The gold standard for assessing pubertal activation is the gonadotropinreleasing hormone (GnRH) stimulation test (GnRHST), which is invasive, time-consuming, and inconvenient. This study evaluated whether a single random measurement of urinary luteinizing hormone (LH) concentration could substitute for the GnRHST in diagnosing and monitoring central precocious puberty (CPP) in girls.
Fifty-five girls with breast buds before 8 years of age were assessed by both the GnRHST and urinary gonadotropin assays. Based on the GnRHST results, 29 girls were assigned to the CPP group (peak LH≥5 IU/L), and 26 were placed in the premature thelarche (PT) group (peak LH<5 IU/L). Auxological data and urine and serum samples were collected at baseline and after treatment with a GnRH agonist for 12 and 24 weeks.
Although the auxological data did not differ between the 2 groups, the serum levels of insulin-like growth factor-1, basal LH, follicle-stimulating hormone (FSH), estradiol, and peak LH; urinary LH; and peak serum LH/FSH and urinary LH/FSH ratios were higher in the CPP group than in the PT group. Pearson correlation analysis showed a positive correlation between the urinary and serum LH concentrations (
A single, random measurement of urinary gonadotropin concentration could be a reliable tool for initial screening and therapeutic monitoring of CPP in girls.
· There are positive correlations between serum gonadotropins and urine gonadotropins.
· Urinary gonadotropin levels reflected the H-P-G axis status in girls with CPP.
· Single, random urinary measurements of gonadotropin concentrations are noninvasive, convenient method for CPP patients.
The initiation of puberty requires an increase in the pulsatile release of gonadotropin-releasing hormone (GnRH) from the hypothalamus, which activates pulse generation of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in the anterior pituitary [
Precocious puberty (PP) is historically defined as the onset of developmental signs of sexual maturation before age 8 years in girls and 9 years in boys [
The GnRH stimulation test (GnRHST) is considered the gold standard for identifying pubertal activation of the HPG axis and diagnosing CPP. GnRH agonists (GnRHa), which suppress the HPG axis, have been shown to be effective and safe treatments for CPP patients [
Recent technological improvements in hormone assays have increased interest in using urine or saliva samples to assess human pituitary function. The collection and analysis of urine samples is more convenient and better tolerated than blood sampling in populations such as neonates and children with needle phobia and for long-term studies. Urinary gonadotropin concentrations have been shown to distinguish among pubertal stages [
To determine whether measuring the gonadotropin concentration in a single, random urinary sample can be used as an initial screen for diagnosing CPP and monitoring the treatment response in girls with CPP, we compared the concentrations of urinary gonadotropins and with the basal and peak serum gonadotropins in girls with CPP and PT.
Girls showing signs of early puberty who had already undergone both the GnRHST and urinary gonadotropin assays were recruited from the growth clinic in the Department of Pediatrics, Chungnam National University Hospital, between August 2017 and July 2018. CPP was defined as breast budding as a first sign of puberty before 8 years of age accompanied by an at least 1-year greater bone age than chronological age and a peak LH level≥5.0 IU/L during the rapid-acting GnRHST (0.1 mg Relefact LHRH; Sanofi-Aventis, Frankfurt am Main, Germany). PT was defined as a peak LH<5.0 IU/L in the GnRHST. Subjects were excluded if they had an underlying disorder or history of medication associated with altered pubertal timing or gonadal development, such as steroids, growth hormone, or GnRHa, or if they were found to have an abnormality on brain MRI or pelvis sonography.
The age, height, weight, body mass index (BMI), parental height, pubertal status, bone age, thyroid function, concentrations of insulin-like growth factor-1 (IGF-1) and estradiol, and medical history were collected from the patients’ medical records. The standard deviations (SDs) for height, weight, and BMI were calculated using the 2017 growth reference for Korean children and adolescents [
The concentrations of serum and urinary LH and FSH were measured using a 2-site sandwich immunoassay involving a direct chemiluminometric technique on an ADVIA Centaur instrument (Siemens Healthcare Diagnostics Inc., New York, NY, USA). The LH assay had a range of 0.07–200 mIU/mL, with an interassay correlation coefficient of 0.99 and a total coefficient of variation (CV) of 2.7%–3.8%. The FSH assay had a range of 0.3–200 mIU/mL, an interassay correlation coefficient of 0.99, and a total CV of 2.2%–3.9%.
Continuous variables are presented as mean±SD. The presence of a normal distribution was determined using the Kolmogorov–Smirnov and Shapiro-Wilk tests. The means of auxological and clinical laboratory data in the CPP and PT groups were compared using independent t-tests. Relationships between serum and urinary variables and positive results on the GnRHST were determined using Pearson bivariate correlation, receiver-operating characteristic (ROC) curve, and logistic regression analyses. Associations between the serum and urinary LH concentrations were evaluated using a linear regression analysis. Mean serum and urinary LH levels in response to GnRHa treatment were compared using paired-sample t-tests. Graphs were drawn using GraphPad Prism 6 (GraphPad Software Inc., San Diego, CA, USA). Data were analyzed using IBM SPSS Statistics ver. 22.0 (IBM Co., Armonk, NY, USA), with P<0.05 considered statistically significant.
Of the 55 girls who showed signs of early puberty and had undergone both the GnRHST and urinary gonadotropin assays, 29 were assigned to the CPP group (peak LH≥ 5 IU/L) and 26 to the PT group (peak LH<5 IU/L). Their anthropometric and laboratory characteristics are summarized in
The associations between pairs of urinary variables and serum variables obtained in the GnRHST were calculated using a Pearson bivariate correlation analysis (
To determine whether measuring the urinary LH level can be used to diagnose CPP, ROC curves were drawn, and the areas under the curves (AUCs) were calculated (
Girls with CPP who were treated with GnRHa for 12 and 24 weeks saw significant reductions from baseline in their serum and urinary LH and FSH concentrations (
Urinary concentrations of gonadotropins can be easily, noninvasively, and conveniently measured, which could make them a good substitute for a positive GnRHST in diagnosing girls with CPP. In this study, we found that randomly measured urinary concentrations of gonadotropins correlated strongly with the serum concentrations of gonadotropins and that urinary gonadotropins were predictive of a positive GnRHST. Thus, this method could be useful for monitoring girls with CPP during GnRHa treatment.
Although the GnRHST is the gold standard test for diagnosing CPP, it is time-consuming, expensive, invasive, and labor intensive for both patients and physicians in outpatient clinics. Previous studies have sought methods to reduce the discomfort and improve the convenience of diagnosing CPP. One study tested the efficacy of using a single unstimulated LH measurement of ≥0.83 IU/L as a diagnosis of CPP, but that study did not test the efficacy of the LH/FSH ratio [
Compared with blood sampling, urine sampling is convenient, painless, and more tolerable for young patients. In the 1960s, scientists first detected the urinary excretion of LH and FSH in adults and reported the usefulness of urine samples in differentiating sex hormone disorders [
Gonadotropins can be measured in 24-hour urine samples obtained from healthy prepubertal children, but that method is cumbersome, requiring many technical procedures and much effort [
Our results are consistent with those of previous studies. We found that urinary LH levels and urinary LH/FSH ratios, but not urinary FSH/Cr ratios, differed significantly between girls with CPP and those with PT. We did not evaluate Cr-adjusted urinary concentrations because that can result in over-or underestimation of urinary LH and FSH concentrations. Similar to previous studies, we found that urinary gonadotropin concentrations correlated strongly with serum gonadotropin levels without adjustment for Cr. However, urinary LH/Cr and FSH/Cr ratios did not correlate positively with their serum basal and peak levels. Singh et al. [
FMV urine samples are better indicators of gonadotropin excretion than random samples because they avoid nocturnal episodic variation. The use of FMV urine to monitor pubertal changes was introduced more than a decade ago [
To date, no consensus has been reached about monitoring the effects of treatment in patients with CPP, other than assessing physical changes and bone age. HPG suppression is usually evaluated by measuring GnRH-stimulated or unstimulated serum gonadotropin and sex hormone concentrations [
This study had several limitations. Our sample size was small, and we did not evaluate changes in urinary gonadotropin concentrations after the girls stopped taking GnRHa.
In conclusion, this study describes a reliable and convenient method for initially screening and monitoring young patients with signs of early puberty. The single random urinary LH concentration and urinary LH/FSH ratio were strong noninvasive reflectors of a positive GnRHST result. Urinary LH and FSH levels could also be useful markers for monitoring HPG suppression during GnRHa treatment of children with CPP. Longer-term studies with more participants are needed to determine the optimal strategies and guidance for obtaining urine samples to assess hormone status in children and adolescents.
These study protocols were approved by the Institutional Review Board (IRB) of Chungnam National University Hospital, Daejeon, South Korea, and written informed consent was obtained from all participating patients and their legitimate guardians (IRB No. 2020-09-095).
No potential conflict of interest relevant to this article was reported.
ROC curves for a positive GnRH stimulation test. ROC, receiver-operating characteristics; GnRH: gonadotropin-releasing hormone; LH, luteinizing hormone; FSH, follicle-stimulating hormone.
Changes in serum and urinary gonadotropin levels in girls with CPP before and after treatment with GnRHa. (A1) Urinary LH level, (A2) urinary FSH level. (B1) serum LH level, and (B2) serum FSH level. Values are presented as mean±standard deviation. CPP, central precocious puberty, GnRHa: gonadotropin-releasing hormone agonist; LH, luteinizing hormone; FSH, follicle-stimulating hormone.
Auxological and clinical characteristics of the participants
Characteristic | Central precocious puberty (n=29) | Premature thelarche (n=26) | |
---|---|---|---|
Age (yr) | 8.2±0.6 | 8.1±0.7 | 0.558 |
Height (cm) | 131.7±4.6 | 129.8±5.2 | 0.2 |
Weight (kg) | 30.3±3.8 | 30.9 ±5.9 | 0.747 |
BMI (kg/m2) | 17.4±1.52 | 18.1±2.5 | 0.256 |
Height SDS | 1.01±0.76 | 0.81±0.76 | 0.355 |
Weight SDS | 0.91±0.96 | 0.86±0.79 | 0.828 |
BMI SDS | 0.45±0.66 | 1.0±1.9 | 0.173 |
MPH (cm) | 160.5±3.9 | 160.6±3.7 | 0.934 |
PAH (cm) | 150.2±5.5 | 151.0±3.9 | 0.566 |
MPH-PAH | 10.3±5.4 | 9.6±4.3 | 0.625 |
Bone age (yr) | 10.4±0.9 | 9.9±0.8 | 0.059 |
BA–CA (yr) | 2.2±0.8 | 1.8±0.8 | 0.087 |
Tanner stage | 2.2±0.4 | 2.1±0.3 | 0.375 |
IGF-1 (ng/mL) | 272.5±96.2 | 222.4±33.4 | 0.021 |
Basal LH (IU/L) | 1.2±0.7 | 1.0±0.5 | 0.005 |
Basal FSH (IU/L) | 2.1±1.2 | 1.9±1.7 | 0.002 |
Estradiol (pg/mL) | 22.1±10.5 | 15.2±8.1 | 0.018 |
Peak LH (IU/L) | 12.3±7.9 | 3.0±1.1 | <0.001 |
Peak FSH (IU/L) | 9.5±3.2 | 9.4±3.2 | 0.889 |
Peak LH/FSH ratio | 1.3±0.6 | 0.4±0.2 | <0.001 |
Urine LH (IU/L) | 1.2±0.6 | 0.7±0.3 | 0.004 |
Urine FSH (IU/L) | 14.6±3.4 | 13.4±2.3 | 0.168 |
Urine Cr. (mg/dL) | 113.5±56.6 | 93.1±51.1 | 0.247 |
Urine LH/FSH ratio | 0.08±0.03 | 0.05±0.02 | 0.009 |
Urine LH/Cr | 0.014±0.01 | 0.01±0.01 | 0.577 |
Urine FSH/Cr | 0.19±0.2 | 0.24±0.03 | 0.508 |
Values are presented as mean±standard deviation.
BMI, body mass index; SDS, standard deviation score; MPH, mean parenteral height; PAH, predicted adult height; BA–CA, bone age minus chronological age; IGF-1, insulin-like growth factor-1; LH, luteinizing hormone; FSH, follicle-stimulating hormone; Cr, creatinine.
Independent
Correlations between urinary and serum gonadotropin levels
Variable | bLH | bFSH | pLH | pFSH | uLH |
---|---|---|---|---|---|
bFSH | 0.622 |
||||
pLH | 0.644 |
0.464 |
|||
pFSH | 0.057 | 0.158 | 0.383 |
||
uLH | 0.660 |
0.618 |
0.280 |
0.168 | |
uFSH | 0.543 |
0.625 |
0.192 | 0.088 | 0.523 |
bLH, basal serum luteinizing hormone; bFSH, basal serum folliclestimulating hormone; pLH, peak serum luteinizing hormone on gonadotropin-releasing hormone (GnRH) stimulation test; pFSH, peak serum follicle-stimulating hormone on GnRH stimulation test; uLH, urine luteinizing hormone; uFSH, urine folliclestimulating hormone.
All variables were calculated by Pearson bivariate correlations analysis.
Area under the receiver-operating characteristic curve for a positive gonadotropin-releasing hormone stimulation test
Variable | Area | SE | |
---|---|---|---|
Peak serum LH (≥5 IU/L) | 1.000 | 0.000 | <0.001 |
Peak serum LH/FSH ratio | 0.958 | 0.030 | <0.001 |
Basal serum LH (IU/L) | 0.749 | 0.072 | 0.003 |
Urine LH/FSH ratio | 0.731 | 0.075 | 0.007 |
Urine LH (IU/L) | 0.721 | 0.077 | 0.010 |
SE, standard error; LH, luteinizing hormone; FSH, follicle-stimulating hormone.