4 Trends
Genetic, epigenetic and environmental
influences on dental development
 OT page 1
there is a group of “dental genes” on assumptions that may not always
that not only influences the size and be valid.
shape of teeth but also the expres- There is a simpler research
sion of missing or extra teeth. In model involving twins that can also
other words, there are both be employed by practising dentists.
pleiotropic genetic effects operating This model is referred to as the MZ
on the human dentition and spatial co-twin design and it essentially
and/or temporal variations in local involves studying pairs of MZ twins
epigenetic events during odontogen- who may have different habits,
esis that lead to distinct phenotypic receive different treatments, or dif-
differences in the dentition, even in fer in expression for one or more
genetically identical twin pairs. features of interest. Because each
MZ pair is matched for sex, age and
genetic make-up, the co-twins pro-
vide an extremely valuable research
model. For example, just one pair of
MZ twins displaying differences in
their dentitions offers a great oppor-
tunity to explore the underlying bio-
logical processes of tooth formation.
Given that MZ twin pairs almost
always share the same genes, any
differences observed between the
members of an MZ pair are assumed
to be due to differences in environ-
mental effects between the co-twins
or to the way in which their genes
are expressed, that is, epigenetic
effects. The MZ pairs described in
Fig. 2. MZ co-twins showing different this article have a very high proba-
expressions of missing, tapering and small
bility of being genetically identical
maxillary lateral incisors.
based on DNA analysis. Further-
Fig. 3. MZ co-twins showing different expression of missing and small maxillary lateral inci-
sors. Twin A displays agenesis of the maxillary right lateral incisor and a small maxillary left
lateral incisor, whereas Twin B has two small maxillary lateral incisors. The mandibular pre-
molars had been extracted for orthodontic reasons.
Fig. 4. MZ co-twins showing different expression of missing and tapering maxillary lateral
incisors. Twin A displays bilateral agenesis of the maxillary lateral incisors, whereas Twin B has
a tapering maxillary right lateral incisor and a missing maxillary left lateral incisor.
more, there is no evidence in their
Tw in m odels records of major differences in envi-
We agree completely with Profes- ronmental influences, either pre-
sor Carels that studies of twins have natally or post-natally. Therefore,
contributed greatly to our under- the differences between these
standing of the role of genetic and co-twins could have been caused by
environmental influences on dental epigenetic differences in the control
development. However, the tradi- of their DNA and/or by relatively
tional twin approach involving com- minor variations in local environ-
parisons between monozygotic (MZ) mental conditions during the
and dizygotic (DZ) twin pairs process of odontogenesis.
requires large samples and is based
ORTHO TRIBUNE | APRIL 2008
Fig. 5. MZ co-twins showing different
expressions of missing lower second pre-
molars and third molar development.
Dif f erences in toot h size in
MZ t w in pairs
We have found many examples of
MZ twin pairs who show quite
marked differences in tooth size and
an example of one pair who show
marked differences in the size of their
upper right central incisors is provid-
ed (Fig. 1). The crowns of these teeth
differ in mesiodistal size between the
co-twins by 0.5 mm, the measure-
ment in Twin A being 7.9 mm and
that in Twin B being 8.4 mm. Given
that the error of measurement for
dental crown diameters is around 0.1
to 0.2 mm, the discrepancy observed
represents a “real” difference in the
size of these teeth. Around 5% of
nearly 200 MZ pairs examined as
part of our studies have shown size
differences in upper central incisors
that are at least as large, or larger, as
that shown in the example.
Dif f erences in ex pression of
m issing t eet h in MZ tw in pairs
We have also found many exam-
ples of MZ twin pairs who show dif-
ferences in the expression of miss-
ing teeth, both in terms of number
and location. Panoramic radio-
graphs of a pair of MZ twins aged 14
years are provided (Fig. 2). Twin A
has a missing maxillary right lateral
incisor and a tapering maxillary left
lateral incisor (arrowed), whereas
Twin B has two small maxillary lat-
eral incisors (arrowed). Several
other examples of different expres-
sion of missing, tapering and small
lateral incisors in MZ twin pairs
have been observed in our studies
(Figs. 3, 4), supporting the idea that
there is a strong relationship
between these features (Townsend
et al., 1995).
The panoramic radiographs of
another pair of MZ twins aged 12.5
years highlight the fact that dental
development can differ quite marked-
ly between MZ co-twins (Fig. 5).
Twin A has a missing lower right
second premolar (arrowed),
whereas the corresponding tooth is
present in Twin B (arrowed). There
is also no evidence of the lower
right third molar in Twin A but the
corresponding tooth is present in
Twin B (arrowed). The upper third
molars are evident in Twin A
(arrowed) but not in Twin B
(arrowed).
Dif f erences in ex pression of
ex tra t eet h in MZ tw in pairs
Not only have we observed differ-
ences of expression of missing teeth
in MZ twin pairs, there are also
examples of different expression of
extra teeth in our samples. For
example, panoramic radiographs of
a pair of MZ twins aged 9.5 years
show different expressions of super-
numerary teeth (Fig. 6). Twin A has
one mesiodens (arrowed), whereas
Twin B has two (arrowed). Futher-
more, the development of the
unerupted teeth is more advanced in
Twin B than Twin A.
The examples shown of missing
and extra teeth in our twin samples
are not rare. Of 24 MZ pairs who
were selected from our records
because they showed at least one
missing lateral incisor or second
premolar, 21 pairs showed differ-
ences in the number or position of
the affected teeth between co-twins.
A similar pattern emerged for super-
numerary teeth. Of nine MZ twin
pairs who showed evidence of
mesiodentes, eight pairs were dis-
cordant for the number of supernu-
meraries (Townsend et al., 2005a,b).
Fig. 6. MZ co-twins showing different
expressions of supernumerary teeth.
Asymm et rical ex pression of
d en t a l f ea t u r es
It is generally assumed that the
genetic information controlling the
development of bilateral structures,
such as teeth, is the same on both
sides. However, teeth rarely display
complete symmetry in their mor-
phology or their development.
Asymmetrical expression of fea-
tures can be either random in its
expression, so-called fluctuating
asymmetry, or favour one side over
the other, so-called directional
ORTHO TRIBUNE | APRIL 2008 Trends 5

asymmetry. By examining the pat-
terns and degrees of expression of
asymmetry between contra-lateral
teeth, it is possible to obtain further
insights into the roles of genetic, epi-
genetic and environmental influ-
ences on dental development (Kha-
laf et al., 2005a).
One particularly interesting
expression of asymmetry that can be
observed in MZ twin pairs is the
phenomenon of mirror imaging,
where one twin “mirrors” the other
for one or more features. Figure 7
shows panoramic radiographs of a
pair of MZ male twins, aged 15
years, who show mirror-imaging for
hypodontia of the lower second pre-
molars. The left premolar is missing
in Twin A (arrowed), whereas the
right premolar is missing in Twin B
(arrowed). In addition, a developing
lower right third molar is evident in
Twin A (arrowed) but not in Twin B
(arrowed). The biological basis of
mirror imaging is still not well
understood, although it apparently
reflects an underlying alteration in
the determination of body symmetry
at an early stage of development.
teeth will occur, and the prevalence
of missing teeth is greater in females
who have smaller teeth, on average,
than males. At the other extreme of
the distribution, with increasing
tooth size, another threshold is
reached above which extra teeth will
occur. The prevalence of extra teeth
is greater in males who have larger
teeth, on average, than females (Fig 8.).
Int erpret ing phenot ypic st udies in the
light of findings at a mol ecular lev el
To date, molecular studies in
humans have concentrated mainly
on locating the genes associated
with missing teeth. As Carels has
pointed out, mutations in two genes,
MSX1 and PAX9, have been shown to
be associated with familial cases of
severe hypodontia (where many
teeth are missing) and the pedigrees
have been consistent with an autoso-
mal dominant mode of inheritance
although showing variations in the
number and position of teeth miss-
ing. However, there are some 300
genes that appear to be involved in
dental development and a number of
them could be candidates for miss-
ing teeth. Furthermore, the most
common clinical presentation relat-
ing to missing teeth is hypodontia
where only a small number of teeth
are missing.
Given that there appears to be a
link between the size and shape of
teeth, and hypodontia or supernu-
merary teeth, we propose that there
is likely to be a group of “dental
genes” that exert pleiotropic effects
on all of these dental phenotypes,
accounting for their observed co-
variation. Just how many genes are
involved remains to be seen, but it is
possible that it may be a relatively
small number. Support for this view
is provided by a paper in Nature by
researchers from Finland (Kangas et
al., 2004) showing that dental char-
acters seem to be non-independent
and that increasing the levels of
expression of just one gene can lead
to increases in cusp number, altered
cusp shape and position, develop-
ment of longitudinal crests on teeth,
and increases in tooth number in
experimental mice.
Rather than a monogenic mode of
inheritance, we believe that a multi-
factorial model (with genetic, epige-
netic and environmental influences)
provides the best explanation for our
observations involving hypodontia
and supernumerary teeth in MZ twin
pairs. Such a model, with superim-
posed thresholds linking tooth size,
morphology and number, enables us
to explain why MZ co-twins, who
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How can t he dif ferences bet w een

MZ co-tw ins be ex plained?
The examples provided in this
article and in other studies we have
performed support the view that,
even though there is a relatively
strong genetic basis to missing or
extra teeth, the number or position
of affected teeth can be influenced
by epigenetic factors. The exact
nature of these influences is still
unclear but they do not need to be
due to differences in methylation of
DNA or acetylation of histones.
Rather, we suggest that they may
reflect different responses of odonto-
genic cells to minor variations in the
spatial and temporal expression of
local signalling molecules passing
between cells during development.
In other words, we are proposing
that “disturbances” in epigenetic
events at the local level during tooth
formation can lead to quite major
differences in the final appearance
of the dentitions of MZ co-twins.
A multi-factorial model linking
tooth size and number has been pro-
posed to account for the different
patterns of expression of missing
and extra, and also large and small
teeth observed in males and females
(Brook, 1984; Brook et al., 2002;
McKeown et al., 2002; Khalaf et al.,
2005b). The relatives of individuals
with missing or extra teeth have
been found to be more likely to also
display missing or extra teeth,
supporting the concept of an under-
lying genetic predisposition to
hypodontia or supernumerary teeth.
We believe that such a multifactorial
model, with multiple genetic and
environmental influences, provides
the best explanation for our observa-
tions involving missing and extra
teeth in MZ twin pairs. With the
superimposition of thresholds on
this underlying distribution, it is pos-
sible to explain the relationship
between tooth size and the presence
or absence of teeth. Below a certain
threshold for tooth size, missing
6 Trends
OT
 page 5
have the same genotypes, may dis-
play different expressions of miss-
ing, tapering and microdont inci-
sors. Presumably these MZ twin
pairs have a genetic predisposition
for hypodontia that places them near
the threshold for agenesis, but minor
variations in local epigenetic events
during odontogenesis may lead to
different phenotypic expression
between co-twins. A similar expla-
nation may also account for the dis-
Fig. 7. MZ co-twins showing mirror-imaging
for missing lower second premolars and differ-
ent expressions of third molar development.
cordant patterns of missing premo-
lars or supernumerary teeth within
MZ co-twins. Presumably, these MZ
twin pairs have a genetic make-up
that places them near to a threshold
for either missing or extra teeth, but
variations in local epigenetic events
during odontogenesis, probably
relating to the spatial arrangement
of cells or temporal events, deter-
mine on which side of the threshold
they fall.
Molenaar’s concept of develop-
mental systems with emergent self-
OT About the authors
Professor Townsend
has research interests
in craniofacial biology
and dental education.
His research involv-
ing twins has been
supported by the
National Health and
Medical Research Council of Australia. He
spent eight months during 2007-2008 on
study leave in Liverpool working with Pro-
fessor Brook and Professor Alvesalo from
Finland to establish an International
Collaborating Research Centre in
Orofacial Genetics and Development.
OT Contact
Grant Townsend, BDS, BScDent,
PhD, DDSc, FICD, FADI
Professor of Dental Science
School of Dentistry
University of Adelaide
South Australia, 5005
Professor of Basic Dental Sciences
School of Dental Sciences
University of Liverpool
UK, L69 3GN
E-mail: grant.townsend@adelaide.edu.au
organizing properties is consistent
with our current understanding of the
molecular basis of tooth development.
The various stages of odontogenesis,
including initiation, morphogenesis
and differentiation, result from a
series of epithelial-mesenchymal
interactions between oral epithelial
and ecto-mesenchymal tissues that
are facilitated by the exchange of var-
ious signalling molecules. The work
of Jernvall and colleagues in Helsinki
has shown how the same genes are
expressed and the same signalling
molecules released in a reiterative
fashion to produce each of the cusps
of a molar tooth (Jernvall and Jung,
2000). In fact, these genes seem to be
highly conserved in an evolutionary
sense and once the process of odonto-
genesis has been initiated, it tends to
proceed as a continuous self-organiz-
ing process as described by Molenaar
and colleagues (Molenaar et al., 1993).
Our studies of intra-coronal
dimensions of molar teeth in twins
are also consistent with the concept
of a dynamically developing crown
pattern during odontogenesis,
linked to the formation of signalling
centres referred to as enamel knots
(Townsend et al., 2003). We suggest
that variations in dental crown form
between species probably result
from regulation of a relatively small
number of highly conserved genes
that control tooth formation in verte-
brates, whereas variations observed
within a species, for example in
humans, probably result from alter-
ations in the timing of interactions
between cells during odontogenesis,
as well as the positions of cells rela-
tive to each other.
Using our broad definition of epi-
genetics, both of these processes can
Professor Brook’s
research interests
are in craniofacial
biology, particular-
ly dental develop-
ment. He is also
currently the Royal
Society of Medi-
cine Frohlich Visiting Professor to estab-
lish collaborative international research
links and is Clinical Principal Investiga-
tor in a five-year Wellcome Programme
Award on Biomineralisation.
Alan Brook, BDS, LDS, MDS, FDS, ILTM
Professor of Paediatric Dentistry
Director, International Collaborating
Research Centre in Orofacial Genetics
and Development
School of Dental Sciences
University of Liverpool
UK, L69 3GN
Email: A.H.Brook@liv.ac.uk
ORTHO TRIBUNE | APRIL 2008
Fig. 8. Multifactorial model with superimposed thresholds that explains the relationship
between tooth size and missing or extra teeth in males and females. (Brook, 1984)
be considered to be examples of
epigenetic control. In the case of
variation between species the con-
trol is likely to reside at the level of
DNA, whereas in variation within a
species the epigenetic influences are
likely to occur at the local tissue level.
Finding the genes f or dental
developm ent
Genome-wide association studies
(GWAS) are currently being used to
identify genes linked to various com-
mon diseases, including coronary
heart disease, hypertension, diabetes
and arthritis. We plan to use a similar
approach to identify the key genes
involved in dental development.
While the identification of key
genes for dental development in
humans will undoubtedly be a major
step forward, there will still be much
work to do. Merely identifying the
genes will not necessarily mean that
we will be able to explain fully how
various dental anomalies arise in
OT References
1. Brook AH. A unifying aetiological explana-
tion for anomalies of tooth number and size.
Archs Oral Biol 1984;29:373–378.
2. Brook AH, Elcock C, Al-Sharood MH, McKe-
own HF, Khalaf K, Smith RN. Further studies
of a model for the etiology of anomalies of
tooth number and size in humans. Conn
Tiss Res 2002; 43:289–295.
3. Carels C. The impact of genes on facial mor-
phology. Ortho 2007;1:38–46.
4. Jernvall J, Jung HS. Genotype, phenotype and
developmental biology of molar tooth charac-
ters. Yrbk Phys Anthropol 2000;43:171–190.
5. Kangas AT, Evans AR, Thesleff I, Jernvall J.
Nonindependence of mammalian dental
characters. Nature 2004;432:211–214.
6. Khalaf K, Elcock C, Smith RN, Brook AH.
Fluctuating dental asymmetry of multiple
crown variables measured by an image
analysis system. Arch Oral Biol
2005a;50:249–253.
7. Khalaf K, Robinson DL, Elcock C, Smith RN,
Brook AH. Tooth size in patients with super-
numerary teeth and a control group meas-
ured by image analysis system. Arch Oral
Biol 2005b;50:243–248.
8. McKeown HF, Robinson DL, Elcock C,
Al-Sharood M, Brook AH. Tooth dimensions
in hypodontia patients, their unaffected
relatives and a control group measured with
a new image analysis system. Europ J
individuals. This is where further
exploration of epigenetic factors will
be essential. Already researchers are
beginning to study epigenetic bio-
markers in an attempt to explain the
reasons for observed differences
between MZ twin pairs (Wong et al.,
2007). At this stage, the focus is on try-
ing to determine the extent of differ-
ences in global genomic DNA methy-
lation levels but it is likely that more
specific analyses will be developed
soon. Once these approaches aimed
at the level of DNA are refined further,
and our understanding of the nature
of the epigenetic influences at a local
tissue level improves, we should be
able to provide a clearer picture of
how genetic, epigenetic and environ-
mental factors influence human den-
tal development. With this knowl-
edge, we will be in a better position to
consider preventive and therapeutic
approaches to many of the common
developmental problems affecting
the human dentition.
Orthodont 2002;24:131–141.
9. Molenaar PCM, Boomsma DI, Machin G. A
third source of developmental differences.
Behav Genet 1993; 23:519–524.
10. Townsend GC, Rogers J, Richards L, Brown
T. Agenesis of permanent maxillary lateral
incisors in South Australian twins. Aust
Dent J 1995;40:186–192.
11. Townsend GC, Richards LC, Hughes TE.
Molar intercuspal dimensions: genetic
input and phenotypic variation. J Dent Res
2003;82:350–355.
12. Townsend G, Hughes T and Richards L.
The dentitions of monozygotic twin pairs:
focusing on the differences rather than the
similarities. In: E. Zadzinska E. Current
trends in dental morphology research. Ref-
ereed proceedings, 13th International Sym-
posium on Dental Morphology, University
of Lodz, Poland, 2005a;337–352.
13. Townsend G, Richards L, Hughes T,
Pinkerton S, Schwerdt W. Epigenetic influ-
ences may explain dental differences in
monozygotic twin pairs. Aust Dent J
2005b;50: 95–100.
14. Wong NC, Joo EJ, Weinrich B, Mossman D,
Scott RJ, Morely R, Craig JM and Saffery R.
Investigating epigenetic biomarkers
underlying phenotypic discordance in
monozygotic twins. Twin Res Hum Genet
2007;10 (Supplement): 58.