
==== Front
J Mother Child
J Mother Child
jmotherandchild
jmotherandchild
Journal of Mother and Child
2719-6488
2719-535X
Sciendo

38639100
jmotherandchild.20242801.d-23-00105
10.34763/jmotherandchild.20242801.d-23-00105
Original Research
Assessment of Biochemical Bone Turnover Markers in Polish Healthy Children and Adolescents
https://orcid.org/0000-0002-1349-0155
Gajewska Joanna joanna.gajewska@imid.med.pl

https://orcid.org/0000-0002-6174-6813
Chełchowska Magdalena
https://orcid.org/0000-0002-0560-9435
Rowicka Grażyna
https://orcid.org/0000-0002-7088-0117
Klemarczyk Witold
https://orcid.org/0000-0002-8061-0049
Głąb-Jabłońska Ewa
https://orcid.org/0000-0001-7320-7561
Ambroszkiewicz Jadwiga
Department of Screening Tests and Metabolic Diagnostics, Institute of Mother and Child, Warsaw 01-211, Kasprzaka 17a, Poland
Department of Nutrition, Institute of Mother and Child, Warsaw 01-211, Kasprzaka 17a Poland
19 4 2024
2 2024
28 1 1422
27 11 2023
9 2 2024
© 2024 Joanna Gajewska et al., published by Sciendo
2024
Joanna Gajewska et al., published by Sciendo
https://creativecommons.org/licenses/by/4.0/ This work is licensed under the Creative Commons Attribution 4.0 International License.
Abstract

Background

Assessing bone turnover in paediatric populations is crucial for understanding the physiological changes occurring during skeletal development and identifying potential abnormalities. The objective of this study was to assess osteocalcin (OC), bone alkaline phosphatase (BALP), and C-terminal telopeptide of type I collagen (CTX-I) levels reflecting bone formation and resorption for age and sex in Polish healthy children and adolescents.

Materials and Methods

A total of 355 healthy normal-weight children and adolescents (46.5% girls) aged 1–18 years old were recruited. Total body less head (TBLH) and spine L1-L4 were used in children to assess bone mineral density (BMD) by dual-energy X-ray absorptiometry (DXA). Bone marker concentrations were determined by immunoenzymatic methods.

Results

Bone marker levels in girls and boys started with higher values in the first year of life and subsequently decreased until reaching a nadir during the prepubertal period. The pubertal peak values of bone markers were reached at 11–13 years old in boys and at 9–11 years old in girls. After puberty, the adolescents showed a gradual decline in bone marker concentrations to the values observed in adults. We found positive correlations between OC level and TBLH-BMD (r = 0.329, p = 0.002), TBLH-BMD Z-score (r = 0.245, p = 0.023), and L1-L4 BMD (r = 0.280, p = 0.009) in the prepubertal group.

Conclusions

We showed serum levels of bone turnover markers—BALP, OC, and CTX-I—in relation to age and sex in healthy Polish children and adolescents. The age intervals of these markers for girls and boys aged 1–18 years old may be clinically useful in the assessment of bone metabolism in individuals with skeletal disorders.

Keywords

osteocalcin
bone alkaline phosphatase
C-terminal telopeptide of type I collagen
children
adolescents
age intervals
==== Body
pmc1. Introduction

Skeletal health is of paramount importance during childhood and adolescence, as it directly affects growth, development, and overall quality of life. Optimal bone development and maintenance require a proper balance between bone formation and resorption [1]. The process of bone remodelling, which involves the continuous removal of old bone tissue and the subsequent formation of new bone, is pivotal for maintaining skeletal integrity, strength, and mineralization. Imbalances in bone metabolism can lead to growth retardation and a range of skeletal disorders, including osteoporosis and fractures [2].

Assessing bone turnover in paediatric populations is crucial for understanding the physiological changes occurring during skeletal development and identifying potential abnormalities. Traditionally, bone mineral density (BMD) measurements using dual-energy X-ray absorptiometry (DXA) have been widely used to evaluate bone health in children and adolescents [3]. However, BMD alone does not provide a comprehensive understanding of bone turnover dynamics.

In recent years, biochemical markers have emerged as valuable tools for assessing bone metabolism and turnover [4]. These markers—measured in blood or urine samples—provide insights into the balance between bone formation and resorption processes. Among the various bone turnover markers, osteocalcin (OC), bone alkaline phosphatase (BALP), and C-terminal telopeptide of type I collagen (CTX-I) have gained significant attention due to their association with specific aspects of bone remodelling [5].

OC, a non-collagenous protein synthesized by osteoblasts, plays a crucial role in bone mineralization and is considered a marker of the bone formation process [6]. Studies have suggested that OC levels are influenced by factors such as age, sex, puberty, hormonal, and nutritional status. Similarly, bone alkaline phosphatase, an enzyme primarily synthesized by osteoblasts, is involved in bone matrix mineralization [7]. BALP levels have been shown to correlate with bone formation rates and exhibit variations during different stages of growth and development. On the other hand, CTX-I is a degradation product of collagen released during the bone resorption process [8]. As bone resorption occurs, CTX-I is released into the bloodstream, making it a reliable marker of the bone resorption activity of osteoclasts. CTX-I levels have been found to increase during pubertal growth spurts and decrease following interventions that suppress bone resorption, such as bisphosphonate therapy [9].

Although numerous studies have explored bone turnover markers in various populations, limited research has focused specifically on healthy children and adolescents [10,11,12,13,14]. Establishing reference ranges for these markers in healthy paediatric populations is crucial for understanding normal bone metabolism during growth and development. Moreover, reference ranges can be useful to improve clinical practices by aiding in the early detection of bone health disorders and diseases in children and adolescents. Therefore, the primary objective of this study was to assess the levels of OC, BALP, and CTX-I reflecting bone formation and resorption for each age interval, separately for boys and girls, in Polish healthy children and adolescents.

2. Materials and methods

2.1. Patients

A total of 355 healthy children and adolescents (165 girls, 190 boys) aged 1–18 years old with a BMI Z-score <−1 + 1> were recruited to the study group. Densitometric examination was performed in the group of 88 children aged 5 to 10 years old. On the basis of data from the history and medical examination, these children were: (a) without either acute or chronic disorders, among them obesity; (b) without bone fractures in the period up to one year before the study; (c) not taking any medications that could affect their growth, pubertal development, nutritional or dietary status; (d) whose parents signed the informed consent form. All of the participants were Caucasian. Pubertal stage was determined according to the Tanner scale by the physician during the medical appointment.

Physical examinations, including body height and weight measurements, were performed in prepubertal children. The Body Mass Index (BMI) was calculated as body weight divided by height squared (kg/m2). The BMI of each individual was converted to a standard BMI Z-score for the child’s age and sex using Polish reference tables [15]. Densitometry examination was performed in the studied children in two projections: total body and spine L1-L4. Total body less head (TBLH) were used for performing bone mineral content (BMC) and BMD by DXA, (Lunar Prodigy, General Electric Healthcare, Madison, WI, paediatric software 9.30.044). All subjects were measured on the same machine. The measurements were performed using standard positioning techniques.

Written informed consent was obtained from the parents of all the examined children. The study was conducted between September 2012 and December 2018 in accordance with the Helsinki Declaration for Human Research, and the study protocol was approved (protocol code no. 03/09) by the Ethics Committee of the Institute of Mother and Child in Warsaw, Poland.

2.2. Biochemical methods

Venous blood samples were collected between 8:00 and 10:00 a.m. after an overnight fast and centrifuged at 1000 × g for 10 min at 4°C. Serum specimens were stored at −70°C prior to assay (no longer than for six months). Biochemical parameters were determined by immunoenzymatic methods. BALP activity was estimated using the BAP EIA kit from Quidel (Athens, USA) with a within-assay variability of less than 5.8% and a between-assay variability of less than 7.6%. OC and CTX-I concentrations were measured with the N-MID OC ELISA kit and serum CrossLaps ELISA kit (IDS, Bolton, UK), respectively. The intra- and inter-assay coefficients of variation were less than 2.2% and 5.1% for OC, and 3.0% and 10.9% for CTX-I. The detection limit was 0.7 U/L for BALP, 0.5 ng/mL for OC, and 0.02 ng/mL for CTX-I.

2.3. Statistical analyses

Statistical analysis was performed using Statistica 6.0 (StatSoft Inc.) software. The results are presented as medians and interquartile ranges (25th–75th percentiles) for non-normally distributed variables. The Kolmogorov–Smirnov test and graphical inspections of data were used to evaluate distribution for normality. Differences in the anthropometric characteristics and biochemical parameters of healthy girls and boys were assessed using the non-parametric Mann–Whitney U test. Spearman correlations between the anthropometric and biochemical parameters were calculated. Differences were regarded as statistically significant at p < 0.05.

3. Results

The general distribution of bone markers in 355 subjects (165 girls and 190 boys) are shown in Table 1. The group of girls and group of boys were of similar age and did not differ in terms of OC and CTX-I levels. However, BALP activity was higher by about 10% in boys than girls (p = 0.048).

Table 1. Bone markers in the studied population of children and adolescents.

Variable	Girls	Boys	Total	P *	
Participants, n (%)	165 (46.5)	190 (53.5)	355 (100)	0.189	
Age (years)	8.5 (5.5–12.0)	8.4 (5.5–13.0)	8.4 (5.5–12.3)	0.981	
BALP (U/L)	106.6 (84.6–128.4)	115.4 (92.0–136.5)	112.1 (87.9–132.5)	0.048	
OC (ng/mL)	84.0 (52.6–114.4)	80.9 (55.5–116.9)	83.2 (52.6–114.4)	0.518	
CTX-I (ng/mL)	1.72 (1.12.–2.22)	1.67 (1.22.–2.09)	1.70 (1.19–2.12)	0.805	
Medians and interquartile ranges (25th–75th percentiles),

* p-values between girl and boy groups. BALP–bone alkaline phosphatase, OC–osteocalcin, and CTX-I–C-terminal telopeptide of type I collagen.

BALP, CTX-I, and OC reference curves for children and adolescents aged 1–18 years old are shown in the Fig. 1 (A,B,C). In general, all the studied bone markers in girls and boys started with higher values in the first year of life and subsequently decreased until reaching a nadir during the prepubertal period, earlier in girls than boys. The pubertal peak values of bone markers were reached at 9–11 years old in girls and at 11–13 years old in boys. In girls, after the age of 11 years old, a gradual decrease in marker values was observed, until the lowest values were reached at the age of 18 years old. In boys, after the age of 13 years old, continuous decreases in the values of these parameters until age 18 were also observed.

Figure 1. BALP (A), OC (B), and CTX-I (C) trend curves (polynomial trendlines: girls, boys -----) with individual data points for girls and boys aged 1–18 years old.

Characteristics of the study population with age and sex stratification are presented in Table 2. Reference ranges of bone markers in children and adolescents were defined as medians, between 25th and 75th percentiles of serum concentrations. From our results, the observed bone markers varied significantly in adolescents, with sex differences. Median values of BALP at ages 14–15.9 years old and 16–18 years old were higher in boys than girls (p < 0.001; p = 0.022, respectively). Median values of OC at ages 12–13.9 years old, 14–15.9 years old, and 16–18 years old were higher in boys than in girls (p = 0.019; p = 0.010; p = 0.018, respectively). The median values of CTX-I at ages 14–15.9 were higher in boys than in girls (p = 0.004).

Table 2. Bone markers in children and adolescents aged 1–18 years old stratified by sex and age.

Parameter/years	Girls (n=165)	Boys (n=190)	p	
		
n	Median	Range	n	Median	Range	
BALP (U/L)	
1.0–2.9	10	139.5	122.8–143.9	14	126.7	116.3–170.2	0.861	
3.0–4.9	20	112.8	86.7–138.3	19	125.5	100.7–142.4	0.227	
5.0–7.9	43	117.9	102.5–132.1	53	118.7	103.6–138.8	0.945	
8.0–9.9	32	118.8	98.4–128.0	33	109.7	94.8–125.7	0.568	
10.0–11.9	16	109.4	97.2–129.9	14	118.6	88.3–152.7	0.618	
12.0–13.9	10	120.7	101.0–138.8	18	129.1	101.8–190.1	0.598	
14.0–15.9	12	67.2	49.5–84.4	18	120.9	99.3–134.4	<0.001	
16.0–18.0	22	34.2	20.7–39.7	21	44.1	30.8–71.6	0.022	
OC (ng/mL)	
1.0–2.9	10	72.0	62.6–118.0	14	57.8	41.3–81.8	0.160	
3.0–4.9	20	89.7	53.7–113.5	19	62.5	52.4–77.1	0.092	
5.0–7.9	43	77.3	58.0–105.7	53	77.4	62.8–116.6	0.749	
8.0–9.9	32	95.6	69.9–126.1	33	92.2	74.0–110.0	0.436	
10.0–11.9	16	114.8	95.7–127.1	14	101.0	66.2–132.3	0.406	
12.0–13.9	10	88.8	77.9–111.2	18	122.5	96.0–135.8	0.019	
14.0–15.9	12	73.5	59.0–110.0	18	122.0	110.5–134.0	0.010	
16.0–18.0	22	30.4	23.2–41.3	21	38.8	30.3–49.5	0.018	
CTX-I (ng/mL)	
1.0–2.9	10	1.58	1.05–2.22	14	1.40	0.90–1.59	0.266	
3.0–4.9	20	1.94	1.23–2.37	19	1.66	1.21–2.02	0.369	
5.0–7.9	43	1.80	1.25–2.17	53	1.85	1.33–2.14	0.651	
8.0–9.9	32	1.80	1.43–2.25	33	1.67	1.49–1.98	0.240	
10.0–11.9	16	2.23	1.72–2.43	14	1.91	1.79–2.19	0.383	
12.0–13.9	10	1.74	1.47–2.64	18	2.13	1.41–2.66	0.905	
14.0–15.9	12	1.37	0.95–1.79	18	2.19	1.96–2.50	0.004	
16.0–18.0	22	0.72	0.41–0.97	21	0.86	0.64–0.95	0.375	
Medians and interquartile ranges (25th–75th percentiles),

* p-values between girl and boy age groups. BALP–bone alkaline phosphatase, OC–osteocalcin, and CTX-I–C-terminal telopeptide of type I collagen.

In the group of girls, we found negative correlations between age and BALP activity (r = −0.475, p < 0.001), OC (r = −0.202, p = 0.01), and CTX-I levels (r = −0.248, p = 0.001). In the group of boys, a negative correlation was found between age and BALP activity (r = −0.348, p < 0.001), but a positive correlation between age and OC level (r = 0.155, p = 0.033). In the whole study group of children and adolescents, we observed negative correlations between age and BALP activity (r = −0.402, p < 0.001), and CTX-I level (r = −0.118, p = 0.026).

Positive correlations were found in the whole study group as well as in the girl and boy groups separately between CTX-I and OC levels (r = 0.664, p < 0.001; r = 0.687, p < 0.001; r = 0.645, p < 0.001, respectively), and between CTX-I level and BALP activity (r = 0.207, p < 0.001; r = 0.256, p = 0.001; r = 0.173, p = 0.017, respectively). Positive correlations were also found between OC level and BALP activity in the whole study group and the girls subgroup (r = 0.170, p = 0.001; r = 0.219, p = 0.005, respectively).

Characteristics of the prepubertal children (45 girls, 43 boys) group are presented in Table 3. Age, anthropometric parameters, and bone markers did not show any differences between the two sex groups. However, in the whole group of prepubertal children, we found positive correlations between BMI Z-score and TBLH-BMD (r = 0.282, p = 0.008), TBLH-BMD Z-score (r = 0.366, p < 0.001), TBLH-BMC (r = 0.266, p = 0.012), L1-L4 BMD (r = 0.248, p = 0.021), and L1-L4 BMD Z-score (r = 0.265, p = 0.014) values (Table 4). In addition, we observed positive correlations between OC level and TBLH-BMD (r = 0.329, p = 0.002), TBLH-BMD Z-score (r = 0.245, p = 0.023), TBLH-BMC (r = 0.329, p = 0.002), and L1-L4 BMD (r = 0.280, p = 0.009) values in this group.

Table 3. Serum bone marker concentrations and anthropometric parameters in prepubertal healthy children aged 5–10 years old.

Variable	Girls n=45	Boys n=43	Total n=88	p *	
Age (years)	8.2 (6.8–9.1)	7.5 (5.6–9.0)	8.0 (6.0–9.1)	0.081	
Weight (kg)	27.0 (21.3–29.4)	23.9 (20.3–29.9)	25.1 (20.4–29.7)	0.361	
Height (cm)	128.5 (119.0–133.8)	125.6 (117.8–132.6)	126.4 (118.6–133.5)	0.278	
BMI (kg/m2)	15.5 (14.5–17.0)	15.6 (14.6–16.6)	15.5 (14.6–16.8)	0.910	
BMI Z–score	−0.36 (−0.70–0.24)	−0.37 (−0.75–0.04)	−0.37 (−0.74–0.18)	0.841	
TBLH-BMD (g/cm2)	0.686 (0.642–0.737)	0.644 (0.622–0.720)	0.672 (0.627–0.730)	0.246	
TBLH-BMD Z-score	−0.40 (−0.70–0.20)	−0.10 (−0.60–0.40)	−0.30 (−0.63–0.20)	0.101	
TBLH-BMC (g)	600.6 (452.8–764.5)	526.2 (419.4–751.3)	582.8 (436.2–760.3)	0.258	
L1-L4 BMD (g/cm2)	0.666 (0.631–0.731)	0.624 (0.569–0.707)	0.659 (0.595–0724)	0.057	
L1-L4 BMD Z-score	−0.40 (−0.80–0.30)	−0.40 (−1.20–0.20)	−0.4 (−1.00–0.20)	0.296	
BALP (U/L)	119.8 (100.3–131.3)	116.7 (102.0–130.5)	117.9 (100.3–131.4)	0.799	
OC (ng/mL)	81.2 (63.8–102.7)	75.2 (63.5–94.4)	77.5 (63.7–100.5)	0.450	
CTX-I (ng/mL)	1.78 (1.37–2.16)	1.66 (1.44–2.00)	1.73 (1.39–2.02)	0.419	
Medians and interquartile ranges (25th–75th percentiles),

* p-values between girl and boy groups.

BMI–body mass index, TBLH–total body less head, BMD–bone mineral density, BMC–bone mineral content, BALP–bone alkaline phosphatase, OC–osteocalcin, and CTX-I–C-terminal telopeptide of type I collagen.

Table 4. Relations between BMI Z-score, bone markers and densitometric parameters in prepubertal healthy children aged 5–10 years old.

Parameters	TBLH-BMD r (p)	TBLH-BMD Z-score r (p)	TBLH-BMC r (p)	L1-L4 BMD r (p)	L1-L4 BMD Z-score r (p)	
BMI Z-score	0.282 (0.008)	0.366 (<0.001)	0.266 (0.012)	0.248 (0.021)	0.265 (0.014)	
BALP	−0.049 (0.651)	−0.092 (0.398)	−0.032 (0.769)	−0.019 (0.861)	0.006 (0.95)	
OC	0.329 (0.002)	0.245 (0.023)	0.329 (0.002)	0.280 (0.009)	0.192 (0.077)	
CTX-I	0.078 (0.476)	0.048 (0.661)	0.101 (0.356)	0.097 (0.374)	0.051 (0.051)	
BMI–body mass index, TBLH–total body less head, BMD–bone mineral density, BMC–bone mineral content, BALP–bone alkaline phosphatase, OC–osteocalcin, and CTX-I–C-terminal telopeptide of type I collagen.

4. Discussion

The application and interpretation of the results regarding bone turnover markers is more complicated in children than in adults. Children and adolescents have elevated circulating concentrations of biochemical bone markers, reflecting high growth velocity and rapid bone turnover [8].

In adults, osteoblasts and osteoclasts participate only in bone remodelling, while in children they play a role in both the bone remodelling process and bone growth, including longitudinal growth and increases in bone circumference and thickness [16]. The use of bone markers in children and adolescents is increasing, for example—in Paget’s disease, treatment-induced bone diseases such as glucocorticoid-induced osteoporosis and other chronic bone disorders including those accompanying autoimmune diseases in children, such as inflammatory bowel disease, food allergies, or chronic kidney diseases [17,18,19,20,21]. Therefore, there is a need to establish reference values of bone turnover markers for developmental age for boys and girls separately.

Some authors described the use of bone marker values for children and adolescents from the age of a few months to 17–18 years old or only for adolescence [22,23,24,25,26]. Age and sex are important variables that affect bone formation and resorption marker concentrations in childhood and adolescence [27–28]. Usually, two peaks of bone growth were observed with the highest bone marker levels during the developmental period. The first peak was in infants and children during the first four years of life [10,29] and the second was in children during the pubertal period [14,23]. Bone marker concentrations were slightly lower and rather stable between these two peaks. After the pubertal period, girls and boys showed a gradual decline in bone marker concentrations to the values observed in young adults [26].

In our study, BALP values were similar for both sexes in the group of small children, but the peak in puberty was slightly higher in boys and occurred later than in girls. During puberty, BALP showed a peak at 9–11 years of age in girls, while this was 2–3 years later in boys. These results are in agreement with data obtained in German and multi-ethnic populations by other authors [30,31]. Our study also showed elevated OC—second marker of bone formation—during infancy but more in girls than boys. However, the peak of OC during puberty was higher and occurred 2–3 years later in boys than in girls.

The literature data on serum OC concentrations are ambiguous. Rauchenzauner et al. [32] found rather stable results in the Austrian population. Other authors observed, similar to us, increased OC concentrations during infancy which decreased before reaching a peak during early puberty in Central European and Danish populations [14,33]. In terms of the bone resorption marker CTX-I, we observed a peak during the pubertal period only and it was slightly higher and occurred about two years later in boys than in girls. Other authors also did not show a peak of CTX-I in early infancy, but found that it was only slightly increasing to reach a peak of this marker in early puberty in Canadian and European populations [29,34].

In the presented study, we analysed bone markers and anthropometric parameters in prepubertal healthy children. We used TBLH as a scanning site when assessing BMD according to The International Society for Clinical Densitometry (ISCD) recommendations [35]. We observed similar values of height, weight, BMI and BMI Z-score, TBLH-BMD, TBLH-BMC, L1-L4 BMD, and L1-L4 BMD Z-score as well as bone markers in girls and boys. We found positive relations between BMI Z-score and TBLH-BMD, TBLH-BMD Z-score, TBLH-BMC, L1-L4 BMD, and L1-L4 BMD Z-score in prepubertal children.

The influence of obesity on bone turnover and bone quality in children and adolescents remains controversial [36]. Some researchers observed a significantly greater whole-body bone area and BMC for age and for height in obese children and adolescents [37], while others described a lower bone mass and bone area in obese children compared with normal weight subjects [38]. Our earlier and present studies in prepubertal children confirm the relations between greater values of BMI Z-score and greater bone mass (BMC, BMD, and BMD Z-score) [39].

We also found positive relations between OC and TBLH-BMD, TBLH Z-score, TBLH-BMC, and spine L1-L4 BMD in prepubertal children. However, we did not observe any relations between BALP, CTX-I levels, and DXA measurement values in this study group. Paldanius et al. [24] found no associations between BMD with OC values in healthy Finnish children and adolescents in an older age group (7 to 19 years old) than the group examined in this study. Zurcher et al. [40] and Monjardino et al. [41] also found no associations of CTX-I with DXA measurements in Swiss and Portuguese populations of children and adolescents.

According to Jurimae [42], the interpretation of bone metabolism in children and adolescents should not be based on one bone turnover marker but several markers reflecting different steps in the bone formation and resorption processes. In our study, we showed the concentrations of three markers concerning bone turnover stratified by age and sex in healthy children and adolescents. The use of bone markers requires compliance with a number of pre-analytical and analytical procedures.

Various reagent kits of bone turnover markers are available and may not give identical results for these markers. OC is an unstable protein, hence there is a problem with selecting appropriate kits for determining this marker in serum. According to Rosenquist et al. [43], immunoassays targeting the NH2-terminal middle fragment (N-middle OC) are preferred for routine testing, because the C-terminal OC sequence is the least stable. BALP was defined as the marker of choice in haemodialyzed patients [44]. This marker is almost not impacted by renal function, fasting, or circadian rhythm [45]. However, there are reported cases of transient hyperphosphatasaemia in infants under five years old [46]. In these cases, BALP activity was transiently increased but returned to normal values after a few weeks without any symptoms. CTX-I, analysed in our study using the CrossLaps kit, was influenced by fasting, circadian rhythm, and renal failure [45]. To reduce preanalytical biochemical bone marker variance, blood samples should be collected after an overnight fast and within the same four-hour time frame for all participants. Thus, the effect of circadian rhythm and food intake on tested marker levels will be limited [45].

The ISCD indicates that early identification and treatment of children at risk of fractures is necessary due to the possibility of restoring bone strength to growing children [47]. Therefore, the use of additional diagnostic and treatment monitoring methods, such as biochemical markers of bone turnover, would be useful to better identify patients affected by paediatric osteoporosis or those at highest risk of other skeletal disorders. In particular, reference values of bone turnover markers could be useful in monitoring bisphosphonate treatment in children with Juvenile Paget’s disease, osteogenesis imperfecta, and osteoporosis [48,49,50]. These markers may be helpful in estimating an effective inhibition of osteoclast function with reduction of bone resorption during therapy.

In conclusion, we showed serum levels of bone turnover markers BALP, OC, and CTX-I in relation to age and sex in healthy Polish children and adolescents. In this study, associations between OC and DXA measurements in prepubertal children were also found. The age intervals of the studied markers for healthy girls and boys aged 1–18 years old may be clinically useful in the assessment of bone metabolism in children and adolescents with primary skeletal disorders and those accompanying other diseases.

Author Contributions

Joanna Gajewska: conceptualization, methodology, formal analysis, writing—original draft preparation; Magdalena Chełchowska: investigation, writing—review and editing; J.A.; Grażyna Rowicka: resources, writing—review and editing; Witold Klemarczyk: resources, writing—review and editing; Ewa Głąb-Jabłońska: software, formal analysis; and Jadwiga Ambroszkiewicz: investigation, methodology, writing—review and editing.

Funding

This research received no external funding.

Conflicts of Interest

The authors declare no conflict of interest.
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References

1. Nandiraju D Ahmed I Human skeletal physiology and factors affecting its modelling and remodelling Fertil Steril 2019 112 5 775 781 10.1016/j.fertnstert.2019.10.005 31731931
2. Gómez-Alonso C Paediatric metabolic bone disease: A lifetime ahead Adv Ther 2020 37 Suppl 2 38 46 10.1007/s12325-019-01174-3 32236872
3. Guss CE McAllister A Gordon CM DXA in children and adolescents J Clin Densitom 2021 24 1 28 35 10.1016/j.jocd.2020.01.006 32111573
4. Ladang A Rauch F Delvin E Cavalier E Bone turnover markers in children: from laboratory challenges to clinical interpretation Calcif Tissue Int 2023 112 2 218 232 10.1007/s00223-022-00964-2 35243530
5. Hart NH Newton RU Tan J Rantalainen T Chivers P Siafarikas A Biological basis of bone strength: anatomy, physiology and measurement J Musculoskelet Neuronal Interact 2020 20 3 347 371 32877972
6. Neve A Corrado A Cantatore FP Osteocalcin: skeletal and extra-skeletal effects J Cell Physiol 2013 228 1149 1153 10.1002/jcp.24278 23139068
7. Golub EE Boesze-Battaglia K The role of alkaline phosphatase in mineralization Curr Opin Orthop 2007 18 444 448 10.1097/BCO.0b013e3282630851
8. Szulc P Bone turnover: biology and assessment tools Best Pract Res Clin Endocrinol Metab 2018 32 5 725 738 10.1016/j.beem.2018.05.003 30449551
9. Hwang JS Lee S Gong HS The impact of acute fracture on interpretation of bone turnover marker measurements for patients starting anti-resorptive therapies Bone 2022 154 116199 10.1016/j.bone.2021.116199 34534710
10. van Coeverden SC Netelenbos JC de Ridder CM Roos JC Popp-Snijders C Delemarre-van de Waal HA Bone metabolism markers and bone mass in healthy pubertal boys and girls Clin Endocrinol (Oxf) 2002 57 1 107 16 10.1046/j.1365-2265.2002.01573.x 12100078
11. Ambroszkiewicz J Gajewska J Laskowska-Klita T Serum osteocalcin and bone alkaline phosphatase in healthy children in relation to age and gender Med Wieku Rozwoj 2002 6 3 257 65 12637780
12. Gajewska J Ambroszkiewicz J Laskowska-Klita T Osteoprotegerin and C-telopeptide of type I collagen in Polish healthy children and adolescents Adv Med Sci 2006 51 269 72 17357324
13. Geserick M Vogel M Eckelt F Schlingmann M Hiemisch A Baber R Children and adolescents with obesity have reduced serum bone turnover markers and 25-hydroxyvitamin D but increased parathyroid hormone concentrations - results derived from new pediatric reference ranges Bone 2020 132 115124 10.1016/j.bone.2019.115124 31830529
14. Diemar SS Lylloff L Rønne MS Møllehave LT Heidemann M Thuesen BH Reference intervals in Danish children and adolescents for bone turnover markers carboxy-terminal cross-linked telopeptide of type I collagen (β-CTX), pro-collagen type I N-terminal propeptide (PINP), osteocalcin (OC) and bone-specific alkaline phosphatase (bone ALP) Bone 2021 146 115879 10.1016/j.bone.2021.115879 33561588
15. Kułaga Z Różdżyńska-Świątkowska A Grajda A Gurzkowska B Wojtyło M Góźdź M Percentile charts for growth and nutritional status assessment in Polish children and adolescents from birth to 18 year of age Stand. Med 2015 12 119 135
16. Maggioli C Stagi S Bone modeling, remodeling, and skeletal health in children and adolescents: mineral accrual, assessment and treatment Ann Pediatr Endocrinol Metab 2017 22 1 1 5 10.6065/apem.2017.22.1.1 28443253
17. Cook SJ Wall C Paget’s disease of bone: A clinical update Aust J Gen Pract 2021 50 1–2 23 29 10.31128/AJGP-10-20-5690 33543158
18. Okten A Cakir M Makuloglu M Bone mineral status, bone turnover markers and vitamin D status in children with congenital adrenal hyperplasia Minerva Endocrinol 2012 37 3 275 82 22766894
19. Karava V Dotis J Christoforidis A Kondou A Printza N Muscle-bone axis in children with chronic kidney disease: current knowledge and future perspectives Pediatr Nephrol 2021 36 12 3813 3827 10.1007/s00467-021-04936-w 33534001
20. Guandalini S Assiri A Celiac disease: a review JAMA Pediatr 2014 168 3 272 278 10.1001/jamapediatrics.2013.3858 24395055
21. Wei H Zhao Y Xiang L Bone health in inflammatory bowel disease Expert Rev Gastroenterol Hepatol 2023 17 9 921 935 10.1080/17474124.2023.2248874 37589220
22. Gajewska J Ambroszkiewicz J Laskowska-Klita T Some bone turnover markers in serum of healthy children and adolescents in relation to age and gender Wiad Lek 2005 58 9–10 476 80 16529055
23. Wang X Liu L Li P Ma J Jiang R Wang R Reference and influential factors of serum bone markers in Chinese adolescents Sci Rep 2017 7 1 17340 10.1038/s41598-017-17670-x 29229962
24. Paldánius PM Ivaska KK Mäkitie O Viljakainen H Serum and urinary osteocalcin in healthy 7- to 19-year-old Finnish children and adolescents Front Pediatr 2021 9 610227 10.3389/fped.2021.610227 34504811
25. Rand MS Diemar SS Møllehave LT Heidemann M Thuesen BH Petersen JH Z-scores of bone turnover markers calculated from new established sex- and age-specific reference curves are associated to future change in BMD in children and adolescents Bone 2023 167 116641 10.1016/j.bone.2022.116641 36481707
26. Chubb SAP Vasikaran SD Gillett MJ Reference intervals for plasma β-CTX and P1NP in children: A systematic review and pooled estimates Clin Biochem 2023 118 110582 10.1016/j.clinbiochem.2023.05.001 37187224
27. Federico G Baroncelli GI Vanacore T Fiore L Saggese G Pubertal changes in biochemical markers of growth Horm Res 2003 60 Suppl1 46 51 10.1159/000071225 12955017
28. Diemar SS Møllehave LT Quardon N Lylloff L Thuesen BH Linneberg A Effects of age and sex on osteocalcin and bone-specific alkaline phosphatase-reference intervals and confounders for two bone formation markers Arch Osteoporos 2020 15 1 26 10.1007/s11657-020-00715-6 32095898
29. Huang Y Eapen E Steele S Grey V Establishment of reference intervals for bone markers in children and adolescents Clin Biochem 2011 44 10–11 771 778 10.1016/j.clinbiochem.2011.04.008 21531216
30. Colantonio DA Kyriakopoulou L Chan MK Daly CH Brinc D Venner AA Closing the gaps in pediatric laboratory reference intervals: a CALIPER database of 40 biochemical markers in a healthy and multiethnic population of children Clin Chem 2012 58 5 854 868 10.1373/clinchem.2011.177741 22371482
31. Fischer DC Mischek A Wolf S Rahn A Salweski B Kundt G Paediatric reference values for the C-terminal fragment of fibroblast-growth factor-23, sclerostin, bone-specific alkaline phosphatase and isoform 5b of tartrate-resistant acid phosphatase Ann Clin Biochem 2012 49 Pt 6 546 553 10.1258/acb.2012.011274 22984195
32. Rauchenzauner M Schmid A Heinz-Erian P Kapelari K Falkensammer G Griesmacher A Sex- and age-specific reference curves for serum markers of bone turnover in healthy children from 2 months to 18 years J Clin Endocrinol Metab 2007 92 2 443 449 10.1210/jc.2006-1706 17105843
33. Bayer M Reference values of osteocalcin and procollagen type I N-propeptide plasma levels in a healthy Central European population aged 0–18 years Osteoporos Int 2014 25 2 729 736 10.1007/s00198-013-2485-4 23974858
34. Herrmann D Intemann T Lauria F Mårild S Molnár D Moreno LA Reference values of bone stiffness index and C-terminal telopeptide in healthy European children Int J Obes (Lond) 2014 38 Suppl 2 S76 85 10.1038/ijo.2014.138 25376222
35. Crabtree NJ Arabi A Bachrach LK Fewtrell M El-Hajj Fuleihan G Dual-energy X-ray absorptiometry interpretation and reporting in children and adolescents: the revised 2013 ISCD Pediatric Official Positions J Clin Densitom 2014 17 2 225 242 10.1016/j.jocd.2014.01.003 24690232
36. Dimitri P Wales JK Bishop N Fat and bone in children: differential effects of obesity on bone size and mass according to fracture history J Bone Miner Res 2010 25 3 527 536 10.1359/jbmr.090823 19778184
37. Vandewalle S Taes Y Van Helvoirt M Debode P Herregods N Ernst C Bone size and bone strength are increased in obese male adolescents J Clin Endocrinol Metab 2013 98 7 3019 28 10.1210/jc.2012-3914 23666962
38. Nagasaki K Kikuchi T Hiura M Uchiyama M Obese Japanese children have low bone mineral density after puberty J Bone Miner Metab 2004 22 4 376 81 10.1007/s00774-004-0498-y 15221498
39. Gajewska J Ambroszkiewicz J Klemarczyk W Chełchowska M Weker H Szamotulska K The effect of weight loss on body composition, serum bone markers, and adipokines in prepubertal obese children after 1-year intervention Endocr Res 2018 43 2 80 89 10.1080/07435800.2017.1403444 29192796
40. Zürcher SJ Borter N Kränzlin M Neyer P Meyer U Rizzoli R Relationship between bone mineral content and bone turnover markers, sex hormones and calciotropic hormones in pre- and early pubertal children Osteoporos Int 2020 31 2 335 349 10.1007/s00198-019-05180-7 31784786
41. Monjardino T Silva P Amaro J Carvalho O Guimarães JT Santos AC Bone formation and resorption markers at 7 years of age: Relations with growth and bone mineralization PLoS One 2019 14 8 e0219423 10.1371/journal.pone.0219423 31437153
42. Jürimäe J Interpretation and application of bone turnover markers in children and adolescents Curr Opin Pediatr 2010 22 4 494 500 10.1097/MOP.0b013e32833b0b9e 20508524
43. Rosenquist C Qvist P Bjarnason N Christiansen C Measurement of a more stable region of osteocalcin in serum by ELISA with two monoclonal antibodies Clin Chem 1995 41 10 1439 45 7586514
44. Cavalier E Delanaye P Collette J Krzesinski JM Chapelle JP Evaluation of different bone markers in hemodialyzed patients Clin Chim Acta 2006 371 1–2 107 11 10.1016/j.cca.2006.02.029 16600202
45. Diemar SS Dahl SS West AS Simonsen SA Iversen HK Jørgensen NR A systematic review of the circadian rhythm of bone markers in blood Calcif Tissue Int 2023 112 2 126 147 10.1007/s00223-022-00965-1 35305134
46. Oladipo OO DeCrescenzo AJ Marquez CP Okorodudu AO Increased alkaline phosphatase in a child Clin Chem 2017 63 6 1174 1175 10.1373/clinchem.2016.268904 28550128
47. Bianchi ML Leonard MB Bechtold S Högler W Mughal MZ Schönau E Bone health in children and adolescents with chronic diseases that may affect the skeleton: the 2013 ISCD Pediatric Official Positions J Clin Densitom 2014 17 2 281 94 10.1016/j.jocd.2014.01.005 24656723
48. Höppner J Steff K Misof BM Schündeln MM Hövel M Lücke T Clinical course in two children with Juvenile Paget’s disease during long-term treatment with intravenous bisphosphonates Bone Rep 2021 14 100762 10.1016/j.bonr.2021.100762 33850973
49. Arundel P Borg SA Early life management of osteogenesis mperfecta Curr Osteoporos Rep 2023 21 6 779 786 10.1007/s11914-023-00823-5 37752354
50. Weber DR Bone health in childhood chronic disease Endocrinol Metab Clin North Am 2020 49 4 637 650 10.1016/j.ecl.2020.07.002 33153671
