ORIGINAL ARTICLE

Insulin-like growth factor I: A biologic

maturation indicator
Ramy Abdul Rahman Ishaq,a Sanaa Abou Zeid Soliman,b Manal Yehya Foda,b and Mona Mohamed Salah Fayedc
Cairo, Egypt

Introduction: Determination of the maturation level and the subsequent evaluation of growth potential during
preadolescence and adolescence are important for optimal orthodontic treatment planning and timing. This
study was undertaken to evaluate the applicability of insulin-like growth factor I (IGF-I) blood level as
a maturation indicator by correlating it to the cervical vertebral maturation index. Methods: The study was con-
ducted with 120 subjects, equally divided into 60 males (ages, 10-18 years) and 60 females (ages, 8-16 years). A
lateral cephalometric radiograph and a blood sample were taken from each subject. For each subject, cervical
vertebral maturation and IGF-I serum level were assessed. Mean values of IGF-I in each stage of cervical
vertebral maturation were calculated, and the means in each stage were statistically compared with those of
the other stages. Results: The IGF-I mean value at each cervical vertebral maturation stage was statistically
different from the mean values at the other stages. The highest mean values were observed in stage 4,
followed by stage 5 in males and stage 3 in females. Conclusions: IGF-I serum level is a reliable maturation
indicator that could be applied in orthodontic diagnosis. (Am J Orthod Dentofacial Orthop 2012;142:654-61)

T
he timing and the amount of remaining facial
growth are important factors that play major roles
in treatment planning and retention in orthodon-
tics and dentofacial orthopedics. Hence, the determina-
tion of biologic maturation level and the subsequent
evaluation of growth potential during preadolescence
and adolescence are extremely important.1
Chronologic age, dental development, body weight,
height, menarche, and voice and breast changes have
been shown to be unreliable and impractical for estimat-
ing the pubertal growth spurt.2-5
Skeletal age assessments with hand-wrist and lateral
cephalometric radiographs have been shown to be corre-
lated with skeletal growth changes during puberty.2-7
The hand-wrist maturation indicator is classically the
most reliable skeletal maturation indicator. The use of
the cervical vertebral maturation indicator has the advan-
tage of not requiring an additional radiograph. Studies of
From the Department of Orthodontics and Dentofacial Orthopedics, Faculty of
Oral and Dental Medicine, Cairo University, Cairo, Egypt.
a
Senior resident.
b
Professor.
c
Associate professor.
The authors report no commercial, proprietary, or financial interest in the prod-
ucts or companies described in this article.
Reprint requests to: Mona Mohamed Salah Fayed, Department of Orthodontics
and Dentofacial Orthopedics, Faculty of Oral and Dental Medicine, Saraya El-
Manial St, Manial, Cairo, Egypt; e-mail, monasfayed@yahoo.com.
Submitted, February 2012; revised and accepted, June 2012.
0889-5406/$36.00
Copyright Ó 2012 by the American Association of Orthodontists.
http://dx.doi.org/10.1016/j.ajodo.2012.06.015
654
the cervical vertebral maturation stages have shown cor-
relations with the hand-wrist stages of maturation, thus
providing a practical means for estimating facial and
mandibular growth.8 The original method, proposed by
Lamparski,9 incorporates vertebrae that might be ob-
scured by the thyroid collar. The method of Hassel and
Farman1 uses the second through the fourth vertebrae
to avoid the defect in the original method. The 5-stage
method of Baccetti et al10 merges the first 2 stages. The
6-stage method of Baccetti et al11 also uses the second
through the fourth vertebrae; however, it was reported
to have poor reproducibility and predictability.12-14
Insulin-like growth factor I (IGF-I) is a polypeptide
hormone synthesized mainly by the liver. It is a member
of a group of hormones termed insulin-like growth fac-
tors. It is considered a mediator of growth-hormone
function. It is involved in the growth of almost every or-
gan and plays a major role in postnatal growth and pre-
cisely in the process of longitudinal bone growth.15,16
Salmon and Daughaday17 were the first to discover
IGF-I as a mediator of growth-hormone function, which
was termed the sulphation factor. Several studies con-
ducted on IGF-I have reported that its serum levels in
children and adolescents followed a pattern that was
closely related to the pubertal growth curve: low in the
prepubertal stages followed by a sharp increase at pu-
berty and, after pubertal growth had ceased, returning
to lower baseline values.18,19
In a sample of 807 healthy Turkish children, age-
related and sex-related reference ranges for serum IGF-I
Ishaq et al
levels were established. Peak IGF-I concentrations were
reached 1 year earlier and in 1 pubertal stage earlier in
girls than boys, and they started to decline thereafter.20
Reference values in a group of Chinese adolescents for se-
rum levels of IGF-I have been established; it was con-
cluded that serum IGF-I levels peaked at typical
pubertal ages (12-14 years in girls; 14-16 years in boys).21
IGF-I blood levels have been proposed as an alterna-
tive method to detect pubertal growth-spurt timing.
IGF-I blood levels have been correlated with the cervical
vertebral maturation stages in a sample of 84 Saudis; it
was concluded that IGF-I could be used as a skeletal ma-
turity indicator and might be useful in detecting residual
mandibular growth in young adults.22 IGF-I was also
correlated with the hand-wrist skeletal maturation pat-
tern, and it was concluded that mean IGF-I levels
followed at various skeletal stages mirrored the mandib-
ular growth velocity pattern that Fishman23 and Masoud
et al24 observed.
The importance of growth determination in ortho-
dontics is the driving force of the quest for the most accu-
rate and least invasive methodology to track the pubertal
growth spurt. The aim of this study was to evaluate IGF-I
as a maturation indicator in correlation to cervical verte-
bral maturation in a group of circumpubertal adolescents.
MATERIAL AND METHODS
One hundred twenty subjects were selected from the
outpatient clinic at the Department of Orthodontics,
Faculty of Oral and Dental Medicine, at Cairo University
in Egypt. All subjects were either patients under active
orthodontic treatment or new patients requiring ortho-
dontic treatment at the clinic.
The age ranges were 10 to 18 years for male subjects
and 8 to 16 years for female subjects. Exclusion criteria
were systemic disease, bleeding disorders, chronic med-
ication uptake, and trauma or operation in the area of
the cervical vertebrae.
Subject selection for sample collection was organized
by creating age subgroups for both male and female
groups that would result in a uniformly distributed sam-
ple among the different stages of the cervical vertebral
maturation. The distribution of various groups and sub-
groups is illustrated in Figure 1.
Personal information and history were recorded for
every subject to rule out the exclusion criteria. The
parents and each subject were then informed about
the research plan for approval of taking a blood sample
and using a lateral cephalometric radiograph that was
already available as part of the patient's records file.
Approval was obtained from the research ethical com-
mittee of the Faculty of Oral and Dental Medicine of
Cairo University.
American Journal of Orthodontics and Dentofacial Orthopedics
655
Blood samples were collected for analysis of IGF-I by
venipuncture between 9:00 AM and 11:00 AM. The samples
were transferred to the laboratory of the Clinical Pathol-
ogy Department at the Faculty of Medicine of Cairo
University. The samples were analyzed with the enzyme-
linked immunosorbent assay (ELISA) technique. Once
received in the laboratory, the samples were placed on
a centrifuge for separation of plasma from blood cells.
The plasma was then placed in plastic Eppendorf dispos-
able centrifuge tubes (Eppendorf, Zhijiang, China) and
stored in a freezer at –20 C according to the instructions
of the blood analysis kit manufacturer (DIAsource Immu-
noAssays, Nivelles, Belgium). A pretreatment step was in-
troduced to enhance the clinical performance of the assay
with the acid-ethanol procedure of Daughaday et al.25 A
fixed amount of IGF-I was labeled with horseradish perox-
idase, compete with unlabelled IGF-I in the calibrators,
controls, and samples for a limited number of binding
sites on a specific antibody. After a 1-hour incubation
at room temperature, the microtiter plate was washed to
stop the competition reaction. The chromogenic solution
(tertamethylbenzydine) was then added and incubated for
30 minutes. The reaction was stopped with the addition of
a stop solution, and the microtiter plates were read at the
appropriate wavelength. The amount of substrate turn-
over was determined colorimetrically by measuring the
absorbance, which is inversely proportional to the IGF-I
concentration. A calibration curve was plotted, and the
IGF-I concentration in the samples was determined by in-
terpolation from the calibration curve.
Subjects were referred to obtain a digital lateral ceph-
alometric radiograph on the same day as the blood sam-
ple was taken. All subjects were oriented in natural head
position.26-29 All lateral cephalometric radiographs were
taken by the same operator on the same machine.
Upon retrieval of the radiographic films, the area of
the cervical vertebrae was traced on matte acetate paper
by 1 examiner (R.I.). The superior, inferior, posterior, and
anterior borders of the second, third, and fourth cervical
vertebrae were traced. We used a standardized stepwise
technique to facilitate the assessment of the cervical ver-
tebral maturation index stages.
1. Point identification and line construction: a sche-
matic representation of the traced vertebrae show-
ing the points identified and the lines constructed;
the description of each is presented in Figure 2.
2. Cephalometric measurements included the follow-
ing. Measurement of concavity depth at the inferior
border of the vertebral body: the linear distance be-
tween point CD and the line connecting points CLP
and CLA (see Fig 2 for definitions) dictates the pres-
ence and extent of concavity at the inferior border of
November 2012  Vol 142  Issue 5
656
Fig 1. Schematic representation of the groups and subgroups in the sample.
the body of the vertebrae as described in Table I.
Measurements for assessment of the shape of the
body of the cervical vertebrae: the ratio of line W to
line H indicated the shape of the vertebral body as de-
scribed in Table I. All measurements were repeated by
2 other examiners on 2 different occasions.
3. The cervical vertebral maturation index was then
evaluated according to the method described by Has-
sel and Farman.1 It describes the 6 cervical maturation
stages according to the morphologic characteristics
of the second, third, and fourth vertebrae (Fig 3).
Statistical analysis
Statistical analysis was performed with SPSS for Win-
dows software (version 16.0; SPSS, Chicago, Ill). The sig-
nificance level was set at P #0.05. The Kruskal-Wallis
test was used to compare the mean IGF-I values in the
different cervical vertebral maturation stages. The
Mann-Whitney U test was used to compare the mean
IGF-I values between the sexes in each cervical vertebral
maturation stage. Interexaminer reliability in identifying
cephalometric landmarks was evaluated by using the
Cronbach alpha reliability coefficient.
RESULTS
Table II and Figure 4 present mean and standard de-
viation values of IGF-I for each stage of the cervical ver-
tebral maturation index in the male group, the female
group, and the whole sample. Mean IGF-I serum level in-
creased gradually from its lowest value at stage 1 toward
stage 2. A sharper increase was observed between stages
2 and 3, and the peak value was reached at stage 4. It
November 2012  Vol 142  Issue 5 American Journal of Orthodontics and Dentofacial Orthopedics
Ishaq et al
then declined toward stage 5 to reach its baseline level
at stage 6 (Fig 5). Mean IGF-I values recorded at each
stage of the cervical vertebral maturation index were sta-
tistically different from the values recorded at the other
stages.
In the male subjects, the highest mean IGF-I value
was observed in stage 4 at a mean age of 14.5 years
and a mean value of 893 6 171 ng/mL. The second
highest mean IGF-I value was observed in stage 5, fol-
lowed by stage 3. The lowest mean values of IGF-I
were observed in stage 1, followed by stage 6. Mean
IGF-I values recorded at each stage of the cervical verte-
bral maturation index were statistically different from
the values recorded at the other stages, as denoted by
the letters in Table II, from a for the highest value to f
for the lowest value.
In the female group, the highest mean IGF-I value was
observed in stage 4 at a mean age of 14 years and a mean
value of 794 6 217 ng/mL. The second highest mean
IGF-I value was observed in stage 3, followed by stage
5. The lowest mean values were observed in stage 1, fol-
lowed by stages 2 and 6, which showed no statistical dif-
ferences. Mean IGF-I values recorded at each stage of the
cervical vertebral maturation index were statistically dif-
ferent from the values recorded at the other stages, as de-
noted by the letters in Table II, from a for the highest
value to e for the lowest value at stage 1.
Sex differences between the male and female groups
were only observed in the mean values of IGF-I at stages
3 and 5. The second highest stage in the male group was
stage 5, followed by stage 3, whereas in the female
group the opposite was true.
Ishaq et al 657

Fig 2. Diagram showing the various points and lines used in the cephalometric analysis of the cervical
vertebrae.

Interexaminer reliability tests between the observers

Table I. Characteristics of the various stages of the
showed good agreement (kappa, 0.944-0.960).
cervical vertebral maturation index and the values of
each DISCUSSION
Criteria Description Value This study was undertaken to test the hypothesis that
Vertebral body shape Wedge shaped \65% the IGF-I serum level could be used as a skeletal matura-
Horizontally rectangular 65-80% tion indicator in orthodontics.
Almost square 80-95%
The study included 120 subjects equally divided into
Square 95-105%
Vertically rectangular .105% males (ages 10-18 years) and females (ages 8-16 years).
Concavity depth Flat 0 mm Those age ranges were assigned to the groups to include
Beginning \1 mm subjects in the circumpubertal period (prepubertal, pu-
Distinct 1-2 mm bertal, and postpubertal), in whom skeletal maturation
Accentuated 2-3 mm
has a diagnostic value in orthodontics. Age subgrouping
Deep .3 mm
was implemented to have a sample that was almost uni-
formly distributed among the stages of cervical vertebral
maturation and to make comparisons between male and
In the whole sample, the highest mean IGF-I value female groups possible (Table II). Previous studies con-
(835.6 6 201 ng/mL) was observed in stage 4 at ducted on IGF-I with the same sample had a smaller
a mean age of 14 years. The second highest mean IGF- sample size (83 subjects) and a wider age range (5-25
I value was observed in stage 5, followed by stage 3. years), and the distribution of subjects among the stages
The lowest mean value was observed in stage 1, followed was not even.22,24 Furthermore, no sex discrimination
by stage 6. Mean IGF-I values recorded at each stage of was implemented.
the cervical vertebral maturation index were statistically Our study included only physiologically balanced
different from the values recorded at the other stages as subjects because certain diseases have a direct effect
denoted by the letters in Table II, from a for the highest on IGF-I metabolism and serum levels: eg, diabetes
value to f for the lowest value. mellitus and liver diseases.15,30 Subjects with

American Journal of Orthodontics and Dentofacial Orthopedics November 2012  Vol 142  Issue 5
658 Ishaq et al

Fig 3. The 6 stages of cervical vertebral maturation according to the method of Hassel and Farman1
cropped from lateral cephalometric radiographs of the subjects.

bleeding disorders were excluded so that no
complications would occur during collection of the
blood samples.22 The integrity of the cervical vertebral
area was essential for the proper evaluation of its mor-
phology on which the stage of cervical maturity is de-
termined.
The IGF-I assay was performed with an ELISA IGF-I
technique that was advocated by other researchers.31,32
Other researchers adopted different techniques such as
radioimmunoassays18,22,24 and immunoradiometric
assays.19 Different techniques including ELISA, radioim-
munoassays, and immunoradiometric assays used in
IGF-I analysis were compared for accuracy and were
also compared by using samples from normal and dia-
betic patients. It was concluded that the different assays
were comparably accurate, especially in healthy sub-
jects.33 The radioimmunoassay technique requires spe-
cial laboratories that should have been equipped for
radiation control. The ELISA technique was used in
this research because of the availability of the kit, and
the applicability and accuracy of the technique.
Cervical vertebral maturation had been extensively
studied and proven to be a reliable diagnostic tool that
is routinely applied in orthodontics as an indicator of
the status of maturation.1,5,34-36 This method was also
found advantageous compared with the hand-wrist
skeletal maturation indicator, which requires additional
radiographic exposure.34-37
Several methods for the assessment of cervical verte-
bral maturation are available. In this study, the IGF-I se-
rum levels were evaluated relative to the cervical
vertebral maturation index as described by Hassel and
Farman.1 It was evaluated by several authors in correla-
tion with the hand-wrist method and determined to be
reliable.34-37
To have an accurate assessment of the morphologic
changes of the cervical vertebrae, a standardized step-
wise method was introduced in this research. These

November 2012  Vol 142  Issue 5 American Journal of Orthodontics and Dentofacial Orthopedics
Ishaq et al 659

Table II. Descriptive statistics and comparisons between IGF-I values in the different cervical vertebral maturation
stages with the Kruskal-Wallis test
IGF-I

Whole sample Males females

Mean value Mean value Mean value

Stage n Mean age (y) (ng/mL) SD n Mean age (y) (ng/mL) SD n Mean age (y) (ng/mL) SD P value
Stage 1 22 10.3 209.3f 61 10 11 187f 30 12 9.8 227e 74 \0.001*
Stage 2 23 11.7 298.6d 85 13 12.2 309d 93 10 11.2 284d 75 \0.001*
Stage 3 19 13.1 519.7c 175 10 13 463c 164 9 13.1 582b 175 \0.001*
Stage 4 24 14 835.6a 201 10 14.5 893a 171 14 13.7 794a 217 \0.001*
Stage 5 15 15.3 617.3b 175 8 15.6 681b 122 7 14.9 544c 207 \0.001*
Stage 6 17 16.6 253.5e 111 9 17.3 230e 119 8 15.8 279d 104 \0.001*

*Significance level was set at P #0.05. Means with different letters are statistically significantly different according to Mann-Whitney U test.

Fig 4. Bar chart representing mean values of IGF-I with different cervical vertebral maturation stages in
the whole sample (blue), in males (red), and in females (green).

measurements made observations more consistent and
accurate.
The results of our study demonstrated the pattern of
IGF-I serum levels in the circumpubertal period. The
levels were lowest at the stages of initiation and acceler-
ation and gradually rose until peak values were reached
in the stage of deceleration. The levels then gradually de-
clined as pubertal growth ceased, to reach baseline levels
at the stages of maturation and completion. IGF-I serum
levels peaked at stage 4 of the cervical vertebral matura-
tion index with a mean value of 835.6 ng/mL.
Mean IGF-I values recorded at each stage of cervical
vertebral maturation were statistically different from the
values at the other stages as denoted by the letters in
Table II from a for the highest value to f for the lowest
values. The values were specific for each stage, denoting
that they can be used to differentiate one stage from
another. In the female group, stages 2 and 6 showed
no statistical differences.
Previous research correlated IGF-I with cervical verte-
bral maturation in a group of 83 subjects aged from 5 to
25 years.22 The authors of that study reported that serum
levels of IGF-I peaked at stage 5 of cervical vertebral
maturation, and the highest mean IGF-I value was
406.8 mg/L (equivalent to 406.8 ng/mL). The differences
might be attributed to (1) the difference in the popula-
tion studied, since our study was conducted with Egyp-
tians, and the other study was conducted with Saudis;
(2) 2 laboratory techniques were implemented in the
studies to measure the IGF-I levels; and (3) our study
had a larger sample size.
Masoud et al24 correlated mean IGF-I values to the
stages of the skeletal maturation index as described by
Fishman.23 They reported that mean IGF-I values were

American Journal of Orthodontics and Dentofacial Orthopedics November 2012  Vol 142  Issue 5
660
Fig 5. Pattern of IGF-I in relation to the stages of the cervical vertebral maturation index.
low at the prepubertal stages (stages 1-3 of the skeletal
maturation index) and highest at the peak growth velocity
(stages 6-8 of the skeletal maturation index), with a mean
value of 359 mg/L (equivalent to 359 ng/mL), and de-
creased gradually as maturation progressed to approach
its prepubertal values. According to Hassel and Farman,1
stages 6 to 8 of the skeletal maturation index occurred in
concordance with stages 3 and 4 of cervical vertebral mat-
uration index. The peak growth velocities in height and
mandibular growth have been shown to occur between
stages 3 and 4 of the cervical vertebral maturation in-
dex.5,14 This supported the findings of our study, in
which the rise in mean values of IGF-I started in subjects
in stage 3, peaked at stage 4, and declined gradually in
stage 5 as pubertal growth approached its final stage.
Upon comparing the male and female groups, we
found a statistical difference in the mean IGF-I values
in stages 3 and 5 of cervical vertebral maturation. Stage
3 in girls had higher mean values than in boys, whereas
stage 5 in boys had higher mean values than in girls.
Such sex differences were not studied in previous studies
that correlated IGF-I to skeletal maturation.22,24 Bereket
et al20 found that peak IGF-I concentrations were
reached 1 year earlier and in 1 pubertal stage earlier in
girls than in boys, and they started to decline thereafter.
This agreed with our findings.
Furthermore, the chronologic ages for peak IGF-I
values in this study for males, females, and the whole
sample were at ages 14.5, 14, and 14 years, respectively.
This agrees with the study of Ball et al,14 who reported
that the mean age for peak mandibular growth velocity
was 14 years for boys. They also reported that cervical
vertebral maturation stages should be used with other
methods of biologic maturity assessment when
November 2012  Vol 142  Issue 5 American Journal of Orthodontics and Dentofacial Orthopedics
Ishaq et al
considering both dentofacial orthopedic treatment and
orthognathic surgery to ensure accuracy.
CONCLUSIONS
1. IGF-I mean values can be used in orthodontic diag-
nosis as a reliable maturation indicator that is com-
patible with the cervical vertebral maturation index.
2. IGF-I mean values increased gradually from stage 1
(initiation) of cervical vertebral maturation to the
peak level at stage 4 (deceleration) and then de-
clined gradually to approach baseline levels at stage
6 (completion).
3. Stages 3 and 4 showed a difference between boys
and girls. Girls had higher values in stage 3 (transi-
tion), indicating their earlier onset of puberty, and
boys showed higher values in stage 5 (maturation),
indicating their more delayed pubertal growth spurt.
We recommend that a longitudinal study should be
conducted to confirm the usefulness of the variations
in IGF-I serum levels as a predictor of skeletal maturation
and the timing of the pubertal growth spurt and to cor-
relate the mandibular growth pattern with IGF-I serum
levels at the different stages of maturation.
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