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Adrenarche and the Emotional and Behavioral Problems of Late Childhood
Journal of Adolescent Health 57 (2015) 608e616
www.jahonline.org Original article
Adrenarche and the Emotional and Behavioral Problems of Late Childhood Lisa K. Mundy, Ph.D. a, b, c, *, Helena Romaniuk, Ph.D. a, b, c, d, Louise Canterford, M.Biostat. a, b, Stephen Hearps, P.G.Dip.(Psych) a, b, Russell M. Viner, Ph.D. e, Jordana K. Bayer, Ph.D. a, c, f, Julian G. Simmons, Ph.D. a, g, John B. Carlin, Ph.D. a, c, d, Nicholas B. Allen, Ph.D. g, h, and George C. Patton, M.D. a, b, c a
Murdoch Childrens Research Institute, Melbourne, Victoria, Australia Centre for Adolescent Health, The Royal Children’s Hospital, Melbourne, Victoria, Australia c Department of Paediatrics, The University of Melbourne, Melbourne, Victoria, Australia d Clinical Epidemiology and Biostatistics Unit, The Royal Children’s Hospital, Melbourne, Victoria, Australia e UCL Institute of Child Health, University College London, London, UK f Department of Psychology and Counseling, La Trobe University, Melbourne, Victoria, Australia g Melbourne School of Psychological Sciences, The University of Melbourne, Melbourne, Victoria, Australia h Department of Psychology, University of Oregon, Eugene, Oregon b
Article history: Received April 21, 2015; Accepted September 1, 2015 Keywords: Adrenarche; Peer relationships; Behavior problems; Emotional problems; Puberty; Hormones; CATS study
A B S T R A C T
Purpose: Mental and behavioral disorders increase in prevalence with the passage through puberty. Yet the first symptoms for many children emerge between seven and 11 years, before the pubertal rise in gonadal hormones. A possibility that symptom onset may be linked to the adrenarchal rise in androgens has been little explored. Methods: The Childhood to Adolescence Transition Study recruited a stratified random sample of 1,239 eightenine year olds from primary schools in Melbourne, Australia. Saliva samples were assayed for dehydroepiandrosterone, dehydroepiandrosterone-sulphate (DHEA-S), and testosterone. Emotional and behavioral problems were assessed through parental report on the Strengths and Difficulties Questionnaire. Results: In males, high levels of all androgens were associated with greater total difficulties and peer problems. Higher dehydroepiandrosterone and testosterone were associated with emotional symptoms and DHEA-S with conduct problems. In females, DHEA-S was associated with peer problems. Conclusions: In late childhood, androgens are associated with emotional and behavioral problems in males, raising a possibility that the adrenarchal transition plays a contributing role. If so, the late primary school years may prove to be an important phase for preventing the onset of mental health and behavioral problems in boys. Ó 2015 Society for Adolescent Health and Medicine. All rights reserved.
Conflicts of Interest: The authors report no financial interests or potential conflicts of interest. * Address correspondence to: Dr. Lisa K. Mundy, Ph.D., Centre for Adolescent Health, The Royal Children’s Hospital, Flemington Road, Melbourne, Victoria, 3052, Australia. E-mail address: [email protected] (L.K. Mundy). 1054-139X/Ó 2015 Society for Adolescent Health and Medicine. All rights reserved. http://dx.doi.org/10.1016/j.jadohealth.2015.09.001
IMPLICATIONS AND CONTRIBUTION
This population-based study shows that during late childhood, androgens are associated with emotional and behavioral problems, especially in males. Adrenarche may be an important point for preventive and early intervention.
Late childhood and early adolescence are the peak years of onset for common mental health disorders, with more than half of all disorders having a first onset by 14 years [1]. Puberty transects this risk period and has been implicated in the onset of a range of mental and behavioral disorders that become fully
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manifest in adolescence. Advanced pubertal stage is associated, independently of age, with mental health problems [2]. Furthermore, it is clear that the rise in gonadal hormones is also associated with changes in brain function [3]. These findings have led to suggestions that gonadal hormonal changes play an important role in the onset of common mental health problems [4]. Yet the first symptoms of many of these problems begin somewhat earlier, with symptoms typically emerging between the ages of 7 and 11 years, before the gonadal hormone rise for many children [1,5]. Gonadarche involves the reactivation of the hypothalamic pituitary gonadal axis and results in sexual maturation and reproductive capability [6]. Gonadarche occurs between 8 and 13 years in girls and about 6e12 months later in boys [6]. In contrast to the interest in gonadal maturation, adrenal androgens and adrenarche have attracted less attention [7]. Adrenarche and the maturation of the hypothalamicepituitaryeadrenal axis is the earliest process in a cascade of pubertal hormonal changes. There are few external physical signs of adrenarche, one likely reason for a paucity of studies examining links between adrenal maturation and mental health problems. Adrenarche typically begins between the ages of 6e8 years [6]. The maturation of the zona reticularis in the adrenal gland is accompanied by increasing levels of androgens such as dehydroepiandrosterone (DHEA) and its sulphate DHEA-S (referred together as DHEA(S)) [6]. Testosterone is also secreted from the adrenal glands and levels rise during adrenarche [8]. Throughout we refer to DHEA(S) and testosterone as androgens, and these hormones are almost entirely adrenal in origin before the onset of gonadarche. Adrenal androgens are neurosteroids active in brain regions involved in emotional and behavioral regulation [9]. High levels of adrenal androgens have been associated with affective brain function and symptoms of psychopathology [10]. Understanding the associations between adrenal androgens and emotional and behavior problems may prove important for future interventions to reduce mental health problems [11]. However, current research is inconclusive [7,12,13], with the inconsistencies observed to date likely a result of a number of limitations, such as a focus on single hormones, very small sample sizes, and clinical or high-risk populations. Existing studies have also included wide age ranges including adolescents in puberty, well after the adrenarchal transition, meaning results are confounded by gonadal changes. The present article is the first large population-based study to describe associations between androgens and emotional and behavioral problems in a sample of children assessed around the time of adrenarche. Children were 8e9 years of age, providing a narrow age range in which to examine associations between adrenal hormones and emotional and behavioral problems. Specifically, we address a question of whether children with higher levels of androgens exhibit higher levels of emotional and behavior problems, and we examine this separately for males and females. Methods Study design and participants Data for this study are based on the first wave of the Childhood to Adolescence Transition Study, whose design is described in detail elsewhere [14]. Briefly, children were sampled from a stratified random sample of 43 primary schools (government,
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catholic, independent strata) in metropolitan Melbourne, Australia. School principals provided consent for their school’s participation. If a school did not provide consent to take part then a replacement school from the same stratum was randomly selected and offered participation. In 2012, all grade three children (8e9 years of age; the fourth year of formal schooling) in the selected schools were approached to participate. There were no inclusion/exclusion criteria. Trained graduate research assistants visited each school and guided the children through a brief questionnaire. Children provided a saliva sample between 9 A.M.e10 A.M., when hormonal levels were likely to be at their highest. All components of data collection were voluntary and children could skip any component they did not want to complete. It was not expected that the session would cause distress, but all children were given a resource sheet. A parent (usually the mother) completed two questionnaires (part one [84% completed by mothers] and part two [87% completed by mothers]). Part one was administered as a paper version at the same time as the consent process and took 10 minutes to complete. This included the Strengths and Difficulties Questionnaire (SDQ) and some demographic questions. Part two was a 30-minute questionnaire, which was sent to parents via post or online, after the child’s data collection session. This included questions about the family environment, prenatal and perinatal factors, the child’s health, sleep, nutrition, and pubertal development. Ethics approval was granted by the Royal Children’s Hospital Human Research Ethics Committee (#31089). Permission was granted from the Victorian Department of Education and Early Childhood Development Office and the Catholic Education Office Melbourne to recruit through their schools. Measures Child emotional and behavioral problems. The Australian version of the SDQ was used to assess child emotional and behavioral problems. The SDQ is a brief, well-validated behavioral screening questionnaire used widely in cohort studies. It has moderate to strong internal reliability (a .59e.80) and has strong concurrent validity with diagnostic interviews [15,16]. The SDQ consists of 25 items, which are divided between five scales: emotional symptoms, conduct problems, hyperactivity/inattention, peer relationship problems, and prosocial behavior [17]. A total difficulties score is derived from the first four subscales and is a marker of overall psychopathology. Parents rate each item on the basis of the child’s behavior over the last 6 months. The score for each subscale ranges from 0 to 10 and for total difficulties from 0 to 40. Higher scores indicate higher levels of emotional or behavioral problems, with the exception of the prosocial subscale where higher scores indicate better functioning. Androgen hormone assessment. Salivary measures of DHEA(S) and testosterone were used to provide an index of adrenal maturation. Levels of these hormones in saliva have been shown to be a reliable index of those in blood in children and adolescents [12]. Saliva samples were collected in the class setting using the passive drool method. This was timed for 3 minutes, and the total saliva sample provided was approximately 3 mL. Children who were unable to provide 1 mL were invited to provide a second sample (2%). This sample was collected in a small group setting at the end of the data collection session, approximately 30 minutes after the initial sample was collected. Samples were
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transported, immediately after data collection, in a portable insulated container with a frozen ice brick to the laboratory at Murdoch Childrens Research Institute and stored at 30 C until assayed. Before testing, thawed samples were weighed to obtain the volume of saliva collected, which was used to calculate flow rate. Following the manufacturer’s guidelines and adopting a strategy used in other studies of hormone-behavior associations, DHEA-S was adjusted for flow rate because concentrations in saliva decrease as flow rates increase [18,19]. Thus, results for DHEA-S are presented as a function of time, that is, pg/minute. Samples were centrifuged at 3000 RPM at 4 C for 15 minutes. Samples were assayed in duplicate, within 6 months of collection, using highly sensitive enzyme immunoassay kits specifically designed for use with saliva (Salimetrics, State College, PA). The average of the duplicate tests was used in the analyses. For DHEA, the range of sensitivity was from 10.2 to 1,000 pg/mL; the average intraassay coefficient of variation (CV) was 9.9% and the average interassay CV was 16.5%. For DHEA-S, the range of sensitivity was from 188.9 to 15,300 pg/mL; average intra-assay and interassay CVs were 8.3% and 9.1%, respectively. For testosterone, the range of sensitivity was 6.1e600 pg/mL, with average intra-assay and interassay CVs 7.8% and 13.2%, respectively. Covariates. To assess potential confounding, the following variables were included in regression models because they may be associated with both a rise in androgens and emotional and behavioral problems: Child’s age and family socioeconomic status (SES) [20e22]. Child’s age in months at the time of the child assessment was used. Family SES was assigned from small area deprivation measures calculated from home postcode using the Index of Relative Socioeconomic Disadvantage (population mean [standard deviation {SD}] ¼ 1,000 [100]) from the Australian Bureau of Statistics censusebased local neighborhood SocioEconomic Index for Areas [23]. Higher scores on this measure indicate better socioeconomic status. Additional individual and family demographic characteristics were used to summarize the sample. Child pubertal development was assessed using an adaptation of the Pubertal Development Scale (PDS) for parent report [24]. The PDS is acceptable for use in community samples of children [24] and has been used for parent report in similar cohort studies [25]. Items included breast growth (girls only), skin changes, body odor, body hair, and menarche (girls only). Parents rated their child’s pubertal development as “has not started yet,” “has barely started,” “has definitely started,” or “seems complete,” apart from menarche, which was a dichotomous yes/no response. An overall pubertal development score was created by summing responses to obtain a total score and this score was used to categorize children as “prepubertal,” “early pubertal,” “midpubertal,” or “late pubertal/postpubertal” [24]. Data analysis Hormone measures; pubertal stage; SDQ scores; child, maternal, and family characteristics were summarized and compared by sex. Mean and SD were calculated for all continuous measures. Median and interquartile ranges were also calculated for the hormone measures, as their distributions were positively skewed and compared using ManneWhitneyeWilcoxon test. Percentages were calculated for categorical variables and compared using chi-squared tests.
Linear generalized estimating equations with robust standard errors were used to estimate the effect of hormone levels on SDQ outcomes and to account for the clustering of children in schools [26]. Because of the potential effect modification by sex of the association between SDQ outcomes and hormone levels, separate models were fitted for females and males. Hormone levels were modeled as continuous and categorical measures. Continuous: Each hormone measure was log transformed (to the base of 2) before inclusion in the model; this scaling meant that regression coefficients represented expected change in outcome per doubling of the hormone level. Categorical: Hormone levels were categorized into tertiles after they had been standardized by age to identify hormonal development relative to their peers. Standardization was performed before categorization because there was some statistical evidence that hormone levels were associated with age for males and females, aged 96 and <120 months, consistent with previous studies [27,28]. Hormone levels standardized by age were estimated by calculating residuals after regressing, using a linear fit, each hormone measure on age separately for males and females. These residuals were then categorized into tertiles to identify those with minimal, intermediate and advanced hormonal development relative to their peers of the same age. The effect of each continuous and categorical hormone measure on SDQ scores was estimated after adjustment for child’s age and Socio-Economic Index for Areas score. Marginal means were estimated from the adjusted model at the mean values for these covariates. Sensitivity analysis was performed using only data for those who were identified as prepubescent. All data analyses were undertaken using Stata 13 (Stata Corp, College Station, TX) [29]. Results In total, 1,239 (54%) participants were recruited through the provision of active, informed parent consent (see Figure 1). Of the recruited sample, the mean age was 108 months (SD 5 months, range 94e128 months). Of the children recruited, 1,194 (96%) children and 1,222 (99%) of their parents took part in wave one. The recruited sample contained a slightly smaller proportion of males (46%) compared with census data for eight- to nine-yearold children enrolled in grade three across the state of Victoria in Australia (51% males) [30]. This sample scored very slightly higher on a measure of SES compared with the entire Australian population (mean ¼ 1,012, SD ¼ 67 vs. mean ¼ 1,000, SD ¼ 100) [23]. A higher percentage identified as indigenous compared with all grade three children in Victoria (5% vs. 1%). Of the 1,239 children recruited, 100 were excluded from these analyses because they did not have at least one salivary hormone measure and one SDQ subscale score and a further 15 were excluded because they were <8 and 10 years of age. Of the resulting 1,124 (91%) who were included in the analysis sample: 511 (45%) were males and 613 (55%) were females. Child’s age, sex, and family SES were available for all children in the analysis sample. Child and family characteristics were similar for males and females: born in Australia (87% vs. 89%); Aboriginal Torres Strait Islander (5% vs. 4%); English main language spoken at home (85% vs. 85%; reported for 397 males and 482 females), and single parent household (12% vs. 12%; reported for 398 males and 482 females). Males scored higher on the SDQ total difficulties, conduct, and hyperactivity/inattention scales and scored lower on the prosocial behavior scale (Table 1). Females had higher
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Figure 1. Flowchart showing participant recruitment through to final analysis sample.
DHEA and testosterone levels, with no difference in DHEA-S levels between males and females. Males were more likely to be in the prepuberty category of the PDS. Table 2 shows the association between each SDQ scale and each hormonal index for males and females. In males, greater DHEA levels were associated with higher levels of peer relationship problems and emotional symptoms. Greater DHEA-S levels were associated with higher total difficulties, conduct problems, and peer relationship problems. Higher testosterone levels were associated with increased total difficulties, peer
relationship problems, and emotional symptoms. In females, greater DHEA-S levels were associated with greater peer relationship problems. For males, similar findings were found when only looking at prepubertal children (i.e., excluding children with physical pubertal changes as defined by the PDS; see Supplementary Table 1). However, when looking at prepubertal females additional associations were found. DHEA-S and testosterone were negatively associated with prosocial behavior, and DHEA-S was associated with increased total difficulties.
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Table 1 SDQ scales, hormonal measures, and demographic characteristics for the analytical sample of 8- to 9-year-old grade 3 children (n ¼ 1,124) stratified by sex Measure/characteristic
Females (N ¼ 613) N
Hormonal measure DHEA (pg/mL) DHEA-S (pg/min) Testosterone (pg/mL)
613 603 613
DHEA (pg/mL) DHEA-S (pg/min) Testosterone (pg/mL)
613 603 613
Males (N ¼ 511) Median (IQR)
N
27.2 (36.3) 427 (672) 20.5 (13.9)
510 501 505
Mean (SD)
455 455 455 455
26 92 94 7
510 501 505
(57.6) (20.2) (20.7) (1.5)
612 612 612 612 612 613
7.8 1.2 2.8 1.4 2.2 8.6
613
10 (4.2)
613
19.0 (33.0) 414 (662) 18.8 (13.0)
<.0001 .63 .0001
27.7 (26.5) 788 (1,311) 19.6 (10.1)
<.0001 .69 <.0001
n (%) 386 386 386 386
Mean (SD) SDQ scale score Total difficulties Conduct problems Hyperactivity/inattention Peer relationship problems Emotional symptoms Prosocial behavior Children’s characteristic Age (months) Neighborhood characteristic SEIFA disadvantage score
p value
Mean (SD)
37.9 (36.4) 759 (1,097) 22.2 (11.4) n (%)
Pubertal stage Prepuberty Early pubertal Midpubertal Late pubertal/postpubertal
Comparison Median (IQR)
315 58 10 3
(81.6) (15.0) (2.6) (.78)
<.0001a
Mean (SD)
(5.11) (1.39) (2.23) (1.58) (2.12) (1.53)
1,013 (69.0)
511 511 510 511 510 511
9.0 1.5 3.9 1.5 2.0 8.1
511
10 (4.4)
511
(5.85) (1.58) (2.63) (1.70) (2.02) (1.84)
1,013 (64.2)
.0005 .008 <.0001 .59 .12 <.0001 .49 .25
DHEA ¼ dehydroepiandrosterone; DHEA-S ¼ dehydroepiandrosterone sulphate; IQR ¼ interquartile range; IRSD ¼ index of Relative Socio-Economic Disadvantage; SD ¼ standard deviation; SDQ ¼ Strengths and Difficulties Questionnaire; SEIFA ¼ Socioeconomic Index for Areas. a p value from joint test of null hypothesis of no group differences.
Table 3 shows associations between androgen levels and SDQ scores for males and females. For males, DHEA-S levels were found to be associated with total difficulties, with mean difference in total difficulties of .92 (95% confidence interval: .43 to 2.3) for
those with intermediate levels of DHEA-S and 2.5 (95% confidence interval: 1.3e3.7) for those with advanced levels of DHEA-S compared with those who had minimal DHEA-S development (see Figure 2). DHEA-S levels in males were also found to be
Table 2 Estimated adjusteda associations between three androgensb and SDQ scores in 8- to 9-year-old grade 3 children (n ¼ 1,124) stratified by sex Outcome
Total difficulties
Conduct problems
Hyperactivity/inattention
Peer relationship problems
Emotional symptoms
Prosocial behavior
Hormone
DHEA DHEA-S Testosterone DHEA DHEA-S Testosterone DHEA DHEA-S Testosterone DHEA DHEA-S Testosterone DHEA DHEA-S Testosterone DHEA DHEA-S Testosterone
Females (n ¼ 613)
Males (n ¼ 511)
bc (95% CI)
p value
bc (95% CI)
p value
.03 .17 .09 .03 .05 .06 .02 .04 .04 .02 .09 .06 .05 .006 .008 .05 .06 .06
.84 .23 .69 .49 .19 .38 .74 .52 .70 .63 .01 .34 .31 .91 .94 .28 .16 .38
.39 .46 .65 .04 .11 .08 .01 .12 .05 .16 .15 .24 .19 .07 .27 .03 .02 .04
.06 .003 .02 .50 .04 .25 .87 .09 .70 .01 .001 .002 .005 .08 .002 .58 .56 .74
(.30 to .24) (.10 to .44) (.33 to .51) (.05 to .11) (.03 to .13) (.07 to .19) (.16 to .11) (.08 to .16) (.22 to .15) (.06 to .10) (.02 to .16) (.07 to .19) (.15 to .05) (.11 to .10) (.18 to .20) (.14 to .04) (.13 to .02) (.19 to .07)
(.02 to .81) (.15 to .76) (.13 to 1.2) (.08 to .17) (.005 to .22) (.06 to .23) (.17 to .14) (.02 to .25) (.19 to .29) (.04 to .28) (.06 to .24) (.09 to .40) (.06 to .33) (.01 to .16) (.10 to .45) (.09 to .15) (.05 to .10) (.18 to .25)
CI ¼ confidence interval; DHEA ¼ dehydroepiandrosterone; DHEA-S ¼ dehydroepiandrosterone sulphate; SDQ ¼ Strengths and Difficulties Questionnaire; SEIFA ¼ Socioeconomic Index for Areas. a Adjusted for age (months) and SEIFA disadvantage score. b Transformed (log base 2). c Regression coefficients represent expected change in outcome per doubling of the hormone level.
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Table 3 Estimated adjusteda associations between level of hormonal development relative to peersb and SDQ scores in 8- to 9-year-old grade 3 children (n ¼ 1,124) stratified by sex Outcome
Total difficulties
Hormone SDQ
DHEA DHEA-S Testosterone
Conduct problems
DHEA DHEA-S Testosterone
Hyperactivity/inattention
DHEA DHEA-S Testosterone
Peer relationship problems
DHEA DHEA-S Testosterone
Emotional symptoms
DHEA DHEA-S Testosterone
Prosocial behavior
DHEA DHEA-S Testosterone
Development level relative to peersc
Females (n ¼ 613)
b (95% CI)
p valued
b (95% CI)
Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced
.10 .17 .67 .67 .28 .34 .04 .10 .03 .19 .15 .20 .23 .06 .07 .17 .06 .07 .001 .09 .37 .38 .08 .14 .28 .10 .22 .02 .11 .07 .19 .12 .07 .31 .17 .17
.94
.41 1.2 .92 2.5 .91 1.4 .03 .20 .21 .51 .17 .19 .10 .09 .23 .82 .06 .15 .16 .41 .24 .79 .28 .47 .21 .59 .26 .34 .42 .57 .17 .12 .05 .03 .002 .02
(.97 to .77) (1.1 to .77) (.22 to 1.6) (.47 to 1.8) (.76 to 1.3) (.69 to 1.4) (.24 to .16) (.22 to .41) (.21 to .27) (.12 to .50) (.14 to .44) (.08 to .48) (.25 to .71) (.54 to .42) (.32 to .46) (.38 to .73) (.56 to .43) (.50 to .36) (.29 to .29) (.33 to .15) (.10 to .63) (.10 to .67) (.28 to .45) (.16 to .44) (.73 to .17) (.46 to .25) (.19 to .62) (.46 to .41) (.28 to .49) (.39 to .53) (.49 to .11) (.42 to .18) (.36 to .23) (.64 to .02) (.49 to .15) (.48 to .13)
Males (n ¼ 511)
.32 .78 .68 .42 .33 .31 .83 .95 .74 .007 .67 .46 .47 .86 .44 .18 .48
(.77 to 1.6) (.16 to 2.5) (.43 to 2.3) (1.3 to 3.7) (.54 to 2.4) (.32 to 2.5) (.42 to .36) (.24 to .64) (.13 to .54) (.11 to .91) (.24 to .57) (.13 to .51) (.39 to .60) (.59 to .41) (.47 to .93) (.29 to 1.4) (.62 to .75) (.34 to .65) (.18 to .49) (.01 to .80) (.09 to .57) (.45 to 1.1) (.04 to .59) (.13 to .81) (.22 to .65) (.13 to 1.0) (.18 to .70) (.01 to .69) (.03 to .81) (.16 to .98) (.22 to .56) (.26 to .50) (.34 to .43) (.34 to .29) (.41 to .40) (.44 to .41)
p valued .22 <.0001 .04 .30 .04 .48 .69 .002 .83 .12 <.0001 .02 .04 .16 .01 .68 .94 .99
CI ¼ confidence interval; DHEA ¼ dehydroepiandrosterone; DHEA-S ¼ dehydroepiandrosterone sulphate; SDQ ¼ Strengths and Difficulties Questionnaire; SEIFA ¼ Socioeconomic Index for Areas. a Adjusted for age (months) and SEIFA disadvantage score. b Age standardized hormone levels classified into tertiles. c Baseline category: Minimal development relative to peers. d p value from joint test of null hypothesis of no group differences.
associated with conduct problems, hyperactivity/inattention, and peer relationship problems. For DHEA, some evidence was found of an association only with emotional symptoms. For testosterone, there was some evidence of an association with total difficulties, peer relationship problems, and emotional symptoms. In females, there was only statistical evidence of an association between DHEA-S and peer relationship problems.
Discussion This is the first large population-based study to show that DHEA(S) and testosterone are associated with emotional and behavioral problems in males in late childhood. For males, all androgens tended to be associated with greater total difficulties and peer relationship problems. Higher DHEA and testosterone were associated with emotional symptoms and DHEA-S with conduct problems. In contrast, for females, little evidence was found of relationships between androgens with emotional and
behavioral problems with only DHEA-S associated with peer problems. The narrow age range used in the present study allowed us to examine the associations with adrenarche, independently of age [22,31]. To exclude a possibility that the associations may have been due to early gonadal changes, we carried out sensitivity analyses by removing children rated as having any physical pubertal signs from analyses. Although the specific patterns of association varied slightly in the sensitivity analyses, overall the associations between androgens and emotional and behavioral problems remained for males, suggesting the hormone-behavior associations observed are independent of gonadal changes. This suggests, adrenarche is independently associated with the emergence of emotional and behavioral problems in males. In females, additional associations were found; DHEA-S and testosterone were negatively associated with prosocial behavior, and DHEA-S was associated with increased total difficulties. Strengths of this study include its large size compared with earlier studies, narrow age range focused around adrenarche,
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Figure 2. Estimated marginal means (with 95% confidence intervals) for total difficulties score by sex and level of hormonal development relative to peers (hormone levels were categorized into tertiles after they had been standardized by age to identify hormonal development relative to peers).
inclusion of both males and females in similar proportions, and the use of assays of DHEA(S) and testosterone to gain a measure of adrenal maturation. Some limitations are important to note. An active parental consent process was used to establish this cohort, and as a result, only 54% of parents approached provided written consent for participation (possibly because of the intensive and longitudinal nature of the study). Yet the sample was close to the Australian population mean on SES but did have a higher proportion of Aboriginal Torres Strait Islander children. This suggests that, we have managed to achieve a fairly representative sample but there remains a possibility of some selection bias toward more disadvantaged groups. It seems unlikely, however, that this would result in any major mis-specification of associations between androgens and emotional and behavioral problems. We collected one saliva sample from children at a standardized morning time. Ideally, two or more samples would be collected across days, but this would have been infeasible in this large school-based sample. Although the hormones under study here do show significant intraindividual variation, studies suggest that saliva measures of these hormones are relatively reliable across days and weeks [32]. In assessing DHEA(S) and testosterone at 8e9 years of age to give an index of adrenarchal timing, we are replicating a previously adopted strategy [33]. However, it is possible that androgen levels at this age may not only reflect timing of adrenarche but also individual differences in the rate of adrenal maturation or in baseline (i.e., trait like) levels of these hormones. Levels of testosterone were higher in females than males, consistent with smaller studies in this age group [27]. However, it is possible that we might be seeing very low levels of gonadal production of testosterone in males. Although sensitivity analyses were conducted, pubertal stage was collected by parent report with a possibility of some parents over reporting. Also, gonadal hormonal changes may occur 6e12 months before the onset of breast development in girls [34], so the sensitivity analyses may still have included children in early gonadarche. Finally, there was some inconsistency between hormones in the patterns of association, with the strongest associations generally
being observed for DHEA-S. This may reflect differences in action but is likely also to reflect the fact that DHEA-S is both more abundant in saliva and a more biologically active androgen [35]. Our results provide the clearest evidence to date that androgens are associated with emotional and behavioral problems in males in late childhood [1,5]. Two explanations are important to consider: A possibility that the rise in androgens introduces a period of higher risk for the onset of symptoms particularly in males; or that males on a life course trajectory to earlier adrenarche also have a different pattern of emotional and behavioral development during childhood. Del Giudice et al. [36] have suggested that adrenarche may introduce a period of developmental realignment, with a shift from infant attachment patterns to those of reproductive attachment, resulting in changes in children’s emotional state and behavior. Few studies have investigated the neurobiological effects of DHEA(S) but high DHEA levels have been linked with decreased affect-related brain activity in the midcingulate cortex [10]. Furthermore, an association of high DHEA levels with externalizing symptoms in females appeared partly mediated by posterior insula activation [10]. Adrenal androgens have also been linked to chronic stress and childhood adversity, although findings are inconsistent [37]. However, there is a possibility of adrenarche being “a developmental switch point”; a period during which environmental cues are more likely to affect an individual’s reproductive strategy and related behaviors. This may include patterns of aggression and attachment, characteristics linked to the social, behavioral, and emotional outcomes of this study. If correct, risk factors such as bullying or abuse in the family setting might be expected to have more profound and longer-lasting effects at this age than at later points. The alternative explanation also deserves consideration. Early childhood factors such as small for gestational birth and childhood obesity have been implicated in both the timing of adrenarche and aspects of mental health and behavior [33]. Thus, both the timing of adrenal hormonal change and rates of childhood emotional and behavioral problems may reflect an adaptive strategy set in place early in life [25]. The cross-sectional nature of the data does not allow us to distinguish between these two
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possible explanations. However, the associations with peer problems for both males and females, are similar to those Mensah et al. [25] reported earlier in childhood for children with early puberty. Given that poor peer relationships are often manifest earlier in childhood, it remains possible that early adrenarche is more likely in males with early childhood emotional and behavioral problems. Before puberty, males have generally been found to have higher rates of behavioral problems than females, and emotional problems are generally similar between sexes [38]. The association with DHEA(S) and testosterone suggests that the adrenarchal transition may heighten risks for some boys, providing at least a partial explanation for these sex differences before puberty. This explanation could carry important implications for the timing and focus of early and preventive interventions for emotional and behavioral problems. As an example, given the clear evidence for longitudinal relationships between peer problems and mental health problems, focusing on interventions and preventive interventions to reduce bullying and peer problems during and before the adrenarchal transition may be particularly beneficial [39]. Furthermore, adrenarche may represent a sensitive period for the onset of emotional and behavioral problems in boys [36]. Acknowledgments The authors would like to thank all the families and schools who have participated in this study. The authors would like to thank all staff and volunteers involved in data collection and processing at MCRI. Contributors: G.P., N.A., R.V., J.B. led the overall conception and design of the study. L.M., G.P., and N.A. contributed to study implementation and coordination. L.M. contributed to acquisition of the data. H.R., L.C., S.H., J.C., and G.P. were involved in analysis. All authors were involved in interpretation of data. L.M. drafted the manuscript. All authors were involved in revising the article critically for important intellectual context, and final approval of the version to be published. Funding Sources This research was supported by a project grant from Australia’s National Health and Medical Research Council (NHMRC; #1010018). MCRI research is supported by the Victorian Government’s Operational Infrastructure Program. Dr Mundy is partially supported by a grant from the Invergowrie Foundation. Professor Patton is supported by a Senior Principal Research Fellowship from NHMRC. The funding sources had no further role in study design, in the collection, analysis or interpretation of data, in the writing of the report, and in the decision to submit the article for publication. Supplementary Data Supplementary data related to this article can be found at http://dx.doi.org/10.1016/j.jadohealth.2015.09.001. References [1] Kessler RC, Berglund P, Demler O, et al. Lifetime prevalence and age-ofonset distributions of DSM-IV disorders in the National Comorbidity Survey Replication. Arch Gen Psychiatry 2005;62:593e602.
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[2] Patton GC, Hibbert ME, Carlin J, et al. Menarche and the onset of depression and anxiety in Victoria, Australia. J Epidemiol Community Health 1996;50: 661e6. [3] Goddings AL, Burnett Heyes S, Bird G, et al. The relationship between puberty and social emotion processing. Dev Sci 2012;15:801e11. [4] Angold A, Costello EJ, Erkanli A, et al. Pubertal changes in hormone levels and depression in girls. Psychol Med 1999;29:1043e53. [5] Costello EJ, Mustillo S, Erkanli A, et al. Prevalence and development of psychiatric disorders in childhood and adolescence. Arch Gen Psychiatry 2003;60:837e44. [6] Dorn LD, Biro FM. Puberty and its measurement: A decade in review. J Res Adolesc 2011;21:180e95. [7] Goodyer IM, Park RJ, Netherton CM, et al. Possible role of cortisol and dehydroepiandrosterone in human development and psychopathology. Br J Psychiatry 2001;179:243e9. [8] Grumbach MM, Styne DM. Puberty: Ontogeny, neuroendocrinology, physiology, and disorders. Williams Textbook Endocrinol 1998;9: 1509e625. [9] Maninger N, Wolkowitz OM, Reus VI, et al. Neurobiological and neuropsychiatric effects of dehydroepiandrosterone (DHEA) and DHEA sulfate (DHEAS). Front Neuroendocrinol 2009;30:65e91. [10] Whittle S, Simmons JG, Byrne ML, et al. Associations between early adrenarche, affective brain function and mental health in children. Soc Cogn Affect Neurosci 2015;10:1282e90. [11] Marceau K, Ruttle PL, Shirtcliff EA, et al. Developmental and contextual considerations for adrenal and gonadal hormone functioning during adolescence: Implications for adolescent mental health. Dev Psychobiol 2015;57:742e68. [12] Goodyer IM, Herbert J, Altham PM, et al. Adrenal secretion during major depression in 8- to 16-year-olds, I. Altered diurnal rhythms in salivary cortisol and dehydroepiandrosterone (DHEA) at presentation. Psychol Med 1996;26:245e56. [13] Susman EJ, Inoff-Germain G, Nottelmann ED, et al. Hormones, emotional dispositions, and aggressive attributes in young adolescents. Child Dev 1987;58:1114e34. [14] Mundy L, Simmons J, Allen NB, et al. Study protocol: The childhood to adolescence transition study (CATS). BMC Pediatr 2013;13:160. [15] Goodman R, Scott S. Comparing the Strengths and Difficulties Questionnaire and the Child Behavior Checklist: Is small beautiful? J Abnorm Child Psychol 1999;27:17e24. [16] Hawes DJ, Dadds MR. Australian data and psychometric properties of the Strengths and Difficulties Questionnaire. Aust N Z J Psychiatry 2004;38: 644e51. [17] Goodman R. The Strengths and Difficulties Questionnaire: A research note. J Child Psychol Psychiatry 1997;38:581e6. [18] Vining RF, McGinley RA. The measurement of hormones in saliva: Possibilities and pitfalls. J Steroid Biochem 1987;27:81e94. [19] Rohleder N, Wolf JM, Maldonado EF, et al. The psychosocial stress-induced increase in salivary alpha-amylase is independent of saliva flow rate. Psychophysiology 2006;43:645e52. [20] Euling SY, Herman-Giddens ME, Lee PA, et al. Examination of US pubertytiming data from 1940 to 1994 for secular trends: Panel findings. Pediatrics 2008;121(Suppl 3):S172e91. [21] Sawyer MG, Arney FM, Baghurst PA, et al. The mental health of young people in Australia: Key findings from the child and adolescent component of the national survey of mental health and well-being. Aust N Z J Psychiatry 2001;35:806e14. [22] Dorn LD, Dahl RE, Woodward HR, et al. Defining the boundaries of early adolescence: A user’s guide to assessing pubertal status and pubertal timing in research with adolescents. Appl Dev Sci 2006;10:30e56. [23] Australian Bureau of Statistics. Census of Population and Housing: Socio-Economic Indexes for Areas (SEIFA), Australia; 2011. Available at: http://www.abs.gov.au/ausstats/[email protected]/mf/2001.0. Accessed November 26, 2014. [24] Petersen AC, Crockett L, Richards M, et al. A self-report measure of pubertal status: Reliability, validity, and initial norms. J Youth Adolesc 1988;17: 117e33. [25] Mensah FK, Bayer JK, Wake M, et al. Early puberty and childhood social and behavioral adjustment. J Adolesc Health 2013;53:118e24. [26] Liang KY, Zeger SL. Longitudinal data analysis using generalized linear models. Biometrika 1986;73:13e22. [27] Granger DA, Schwartz EB, Booth A, et al. Salivary testosterone determination in studies of child health and development. Horm Behav 1999;35: 18e27. [28] Granger DA, Schwartz EB, Booth A, et al. Assessing dehydroepiandrosterone in saliva: A simple radioimmunoassay for use in studies of children, adolescents and adults. Psychoneuroendocrinology 1999;24:567e79. [29] StataCorp. Stata Statistical Software: Release 13. College Station, TX: StataCorp LP; 2013. [30] Australian Bureau of Statistics. Available at: http://www.abs.gov.au/ AUSSTATS/[email protected]/DetailsPage/4221.02013. Accessed November 26, 2014.
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[31] Shirtcliff EA, Dahl RE, Pollak SD. Pubertal development: Correspondence between hormonal and physical development. Child Dev 2009;80:327e37. [32] Dabbs JM. Salivary testosterone measurements: Reliability across hours, days, and weeks. Physiol Behav 1990;48:83e6. [33] Ong KK, Potau N, Petry CJ, et al. Opposing influences of prenatal and postnatal weight gain on adrenarche in normal boys and girls. J Clin Endocrinol Metab 2004;89:2647e51. [34] Biro FM, Pinney SM, Huang B, et al. Hormone changes in peripubertal girls. J Clin Endocrinol Metab 2014;99:3829e35. [35] Wolf OT, Kirschbaum C. Actions of dehydroepiandrosterone and its sulfate in the central nervous system: Effects on cognition and emotion in animals and humans. Brain Res Rev 1999;30:264e88.
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www.jahonline.org Original article
Adrenarche and the Emotional and Behavioral Problems of Late Childhood Lisa K. Mundy, Ph.D. a, b, c, *, Helena Romaniuk, Ph.D. a, b, c, d, Louise Canterford, M.Biostat. a, b, Stephen Hearps, P.G.Dip.(Psych) a, b, Russell M. Viner, Ph.D. e, Jordana K. Bayer, Ph.D. a, c, f, Julian G. Simmons, Ph.D. a, g, John B. Carlin, Ph.D. a, c, d, Nicholas B. Allen, Ph.D. g, h, and George C. Patton, M.D. a, b, c a
Murdoch Childrens Research Institute, Melbourne, Victoria, Australia Centre for Adolescent Health, The Royal Children’s Hospital, Melbourne, Victoria, Australia c Department of Paediatrics, The University of Melbourne, Melbourne, Victoria, Australia d Clinical Epidemiology and Biostatistics Unit, The Royal Children’s Hospital, Melbourne, Victoria, Australia e UCL Institute of Child Health, University College London, London, UK f Department of Psychology and Counseling, La Trobe University, Melbourne, Victoria, Australia g Melbourne School of Psychological Sciences, The University of Melbourne, Melbourne, Victoria, Australia h Department of Psychology, University of Oregon, Eugene, Oregon b
Article history: Received April 21, 2015; Accepted September 1, 2015 Keywords: Adrenarche; Peer relationships; Behavior problems; Emotional problems; Puberty; Hormones; CATS study
A B S T R A C T
Purpose: Mental and behavioral disorders increase in prevalence with the passage through puberty. Yet the first symptoms for many children emerge between seven and 11 years, before the pubertal rise in gonadal hormones. A possibility that symptom onset may be linked to the adrenarchal rise in androgens has been little explored. Methods: The Childhood to Adolescence Transition Study recruited a stratified random sample of 1,239 eightenine year olds from primary schools in Melbourne, Australia. Saliva samples were assayed for dehydroepiandrosterone, dehydroepiandrosterone-sulphate (DHEA-S), and testosterone. Emotional and behavioral problems were assessed through parental report on the Strengths and Difficulties Questionnaire. Results: In males, high levels of all androgens were associated with greater total difficulties and peer problems. Higher dehydroepiandrosterone and testosterone were associated with emotional symptoms and DHEA-S with conduct problems. In females, DHEA-S was associated with peer problems. Conclusions: In late childhood, androgens are associated with emotional and behavioral problems in males, raising a possibility that the adrenarchal transition plays a contributing role. If so, the late primary school years may prove to be an important phase for preventing the onset of mental health and behavioral problems in boys. Ó 2015 Society for Adolescent Health and Medicine. All rights reserved.
Conflicts of Interest: The authors report no financial interests or potential conflicts of interest. * Address correspondence to: Dr. Lisa K. Mundy, Ph.D., Centre for Adolescent Health, The Royal Children’s Hospital, Flemington Road, Melbourne, Victoria, 3052, Australia. E-mail address: [email protected] (L.K. Mundy). 1054-139X/Ó 2015 Society for Adolescent Health and Medicine. All rights reserved. http://dx.doi.org/10.1016/j.jadohealth.2015.09.001
IMPLICATIONS AND CONTRIBUTION
This population-based study shows that during late childhood, androgens are associated with emotional and behavioral problems, especially in males. Adrenarche may be an important point for preventive and early intervention.
Late childhood and early adolescence are the peak years of onset for common mental health disorders, with more than half of all disorders having a first onset by 14 years [1]. Puberty transects this risk period and has been implicated in the onset of a range of mental and behavioral disorders that become fully
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manifest in adolescence. Advanced pubertal stage is associated, independently of age, with mental health problems [2]. Furthermore, it is clear that the rise in gonadal hormones is also associated with changes in brain function [3]. These findings have led to suggestions that gonadal hormonal changes play an important role in the onset of common mental health problems [4]. Yet the first symptoms of many of these problems begin somewhat earlier, with symptoms typically emerging between the ages of 7 and 11 years, before the gonadal hormone rise for many children [1,5]. Gonadarche involves the reactivation of the hypothalamic pituitary gonadal axis and results in sexual maturation and reproductive capability [6]. Gonadarche occurs between 8 and 13 years in girls and about 6e12 months later in boys [6]. In contrast to the interest in gonadal maturation, adrenal androgens and adrenarche have attracted less attention [7]. Adrenarche and the maturation of the hypothalamicepituitaryeadrenal axis is the earliest process in a cascade of pubertal hormonal changes. There are few external physical signs of adrenarche, one likely reason for a paucity of studies examining links between adrenal maturation and mental health problems. Adrenarche typically begins between the ages of 6e8 years [6]. The maturation of the zona reticularis in the adrenal gland is accompanied by increasing levels of androgens such as dehydroepiandrosterone (DHEA) and its sulphate DHEA-S (referred together as DHEA(S)) [6]. Testosterone is also secreted from the adrenal glands and levels rise during adrenarche [8]. Throughout we refer to DHEA(S) and testosterone as androgens, and these hormones are almost entirely adrenal in origin before the onset of gonadarche. Adrenal androgens are neurosteroids active in brain regions involved in emotional and behavioral regulation [9]. High levels of adrenal androgens have been associated with affective brain function and symptoms of psychopathology [10]. Understanding the associations between adrenal androgens and emotional and behavior problems may prove important for future interventions to reduce mental health problems [11]. However, current research is inconclusive [7,12,13], with the inconsistencies observed to date likely a result of a number of limitations, such as a focus on single hormones, very small sample sizes, and clinical or high-risk populations. Existing studies have also included wide age ranges including adolescents in puberty, well after the adrenarchal transition, meaning results are confounded by gonadal changes. The present article is the first large population-based study to describe associations between androgens and emotional and behavioral problems in a sample of children assessed around the time of adrenarche. Children were 8e9 years of age, providing a narrow age range in which to examine associations between adrenal hormones and emotional and behavioral problems. Specifically, we address a question of whether children with higher levels of androgens exhibit higher levels of emotional and behavior problems, and we examine this separately for males and females. Methods Study design and participants Data for this study are based on the first wave of the Childhood to Adolescence Transition Study, whose design is described in detail elsewhere [14]. Briefly, children were sampled from a stratified random sample of 43 primary schools (government,
609
catholic, independent strata) in metropolitan Melbourne, Australia. School principals provided consent for their school’s participation. If a school did not provide consent to take part then a replacement school from the same stratum was randomly selected and offered participation. In 2012, all grade three children (8e9 years of age; the fourth year of formal schooling) in the selected schools were approached to participate. There were no inclusion/exclusion criteria. Trained graduate research assistants visited each school and guided the children through a brief questionnaire. Children provided a saliva sample between 9 A.M.e10 A.M., when hormonal levels were likely to be at their highest. All components of data collection were voluntary and children could skip any component they did not want to complete. It was not expected that the session would cause distress, but all children were given a resource sheet. A parent (usually the mother) completed two questionnaires (part one [84% completed by mothers] and part two [87% completed by mothers]). Part one was administered as a paper version at the same time as the consent process and took 10 minutes to complete. This included the Strengths and Difficulties Questionnaire (SDQ) and some demographic questions. Part two was a 30-minute questionnaire, which was sent to parents via post or online, after the child’s data collection session. This included questions about the family environment, prenatal and perinatal factors, the child’s health, sleep, nutrition, and pubertal development. Ethics approval was granted by the Royal Children’s Hospital Human Research Ethics Committee (#31089). Permission was granted from the Victorian Department of Education and Early Childhood Development Office and the Catholic Education Office Melbourne to recruit through their schools. Measures Child emotional and behavioral problems. The Australian version of the SDQ was used to assess child emotional and behavioral problems. The SDQ is a brief, well-validated behavioral screening questionnaire used widely in cohort studies. It has moderate to strong internal reliability (a .59e.80) and has strong concurrent validity with diagnostic interviews [15,16]. The SDQ consists of 25 items, which are divided between five scales: emotional symptoms, conduct problems, hyperactivity/inattention, peer relationship problems, and prosocial behavior [17]. A total difficulties score is derived from the first four subscales and is a marker of overall psychopathology. Parents rate each item on the basis of the child’s behavior over the last 6 months. The score for each subscale ranges from 0 to 10 and for total difficulties from 0 to 40. Higher scores indicate higher levels of emotional or behavioral problems, with the exception of the prosocial subscale where higher scores indicate better functioning. Androgen hormone assessment. Salivary measures of DHEA(S) and testosterone were used to provide an index of adrenal maturation. Levels of these hormones in saliva have been shown to be a reliable index of those in blood in children and adolescents [12]. Saliva samples were collected in the class setting using the passive drool method. This was timed for 3 minutes, and the total saliva sample provided was approximately 3 mL. Children who were unable to provide 1 mL were invited to provide a second sample (2%). This sample was collected in a small group setting at the end of the data collection session, approximately 30 minutes after the initial sample was collected. Samples were
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transported, immediately after data collection, in a portable insulated container with a frozen ice brick to the laboratory at Murdoch Childrens Research Institute and stored at 30 C until assayed. Before testing, thawed samples were weighed to obtain the volume of saliva collected, which was used to calculate flow rate. Following the manufacturer’s guidelines and adopting a strategy used in other studies of hormone-behavior associations, DHEA-S was adjusted for flow rate because concentrations in saliva decrease as flow rates increase [18,19]. Thus, results for DHEA-S are presented as a function of time, that is, pg/minute. Samples were centrifuged at 3000 RPM at 4 C for 15 minutes. Samples were assayed in duplicate, within 6 months of collection, using highly sensitive enzyme immunoassay kits specifically designed for use with saliva (Salimetrics, State College, PA). The average of the duplicate tests was used in the analyses. For DHEA, the range of sensitivity was from 10.2 to 1,000 pg/mL; the average intraassay coefficient of variation (CV) was 9.9% and the average interassay CV was 16.5%. For DHEA-S, the range of sensitivity was from 188.9 to 15,300 pg/mL; average intra-assay and interassay CVs were 8.3% and 9.1%, respectively. For testosterone, the range of sensitivity was 6.1e600 pg/mL, with average intra-assay and interassay CVs 7.8% and 13.2%, respectively. Covariates. To assess potential confounding, the following variables were included in regression models because they may be associated with both a rise in androgens and emotional and behavioral problems: Child’s age and family socioeconomic status (SES) [20e22]. Child’s age in months at the time of the child assessment was used. Family SES was assigned from small area deprivation measures calculated from home postcode using the Index of Relative Socioeconomic Disadvantage (population mean [standard deviation {SD}] ¼ 1,000 [100]) from the Australian Bureau of Statistics censusebased local neighborhood SocioEconomic Index for Areas [23]. Higher scores on this measure indicate better socioeconomic status. Additional individual and family demographic characteristics were used to summarize the sample. Child pubertal development was assessed using an adaptation of the Pubertal Development Scale (PDS) for parent report [24]. The PDS is acceptable for use in community samples of children [24] and has been used for parent report in similar cohort studies [25]. Items included breast growth (girls only), skin changes, body odor, body hair, and menarche (girls only). Parents rated their child’s pubertal development as “has not started yet,” “has barely started,” “has definitely started,” or “seems complete,” apart from menarche, which was a dichotomous yes/no response. An overall pubertal development score was created by summing responses to obtain a total score and this score was used to categorize children as “prepubertal,” “early pubertal,” “midpubertal,” or “late pubertal/postpubertal” [24]. Data analysis Hormone measures; pubertal stage; SDQ scores; child, maternal, and family characteristics were summarized and compared by sex. Mean and SD were calculated for all continuous measures. Median and interquartile ranges were also calculated for the hormone measures, as their distributions were positively skewed and compared using ManneWhitneyeWilcoxon test. Percentages were calculated for categorical variables and compared using chi-squared tests.
Linear generalized estimating equations with robust standard errors were used to estimate the effect of hormone levels on SDQ outcomes and to account for the clustering of children in schools [26]. Because of the potential effect modification by sex of the association between SDQ outcomes and hormone levels, separate models were fitted for females and males. Hormone levels were modeled as continuous and categorical measures. Continuous: Each hormone measure was log transformed (to the base of 2) before inclusion in the model; this scaling meant that regression coefficients represented expected change in outcome per doubling of the hormone level. Categorical: Hormone levels were categorized into tertiles after they had been standardized by age to identify hormonal development relative to their peers. Standardization was performed before categorization because there was some statistical evidence that hormone levels were associated with age for males and females, aged 96 and <120 months, consistent with previous studies [27,28]. Hormone levels standardized by age were estimated by calculating residuals after regressing, using a linear fit, each hormone measure on age separately for males and females. These residuals were then categorized into tertiles to identify those with minimal, intermediate and advanced hormonal development relative to their peers of the same age. The effect of each continuous and categorical hormone measure on SDQ scores was estimated after adjustment for child’s age and Socio-Economic Index for Areas score. Marginal means were estimated from the adjusted model at the mean values for these covariates. Sensitivity analysis was performed using only data for those who were identified as prepubescent. All data analyses were undertaken using Stata 13 (Stata Corp, College Station, TX) [29]. Results In total, 1,239 (54%) participants were recruited through the provision of active, informed parent consent (see Figure 1). Of the recruited sample, the mean age was 108 months (SD 5 months, range 94e128 months). Of the children recruited, 1,194 (96%) children and 1,222 (99%) of their parents took part in wave one. The recruited sample contained a slightly smaller proportion of males (46%) compared with census data for eight- to nine-yearold children enrolled in grade three across the state of Victoria in Australia (51% males) [30]. This sample scored very slightly higher on a measure of SES compared with the entire Australian population (mean ¼ 1,012, SD ¼ 67 vs. mean ¼ 1,000, SD ¼ 100) [23]. A higher percentage identified as indigenous compared with all grade three children in Victoria (5% vs. 1%). Of the 1,239 children recruited, 100 were excluded from these analyses because they did not have at least one salivary hormone measure and one SDQ subscale score and a further 15 were excluded because they were <8 and 10 years of age. Of the resulting 1,124 (91%) who were included in the analysis sample: 511 (45%) were males and 613 (55%) were females. Child’s age, sex, and family SES were available for all children in the analysis sample. Child and family characteristics were similar for males and females: born in Australia (87% vs. 89%); Aboriginal Torres Strait Islander (5% vs. 4%); English main language spoken at home (85% vs. 85%; reported for 397 males and 482 females), and single parent household (12% vs. 12%; reported for 398 males and 482 females). Males scored higher on the SDQ total difficulties, conduct, and hyperactivity/inattention scales and scored lower on the prosocial behavior scale (Table 1). Females had higher
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611
Figure 1. Flowchart showing participant recruitment through to final analysis sample.
DHEA and testosterone levels, with no difference in DHEA-S levels between males and females. Males were more likely to be in the prepuberty category of the PDS. Table 2 shows the association between each SDQ scale and each hormonal index for males and females. In males, greater DHEA levels were associated with higher levels of peer relationship problems and emotional symptoms. Greater DHEA-S levels were associated with higher total difficulties, conduct problems, and peer relationship problems. Higher testosterone levels were associated with increased total difficulties, peer
relationship problems, and emotional symptoms. In females, greater DHEA-S levels were associated with greater peer relationship problems. For males, similar findings were found when only looking at prepubertal children (i.e., excluding children with physical pubertal changes as defined by the PDS; see Supplementary Table 1). However, when looking at prepubertal females additional associations were found. DHEA-S and testosterone were negatively associated with prosocial behavior, and DHEA-S was associated with increased total difficulties.
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Table 1 SDQ scales, hormonal measures, and demographic characteristics for the analytical sample of 8- to 9-year-old grade 3 children (n ¼ 1,124) stratified by sex Measure/characteristic
Females (N ¼ 613) N
Hormonal measure DHEA (pg/mL) DHEA-S (pg/min) Testosterone (pg/mL)
613 603 613
DHEA (pg/mL) DHEA-S (pg/min) Testosterone (pg/mL)
613 603 613
Males (N ¼ 511) Median (IQR)
N
27.2 (36.3) 427 (672) 20.5 (13.9)
510 501 505
Mean (SD)
455 455 455 455
26 92 94 7
510 501 505
(57.6) (20.2) (20.7) (1.5)
612 612 612 612 612 613
7.8 1.2 2.8 1.4 2.2 8.6
613
10 (4.2)
613
19.0 (33.0) 414 (662) 18.8 (13.0)
<.0001 .63 .0001
27.7 (26.5) 788 (1,311) 19.6 (10.1)
<.0001 .69 <.0001
n (%) 386 386 386 386
Mean (SD) SDQ scale score Total difficulties Conduct problems Hyperactivity/inattention Peer relationship problems Emotional symptoms Prosocial behavior Children’s characteristic Age (months) Neighborhood characteristic SEIFA disadvantage score
p value
Mean (SD)
37.9 (36.4) 759 (1,097) 22.2 (11.4) n (%)
Pubertal stage Prepuberty Early pubertal Midpubertal Late pubertal/postpubertal
Comparison Median (IQR)
315 58 10 3
(81.6) (15.0) (2.6) (.78)
<.0001a
Mean (SD)
(5.11) (1.39) (2.23) (1.58) (2.12) (1.53)
1,013 (69.0)
511 511 510 511 510 511
9.0 1.5 3.9 1.5 2.0 8.1
511
10 (4.4)
511
(5.85) (1.58) (2.63) (1.70) (2.02) (1.84)
1,013 (64.2)
.0005 .008 <.0001 .59 .12 <.0001 .49 .25
DHEA ¼ dehydroepiandrosterone; DHEA-S ¼ dehydroepiandrosterone sulphate; IQR ¼ interquartile range; IRSD ¼ index of Relative Socio-Economic Disadvantage; SD ¼ standard deviation; SDQ ¼ Strengths and Difficulties Questionnaire; SEIFA ¼ Socioeconomic Index for Areas. a p value from joint test of null hypothesis of no group differences.
Table 3 shows associations between androgen levels and SDQ scores for males and females. For males, DHEA-S levels were found to be associated with total difficulties, with mean difference in total difficulties of .92 (95% confidence interval: .43 to 2.3) for
those with intermediate levels of DHEA-S and 2.5 (95% confidence interval: 1.3e3.7) for those with advanced levels of DHEA-S compared with those who had minimal DHEA-S development (see Figure 2). DHEA-S levels in males were also found to be
Table 2 Estimated adjusteda associations between three androgensb and SDQ scores in 8- to 9-year-old grade 3 children (n ¼ 1,124) stratified by sex Outcome
Total difficulties
Conduct problems
Hyperactivity/inattention
Peer relationship problems
Emotional symptoms
Prosocial behavior
Hormone
DHEA DHEA-S Testosterone DHEA DHEA-S Testosterone DHEA DHEA-S Testosterone DHEA DHEA-S Testosterone DHEA DHEA-S Testosterone DHEA DHEA-S Testosterone
Females (n ¼ 613)
Males (n ¼ 511)
bc (95% CI)
p value
bc (95% CI)
p value
.03 .17 .09 .03 .05 .06 .02 .04 .04 .02 .09 .06 .05 .006 .008 .05 .06 .06
.84 .23 .69 .49 .19 .38 .74 .52 .70 .63 .01 .34 .31 .91 .94 .28 .16 .38
.39 .46 .65 .04 .11 .08 .01 .12 .05 .16 .15 .24 .19 .07 .27 .03 .02 .04
.06 .003 .02 .50 .04 .25 .87 .09 .70 .01 .001 .002 .005 .08 .002 .58 .56 .74
(.30 to .24) (.10 to .44) (.33 to .51) (.05 to .11) (.03 to .13) (.07 to .19) (.16 to .11) (.08 to .16) (.22 to .15) (.06 to .10) (.02 to .16) (.07 to .19) (.15 to .05) (.11 to .10) (.18 to .20) (.14 to .04) (.13 to .02) (.19 to .07)
(.02 to .81) (.15 to .76) (.13 to 1.2) (.08 to .17) (.005 to .22) (.06 to .23) (.17 to .14) (.02 to .25) (.19 to .29) (.04 to .28) (.06 to .24) (.09 to .40) (.06 to .33) (.01 to .16) (.10 to .45) (.09 to .15) (.05 to .10) (.18 to .25)
CI ¼ confidence interval; DHEA ¼ dehydroepiandrosterone; DHEA-S ¼ dehydroepiandrosterone sulphate; SDQ ¼ Strengths and Difficulties Questionnaire; SEIFA ¼ Socioeconomic Index for Areas. a Adjusted for age (months) and SEIFA disadvantage score. b Transformed (log base 2). c Regression coefficients represent expected change in outcome per doubling of the hormone level.
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613
Table 3 Estimated adjusteda associations between level of hormonal development relative to peersb and SDQ scores in 8- to 9-year-old grade 3 children (n ¼ 1,124) stratified by sex Outcome
Total difficulties
Hormone SDQ
DHEA DHEA-S Testosterone
Conduct problems
DHEA DHEA-S Testosterone
Hyperactivity/inattention
DHEA DHEA-S Testosterone
Peer relationship problems
DHEA DHEA-S Testosterone
Emotional symptoms
DHEA DHEA-S Testosterone
Prosocial behavior
DHEA DHEA-S Testosterone
Development level relative to peersc
Females (n ¼ 613)
b (95% CI)
p valued
b (95% CI)
Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced Intermediate Advanced
.10 .17 .67 .67 .28 .34 .04 .10 .03 .19 .15 .20 .23 .06 .07 .17 .06 .07 .001 .09 .37 .38 .08 .14 .28 .10 .22 .02 .11 .07 .19 .12 .07 .31 .17 .17
.94
.41 1.2 .92 2.5 .91 1.4 .03 .20 .21 .51 .17 .19 .10 .09 .23 .82 .06 .15 .16 .41 .24 .79 .28 .47 .21 .59 .26 .34 .42 .57 .17 .12 .05 .03 .002 .02
(.97 to .77) (1.1 to .77) (.22 to 1.6) (.47 to 1.8) (.76 to 1.3) (.69 to 1.4) (.24 to .16) (.22 to .41) (.21 to .27) (.12 to .50) (.14 to .44) (.08 to .48) (.25 to .71) (.54 to .42) (.32 to .46) (.38 to .73) (.56 to .43) (.50 to .36) (.29 to .29) (.33 to .15) (.10 to .63) (.10 to .67) (.28 to .45) (.16 to .44) (.73 to .17) (.46 to .25) (.19 to .62) (.46 to .41) (.28 to .49) (.39 to .53) (.49 to .11) (.42 to .18) (.36 to .23) (.64 to .02) (.49 to .15) (.48 to .13)
Males (n ¼ 511)
.32 .78 .68 .42 .33 .31 .83 .95 .74 .007 .67 .46 .47 .86 .44 .18 .48
(.77 to 1.6) (.16 to 2.5) (.43 to 2.3) (1.3 to 3.7) (.54 to 2.4) (.32 to 2.5) (.42 to .36) (.24 to .64) (.13 to .54) (.11 to .91) (.24 to .57) (.13 to .51) (.39 to .60) (.59 to .41) (.47 to .93) (.29 to 1.4) (.62 to .75) (.34 to .65) (.18 to .49) (.01 to .80) (.09 to .57) (.45 to 1.1) (.04 to .59) (.13 to .81) (.22 to .65) (.13 to 1.0) (.18 to .70) (.01 to .69) (.03 to .81) (.16 to .98) (.22 to .56) (.26 to .50) (.34 to .43) (.34 to .29) (.41 to .40) (.44 to .41)
p valued .22 <.0001 .04 .30 .04 .48 .69 .002 .83 .12 <.0001 .02 .04 .16 .01 .68 .94 .99
CI ¼ confidence interval; DHEA ¼ dehydroepiandrosterone; DHEA-S ¼ dehydroepiandrosterone sulphate; SDQ ¼ Strengths and Difficulties Questionnaire; SEIFA ¼ Socioeconomic Index for Areas. a Adjusted for age (months) and SEIFA disadvantage score. b Age standardized hormone levels classified into tertiles. c Baseline category: Minimal development relative to peers. d p value from joint test of null hypothesis of no group differences.
associated with conduct problems, hyperactivity/inattention, and peer relationship problems. For DHEA, some evidence was found of an association only with emotional symptoms. For testosterone, there was some evidence of an association with total difficulties, peer relationship problems, and emotional symptoms. In females, there was only statistical evidence of an association between DHEA-S and peer relationship problems.
Discussion This is the first large population-based study to show that DHEA(S) and testosterone are associated with emotional and behavioral problems in males in late childhood. For males, all androgens tended to be associated with greater total difficulties and peer relationship problems. Higher DHEA and testosterone were associated with emotional symptoms and DHEA-S with conduct problems. In contrast, for females, little evidence was found of relationships between androgens with emotional and
behavioral problems with only DHEA-S associated with peer problems. The narrow age range used in the present study allowed us to examine the associations with adrenarche, independently of age [22,31]. To exclude a possibility that the associations may have been due to early gonadal changes, we carried out sensitivity analyses by removing children rated as having any physical pubertal signs from analyses. Although the specific patterns of association varied slightly in the sensitivity analyses, overall the associations between androgens and emotional and behavioral problems remained for males, suggesting the hormone-behavior associations observed are independent of gonadal changes. This suggests, adrenarche is independently associated with the emergence of emotional and behavioral problems in males. In females, additional associations were found; DHEA-S and testosterone were negatively associated with prosocial behavior, and DHEA-S was associated with increased total difficulties. Strengths of this study include its large size compared with earlier studies, narrow age range focused around adrenarche,
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Figure 2. Estimated marginal means (with 95% confidence intervals) for total difficulties score by sex and level of hormonal development relative to peers (hormone levels were categorized into tertiles after they had been standardized by age to identify hormonal development relative to peers).
inclusion of both males and females in similar proportions, and the use of assays of DHEA(S) and testosterone to gain a measure of adrenal maturation. Some limitations are important to note. An active parental consent process was used to establish this cohort, and as a result, only 54% of parents approached provided written consent for participation (possibly because of the intensive and longitudinal nature of the study). Yet the sample was close to the Australian population mean on SES but did have a higher proportion of Aboriginal Torres Strait Islander children. This suggests that, we have managed to achieve a fairly representative sample but there remains a possibility of some selection bias toward more disadvantaged groups. It seems unlikely, however, that this would result in any major mis-specification of associations between androgens and emotional and behavioral problems. We collected one saliva sample from children at a standardized morning time. Ideally, two or more samples would be collected across days, but this would have been infeasible in this large school-based sample. Although the hormones under study here do show significant intraindividual variation, studies suggest that saliva measures of these hormones are relatively reliable across days and weeks [32]. In assessing DHEA(S) and testosterone at 8e9 years of age to give an index of adrenarchal timing, we are replicating a previously adopted strategy [33]. However, it is possible that androgen levels at this age may not only reflect timing of adrenarche but also individual differences in the rate of adrenal maturation or in baseline (i.e., trait like) levels of these hormones. Levels of testosterone were higher in females than males, consistent with smaller studies in this age group [27]. However, it is possible that we might be seeing very low levels of gonadal production of testosterone in males. Although sensitivity analyses were conducted, pubertal stage was collected by parent report with a possibility of some parents over reporting. Also, gonadal hormonal changes may occur 6e12 months before the onset of breast development in girls [34], so the sensitivity analyses may still have included children in early gonadarche. Finally, there was some inconsistency between hormones in the patterns of association, with the strongest associations generally
being observed for DHEA-S. This may reflect differences in action but is likely also to reflect the fact that DHEA-S is both more abundant in saliva and a more biologically active androgen [35]. Our results provide the clearest evidence to date that androgens are associated with emotional and behavioral problems in males in late childhood [1,5]. Two explanations are important to consider: A possibility that the rise in androgens introduces a period of higher risk for the onset of symptoms particularly in males; or that males on a life course trajectory to earlier adrenarche also have a different pattern of emotional and behavioral development during childhood. Del Giudice et al. [36] have suggested that adrenarche may introduce a period of developmental realignment, with a shift from infant attachment patterns to those of reproductive attachment, resulting in changes in children’s emotional state and behavior. Few studies have investigated the neurobiological effects of DHEA(S) but high DHEA levels have been linked with decreased affect-related brain activity in the midcingulate cortex [10]. Furthermore, an association of high DHEA levels with externalizing symptoms in females appeared partly mediated by posterior insula activation [10]. Adrenal androgens have also been linked to chronic stress and childhood adversity, although findings are inconsistent [37]. However, there is a possibility of adrenarche being “a developmental switch point”; a period during which environmental cues are more likely to affect an individual’s reproductive strategy and related behaviors. This may include patterns of aggression and attachment, characteristics linked to the social, behavioral, and emotional outcomes of this study. If correct, risk factors such as bullying or abuse in the family setting might be expected to have more profound and longer-lasting effects at this age than at later points. The alternative explanation also deserves consideration. Early childhood factors such as small for gestational birth and childhood obesity have been implicated in both the timing of adrenarche and aspects of mental health and behavior [33]. Thus, both the timing of adrenal hormonal change and rates of childhood emotional and behavioral problems may reflect an adaptive strategy set in place early in life [25]. The cross-sectional nature of the data does not allow us to distinguish between these two
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possible explanations. However, the associations with peer problems for both males and females, are similar to those Mensah et al. [25] reported earlier in childhood for children with early puberty. Given that poor peer relationships are often manifest earlier in childhood, it remains possible that early adrenarche is more likely in males with early childhood emotional and behavioral problems. Before puberty, males have generally been found to have higher rates of behavioral problems than females, and emotional problems are generally similar between sexes [38]. The association with DHEA(S) and testosterone suggests that the adrenarchal transition may heighten risks for some boys, providing at least a partial explanation for these sex differences before puberty. This explanation could carry important implications for the timing and focus of early and preventive interventions for emotional and behavioral problems. As an example, given the clear evidence for longitudinal relationships between peer problems and mental health problems, focusing on interventions and preventive interventions to reduce bullying and peer problems during and before the adrenarchal transition may be particularly beneficial [39]. Furthermore, adrenarche may represent a sensitive period for the onset of emotional and behavioral problems in boys [36]. Acknowledgments The authors would like to thank all the families and schools who have participated in this study. The authors would like to thank all staff and volunteers involved in data collection and processing at MCRI. Contributors: G.P., N.A., R.V., J.B. led the overall conception and design of the study. L.M., G.P., and N.A. contributed to study implementation and coordination. L.M. contributed to acquisition of the data. H.R., L.C., S.H., J.C., and G.P. were involved in analysis. All authors were involved in interpretation of data. L.M. drafted the manuscript. All authors were involved in revising the article critically for important intellectual context, and final approval of the version to be published. Funding Sources This research was supported by a project grant from Australia’s National Health and Medical Research Council (NHMRC; #1010018). MCRI research is supported by the Victorian Government’s Operational Infrastructure Program. Dr Mundy is partially supported by a grant from the Invergowrie Foundation. Professor Patton is supported by a Senior Principal Research Fellowship from NHMRC. The funding sources had no further role in study design, in the collection, analysis or interpretation of data, in the writing of the report, and in the decision to submit the article for publication. Supplementary Data Supplementary data related to this article can be found at http://dx.doi.org/10.1016/j.jadohealth.2015.09.001. References [1] Kessler RC, Berglund P, Demler O, et al. Lifetime prevalence and age-ofonset distributions of DSM-IV disorders in the National Comorbidity Survey Replication. Arch Gen Psychiatry 2005;62:593e602.
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