Precocious puberty (PP) currently affects 1 in 5000 children and is 10 times more common in girls [1
]. Previous studies [2
] have reported that the overall PP incidence continues to increase and the puberty development age has shown an advancing trend since the 19th century. Advanced progression of secondary sex characteristics occurs in children with PP leading to compromised adult height, poor social adaptability and emotional disorders. Meanwhile, the risk of endocrine disease and even cancer can sharply increase for a child if hormones stay at an abnormal level for a long time [2
]. The cause of precocious puberty may be associated with certain conditions such as infections, tumors, brain abnormalities, histiocytosis, radiation, injuries, or the exposure to environmental endocrine disruptors [2
]. The phthalic acid esters (PAEs) are one of the classes of environmental endocrine disruptors (EEDs) involved in many endocrine diseases, and studies have argued that exposure to PAEs may be associated with PP [4
]. Population studies have indicated that most children are exposed to PAEs. A German study collected the morning urine of 254 children aged 3–14 years from March 2001 to March 2002, and the average concentrations of the primary metabolite MEHP, and secondary metabolites 5 OH-MEHP, and 5 oxo-MEHP were 7.9, 52.1 and 39.9 μg/L, respectively, indicating that children were not free of PAE exposure [5
]. The U.S. NHANES study from 1999 to 2000 found measurable concentrations of MEP, MBP, and MBzP in more than 97% of the 2540 urine samples tested, indication that exposure to PAEs is widespread in the United States [6
]. Animal studies have confirmed that the mechanism of action of PAEs and their metabolites is to act as anti-androgens, but DBP also has comparatively strong estrogenic properties [7
] which may lead to reproductive disruption [8
] in females including reproductive maturation [9
]. However, existing human studies don not seem to have achieved a consensus so far. Colon et al
] observed significantly higher levels of PAEs in 31 Puerto Rican girls with early thelarche compared to controls, which suggested a possible association between PAEs and premature thelarche, whereas, a study on U.S. girls reported phthalate levels in urine were not related with central precocious puberty of pre-pubescent girls [12
]. Therefore, it is necessary to perform a systematic review and meta-analysis to synthesize current study evidence and identifying the association between PAEs and children with PP.
EEDs are chemicals that either mimic or block hormones, thereby altering the normal hormone levels and endocrine function of the body [30
]. EEDs such as dichlorodiphenyltrichloroethane (DDT) [31
] and benzo(a)pyrene (BaP) [34
]were proved to be associated with early puberty in both female rodents and humans. PAEs, as one of the EEDs often found in plastics and many cosmetic products, were confirmed to affect the female reproductive system and accelerate the onset of puberty in female rats [36
]. However, the association between PAEs and children with PP has been somewhat inconsistent [11
To improve limitations and adjust biases of individual studies, we performed a systematic review and meta-analysis to summarize association between PP and target PAEs and their metabolites, including DEHP, DBP, DEP, DiBP, MEHP, MBP, MEHHP, MEOHP, MCPP, MECPP, MiBP, MMP, MBzP, MEP and MBuP. The pooled estimates showed that serum DEHP exposure was associated with PP, and the serum concentrations of DEHP and DBP in PP group were significantly higher than those in the control group. According to the results of our meta-analysis, DEHP is a risk factor for PP and DBP might be related with PP given the instability results of sensitivity analysis. Only one study showed that DEP might be associated with PP. Therefore, we suggested that PAEs may have association with PP. One reason for the results may be that lots of PAEs were produced and used especially in developing countries every year [28
], and DEHP, DBP as well as DEP account for the commonly used ones of plasticizer. That means, children in developing countries such as China and Puerto Rico have more opportunity to come into contact with DEHP, DBP and DEP. Moreover, since children’s metabolic pathways are immature, they are uniquely vulnerable to toxic chemicals in the environment [38
]. Once exposed to PAEs, they may induce LH secretion by the pituitary, altering pubertal development and causing PP.
There is no significant difference in the concentration in both blood and urine of most PAE metabolites, including MEHP, MBP, MEOHP, MECPP, MMP, MBzP and MEP, between the children with PP and the controls. According to existing studies, although some PAEs have weak estrogenic activity in vitro
, the corresponding PAEs metabolites have no estrogenic effects under the same testing conditions [39
]. In addition, another study [41
] also showed that some PAEs were bound to estrogen receptors in vitro
, however, they had no uterotrophic effects and did not affect vaginal epithelial cell cornification in rats. Although there are no significant differences in MEHP, MBP, MEOHP, MECPP,MMP, MBzP and MEP between the children with PP and the controls in our included studies, the controls and the cases had various age, race, weight, height, and BMI, which could lead to the non-significant results in concentration of PAEs metabolites between the two groups. Anyway, we need more and higher quality evidence to prove that. Besides, the serum concentration of MEHHP and MiBP as well as urinary concentration of MBuP were lower in PP group than control group, while, the serum concentration of MCPP, urinary concentration of MEHHP and MEOHP were higher in PP group than control group. However, the differences need further study as all these results were reported in only one study with small sample size.
The lack of consistency between parent PAEs and metabolites associations with PP could be explained as follows: for one thing, there might be some potential for contamination associated with measurement of parent PAEs in serum, however we still tend to draw the conclusion that PAEs might be a potential risk for girls with PP from a statistical point of view. Previous studies showed that parent PAEs, such as DEHP and DBP, found in numerous medical devices, syringes and laboratory devices [42
], may cause contamination during the process from blood sampling in the experiment procedure, leading to a relatively high exposure. Although seven out of nine included studies which measured the parent PAEs in serum used glass devices for all the experiments to avoid any potential contamination except for two studies which did not mention the laboratory device materials, there is still potential for laboratory contamination as well since phthalates are ubiquitous environmental contaminants. For another reason, it would not be possible to state that phthalate metabolites are not associated with PP due to the limited number of studies with small sample sizes included in the meta-analysis for each metabolite. In addition, phthalate metabolite measurements may outweigh the parent PAE measurements for the reasons that the contamination of serum sample of PAEs would be greatly minimized and they represent an integrative measure of exposure to phthalates from multiple sources and routes [43
In the meta-analysis of serum DEHP exposure, the heterogeneity of the pooled estimate decreased after conducting subgroup analysis by country, which suggests that regional diversity is one of the potential sources of heterogeneity. Regarding the association of serum DEHP and DBP exposure and PP risk, consistent results were found in two subgroups, that is to say no regional differences were detected in the association between PP with DEHP or DBP. However, the pooled estimate of association between serum DBP exposure and PP changed from no difference (OR: 3.26, 95% CI: 0.69 to 15.42) to significant difference (OR: 8.31, 95% CI: 3.31 to 20.82) in the sensitivity analysis after removing Yang [29
], which is attributed to it having the largest sample size with the most weight in this study and the fact its results were totally opposite to those of the other studies. One potential reason of the opposite result might be due to the interactions between chemicals. As the girls in Yang were exposed to 14 EEDs with weak estrogenic effects, the interaction of these different EEDs may enhance or weaken the estrogen effect [28
]. As to serum concentration of DBP, the pooled data showed that the concentration of DBP detected in the PP group was higher than that in control group, and when the result of Song 2014 [28
] was removed in the sensitivity analysis, the pooled data changed to no statistical difference. The likely reason for this change may be the highest standard mean difference between two groups the removed trial provided. For urinary concentration of MEHP, the pooled data showed that the PP group had statistically lower MEHP concentration than the control group when removing the result of Chen 2013 [25
]. Except for Chen’s study, the other two studies showed that the control group had a significantly higher intake of seafood and use of plastic cups per month [18
]. This may explain why the MEHP levels in the control group were higher than that in the PP groups.
Despite a rigorous systematic review and meta-analysis approach and relatively high quality of the studies included in the NOS assessment, some limitations should be considered when interpreting and expanding the present results. First, all the involved subjects of included studies were girls, which may affect the generality of research conclusions on PAEs exposure and PP risk for all children. One reason may be that the indicators of puberty in girls (breast bud development, pubic/axillary hair, menses onset) are more easily identified compared with boys (pubic/axillary, nocturnal emissions) and early stages of puberty in boys may be more subtle and require laboratory tests or standardized medical morphological tests. Second, heterogeneity that comes from regional diversity, race diversity, weight difference, sample size difference, age variation and health status, the concentration and duration of exposure of the study subjects or different detection methods, is relatively high for some meta-analysis and might be a potential effect when interpreting those synthesized results. After subgroup analysis conducted by region, the heterogeneity decreased to a large extent, indicating that different exposure levels of PAEs in different countries may be a more important factor in this case, and besides, it could also reflect the influence of race to some extent. Although most studies had shown that weight may be a factor influencing precocious puberty [44
], we could not conduct the meta-analysis considering the girls’ weight as the included studies did not report the weights or BMI of the subjects. Third, the interpretations of association results are not always straightforward, because false-positive results (type I
error) would be introduced when examining the association of phthalate metabolite with PP for children who exposed to multiple different phthalates. Meanwhile, type I error rate might be higher than 5% (using a p
-value of 0.05 as the threshold for significance) because the sample sizes of most included studies were small (less than 100) [45
]. Moreover, available existing evidence was so limited that the present review was restricted to only four most widely studied PAEs and eight metabolites among five countries based on 14 eligible studies, which might introduce potential bias to some extent.