The Effect Of Environmental Endocrine Disruptors On The Precocious Puberty And The Mechanism Of Chinese Herbal Medicine Therapy

Objective: To explore the role of environmental endocrine disruptors(EEDs) inthe onset of precocious puberty(PP) and the therapeutic effect of traditional Chinesemedicine. Living biological samples from aquaculture in the suburbs were collected,human EEDs exposure were measured and evaluated.Mehtods: (1) The blood samples were collected from 110 cases of precociouspuberty patients and 100 cases of normal children. The concentrations ofoctylphenol(OP), bisphenol A(BPA) and di-2-ethylhexyl phthalate(DEHP),di-n-butyl phthalate (DBP) in the blood serum samples were measured by usingreversed-phase high performance liquid chromatography(HPLC) and gaschromatography. The volume of uterus and ovary, and the content of estradiol(E₂) inthe serum were determined at the same time. The contents of EEDs in the bloodserums of precocious puberty and the indices of the target organs were analyzed bythe methods of correlation and regression. The 21 days immature femaleSprague-Dawley rats were given orally(by gavage)OP, BPA, DEHP(low,moderate, high three dosage) and a mixture of BPA and op 14days; Then therats were sacrificed 24 hours after the last administration. Uterine wet weight,uterine/body weight, the height of the luminal epithelium, the thickness of themyometrium and the gene and protein level of estrogen receptor(ER) of rat uterinewere analysed by one-way ANOVA, and evaluate these estrogen like activity and therelationship between dosage-effect. (3)The therapeutic project of Chinese traditionalmedicine was established to treat children of PP and the experimental animals. Thedeveloping parameters of reproductive organs were observed. Using methods of real—time fluorescence quantitative PCR and immunohistochemistry, we detected theexpression level of estrogen receptor (ER), aromatase(AR) and insulin like factor-1(IGF-1). The data were analysed by one-way ANOVA and T test and to evaluate themechanism of Chinese traditional medicine preventing EEDs estrogen likeactivity. (4)The BPA and BPA+Chinese traditional medicine were given orally(bygavage) to 21 days SD rats. The rats were sacrificed on 0.5h, 1h, 1.5h,4h, 8h, 24h, 48h, 72h, 96h, 6 rats in every group and then measured the BPA concentration in the serum,and analysed the effect of Chinese traditional medicine onthe BPA in the body by Gause-newton method.(5)The EEDs(p-p'-DOE, OP, DEHP) ofthe living biology samples on the 10 surburbs of Shanghai were measured byHPLC,and to evaluate the exposure EEDs of human beings.Results: (1) In the normal control group, OP, BPA, DBP and DEHP (ND～7.42μg/l, NO～6.24μg/l, ND～0.94μg/l, ND～0.97μg/l) were detected in theblood samples respectively. In the precocious puberty group OP, BPA, DBP and DEHP(ND～24.52μg/l, ND～35.18μg/l, ND～15.60μg/l, ND～17.53μg/l) weredetected in the blood samples respectively.The levels of EEDs in blood serum of theprecocious puberty group were notably increased compared with that of the controlgroup(P＜0.01). In the precocious puberty group, there were positive correlationsbetween OP, BPA, DBP and DEHP with volume of uterus ((r=0.676, 0.557,0.456,0.382;P＜0.01); OP, DBP and DEHP; and volume of ovary also had positivecorrelation((r=0.456,0.378.0.689;P＜0.01). (2)In animal experiments, uterine wetweight, uterine/body weight and the thickness of the myometrium were significantlyhigher in the BPA and OP group,and there was a dose-response relationship(P＜0.01),and the uterus ER gene andprotein expression were significantly higher in theBPA and OP group((P＜0.05). The uterine wet weight, uterine/body weight andthickness of the myometrium and uterus ER gene and protein mixture of OP andBPA group were significantly higher than that of the single dose group(P＜0.01).(3)The volume of uterus and ovary decreased in patients after therapy using traditionalChinese medicine (P＜0.01 )The gene and protein expression of ER, IGF-â… and AR ofuteruses and ovary were notably decreased in animals treated withChinese traditional medicine(P＜0.05). (5)The animals absorbed less BPAand the peak time in the blood was delayed after given Chinese traditionalmedicine. (5) Measurement of the living biological samples in the surburbsof Shanghai showed that p-p'-DDE was polluted most extensively, and thenfollowed by4-NP, and DEHP. From the field, study Hangzhou bayarea (Nanhui, Fenxian, Jinshan) and suburbs(Minhang, Jiading, Baoshan, Pudong) were polluted by mostly by p-p'-DDE (56.9% and 50%) and island area (Chongmin) was by 4-NP mostly (43.3%and 38.7%).Conclusions: (1) Normal children and children with precocious pubertyare both contaminated by EEDs, and the later are heavily exposed to EEDs. There isa close statistical relationship between EEDs and the onset of precocious puberty.EEDs are important factors onset orin the induction of the disease. Different kindsof EEDs have different potential in influencing the target organs. (2) The estrogeniceffects of EEDs and their dose-response relationship were validated by uterotrophicassay in immature SD rats. Different kinds of EEDs even below the no-effect levelscould evoke combined additive effects which also domomstrated estrogenic activitiy.(3) Chinese traditional medicine could down regulate the expression of ER,AR andIGF-1 on the target organs. Through the action, the medicine could inhibit theestrogenic effects of EEDs and slow down the accelerated development ofreproductive organs and skeleton in PP patients. (4) The animals absorbed less BPAand the peak time of BPA delayed in the blood from aquaculture ponds aftergiven Chinese traditional medicine. (5) Living biological samples in thesurburbs Shanghai demonstrated that they were polluted extensively andthey might able to be an important factor for human exposure...