collagen fibres were not seen to be preferentially
Microscopic study
aligned along the larger vasculature. However, polar-
ised light images of a CDM specimen showed that
Histologically, CDM was composed primarily of
some of the collagen fibres were aligned along the
collagen fibres (Fig. 5). Transverse sections of the
smaller vasculature (Fig. 6). Thus, there was no
tissue showed 2 distinct layers. The inner layer was the
apparent trend between the orientation of the collagen
meningeal layer and was composed of flattened cells
fibres and the larger vasculature in CDM.
as described by Carpenter (1976), Nolte (1988) and
Gray (1993). Some microscopic regions showed highly
aligned collagen fibres (Fig. 5) while others were
discussion
composed of randomly distributed fibres (Fig. 5a).
These observations are consistent with the SALS Previous investigations of CDM fibre structure have
vector plots which showed that CDM had highly depended on qualitative methods such as light and
aligned fibres interspersed with randomly oriented electron microscopy. The SALS technique, on the
fibres. Images taken using polarised light also aided in other hand, allows quantified structural information
visualising the different orientations of the collagen to be obtained over an entire tissue specimen. Since
fibre populations (Fig. 5b). the spatial resolution of SALS is intermediate between
Fibre architecture of dura mater 105
light microscopy and gross visual analysis, it can calvarial bones are approximated during cranial
produce structural information not previously avail- development, tensile forces are transmitted to the fibre
able. In addition, the SALS technique is able to map tracts of CDM in order to initiate the process of
the local fibre architecture back to the global tissue suture morphogenesis. It is possible that these mech-
anatomy. This can lead to a better understanding of anical forces reorient the collagen fibres in the
structure function relationships. direction of tensile pull towards the sutural sites.
In this study, we used SALS to quantify the Previous studies have noted that mechanical forces in
collagen fibre architecture of the calvarial section of the rat s falx cerebri reorient the tissue fibre or-
human dura mater. The primary findings were as ganisation during development (Moss, 1960). The
follows. (1) The collagen fibres over most of the CDM fibre thickness and density also increase with con-
have multiple preferred directions with no consistent comitant preferential alignment of the fibre tracts.
pattern indicating that, in general, CDM is struc- These structural changes have been interpreted to
turally isotropic. (2) An exception to the above was indicate the mechanical role of CDM in the trans-
found in the temporal region, where 80% of the mission of intracranial forces, a concept consistent
samples (n 20) demonstrated highly aligned col- with our findings of structurally anisotropic areas in
lagen fibres oriented at 6 3 0 80 with respect to the CDM.
sagittal plane (i.e. almost parallel to the superior We have found highly aligned collagen fibres
sagittal sinus). (3) The size of the symmetric regions oriented parallel to the superior sagittal sinus in the
did not have any apparent relationship with the midcoronal section of calvarial CDM, corresponding
donor s age or sex. This last finding should be to tissue regions posterior to the coronal suture. These
interpreted with caution since the results were ob- findings suggest why the collagen fibres are oriented as
tained from a small sample size (n 20). if tensile forces had pulled the tissue towards the
suture line (Fig. 7). Thus, during calvarial bone
development, mechanical forces pulling CDM in the
Basis of CDM structural isotropy
calvarial region might be responsible for suture
During fetal development, CDM conforms to the morphogenesis creating symmetrically aligned areas
configuration of the developing brain, therefore its in the tissue.
formation is influenced by brain growth (Smith &
Tondury, 1978). SALS analysis of CDM, which is

Symmetry and donor age and sex
based on a 0 36 mm laser beam area (Sacks et al.
1977), demonstrated that the collagen fibres had Although our sample size was not large and evenly
distinct preferred directions throughout the tissue. distributed, our results showed that the size of the
However, at larger size scales ( 1 cm ), we found symmetric regions in CDM was not correlated with
that the preferred directions of adjacent test locations donor age or sex. The variability of the location, size,
had no consistent pattern indicating that, in general, and presence of the symmetric regions in CDM may
CDM is structurally isotropic. Assuming that in- reflect the anatomical differences in brain size, location
tracranial mechanical forces during fetal growth of the sutures, and developmental patterns, which are
primarily determine the collagen architecture of dura not the same for every person. Although our sample
mater, our results suggest that these forces generally size was small, we found that the fibre structure of
do not have a consistent direction. This lack of infant and adult dura matter were not distinctly
consistent directed mechanical forces leads to a different. This finding suggests that the organisation
variable, isotropic fibre structure. of collagen fibres does not change after birth. Even
though the coronal suture begins to fuse at about 24 y
of age, the fibre architecture is not affected. Separate
Basis of symmetric regions of aligned collagen fibres
factors are likely to be involved in suture patency and
Although CDM is primarily an isotropic tissue, it did eventual fusion and in suture formation (Kokich,
demonstrate localised regions of well aligned collagen 1986; Hobar et al. 1993; Opperman et al. 1993, 1995).
fibres in the vicinity of the coronal sutures. The In conclusion, collagen fibres over most of the
presence of these highly aligned regions may be CDM have multidirectional orientations without a
attributed to the role of CDM during suture mor- distinct pattern. Localised symmetric areas with
phogenesis (Moss, 1960; Smith & Tondury, 1978; highly aligned fibres were found in the majority of the

Kokich, 1986; Drake et al. 1993; Hobar et al. 1993; samples in the temporal region. Our findings suggest
Opperman et al. 1993, 1995; Roth et al. 1996). As the that cranial suture morphogenesis may have induced
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