Single nucleotide polymorphisms in

odontogenesis-related genes associated with

tooth-size discrepancy
Caio Luiz Bitencourt Reis,* Guido Artemio Marañón-Vásquez,†
Mirian Aiko Nakane Matsumoto,* Flares Baratto-Filho,‡
Maria Bernadete Sasso Stuani,* Peter Proff,§ Christian Kirschneck§
and Erika Calvano Küchler§
Department of Pediatric Dentistry, School of Dentistry of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil*
Department of Pediatric Dentistry and Orthodontics, School of Dentistry, Federal University of Rio de Janeiro,
Rio de Janeiro, Brazil†
School of Dentistry, Univille University, Joinville-SC, Brazil‡
Department of Orthodontics, University Medical Centre of Regensburg, Regensburg, Germany §

Introduction: The present study aimed to determine the association between single nucleotide polymorphisms (SNPs) in RUNX2,
SMAD6, BMP2, and BMP4 genes in relation to tooth-size discrepancy (TSD).
Methods: A cross-sectional study of patients undergoing orthodontic treatment measured the mesiodistal width of permanent teeth
from pretreatment dental casts. Sixty-two patients were included in the study and TSD was assessed according to the Bolton
analysis. The patients were allocated into a control group (without a TSD), an anterior excess group and an overall excess group.
Genomic DNA was extracted from saliva samples, and SNPs previously associated with tooth size were evaluated using a real-
time polymerase chain reaction (PCR) system. The Fisher exact test was performed to compare genotype and allele frequencies at
an α = 0.05. An Odds Ratio (OR) and 95% Confidence Interval (95% CI) were calculated.
Results: The rs59983488 SNP in the RUNX2 gene was significantly related to the presence of anterior mandibular tooth-size
excess in allele (T allele: p<0.001; OR = 11.74; 95% CI =2.61–55.80), and genotype models (GT genotype: p = 0.002;
OR = 12.69; 95% CI = 2.47–64.83). The rs3934908 SNP in the SMAD6 gene was significantly associated with the presence
of an overall maxillary tooth-size excess in allele (T allele: p < 0.001) and genotype models (TT genotype: p = 0.010).
Conclusions: The present results suggest that SNPs in RUNX2 (rs59983488) and SMAD6 (rs3934908) genes may be associated
with the presence of tooth-size excess.
(Aust Orthod J 2023; 39: 86 - 95. DOI: 10.2478/aoj-2023-0008)

Received for publication: August, 2022

accepted: January, 2023.

Caio Luiz Bitencourt Reis: [email protected]; Guido Artemio Marañón-Vásquez: [email protected];

Mirian Aiko Nakane Matsumoto: [email protected]; Flares Baratto-Filho: [email protected];
Maria Bernadete Sasso Stuani: [email protected]; Peter Proff: [email protected];
Christian Kirschneck: [email protected]; Erika Calvano Küchler: [email protected]

the dental arches because accepted overbite, overjet

Introduction and interdigitation require a certain proportional
A tooth-size discrepancy (TSD) is characterised by size relationship between the teeth.3,4 Successful
a dimensional disproportion in the mesiodistal size orthodontic treatment with regard to a functional
between the maxillary and mandibular teeth1,2. A and aesthetic occlusion also, in part, depends on an
TSD contributes to an altered relationship between accurate TSD diagnosis and intervention.1,2

86   Australasian Orthodontic Journal Volume 39 No. 1 2023 ☉ Open Access. Published by Sciendo. cc BY 4.0 © 2023 Author(s). This work is licensed under
the Creative Commons Attribution 4.0 License (https://creativecommons.org/licenses/by/4.0/)
 REIS, MARAÑÓN-VÁSQUEZ, MATSUMOTO, BARATTO-FILHO, SASSO STUANI, PROFF, KIRSCHNECK AND KÜCHLER

Tooth size is determined at the commencement of the reduction, occlusal dental wear, fractured or poor-
dental development stage.5 Odontogenesis involves a quality dental casts, and extreme tooth misalignment,
complex mechanism of interaction between signalling were excluded. The sample was previously described
networks and growth factors.6–8 Bone morphogenetic by Marañón-Vásquez et al.13 Sixty-two patients were
protein 4 (BMP4) is an important growth factor included in the current study.
inducing tooth development as it stimulates the
A single dentist evaluated ten random dental casts
differentiation of mesenchymal-derived dental-
twice within a two-week period to conduct an intra-
specific cells from the beginning of tooth formation.6,9
examiner reliability test. The intraclass correlation
BMP4 also regulates Runt-related transcription
coefficient (ICC) estimated a high reproducibility
factor 2 (RUNX2), which is a critical transcriptional
for all teeth (ICC ranging from 0.888 to 0.996).
regulator of tooth formation.7,10 RUNX2 controls the
Mesiodistal tooth width was obtained by the
morpho-differentiation and growth of the embryonic
largest crown distance between the lateral contact
epithelium of the enamel organ which precedes the
point parallel to the occlusal plane (maximum
formation of tooth enamel.8 In addition, RUNX2
side-distal distance). A digital calliper (Digimatic
stimulates BMP2 production which is a key protein
CD-15DCX; Mitutoyo, Kawasaki, Japan) was
involved in odontogenic differentiation and the
used for measurements. All teeth of each maxillary
control of enamel mineralisation.11 Additionally,
and mandibular cast were consecutively measured
SMAD proteins are important mediators affecting
three times and, when the difference between the
this network of signaling pathways.8,12
measurements was more than 0.2 mm, the tooth was
Single nucleotide polymorphism (SNP) is a DNA remeasured.
sequence variation occurring when a single nucleotide
in the genome differs between members or paired TSD was assessed using the Bolton analysis.17
chromosomes of an individual. Recent studies have Bolton ratios were calculated according to the pre-
investigated SNPs in odontogenesis-related genes and existing formula for the establishment of an anterior
identified some as possible relevant factors related to discrepancy ratio and overall discrepancy ratio, as
tooth-size variability in humans.13–15 Therefore, SNPs follows:
associated with growth factors that play an important
role in odontogenesis could be a risk factor for TSD.
Therefore, the current study aimed to investigate the
association between SNPs in BMP4, BMP2, RUNX2
and SMAD6 genes and TSD. TSD was classified according to Bolton17 as
Anterior TSD: Bar < 75.55 and Bar > 78.85 were
Methods considered as maxillary and mandibular tooth-size
anterior excess, respectively. Bar ranging from 75.55
The present cross-sectional study was approved by
to 78.85 was classified as an absence of anterior TSD.
the Local Research Ethics Committee (protocol:
01451418.3.0000.5419/3.150.551) and all included Overall TSD: Bor < 89.39 and Bor > 93.21 were
patients and their legal guardians signed written considered as maxillary or mandibular overall tooth-
informed consent according to the Declaration size excess, respectively. Bor ranging from 89.39 to
of Helsinki. The STREGA (STrengthening the 93.21 was classified as an absence of overall TSD.
REporting of Genetic Association Studies) checklist DNA was extracted from saliva according to the
was used to design and report this study.16 protocol previously published by Küchler et al.18
The study included pretreatment dental casts and and was used for the molecular analysis. SNPs in
genomic DNA samples from self-reported white RUNX2, SMAD6, BMP2, and BMP4 genes that
and biologically unrelated patients undergoing were previously associated with permanent tooth
orthodontic treatment at School of Dentistry of size14 were selected for genotyping analysis. The
Ribeirão Preto, University of São Paulo from 2016 to SNP characteristics are shown in Table I. Probes
2018. Patients with a previous history of orthodontic (Applied Biosystems, Foster City, CA, USA) were
treatment, congenital syndromes, tooth anomalies, used for genotyping in a real-time polymerase
interproximal caries, restorations or enamel chain reaction (PCR) system (Applied Biosystems,

Australasian Orthodontic Journal Volume 39 No. 1 2023   87

SINGLE NUCLEOTIDE POLYMORPHISMS IN ODONTOGENESIS-RELATED GENES ASSOCIATED WITH TOOTH-SIZE DISCREPANCY

Table I. Characteristics of SNPs.

Genes SNPs Type of Alteration Base Change Global MAF* Genotyping success rate
RUNX2 rs59983488 Upstream Variant G > T 0.15 88.7%
Rs1200425 Intron Variant G > A 0.44 87.0%
SMAD6 rs3934908 Intron Variant C > T 0.41 91.9%
Rs211261 Intron Variant C > T 0.19 90.3%
BMP2 rs1005464 Intron Variant G > A 0.19 91.9%
rs235768 Missense Variant T > A 0.32 90.3%
BMP4 Rs17563 Missense Variant A > G 0.42 88.7%
A, adenine; C, cytosine; G, guanine; MAF, Minor Allele Frequency; T, thymine.
Data are available in https://www.ncbi.nlm.nih.gov/snp.

Foster City, CA, USA) according to the Taqman anterior mandibular tooth-size excess in allele (T
method.19 All reactions were performed blindly and allele: p<0.001; OR = 11.74; 95% CI =2.61–55.80),
10% of the sample was genotyped twice to test for and genotype models (GT genotype: p = 0.002;
reproducibility, which was 100%. OR = 12.69; 95% CI = 2.47–64.83).
Table III demonstrates the allele and genotype
Statistical analysis distribution between the groups for the anterior Bolton
analysis. The rs3934908 SNP in the SMAD6 gene was
The Hardy–Weinberg equilibrium of each SNP was significantly associated with the presence of overall
evaluated by the chi-square test. Genotype and allele maxillary tooth-size excess in allele (T allele: p < 0.001)
distributions were compared using Fisher’s exact test. and genotype models (TT genotype: p = 0.010).
Odds Ratios (OR) and 95% Confidence Intervals
(95% CI) were also calculated. A Bonferroni
adjustment was applied for the total number of SNPs Discussion
(0.05/7 = 0.007). IBM SPSS Version 25.0 (IBM
Predictions of skeletal and tooth patterns is a challenging
Corp, Armonk, USA) software was used for all
aspect of orthodontic practice.20 Knowledge regarding
analyses.
the genes involved in tooth morphology that could be
used to predict tooth-size excess in each patient will
Results assist an individual orthodontic treatment plan and
prognosis as well as in the screening of patients. In the
The number of excluded patients from the study and
present study, the association between SNPs in RUNX2
associated reasons are shown in Figure 1. Sixty-two
and SMAD6 genes and TSD, is identified for the first
patients participated, 33 females (53.2%) and 29
time which may, in the future, be used in orthodontic
(46.8%) males. The mean age of the patients was
finishing strategies to optimise TSD-dependent
15.65 years (Standard Deviation = 6.82 years).
treatment outcomes.21
The genotyping success rate of each SNP is presented
RUNX2 was selected in the study due to its
in Table I. The Hardy–Weinberg equilibrium (HWE)
important role in tooth development and, potentially,
was assessed by a chi-square test (clinicalc.com),
in final tooth morphology.8 Besides regulating
and all SNPs were within the HWE (p>0.05). This
odontoblast differentiation, RUNX2 also plays a
indicated that allele frequencies in this population
role in the later stages of odontogenesis, mainly
did not change from generation to generation.
before crown formation. RUNX2 mRNA is strongly
Table II demonstrates the allele and genotype expressed in immature and mature ameloblasts and
distribution between the groups for the overall TSD is also known to regulate the transcription of the
analysis. The rs59983488 SNP in the RUNX2 gene Amelobastin (Ambn) gene, one of the major proteins
was significantly associated with the presence of an of enamel matrix.8 SNPs in RUNX2 have been

88   Australasian Orthodontic Journal Volume 39 No. 1 2023

 REIS, MARAÑÓN-VÁSQUEZ, MATSUMOTO, BARATTO-FILHO, SASSO STUANI, PROFF, KIRSCHNECK AND KÜCHLER

Figure 1. Patient flow of the study.

previously associated with tooth development and suggesting that these different phenotypes share a
size.14 In the present study, the recessive allele (T) common genetic background. The present results
of rs59983488 was associated with the presence of suggest an association between rs3934908 in SMAD6
anterior mandibular tooth-size excess. This SNP and overall maxillary tooth-size excess. Although
is located in a binding site of an intronic region of Gerber et al.14 did not find an association between this
the RUNX2 gene and has already been associated SNP and mesiodistal and buccal-lingual tooth sizes, it
with cleidocranial dysplasia,22 and also with tooth- is important to consider that previous studies focused
size variability.14 The exact functional effect of this on the evaluation of TSD/tooth excess and therefore
SNP on Runx2 is still unknown; however, Napierala differed from the present study and possibly explains
et al.22 showed that the recessive allele prevents this divergence of results. Further research is indicated
binding between the RUNX2 gene and the Myeloid to clarify the exact role of SMAD6 in odontogenesis
Zinc-Finger transcription factor 1 (MZF1). MZF1 and tooth-size determination.
is a factor that acts as a transcriptional repressor in
stem cells, which may subsequently modify RUNX2 The findings of the present study indicated that
expression and affect tooth development. rs59983488 in RUNX2 was associated with anterior
SMAD6 is characterised by an inhibitory SMAD tooth-size excess, but not with overall tooth-size
protein that restricts the cellular response to BMPs and excess. Similarly, the rs3934908 in SMAD6 was
transforming growth factor β  (TGFβ ).6,23 This mediator associated with overall tooth-size excess, but not with
is present in all tooth-formation stages, but mainly at anterior tooth-size excess. Furthermore, the SNPs
the initiation stage of tooth development.12 Therefore, were associated with tooth-size excess in only one of
it is hypothesised that SNPs in SMAD6 are involved the arches, and not in both arches. Previous studies
in tooth morphogenesis. In addition, SNPs in SMAD6 have indicated that the morphogenesis of each type
have already been associated with other craniofacial of tooth is regulated by different growth factors and
traits, as craniosynostosis,24 skeletal patterns,25 palatal different sets of genes.6,8,12 Therefore, it is reasonable
rugae,26 and tooth agenesis.27 Of note, some of these to suggest that SNPs in growth factor decoder genes
factors have been associated with tooth size variation,14 can influence the tooth size of only one type, one

Australasian Orthodontic Journal Volume 39 No. 1 2023   89

 Table II. Allelic and Genotypic distribution between anterior TSD groups and comparison by Fisher Test.

90  
Control vs. Maxillary tooth-size anterior excess Control vs. Mandibular tooth-size anterior excess

95%
Comparison Alleles or Maxillary Mandibular Odds Confidence
Gene SNP Type Genotypes Control (%) (%) p-value OR 95% CI (%) p-value Ratio Interval
RUNX2 rs59983488 Allelic G 60 (96.8) 14 (87.5) Ref. 23 (71.9) Ref.
T 2 (3.2) 2 (12.5) 0.184 4.28 0.61–28.44 9 (28.1) <0.001** 11.74 2.61–55.80
Genotypic GG 29 (93.5) 6 (75.0) Ref. 8 (50) Ref.
GT 2 (6.5) 2 (25.0) 0.180 4.83 0.62–33.70 7 (43.8) 0.002** 12.69 2.47–64.83
TT 0 (0.0) 0 (0.0) >0.999 - - 1 (6.3) 0.236 - -
rs1200425 Allelic G 37 (63.8) 13 (81.3) Ref. 21 (61.8) Ref.
A 21 (36.2) 3 (18.7) 0.237 0.40 0.11–1.56 13 (38.2) >0.999 1.09 0.47–2.64

Australasian Orthodontic Journal Volume 39 No. 1 2023

Genotypic GG 14 (48.3) 6 (75.0) Ref. 6 (35.3) Ref.
GA 9 (31.0) 1 (12.5) 0.371 0.25 0.02–2.47 9 (52.9) 0.320 2.33 0.61–7.90
AA 6 (20.7) 1 (12.5) 0.633 0.38 0.02–2.87 2 (11.8) 0.268 4.66 0.44–71.52
SMAD6 rs3934908 Allelic C 33 (53.2) 7 (43.7) Ref. 19 (52.8) Ref.
T 29 (46.8) 9 (56.3) 0.580 1.46 0.47–4.12 17 (47.2) >0.999 1.01 0.45–2.25
Genotypic CC 9 (29.0) 2 (25.0) Ref. 4 (22.2) Ref.
CT 15 (48.4) 3 (37.5) >0.999 0.90 0.15–5.84 11 (61.1) 0.728 1.65 0.38–5.76
TT 7 (22.6) 3 (37.5) 0.635 1.92 0.31–12.80 3 (16.7) 0.134 0.25 0.06–1.12
rs211261 Allelic C 29 (55.8) 10 (62.5) Ref. 19 (55.9) Ref.
T 23 (44.2) 6 (37.5) 0.774 0.75 0.22–2.42 15 (44.1) >0.999 0.99 0.39–2.41
Genotypic CC 10 (32.3) 2 (25.0) Ref. 3 (17.6) Ref.
CT 19 (61.3) 6 (75.0) >0.999 1.57 0.27–8.71 13 (76.5) 0.322 2.28 0.53–8.75
SINGLE NUCLEOTIDE POLYMORPHISMS IN ODONTOGENESIS-RELATED GENES ASSOCIATED WITH TOOTH-SIZE DISCREPANCY

TT 2 (6.5) 0 (0.0) >0.999 - - 1 (5.9) >0.999 1.66 0.08–18.07

BMP2 rs1005464 Allelic G 52 13 Ref. 29 Ref.
A 10 3 0.723 1.20 0.31–4.59 5 >0.999 0.89 0.31–2.82
Genotypic GG 23 (74.2) 6 (75.0) Ref. 12 (70.6) Ref.
GA 6 (19.4) 1 (12.5) >0.999 0.63 0.04–4.54 5 (29.4) 0.721 1.59 0.41–5.53
*p < 0.05. **p < 0.007.
 Table II. Allelic and Genotypic distribution between anterior TSD groups and comparison by Fisher Test.

Control vs. Maxillary tooth-size anterior excess Control vs. Mandibular tooth-size anterior excess

95%
Comparison Alleles or Maxillary Mandibular Odds Confidence
Gene SNP Type Genotypes Control (%) (%) p-value OR 95% CI (%) p-value Ratio Interval
AA 2 (6.5) 1 (12.5) 0.536 1.91 0.11–18.28 0 (0.0) >0.999 - -
rs235768 Allelic T 47 (75.8) 11 (68.7) Ref. 26 (72.2) Ref.
A 15 (24.2) 5 (31.3) 0.539 1.42 0.46–4.88 10 (27.8) 0.810 1.20 0.44–2.92
Genotypic TT 16 (51.6) 4 (50.0) Ref. 9 (50.0) Ref.
TA 15 (48.4) 3 (37.5) >0.999 0.80 0.17–3.41 8 (44.4) >0.999 0.94 0.30–2.84
AA 0 (0.0) 1 (12.5) 0.23 - - 1 (5.6) 0.384 - -
BMP4 rs17563 Allelic A 39 (62.9) 9 (64.3) Ref. 21 (61.8) Ref.
G 23 (37.1) 5 (35.7) >0.999 0.94 0.31–3.11 13 (38.2) >0.999 1.05 0.46–2.47
Genotypic AA 13 (41.9) 2 (28.6) Ref. 7 (41.2) Ref.
AG 13 (41.9) 5 (71.4) 0.413 2.50 0.37–13.97 7 (41.2) >0.999 1.00 0.25–3.96
GG 5 (16.1) 0 (0.0) >0.999 - - 3 (17.6) >0.999 1.11 0.23–5.39
*p < 0.05. **p < 0.007.

Australasian Orthodontic Journal Volume 39 No. 1 2023  

REIS, MARAÑÓN-VÁSQUEZ, MATSUMOTO, BARATTO-FILHO, SASSO STUANI, PROFF, KIRSCHNECK AND KÜCHLER

91
 Table III. Allelic and Genotypic distribution between overall TSD groups and comparison by Fisher Test.

92  
Control vs. Maxillary tooth-size overall excess Control vs. Mandibular tooth-size overall excess

95% 95%
Alleles or Maxillary Odds Confidence Mandibular Odds Confidence
Gene SNP Model Genotypes Control (%) (%) p-value ratio Interval (%) p-value Ratio Interval
RUNX2 rs59983488 Allelic G 56 (93.3) 8 (80.0) Ref. 33 (82.5) Ref.
T 4 (6.7) 2 (20.0) 0.201 3.50 0.57–17.98 7 (17.5) 0.110 2.97 0.83–9.51
Genotypic GG 26 (86.7) 3 (60.0) Ref. 14 (70.0) Ref.
GT 4 (13.3) 2 (40.0) 0.195 4.33 0.58–25.79 5 (25.0) 0.281 2.32 0.53–8.40
TT 0 (0.0) 0 (0.0) >0.999 - - 1 (5.0) 0.365 - -
rs1200425 Allelic G 35 (60.3) 8 (80.0) Ref. 28 (70.0) Ref.
A 23 (39.7) 2 (20.0) 0.303 0.38 0.07–1.88 12 (30.0) 0.393 0.65 0.28–1.49

Australasian Orthodontic Journal Volume 39 No. 1 2023

Genotypic GG 14 (48.3) 3 (60.0) Ref. 9 (45.0) Ref.
GA 7 (24.1) 2 (40.0) >0.999 1.33 0.19 – 7.73 10 (50.0) 0.337 2.22 0.64–7.51
AA 8 (27.6) 0 (0.0) 0.527 - - 1 (5.0) 0.210 0.19 0.01–1.58
SMAD6 rs33408 Allelic C 38 (59.4) 0 (0.0) Ref. 21 (52.5) Ref.
T 26 (40.6) 10 (100) <0.001** ∞ 3.75 - ∞ 19 (47.5) 0.544 1.32 0.57–2.85
Genotypic CC 10 (31.3) 0 (0.0) Ref. 5 (25.0) Ref.
CT 18 (56.3) 0 (0.0) >0.999 - - 11 (55.0) >0.999 1.22 0.34–4.33
TT 4 (12.4) 5 (100) 0.010* ∞ 1.96–∞ 4 (20.0) 0.657 2.00 0.39–10.88
rs211261 Allelic C 38 (61.3) 6 (60.0) Ref. 24 (60.0) Ref.
T 24 (38.7) 4 (40.0) >0.999 1.05 0.31–3.72 16 (40.0) >0.999 1.05 0.47–2.28
Genotypic CC 9 (29.0) 1 (20.0) Ref. 5 (25.0) Ref.
CT 20 (64.5) 4 (80.0) >0.999 1.80 0.22–24.21 14 (70.0) >0.999 1.26 0.36–4.31
SINGLE NUCLEOTIDE POLYMORPHISMS IN ODONTOGENESIS-RELATED GENES ASSOCIATED WITH TOOTH-SIZE DISCREPANCY

TT 2 (6.5) 0 (0.0) >0.999 - - 1 (5.0) >0.999 0.90 0.05–9.37

BMP2 rs1005464 Allelic G 50 (80.6) 8 (80.0) Ref. 36 (90.0) Ref.
A 12 (19.4) 2 (20.0) >0.999 1.04 0.20–5.72 4 (10.0) 0.269 0.46 0.15–1.44
Genotypic GG 22 (71.0) 3 (60.0) Ref. 16 (80.0) Ref.
GA 6 (19.3) 2 (40.0) 0.573 2.44 0.35–13.91 4 (20.0) >0.999 0.91 0.25–3.29
Notes: *p < 0.05. **p < 0.007.
 Table III. Allelic and Genotypic distribution between overall TSD groups and comparison by Fisher Test.

Control vs. Maxillary tooth-size overall excess Control vs. Mandibular tooth-size overall excess

95% 95%
Alleles or Maxillary Odds Confidence Mandibular Odds Confidence
Gene SNP Model Genotypes Control (%) (%) p-value ratio Interval (%) p-value Ratio Interval
AA 3 (9.7) 0 (0.0) >0.999 - - 0 (0.0) 0.268 - -
rs235768 Allelic T 48 (75.0) 8 (80.0) Ref. 28 (70.0) Ref.
A 16 (25.0) 2 (20.0) >0.999 0.75 0.14–3.84 12 (30.0) 0.651 1.28 0.55–3.18
Genotypic TT 17 (53.1) 3 (60.0) Ref. 9 (45.0) Ref.
TA 14 (43.8) 2 (40.0) >0.999 0.80 0.13–4.44 10 (50.0) 0.771 1.34 0.40–3.89
AA 1 (3.1) 0 (0.0) >0.999 - - 1 (5.0) >0.999 1.88 0.09–37.71
BMP4 rs17563 Allelic A 34 (55.7) 6 (60.0) Ref. 29 (72.5) Ref.
G 26 (43.3) 4 (40.0) >0.999 11 (27.5) 0.139 0.49 0.21–1.13
Genotypic AA 10 (33.3) 2 (40.0) Ref. 10 (50.0) Ref.
AG 14 (46.7) 2 (40.0) >0.999 0.71 0.10–5.21 9 (45.0) 0.547 0.64 0.19–1.99
GG 6 (20.0) 1 (20.0) >0.999 0.83 0.05–8.47 1 (5.0) 0.183 0.16 0.01–1.64
Notes: *p < 0.05. **p < 0.007.

Australasian Orthodontic Journal Volume 39 No. 1 2023  

REIS, MARAÑÓN-VÁSQUEZ, MATSUMOTO, BARATTO-FILHO, SASSO STUANI, PROFF, KIRSCHNECK AND KÜCHLER

93
SINGLE NUCLEOTIDE POLYMORPHISMS IN ODONTOGENESIS-RELATED GENES ASSOCIATED WITH TOOTH-SIZE DISCREPANCY

arch or one group of teeth, and impact the occlusal Ethics approval and consent to participate
relationship,13–15 depending on the group of genotypes Informed consent/assent was obtained from all
and alleles that the patient carries. The investigated participants and/or their legal guardians where age-
SNPs in BMP2 and BMP4 were not associated with appropriate.
tooth-size excess in the present study. Gerber et
al.14 evaluated tooth size in a Brazilian sample and
showed that the same SNPs in BMP2 and BMP4 Availability of data and material
were associated with size variability of some groups The data generated in the present study are included
of permanent teeth and so these genes were selected within the manuscript.
for examination. However, the present results did not
demonstrate that the SNPs in BMP2 and BMP4 were
involved in TSD. More studies are indicated with Funding
larger sample sizes in other populations to analyse This study was financed in part by the Coordenação
these and other SNPs to confirm the findings. de Aperfeiçoamento de Pessoal de Nível Superior–
Brasil (CAPES)–Finance Code 001 and Alexander-
The method of assessing tooth-size discrepancies von-Humboldt-Foundation (Küchler/Kirschneck
proposed by Bolton17 was adopted for the current study accepted in July 4th, 2019). The São Paulo Research
due to its accuracy and long-standing acceptance by Foundation (FAPESP) financed an individual
the dental profession.4 It is possible that conventional scholarship (CLBR, process 2021/02704-1).
plaster casts may not accurately reproduce exact
tooth size, which is a likely limitation of this study.
Nevertheless, carefully prepared impressions and Authors’ contributions
plaster casts should have negligible size errors. The Conceptualisation, E.C.K., P.P., C.K.; M.A.N.M;
absence of positive associations for other analysed SNPs M.B.S.S. Methodology, M.A.N.M; M.B.S.S.;
may also be due to the small sample size. However, the G.A.M.V; Formal Analysis, C.L.B.R, G.A.M.V;
present study contributes to the identification of some Resources, E.C.K., P.P., C.K.; Data Curation,
genes and SNPs that may affect tooth development C.L.B.R.; Writing— Original Draft Preparation,
and impact directly on the occlusal relationship of the C.L.B.R.; Writing—Review & Editing, C.L.B.R.,
individual, and therefore provides valuable information E.C.K., P.P., C.K.; M.A.N.M; M.B.S.S., G.A.M.V.
for future studies. Funding Acquisition, E.C.K., P.P., C.K.

Conclusions References
1. Laino A, Quaremba G, Paduano S, Stanzione S. Prevalence of
The present results suggest that SNPs in RUNX2
tooth-size discrepancy among different malocclusion groups. Prog
and SMAD6 may be associated with the presence of Orthod 2003;4:37–44.
tooth-size excess. 2. Strujić M, Anić-Milošević S, Meštrović S, Šlaj M. Tooth size
discrepancy in orthodontic patients among different malocclusion
groups. Eur J Orthod 2009;31:584–9.
3. Johe RS, Steinhart T, Sado N, Greenberg B, Jing S. Intermaxillary
Corresponding author tooth-size discrepancies in different sexes, malocclusion groups,
Priv.-Doz. Dr. Dr. Christian Kirschneck and Dr. and ethnicities. Am J Orthod Dentofacial Orthop 2010;138:
Erika Calvano Küchler, Department of Orthodon- 599–607.
4. Jabri MA, Wu S, Zhang Y, Ma J, Wang L. A review on comparison
tics, University of Regensburg. Franz-Josef-Strauss- of tooth size discrepancies among angle’s class I, II, and III
Allee 11, 93053 Regensburg, Germany. Tel. +49 malocclusion: is there a significance. J Contemp Dent Pract
941/944 – 6095/6093, Fax. +49 941/944 – 6169 2019;20:994–9.
Email: [email protected]. 5. Kieser JA, Julius K. Human adult odontometrics: the study of
variation in adult tooth size, 1st edn. Cambridge: Cambridge
de and [email protected] University Press; 1990.
6. Zhang YD, Zhi C, Song YQ, Chao L, Chen YP. Making a tooth:
growth factors, transcription factors, and stem cells. Cell Res
Conflict of interest 2005;15:301–16.
7. Gluhak-Heinrich J, Guo D, Yang W, et al. New roles and mechanism
The authors declare that there is no conflict of of action of BMP4 in postnatal tooth cytodifferentiation. Bone.
interest. 2010;46:1533–45.

94   Australasian Orthodontic Journal Volume 39 No. 1 2023

 REIS, MARAÑÓN-VÁSQUEZ, MATSUMOTO, BARATTO-FILHO, SASSO STUANI, PROFF, KIRSCHNECK AND KÜCHLER

8. Camilleri S, McDonald F. Runx2 and dental development. Eur J human genomic DNA suitable for polymorphism genotyping by PCR-
Oral Sci 2006;114:361–73. RFLP and Real-Time PCR. J Appl Oral Sci 2012;20:467–71.
9. Ohazama A, Tucker A, Sharpe P. Organized tooth-specific cellular 19. Lee LG, Connell CR, Bloch W. Alleleic discrimination by nick-
differentiation stimulated by BMP4. J Dent Res 2005;84:603–6. translation PCR with fluorogenic probes. Nucleic Acids Res
10. Åberg T, Cavender A, Gaikwad JS, et al. Phenotypic changes in 1993;21:3761–6.
dentition of Runx2 homozygote-null mutant mice. J Histochem 20. Küchler EC, Nascimento MAd, Matsumoto MAN, et al. Genetic
Cytochem 2004;52:131–9. polymorphism in RANK is associated with mandibular size. J
11. Wang Y, Li L, Zheng Y, et al. BMP activity is required for orthod 2018;45:157–62.
tooth development from the lamina to bud stage. J Dent Res 21. Uysal T, Sari Z, Basciftci FA, Memili B. Intermaxillary tooth size
2012;91:690–5. discrepancy and malocclusion: is there a relation ? Angle Orthod
12. Xu X, Jeong L, Han J, Ito Y, Bringas P, Chai Y. Developmental 2005;75:208–13.
expression of Smad1-7 suggests critical function of TGF-beta/BMP 22. Napierala D, Garcia-Rojas X, Sam K, et al. Mutations and
signaling in regulating epithelial-mesenchymal interaction during promoter SNPs in RUNX2, a transcriptional regulator of bone
tooth morphogenesis. Int J Dev Biol 2003;47:31–9. formation. Mol Genet Metab 2005;86:257–68.
13. Marañón-Vásquez GA, Vieira AR, Dos Santos LV, et al. FGF10 23. Klopcic B, Maass T, Meyer E, et al. TGF-β  superfamily signaling is
and FGF13 genetic variation and tooth-size discrepancies. Angle essential for tooth and hair morphogenesis and differentiation. Eur
Orthod 2021;91:356–62. J Cell Biol 2007;86:781–99.
14. Gerber JT, Dos Santos KM, Brum BK, et al. Odontogenesis-related 24. Calpena E, Cuellar A, Bala K, et al. SMAD6 variants in
candidate genes involved in variations of permanent teeth size. Clin craniosynostosis: genotype and phenotype evaluation. Genet Med
Oral Investig 2021;25:4481–94. 2020;22:1498–506.
15. Cunha AS, Dos Santos LV, Maranon-Vasquez GA, et al. Genetic 25. Küchler EC, Reis CLB, Carelli J, et al. Potential interactions
variants in tooth agenesis–related genes might be also involved in among single nucleotide polymorphisms in bone-and cartilage-
tooth size variations. Clin Oral Investig 2021;25:1307–18. related genes in skeletal malocclusions. Orthod Craniofac Res
16. Little J, Higgins JP, Ioannidis JP, et al. STrengthening the REporting 2021;24:277–87.
of Genetic Association Studies (STREGA)—an extension of the 26. Silva-Sousa AC, Marañón-Vásquez GA, Stuani MBS, et al. Genetic
STROBE statement. Genetic Epidemiology 2009;33:581–98. variants in bone morphogenetic proteins signaling pathway might be
17. Bolton WA. Disharmony in tooth size and its relation to the analysis involved in palatal rugae phenotype in humans. Sci Rep 2021;11:12715.
and treatment of malocclusion. Angle Orthod 1958;28:113–30. 27. Küchler EC, Reis CLB, Silva-Sousa AC, et al. Exploring the association
18. Küchler EC, Tannure PN, Falagan-Lotsch P, Lopes TS, Granjeiro JM, between genetic polymorphisms in genes involved in craniofacial
Amorim LMF. Buccal cells DNA extraction to obtain high quality development and isolated tooth agenesis. Front Physiol 2021;12:723105.

Australasian Orthodontic Journal Volume 39 No. 1 2023   95