Bone Mass Gain During Puberty and Adolescence: Deconstructing Gender Characteristics

Abstract

Primary prevention of osteoporosis must aim at increasing bone mass acquisition before late adolescence. During pubertal years both genders reach peak bone acquisition, though males develop a greater skeletal mass. This dimorphism is largely regulated by endocrine factors, with critical roles played by gonadal steroids, growth hormone and insulin growth factor-1, amongst the most important. Menstrual history is a surrogate for the adequacy of hormonal functioning, nutrition and physical activity that may be a marker of bone status and development in young women. Adequate levels of adrenal, reproductive and pituitary hormones, growth factors and leptin are needed for the initiation and maintenance of regular menstrual cycles as well as for the achievement of peak bone mass. Adequate regular exercise and body composition are also pivotal elements in maintaining normal mechanical bone stimulus during bone growth. Avoidance of carbonated soft drink consumption, or excessive alcohol and any tobacco should be considered as these may interfere reaching adequate bone mass.

Full text

Current Medicinal Chemistry, 2010, 17, ????-???? 1
0929-8673/10 $55.00+.00 © 2010 Bentham Science Publishers Ltd.
Bone Mass Gain During Puberty and Adolescence: Deconstructing Gender
Characteristics
F.R. Pérez-López*, P. Chedraui and J.L. Cuadros-López
1Department of Obstetrics and Gynecology, Universidad de Zaragoza, Hospital Clínico de Zaragoza, Zaragoza, Spain;
2Instituto de Biomedicina, Facultad de Ciencias Médicas, Guayaquil, Ecuador; 3Department of Obstetrics and Gynecol-
ogy, Universidad de Granada, Hospital Clínico San Cecilio, Granada, Spain
Abstract: Primary prevention of osteoporosis must aim at increasing bone mass acquisition before late adolescence. Dur-
ing pubertal years both genders reach peak bone acquisition, though males develop a greater skeletal mass. This dimor-
phism is largely regulated by endocrine factors, with critical roles played by gonadal steroids, growth hormone and insulin
growth factor-1, amongst the most important. Menstrual history is a surrogate for the adequacy of hormonal functioning,
nutrition and physical activity that may be a marker of bone status and development in young women. Adequate levels of
adrenal, reproductive and pituitary hormones, growth factors and leptin are needed for the initiation and maintenance of
regular menstrual cycles as well as for the achievement of peak bone mass. Adequate regular exercise and body composi-
tion are also pivotal elements in maintaining normal mechanical bone stimulus during bone growth. Avoidance of carbon-
ated soft drink consumption, or excessive alcohol and any tobacco should be considered as these may interfere reaching
adequate bone mass.
Keywords: Osteoporosis, bone mass, puberty, adolescence, exercise, nutrition, menstruation.
1. INTRODUCTION
Bone mass content (BMC) and quality are fracture risk
predictors. BMC increases mainly during the first three years
of life and during puberty growth spurt. The process of bone
accretion is genetically determined, with some changing in-
fluences, in the widest sense, due to lifestyle. Peak bone
mass (PBM) is achieved early in the third decade of life [1,
2], with 26% of adult BMC acquired during the 4-year pe-
riod around peak height velocity [3] and up to 60% during
the remaining peripubertal years [4, 5]. Therefore, adoles-
cence is the period when the greatest BMC accrual occurs.
Endogenous estrogens and androgens independently exert
effects on bone acquisition and maintenance [6]. Growth
hormone (GH), insulin growth factor-1 (IGF-1), cortisol,
thyroid hormones, parathyroid hormone (PTH), vitamin D
and leptin may also influence bone metabolism during pu-
berty [7, 8]. These hormones may directly or indirectly affect
osteoclast and osteoblast functions.
Obviously there are gender differences in the mecha-
nisms and changes associated with puberty, which determine
growth velocity and PBM achieved in young individuals.
During female puberty, women acquire some 40% of PBM
[9]. Young women attain 92% of total BMC by age 18, and
99% by age 26 [10]. However, PBM acquisition is not a ho-
mogeneous process; PBM is attained at the femoral neck by
age 16 while in lumbar spine bone mass increases up to age
30 [1, 5, 11]. In girls bone mineral accrual rates reach the
maximum around age 11-14 while in boys there is a 2-3 year
delay due to pubertal gender differences. Maximizing PBM
is advocated as a way to prevent osteoporosis. The present
review aimed to deconstruct modifiable factors that may
interfere with bone mass gain during adolescence with spe-
cial emphasis in the female gender.
*Address correspondence to this author at the Department of Obstetrics and
Gynecology, Universidad de Zaragoza, Facultad de Medicina, Domingo
Miral s/n, Zaragoza 50009, Spain; Tel: +34 976-76-1734; Fax; +34 976-76-
1735; E-mail; faustino.perez@unizar.es
2. BONE MASS ASSESSMENT DURING ADOLESCE-
NCE
Bone mass assessment in children and adolescents is dif-
ficult due to ethical as well as methodological aspects. Dual
X-ray absorciometry (DXA) is the most widely used tech-
nique for measuring bone acquisition during adolescence due
to its low cost and easy use. Bone mass is a composite meas-
ure of bone size and bone mineral density (BMD) which is a
determinant of bone strength and depends on the acquired
mass during skeletal growth and development. Total-body
BMC measured with DXA has been proposed as the best
outcome measure for bone mass status during growth and
maturation [12, 13]. However, BMC evaluation by DXA has
some limitations due to growth-related variations in bone
size and composition. The technique is based on a two-
dimensional projection of the three-dimensional bone struc-
ture, and results are a sum of cortical and trabecular BMC
within the projected bone area. DXA renders BMC in grams
for a determined bone. The quotient of BMC by the bone
projected area renders the areal BMD (g/cm2). High variation
coefficients were obtained when regression equations for
total body and the different skeleton parts were calculated
with either BMC or BMD as the dependent variable, and
age, body height and body weight as independent variables
[14]. World Health Organization diagnostic categories for
normal, osteopenia, and osteoporosis, based on central BMD
T scores, are not applicable to children and adolescents who
have not yet reached PBM.
In women aged 9 to 22 years, distal forearm and calcane-
ous BMD has been measured using DXA, and agreement
with central BMD was established. Higher BMD values were
found in the total skeleton than in calcaneous and the distal
forearm, with significant moderate to high positive correla-
tion coefficients [15]. In children and adolescents, aged 3 to
18, distal radius BMD, as measured by DXA, correlates with
DXA lumbar spine BMD measurements, and their corre-
spondent correlation coefficients did not significantly differ
for chronological age, weight, height, body surface, body
mass index (BMI) and bone age [16].

2 Current Medicinal Chemistry, 2010 Vol. 17, No. 1 Pérez-López et al.
BMC and areal BMD assessed with DXA and vertebral
volume calculated by quantitative computed tomography
(qCT), showed that DXA BMC is a more accurate and reli-
able measure than DXA BMD for assessing bone acquisition
in the early stages of sexual development [17]. Cortical area
of the radius has been measured with peripheral qCT show-
ing that pubertal stage and the interaction of pubertal sex
stage had a significant influence on cortical radius [18].
However, when DXA values for BMD are assessed by Tan-
ner stage and bone age, the results express bone content [17].
Bone status has been assessed using broadband ultra-
sound attenuation techniques at the calcaneus in children and
adolescents [19]. Due to its low cost, device portability, and
safety, broadband ultrasound attenuation has been used in
clinical setting. However, factors influencing its reproduci-
bility in children remain an issue.
Another difficulty concerning bone mass assessment dur-
ing puberty could be attributable, at least in large part, to the
cross-sectional design of studies, imposing difficulty in the
analysis of implicated factors such as gender, ethnicity, age,
and body in relation to bone size [20].
3. ENDOCRINE INFLUENCES ON BONE MASS
ACCRUAL DURING ADOLESCENCE
Puberty is a developmental and biological process that is
integrated to the complex psychosocial maturation of adoles-
cence. Adolescence is a critical time for bone mass accrual,
and bone mass increase through puberty is dependent on
rising levels of gonadal steroids, GH and IGF-1. Leptin and
other hormones also participate in body composition changes
and bone metabolism. Adolescence is a high bone-turnover
state, with increased levels of both bone formation and re-
sorption that decreases to adult levels in late puberty [21,
22].
Bone mass determinants have been studied in female
twins aged 10 to 26 years, using DXA measurements at dif-
ferent central skeletal sites. Mean BMD increased with age
to around 16 years, when it reached a plateau. Only men-
archeal status, height, and lean mass, were independent pre-
dictors of BMD. Total body BMC was independently associ-
ated with menarcheal status, height, lean mass, and fat mass,
being the effects of the latter two variables stronger in pairs
both premenarcheal . However, after adjusting for constitu-
tional factors, no lifestyle factor was independently predic-
tive [23].
3.1. Steroid Hormones, Puberty and Bone Accrual
Endogenous androgens and estrogens are the engine that
positively stimulates bone growth and mineral acquisition
during the transition from child to adulthood in both males
and females. The physiological increase of androgens during
adrenarche is the first significant endocrine step that initiates
growth spurt and rapid bone acquisition. Adrenal androgens,
at physiologic range, stimulate bone mineral metabolism in
prepubertal healthy children before the appearance of pubic
hair which is measurable in the proximal radial diaphyseal
bone strength [24]. Girls with premature adrenarche show a
transient increase in height and bone age as well as hyperin-
sulinism. In children with premature adrenarche aged 5.4-8.6
years mean BMI was higher than that of healthy children of
the same age. IGF-1, IGF-binding protein (IGFBP) 3 and
leptin levels of lean premature adrenarche girls were higher
than controls. Leptin levels of obese premature adrenarche
girls were higher than that of lean premature adrenarche girls
and controls [25].
BMD measured by DXA correlates with pubertal stage in
boys and girls [4, 26]. Martin et al. [26] measured BMD and
estimated calcium accretion using annual DXA measure-
ments for 4 years. Data were used to calculate peak height
velocity and peak BMD and ages at which they occurred
(boys: 13.3 years; girls: 11.4 years). Boys displayed higher
BMC and BMD as compared to girls, and differences in-
creased through puberty. During pubertal maturation differ-
ent BMD-related endpoints were lower in girls than in boys.
Greater male BMC has been postulated to be due to the spe-
cific effects of androgens, although it has also been related to
increases in mechanical loading as opposed to gender [27,
28].
During early puberty, sex steroids stimulate pubertal
growth spurt in conjunction with GH and IGF-1. The fast
increase in height is associated to an increase in bone turn-
over and bone mineral apposition. Girls that start early men-
strual function or receive combined hormone contraceptive,
have a higher BMC as compared to those without menstrual
function. BMC and increases in bone remodeling have been
explained by high estrogen, GH and IGF-1 levels [21, 22, 29,
30]. Chronological age is solely associated with lumbar
BMD, whereas postmenarcheal age is associated to both
lumbar and total hip BMD [30]. Increase in BMC and bone
turnover persists throughout growth spurt in girls, even when
growth velocity declines [21, 22, 29, 30].
A 2 year longitudinal study in healthy pubertal girls (10
to 13 years) correlated sex hormones and bone markers with
bone geometry and density. The latter two were assessed
with peripheral qCT measurements of the left tibial shaft
performed at baseline and at 1- and 2-years follow-up [31].
At the tibia shaft, estradiol and testosterone showed different
effects: estradiol was a predictor of total BMD, cortical
thickness and a negative predictor of endocortical circumfer-
ence whereas testosterone was a positive predictor for total
cross-sectional area and periosteal circumference and a nega-
tive predictor for total BMD. Thus, it seems that at the en-
docortical surface estradiol inhibits bone resorption during
rapid growth, and after menarche high estradiol concentra-
tions promote bone formation. At periosteal surface, testos-
terone induces bone formation while estradiol is devoid of
effect.
Later menarcheal age was associated with low PBM as
assessed in subjects followed up from 7.9 to 20.4 years using
DXA measurements in six anatomical sites. When later
(14.0±0.7 years) and earlier menarcheal aged (12.1±0.7 yr)
subjects were compared, areal BMD was lower in the later
than the earlier menarcheal group at all sites at mean ages
10.0, 12.4 and 16.4 years, and before pubertal maturation.
Therefore, a deficit in areal BMD was detectable in later
menarcheal girls before the onset of pubertal changes [32].
Thus, a postulated genetic cause of low PBM is plausible
rather than shorter estrogen exposure.

Bone Mass Gain and Adolescence Current Medicinal Chemistry, 2010 Vol. 17, No. 1 3
It is not clear if delay in female puberty reduces bone
mass gain and increases fracture incidence. Short-term delay
of puberty on bone mass and structure has been assessed in
experimental conditions by histomorphometry [33]. For in-
stance, pubertal and gonadal retarded development induced
by GnRH antagonist injections in rats produced a transient
reduction in trabecular bone area as assessed by histomor-
phometry. It seems that any "catch-up" growth following the
cessation of the GnRH-antagonist injection protocol affects
trabeculae oriented perpendicular to 0 degrees at the expense
of trabeculae in other orientations. In rats, hypothalamic
suppression before the onset of puberty, as compared to af-
ter, would result in a relatively large bone strength deficit
and hence more compromised. In addition, increases in body
weight during or prior puberty may not be protective of bone
strength [34].
Data regarding the effect of GnRH analogues on BMD
acquisition in healthy children is lacking. In a small series of
14 girls with progressive CPP, GnRH analogue treatment for
an average of 4 years (6.2 to 10.1 years) improved height and
despite being discontinued no impairment of PBM achieve-
ment was observed [35]. Long term effects of GnRH ago-
nists have been assessed in a multicentric study performed
among young women aged 16.7 years (range: 12.9-23.4
years) over a 4.4 years mean period (range: 1.0-9.7 years).
After treatment 78% of all patients reached a final height
falling within the targeted one without any negative effect on
lumbar spine BMD, as assessed with DXA [36]. In children
with central precocious puberty (CPP), available data
indicate that therapy with GnRH analogues can improve
final height within the targeted height and increase BMI
[37]. Despite this, there are some safety concerns regarding
GnRH analogue treatment in children and adolescents [38,
39]. GnRH treatment has been proposed to be administered
in association with calcium or norethindrone acetate add-
back therapy in order to preserve bone mass achievement
[40, 41],
3.2. Growth hormone and IGF-1
GH affects bone size and mass in part through stimulating
IGF-1 production in the liver and bone. Both are pivotal in
achieving longitudinal bone growth and bone mass [42]. GH
enhances longitudinal bone growth by stimulating prechon-
drocytes in the growth plate followed by a clonal expansion
due to GH induced local and circulating IGF-1 production.
GH directly stimulates osteoblasts in different species, in-
cluding humans, at concentrations that are in the physiologi-
cal range. Response is expressed by osteoblast proliferation
and its production of osteocalcin and type I collagen. GH
regulates circulating IGF-1 while bone IGF-1 production is
regulated by estrogen, PTH, cortisol and local growth fac-
tors. In vitro, GH regulates osteoclast formation, showing,
under different circumstances, both stimulatory and inhibi-
tory mechanisms.
IGFs are the most abundant growth factors stored in bone
and produced by osteoblast cells [43]. The understanding of
IGF actions over bone metabolism can be hampered by the
existence of both systemic (mediated by GH effect on skele-
tal growth) as well as their local actions over several growth
factors [44, 45]. The modulating action of the six IGFBPs
has additional complexity since free IGF-1 levels may suffer
substantial changes [43]. At the cellular level, IGF-1 acts
through its ubiquitously expressed receptor to promote cellu-
lar proliferation and to inhibit apoptosis [46]. Experimental
studies in IGF-1 knockout, IGF-2 knockout, and GH-
deficient mice showed that disruption of IGF-1, but not IGF-
2, completely prevented periosteal expansion that occurs
during puberty, whereas it was reduced by 50% in GH-
deficient mice. Therefore, GH/IGF-1, but not IGF-2, is criti-
cal for puberty-induced bone growth [44].
IGF-1 participates in regulating bone turnover. In a cross-
sectional prospective study performed in healthy children
serum IGF-1 levels were positively correlated with bone
formation (bone alkaline phosphatase) and bone resorption
marker (CrossLap) levels before and after puberty. There
was also a weak correlation between IGF-1 and CrossLaps
during puberty. In addition, serum IGFBP-3 levels were
positively correlated with bone alkaline phosphatase and
CrossLap levels before, during (weak correlation) and after
puberty. These results point out the importance of the IGF-
GH axis on bone metabolism with mild differences across
different phases of puberty probably caused by the evolution
of sex steroids [47]. Experimental results using mutant mice
showed that IGF-1 improved BMD, BMC and body compo-
sition during the puberty transition [48].
3.3. Fat Mass and Leptin
In mammals, puberty initiation is coupled to a nutritional
and metabolic state. In prepubertal female rats, all indices of
pubertal maturation-age were significantly delayed in the
pair-fed group but not different between the leptin-treated
group and ad lib-fed controls suggesting that leptin is not the
primary signal that initiates the onset of puberty yet rather
having a permissive effect when adequate metabolic re-
sources are deemed. In addition, other metabolic factors,
besides leptin, influence the timing of puberty onset under
conditions of severe dietary stress [49]. In adolescents aged
13-18 years, the sum of six skin folds and body fat percent-
age are significantly higher in females than in males [50]. Fat
mass and obesity probably influence bone mass in children
and adolescents. Weight and BMI are independent positive
correlates of BMD in postmenarcheal girls [30, 51]. These
associations have been explained by the impact of greater
weight-bearing on BMD of girls with elevated body weight.
Relationships between both fat mass and two genetic
child obesity variants and DXA bone measure scans at a
mean of 9.9 years were assessed in 7,470 children from the
Avon Longitudinal Study of Parents and Children. Total fat
mass was strongly associated to total body, spinal, and upper
and lower limb BMC. After adjusting for puberty, similar
results were reported when trunk fat mass was used instead
of total fat, and when bone was used instead of bone mass.
When the two genetic markers were used as tools of fat
mass, similar associations between BMC and fat mass were
observed [52]. It seems that a causal relationship exists be-
tween fat and bone mass.
In normal children and adolescents, the effects of weight,
lean tissue mass, and fat mass over BMC are unclear. Lean
mass has a strong effect on bone gain during puberty, al-
though fat mass becomes a stronger positive predictor after

4 Current Medicinal Chemistry, 2010 Vol. 17, No. 1 Pérez-López et al.
pubertal growth [53]. In children aged 5-17 years, a positive
association exists between lean tissue and fat mass and BMC
[54]. Leptin may be the biochemical mediator that explains
the association between fat mass and BMD [55, 56]. Leptin
may have a dual effect on bone: it may decrease bone forma-
tion by a central nervous effect and may stimulate both bone
formation and resorption by direct effects on stroma precur-
sor cells [56]. Weight loss in post-pubertal obese adolescents
is associated to an improvement of metabolic parameters
[57].
Late menarche observed among elite adolescent athletes
has been attributed to change in body composition as com-
pared to girls that do not participate in such training. How-
ever, Sherar et al. [58] assessed body composition by DXA
in 61 girls between -2 and +2 years from menarche. There
was no reduction at menarche in the range of body mass,
body fat percent and total body fat which is contrary for a
critical body or fat mass around menarche. It remains to be
demonstrated which nutritional alterations may contribute to
the later age of menarche in these girls.
4. INFLUENCE OF PHYSICAL ACTIVITY DURING
PUBERTY AND BONE METABOLISM
In growing rats, regular exercise increases serum osteo-
calcin and 1,25-dihydroxyvitamin D3 [1,25(OH)2D] while
decreasing serum PTH levels. While 7 weeks of exercise
reduced urinary deoxypyridinoline levels, 11 weeks of exer-
cise increased serum alkaline phosphatase levels and de-
creased serum tartrate-resistant acid phosphatase levels. Both
durations of physical activity increased femoral length and
tibial BMC, while lumbar BMC was not altered. Therefore,
exercise induced metabolic changes and increases bone mass
and longitudinal bone growth, especially at weight-bearing
sites [59].
Physical activity during puberty and in young women
plays a relevant role in bone acquisition. Regular exercise
has a positive effect in gaining and maintaining bone during
adolescence and later [60-62]. Girls and boys and young
adults who exercise regularly achieve greater PBM than
those who do not. It is well known that immobilization rap-
idly produces bone mass loss, suggesting the importance of
skeletal loading to maintain or gain bone mass. Experimental
observations have demonstrated that bone acquisition re-
quires bone loading with changes in intensity and dynamics
[63]. In fact, adult bone mass is positively associated with
childhood activity and exercise interventions, confirming a
positive association between exercise and bone health across
human life [64, 65].
Sedentary activity in prepuberty is inversely associated
with BMD in young adults while a dietary calcium intake >
1,000 mg/day in adolescence is associated with higher BMD
in women aged 21-24 years [66]. Thus, it is recommended to
ensure adequate calcium intake (> 1,000 mg/day) and physi-
cal activity in order to gain enough bone mass during female
puberty. Exercise exerts its effects on the skeleton through
mechanical strain and remodeling action (Fig. 1). Thus, both,
higher intensity level and ground force impact are associated
with higher BMD in young adults [67, 68]. A key character-
istic of loading is that very few loading cycles are actually
required to elicit new bone formation [69]. Therefore, chil-
dren and adolescents should not have to participate in labori-
ous or complex exercise to disrupt their activities. In addi-
tion, a given number of loading cycles will also be more os-
teogenic if they are broken up into shorter bouts including
rest periods in between [70]. This is due to bone cell sensi-
tivity to loading which returns after a rest period. This bone
mechanical sensitivity should be translated to daily life,
changing from sedentary status to outdoor leisure as it once
was (before TV and the computer era).
Muscle force is the largest loading source applied to
bones. In healthy girls aged 10-13 years BMC and total body
lean and fat mass were assessed by DXA at baseline and 2
years later. Local muscle contraction and weight-bearing
exert an additive effect on bone mass accretion in the lower
limbs in growing children [71]. Bone mass strength is influ-
enced by lean tissue mass accrual during the adolescence
growth spurt. In individuals aged 8-18 years, total body lean
mass was assessed for 6 consecutive years using DXA and
femoral characteristics by hip structural analysis. Peak lean
tissue mass accrual was a significant predictor of the magni-
tude of two variables of bone strength assessed at femoral
sites, supporting the fact that muscle development is an im-
portant factor in affecting bone strength during adolescence
growth spurt [72].
Physical fitness has been proposed as a major marker of
health status. Children and adolescents, especially girls,
should be encouraged to participate in any type of physical
activity, giving preference to metabolically intense activities,
such as basketball, soccer or jogging. Playing sports that
generate high ground reaction appear to stimulate a more
powerful osteogenic response than practicing an activity that
has low ground reaction forces, like yoga or swimming [73,
74]. Prepubertal and early pubertal exercise is more advanta-
geous than post-pubertal exercise in inducing mechanical
response among growing bone [55, 75-77].
5. NUTRITION AND BONE MASS
During adolescence several nutritional factors play a ma-
jor role in the bone mass gain process. Some of the nutrients
and food components consumed as part of a Westernized diet
can potentially impact bone accrual during adolescence. Die-
tary factors that may affect bone metabolism include inor-
ganic minerals (e.g., calcium, magnesium, phosphorus, and
various trace elements), vitamins (D, K, C, and certain B
vitamins), and macronutrients, such as proteins (Fig. 2).
Nutritional status, physical fitness and physical activity
among adolescents influence body composition, metabolism
and immunological status [78] which alter bone metabolism
and mass gain.
5.1. Calcium
Calcium is an essential nutrient for bone health in all age
groups. Adequate calcium intake is critical to achieve opti-
mal PBM and for modifying the rate of bone loss associated
with aging [79]. Calcium must be released in a soluble, and
probably ionized, form before it can be absorbed. The solu-
ble form of calcium is absorbed by transcellular and paracel-
lular transport [80]. Dairy products have been shown to pro-

Bone Mass Gain and Adolescence Current Medicinal Chemistry, 2010 Vol. 17, No. 1 5
vide 77% of dietary calcium among teenage girls [81], but
girls as young as 10 to 12 years of age have low calcium in-
takes [82].
Calcium deficiencies in young individuals can account
for a 5 to 10% difference in PBM and can increase the risk
for hip fracture later in life. Teenage girls of the Western
world are less likely than teenage boys to get enough cal-
cium. In fact, fewer than 10% of girls aged 9 to 17 actually
obtain the calcium they need each day. On the other hand
there is no consensus regarding the minimal calcium needed
dose to obtain a maximal PBM [83]. The US Institute of
Medicine, Food and Nutrition Board [79] recommends a
calcium dosage of 1,300 mg/day in children and adolescents
aged 9 to 18 years.
Milk avoidance may contribute to lower BMC through a
reduction in calcium and limited dietary protein intake. Low
milk diet and milk avoiding girls may have smaller skeletal
size due to the loss of the mild-induced increase of IGF-1
[84]. In addition, self-imposed restriction of dairy foods is
associated with lower spinal BMC values (and non-
significant similar patterns for total body, total hip, and
femoral neck BMC) as compared with girls without milk
intake restriction [85].
Since the effect of calcium intake varies by skeletal site
(cortical responding more than trabecular bone), there is no
consensus regarding the minimal amount of calcium required
to achieve maximal bone accretion [83]. In addition skeletal
calcium needs are also influenced by the degree of physical
activity [86]. In a cross-sectional multicenter investigation
performed among six European countries, healthy Caucasian
girls aged 11-15 years were selected from larger population
samples to represent a large range in calcium intake. Mean
calcium intake among girls and adult women varied between
some 600 mg/day in Italy and 1,200 mg/day in Finland. Af-
ter adjustment for age, height, weight, Tanner stage, and
bone area for the girls, radial BMD measured with DXA did
not significantly vary among quartiles of calcium intake.
There was no evidence for a different relation between cal-
cium and BMD at different levels of intake, suggesting that
dietary calcium is not a determinant of peak BMD in Euro-
pean girls [87].
In a 2 year randomized placebo-controlled study per-
formed among healthy 10 to 12 year old girls at Tanner stage
I-II, the effect of calcium supplementation on bone mass
accrual and body composition has been determined. Treat-
ments were calcium (1,000 mg) plus vitamin D3 (200 IU),
calcium (1,000 mg), cheese (1,000 mg calcium), and pla-
cebo. Outcomes were hip, spine, and whole body bone indi-
ces assessed by DXA and of the radius and tibia by periph-
eral qCT. When treatment compliance was > 50%, calcium
supplementation with cheese was associated with a signifi-
cantly higher percentage change in cortical thickness of the
tibia than did placebo, calcium, or calcium plus vitamin D
treatment and in higher whole-body BMD than did placebo
treatment [88]. It seems that some cheese constituents act as
calcium absorption enhancers. In fact, individual milk com-
ponents, such as lactose, lactulose, and casein phosphopep-
tides may enhance calcium bioavailability [89, 90].
The effect of calcium supplementation on BMC and
BMD, measured at the midshaft and distal radius, has been
studied in a randomized double blind, placebo-controlled
study carried out in rural Gambian children who received
1,000 mg calcium carbonate/day or placebo for 12 months.
There were significant higher BMC and BMD in children
that received calcium supplement, although there were no
effects on height, weight, or bone width at the midshaft or
distal radius. At the end of the study osteocalcin levels were
lower in the treated group as compared to the placebo one
[91].
Fig. (1). Effect of physical activity on bone mass gain during puberty and adolescence.

6 Current Medicinal Chemistry, 2010 Vol. 17, No. 1 Pérez-López et al.
5.2. Magnesium
Magnesium (Mg) is a determinant of BMD in adults. In-
terval BMC change was assessed at different skeletal sites in
healthy 8 to 14 year old Caucasian girls with Mg intake <
220 mg/day that received Mg (300 mg/day in two divided
doses) or placebo for 12 months. A significant increase in
hip BMC accrual was observed in Mg-treated girls as com-
pared to the placebo group. A non significant spinal BMC
increase was also seen. There were no differences in serum
mineral, calciotropic hormones and bone markers during
treatment [92].
5.3. Vitamin D
Vitamin D is both a nutrient and a hormone. Vitamin D
maintains serum calcium and phosphorus levels in the nor-
mal range, thus promoting bone mineralization. Vitamin D is
an independent bone building factor during young ages. Vi-
tamin D is largely produced in the skin after ultraviolet light
exposure while vitamin D ingestion is a secondary source to
maintain endogenous levels [93, 94]. Serum 25-hydroxy-
vitamin D3 25(OH)D is the best indicator of vitamin D status
because it reflects both digestive absorption and cutaneous
photosynthesis.
In young rats under mild calcium deficiency, increased
serum PTH and 1,25(OH)2D with decreased serum 25(OH)D
levels stimulated intestinal calcium absorption and renal cal-
cium reabsorption, and reduced maturation-related cortical
bone gain but did not significantly influence maturation-
related cancellous bone gain. Vitamin D supplementation
stimulates intestinal calcium absorption and prevents the
reduction in maturation-related periosteal bone gain by in-
ducing calcium accumulation from cancellous and endocor-
tical bone [95]. In humans, calcium absorption efficiency
increases with increasing levels of serum 25(OH)D up to the
threshold of approximately 80 nmol/L (32 ng/ml) [93, 96,
97], while PTH levels rise when 25(OH)D levels are be-
tween 30 and 50 nmol/L [98, 99]. These levels are associated
to adequate levels of the bioactive hormone 1,25(OH)2D
[100]. Therefore, the maximal fractional calcium absorption
cut-off should be a primary end-point to maintain health and
cell functions which are at a higher level than that associated
with rickets in children and osteomalacia in adults (25(OH)D
< 20 nmol/L).
Vitamin D insufficiency is very common among children
and adolescents. Sustained very low serum 25(OH)D levels
are associated with rickets, although insufficient levels (< 50
nmol/L) may alter calcium absorption and bone mineraliza-
tion. Adolescent vitamin D serum level is an independent
factor for bone mass gain. Serum calcium and vitamin D
levels are associated with total body and hip BMD, although
associations are reduced when adjusted for age, ethnicity and
swimming status [101]. Outila et al. [102] reported in female
adolescents a significant higher radial BMD using a serum
25(OH)D concentration cut-off of 40 nmol/L. Using periph-
eral qCT, Cheng et al. [84] demonstrated a progressive in-
crease in cortical BMD at both the distal radius and the tibia
shaft, although there were no differences based on 25(OH)D
levels in total femur, lumbar spine, or the whole body as
evaluated by DXA. In a prospective study, a relationship was
found between 25(OH)D levels and BMD at femoral neck
and spine when time prior to the onset of menarche was con-
sidered [103].
Relationships between serum 25(OH)D levels and turn-
over markers or BMD are controversial [104-107]. Low se-
rum 25(OH)D has been reported in 68% of adolescent boys,
with an inverse association found between 25(OH)D levels
and both pyridinoline and serum bone alkaline phosphatase
[108]. Although bone markers are increased in patients with
low 25(OH)D levels the markers cannot be used in individ-
ual patients to infer their 25(OH)D levels.
The effect of low-vitamin D status on BMC and BMD
for whole body and distal and proximal forearm measured by
DXA, bone turnover, and muscle strength has been reported
in adolescent girls. Deficient (<50 nmol/L) and severely de-
ficient (< 25 nmol/L) 25(OH)D levels affected 57.8% and
Fig. (2). Effect of nutrition on bone mass gain during puberty and adolescence.

Bone Mass Gain and Adolescence Current Medicinal Chemistry, 2010 Vol. 17, No. 1 7
31.2%, respectively. Girls with adequate vitamin D status
had higher size-adjusted BMC for the whole body and distal
and proximal forearm than those with insufficient vitamin D
levels [109]. Therefore, it is reasonable to think that suffi-
cient circulating vitamin D levels are biologically relevant in
obtaining optimal bone mass during childhood and adoles-
cence. In addition, girls with adequate 25(OH)D levels sig-
nificantly had higher muscle strength and lower plasma bone
alkaline phosphatase and urine deoxypyridinoline:creatinine
ratios than those with deficient vitamin D levels. Recom-
mendation of the American Academy of Pediatrics is that
children and adolescents receive a 400 IU vitamin D daily
supplementation [110]. Larger vitamin D doses, such as 800-
1,000 IU/day, have been proposed for high risk individuals,
especially during winter [111, 112]. These doses may be
especially low for adolescents who are dark skinned, obese
or sunscreen users. Many children and adolescents would
need at least 1,000 to 2,000 IU/day or even higher doses.
5.4. Vitamin K
Vitamin K is involved in bone growth and development,
probably through its role as a co-factor in the carboxylation
of osteocalcin. In calcium-deficient young rats, vitamin K
supplementation stimulates renal calcium reabsorption, in-
creases maturation-related cancellous bone gain, and delays
the reduction in maturation-related cortical bone gain [95]. In
children and adolescents aged 3-16 years, vitamin K intake
and status are associated with decreased bone turnover in
girls consuming the typical American diet [113]. However,
vitamin K is not significantly associated with BMC in pe-
ripubertal girls [114].
In a longitudinal study, performed in healthy peripubertal
children (mean age 11.2 years), vitamin D status was deter-
mined along with other biochemical indicators and total
body, lumbar spine and femoral neck BMD values at base-
line and 2 years later. There was a wide range of interindi-
vidual vitamin K values at baseline, while improvement of
vitamin K over two years was associated with an increase in
total body BMC. The quotient undercarboxilated/carboxi-
lated osteocalcin was associated with pubertal stage, sex
hormones and vitamin D status [115].
5.5. Macronutrients
Diet proteins have a major role in bone mass status in
children and adolescents as occurs in older individuals. In
subjects aged 6-18 years (yearly controlled for 4 years), pro-
tein intake was positively associated with periosteal proximal
forearm circumference and cortical area, and BMC whereas
calcium intake had no significant effect on the parameters. In
addition, children with higher potential renal acid load had
less cortical area and BMC [116].
Fruit and vegetable intake may influence bone mass gain
during adolescence. In adults, several evidence point out to
some links between fruit and vegetable rich diets and bone
health [117, 118]. The protective effect of fruit and vegetable
over bone growth has been reported in prepubertal girls [119,
120].
In a Canadian cohort followed for 7 years from childhood
to adolescence, the effects of milk products and fruit and
vegetables on bone growth (measured by total body BMC
using DXA) have been assessed defining biologic maturity
by years from age at peak height velocity. In this cohort less
than 30% of subjects consumed the Canadian recommended
quantity of fruit and vegetables [121]. While in boys these
diet components have beneficial effects, in girls there were
no significant results [122].
5.6. Anorexia Nervosa
Anorexia nervosa is common in adolescent girls at the
time of initial PBM formation. Low bone mass content may
been seen in nearly 50% of adolescent girls with anorexia
nervosa, even in short duration cases and may persist after
despite recovery [123, 124]. Moreover, adult women with
anorexia nervosa have a high prevalence of osteopenia and
osteoporosis due to bone loss [125, 126]. Adult women with
anorexia nervosa initiated during adolescence have lower
bone mass than those with adult onset anorexia nervosa
[127]. Both hypogonadism and the cortisol excess associated
to anorexia nervosa may contribute to the development of
osteopenia and osteoporosis. However, anorexia nervosa
includes also alterations of the GH-IGF-1 axis, insulin and
ghrelin secretion [128. 129].
Bone mass loss related to anorexia nervosa can be de-
tected with DXA and should be included during its clinical
management. There are controversial results concerning the
effect of estrogen-progestin treatment in the prevention of
bone mass loss in patients with anorexia nervosa [130-132].
Dehydroepiandrosterone treatment seems to be more effec-
tive than ovarian steroids [133]. Bisphosphonate treatment
has also been studied, although more information is needed
concerning their safety and metabolic consequences [134].
6. LIFESTYLE INFLUENCES
Different toxic substances and behaviors may alter the
biologic pubertal process of bone accrual.
6.1. Carbonated Soft Drinks
Soft drink intake negatively impacts bone mass gain of
adolescent girls but not adolescent boys [135, 136]. High
consumption of carbonated soft drinks (CSD) during child-
hood and adolescence may reduce bone mineral accrual and
increase fracture risk. Low nutrient dense sugar-based bever-
ages (carbonated and non-carbonated) have negative conse-
quences on milk and milk-derived consumption, showing a
negative correlation between BMC gain and dense beverage
intake solely in girls [135]. In a cross-sectional study includ-
ing children aged 12-15 years, BMD was measured by DXA
and beverage consumption was assessed at two ages (12 and
15 years) although pubertal stage was not adjusted for. In
girls, significant inverse associations were reported between
total CSD and dominant heel BMD, non-cola CSD consump-
tion and dominant heel BMD, and diet drinks and heel BMD.
However, no consistent relationships were observed between
CSD intake and BMD in boys [136]. Therefore, it seems that
bone accrual mechanisms in adolescent girls have more vul-
nerable conditions. The amount of calcium and physical ac-
tivity should be considered among factors responsible for
gender difference.

8 Current Medicinal Chemistry, 2010 Vol. 17, No. 1 Pérez-López et al.
Relationships between soft drink and milk consumption,
physical activity, bone mass, and upper limb fractures were
investigated in an Australian population-based case-control
study including children aged 9-16 years. The study included
a questionnaire to assess last year physical activity and sed-
entary habits, and the consumption of milk, colas, and total
carbonated drinks. Bone mass was assessed by DXA and
metacarpal morphometry. Cola CSD, but not total carbon-
ated beverage consumption, was associated with increased
wrist and forearm fracture risk in children. However, this
association is not independent of other factors and appears to
be mediated by television watching and BMD but not by
decreased milk intake [137]. Gender differences may be bio-
logical or the result of physical activities and/or calcium in-
take or both [138, 139].
In 3 to 7 years old children, increased sweetened bever-
age intake is associated with reduced milk and calcium in-
take [140]. However, it is not clear if milk displacement has
been involved as the cause of CSD interference with bone
mass accrual in children and adolescents. A recent study
reported the association between long term (4 year) soft
drink consumption and adolescent bone health as assessed by
radius peripheral qCT [141]. After adjustments, long-term
consumption of all soft drinks and un-caffeinated soft drinks
was negatively and significantly associated with BMC, corti-
cal area, and polar strength strain index, all which reflect a
combination of bone modeling and remodeling. In addition,
long-term consumption of caffeinated soft drinks was nega-
tively and significantly associated with polar strength strain
index and periosteal circumference, which reflect bone mod-
eling. Contrarily, milk intake was positively and significantly
associated with polar strength strain index. In this scenario,
soft drinks appear to have bone catabolic effects in both gen-
ders which are not primarily based on milk displacement
[141].
6.2. Smoking
In growing female rats, nicotine treatment for 2 or 3
months did not produce significant effects over bone mineral
content and density, bone histomorphometry or bone
strength [142]. In adult rats, nicotine was detrimental to bone
due to an increase in bone resorbing cytokine and cotinine
levels, hence altering trabecular histomorphometry [143].
Smoking has been linked to low bone density in adolescents
and is associated with other unhealthy behaviors, such as
alcohol use and a sedentary lifestyle. People who begin
smoking at a younger age are more likely to be heavier
smokers later in life. This fact worsens the negative impact
of smoking over PBM, and places older smokers at addi-
tional risk for bone loss and fracture.
6.3. Alcohol Intake and Bone
In animal studies, mild to moderate alcohol consumption
had negative consequences for female reproductive function,
disrupt female puberty, and affecting growth and bone health
[144]. The impact of alcohol intake on PBM is not clear. The
effects of alcohol on bone have been studied more exten-
sively in adults, and the results indicate that high consump-
tion of alcohol has been linked to low bone density. Experts
assume that high alcohol consumption in youth would have a
similar adverse effect on skeletal health.
6.4. High Trained Athletes
Although exercise in adolescents has benefits on bone
mass accrual, other aspects such as menstrual function (an
indirect measure of sex steroid production), nutrition and
body composition (a balance between fat and lean mass)
should be preserved in the normal range which suggest the
complex interactions between factors during bone growth
and accrual and body maturation during adolescence [145].
The fact that athletes with amenorrhea have lower BMD
raises the possibility that PBM may be affected among them.
In the US, some 20% of adolescent girl athletes have
been reported to develop amenorrhea which is associated to
bone accrual problems [146]. The prevalence varies accord-
ing to the type, intensity, and duration of exercise, and also
due to the nutritional status of the athlete [147, 148]. Since
maximal girl bone mass accrual occurs between 11 and 14
years and 90% of PBM is achieved by the end of the second
decade [5, 149], disruption of the normal endocrine function,
nutrition and energy consumption, and body mass would
have negative consequences for bone health including frac-
ture risk. Consequences of amenorrhea during the second
decade of life could be worse or irreversible as compared to
that occurring in later life [150].
Female hypogonadal status in adolescent athletes with
amenorrhea has a deleterious effect on bone metabolism as
compared with eumenorrheic athletes, showing lower bone
formation and bone resorption markers as well as lower
BMD at the whole body, spine and hip. In addition, duration
of amenorrhea was an inverse predictor of bone density
[151]. Athletic girls with normal menstrual function have
slightly higher BMD compared to non-athletic girls [152].
Athletic girls with amenorrhea may also have higher cortisol
levels as occur with adult athletes with exercise-amenorrhea
[153], that would contribute to a negative bone growth or
even resorption due to estrogen deficit [154].
Low IGF-1 levels in athletic girls with amenorrhea indi-
cate suboptimal nutritional status and a lower bone turnover;
hence reaching late adolescence with decreased bone turn-
over when very little potential of growth is left [152]. Causes
of hypogonadism among athletic girls seem to be related to
low fat mass and a negative energy balance due to exagger-
ated expenditure or balanced diet deviations, rather than ex-
ercise stress [159]. Fat mass is important for preservation of
normal function during malnutrition as compared to amenor-
rheic women with a similar weight which suggests that leptin
secretion may contribute in maintaining bone mass [160].
6.5. Hormonal Contraception
Hormonal contraceptives influence bone mass during
adolescence. Long-term estrogen-progestin contraceptive
with low-dose estrogen preparations seems to suppress nor-
mal bone mineral accrual in adolescent girls. Thus, there is a
significant trend showing less mean adjusted BMC of lumbar
spine in the group of adolescent women who had used oral
formulations with estrogen dosages of ≤ 35 µg for more than
2 years compared with non-users and those who received

Bone Mass Gain and Adolescence Current Medicinal Chemistry, 2010 Vol. 17, No. 1 9
treatment for one year [161]. Treatment for two years with
depot medroxyprogesterone acetate produces a 1.5% reduc-
tion in BMD as compared to 4.2% gain during the same pe-
riod in adolescents who used an oral compound consisting of
daily dose of 20 µg of ethynil estradiol and 100 µg levonorg-
estrel and compared to 6.3% BMC increase in an untreated
control group [162].
6.6. Pregnancy During Adolescence
The negative impact of pregnancy during adolescence
over BMC seems to be higher than that produces at an older
age [3]. A reasonable explanation seems reside upon grow-
ing requirements of calcium and vitamin D related to the
pubertal process itself and the fetal development. Addition-
ally breastfeeding reduces BMD especially during the first
few months [163]. Although BMD of an adult woman usu-
ally recuperates after breastfeeding [163, 164], it is not clear
if gestational bone mass loss is completely reversible after
breastfeeding in adolescence [30].
7. ADOLESCENCE BONE MASS GAIN AND FRAC-
TURES
Risk factors for healthy children and adolescent fractures
include bone mineral content, size and accrual which are all
lower. Other factors that affect pediatric individuals are ge-
netic factors, poverty, lack of regular physical activity, obe-
sity and high exposure to trauma [165]. Fractures after the
sixth decade have been related to structural and biomechani-
cal alterations of the acquired bone during the second and
third decades of life [166, 167]. The importance of reaching
an optimal PBM has been calculated by determining the risk
of osteoporotic fracture in climacteric women. Thus, a 10%
increase in PBM could reduce the risk of fracture by 50% in
postmenopausal women [168]. Therefore, bone health and
gaining an optimal PBM are extremely important to prevent
fractures and postmenopausal osteoporosis. Lifestyle factors
have also been associated with volumetric bone density
[169].
8. FINAL REMARKS
Growth during early life is associated with bone size and
strength in older individuals. The strength of adult bone re-
flects factors that regulate bone quality (architecture) and
density (bone mass) acquired during childhood and adoles-
cence. Building PBM during childhood and adolescence is
an investment to prevent women against developing osteopo-
rosis later in life. To reach optimal bone health, as allowed
by genetic background, it is necessary that maturation from
childhood to adulthood occur with different physiological
inputs (Fig. 3). Genetic factors may account for up to 75% of
bone mass, while environmental factors account for the re-
maining 25%. Genetic susceptibility to osteoporosis would
be detectable in childhood and adolescence [170, 171]. Re-
duced bone mass in daughters of women with osteoporosis is
present in daughter bone characteristics before puberty.
Some genes associated with low bone mass in older women
may be also detectable in children [172, 173]. Among ge-
netic influences there are polygenetic factors [174]. Al-
though evidence is incomplete receptor polymorphisms for
vitamin D, estrogen, type I collagen, IGF-1, transforming
growth factor-β, and interleukin-6 should be mentioned [175,
176]. However, much of the genetic variance in BMD is
presently unexplained by these variables, which suggests that
Fig. (3). Endogenous and environmental factors that influence bone mass acquisition during puberty and adolescence.

10 Current Medicinal Chemistry, 2010 Vol. 17, No. 1 Pérez-López et al.
either most of the genes that regulate BMD remain to be
identified.
Interferences with the endogenous endocrine changes,
nutritional alterations, sedentarism or extreme exercise, toxic
substance consumption, hormone contraception and preg-
nancies during adolescence will reduce bone mass accrual. A
healthy lifestyle, regular moderate exercise, a balanced diet
prevent bone mineral metabolic alterations while promoting
healthy bone development. Lifestyle interventions can im-
prove dietary intake and increase bone mass gain in girls
[177]. Good eating and exercise habits may be established
during the adolescence with aims to persist in adulthood
[178, 179]. These interventions are particularly important in
girls, since women are at higher risk for developing osteopo-
rosis than males.
ACKNOWLEDGEMENT
This study was partially supported by the B/017543/08
AECID grant from the Spanish Ministerio de Asuntos Exte-
riores y Cooperación.
ABBREVIATIONS
1,25(OH)2D = 1,25-dihydroxyvitamin D3
25(OH)D = 25-hydroxyvitamin D3
BMC = bone mass content
BMD = bone mineral density
BMI = body mass index
CPP = central precocious puberty
CSD = carbonated soft drinks
DXA = dual X-ray absorciometry
GH = growth hormone
GnRH = gonadotropin-releasing hormone
IGF = insulin growth factor
IGFBP = IGF-binding protein
PBM = peak bone mass
PTH = parathyroid hormone
qCT = quantitative computed tomography
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