| Objective The objective of the dissertation is to explore the effects of prenatal di-(2-ethylhexyl)phthalate(DEHP) exposure on the development of reproductive systems and related mechanisms in female rats.Methods60-day-old Sprague-Dawley(SD) female rats were mated with sexually matured male rats. The day of virginal plug found was termed gestation day 0(G0). From that day the body weight and the dietary status of the pregnant rats were recorded. Then the pregnant rats were randomly assigned to 2mg/kg, 10mg/kg, 50mg/kg treatment groups and the control group(corn oil) and administrated by gavage from G14 to G19.On the postnatal day 0(PND0), the body weight and anogenital distance(AGD) of new born rats were measured, the condition of foetal deformity and stillbirth were observed. The F1 rats were weaned on PND21 and the female rats were separated from the males. The status of development in F1 female rats were recorded every week. From PND21 each F1 female rat was checked daily for virginal opening, which is the mark of puberty onset. Once the virginal opening, the estrous cycle of female F1 rats were observed based on virginal exfoliative cytology every day. When the female F1 rats were about 70-day-old, they were sacrificed after the body weighting and AGD measuring on anestrus day. Terminal trunk blood samples were collected after the animals were euthanizes by decapitation. The uterus and ovaries were rapidly removed and weighed, and the organ coefficients were calculated. The serum levels of estrodiol(E2), progesterone(P), testosterone(T), Luteinizing hormone(LH)were determined using radioimmunoassay. The relative expression levels of message RNA of aromatase cytochrome P450(P450arom), cholesterol side-chain cleavage enzyme(P450scc), peroxisome proliferator activated receptor γ(PPARγ),steroidogenic acute regulatory protein(St AR), 3β-hydroxysteroid dehydrogenase(3β-HSD), 17β-hydroxysteroid dehydrogenase I(17β-HSDI) and 17β-hydroxysteroid dehydrogenase II(17β-HSDII) of ovaries were determined by real-time fluorescentquantitative polymerase chain reaction(RT-PCR).Results(1) There were no effects of prenatal exposure to different dosage of DEHP on the body weight gain and term of gestation in pregnant rats. No foetal deformity and stillbirth were observed.(2) On postnatal day 0(PND0), the body weight of female F1 rats among treatment and control groups was statistically different(P<0.01).Compared with the control group, the body weight of 10mg/kg and 50mg/kg groups were lower significantly(P<0.05). The body weight of 2mg/kg was significantly higher than that of 10mg/kg and 50mg/kg groups respectively. There were no significant differences of AGD among treatment and control groups(P>0.05).(3)From PND1 to PND21(lactation period), the body weight of female F1 rats among treatment and control groups was statistically different(P<0.001). From PND21 to PND49(prepuberty period), the body weight of female F1 rats among treatment and control groups was statistically different(P<0.05). Compared with the control group,the body weight of 2mg/kg group was higher significantly(P<0.05), while that of50mg/kg was lower significantly(P<0.05). From PND49 to PND70, the periods from puberty to sexual maturity, the body weight of female F1 rats among treatment and control groups was statistically different(P<0.05). Compared with the control group,the body weight of 50mg/kg group was lower significantly(P<0.05). There were no significant differences of body weight among treatment and control groups on PND70(P>0.05).(4) There were no significant differences of virginal opening days among control and treatment groups(P>0.05). The rate of female F1 rats which are always on the anestrus among control and treatment groups were statistically different(P<0.05). Compared with the control group, the rate of that in 50mg/kg group was higher significantly(P<0.05).(5) There were no significant differences of the organ coefficients of uterus and ovaries among control and treatment groups(P>0.05).(6)The serum levels of progesterone(P) of female F1 rats among treatment and control groups were statistically different(P<0.05). Compared with the control group, the serum levels of P of 10mg/kg group was higher significantly(P<0.05). The serum levels of P of 10mg/kg group was significantly higher than that of 2mg/kg group(P<0.05). There was no significant difference of the serum levels of estrodiol(E2)among treatment and control groups(P>0.05). The serum levels of testosterone(T)among treatment and control groups were statistically different(P<0.05). Compared with the control group, the serum level of T of 10mg/kg group was higher significantly(P<0.05). The serum level of T in 10mg/kg group was significantly higher than that of 2mg/kg group(P<0.05). There was no significant difference of the serum levels of luteinizing hormone(LH) among treatment and control groups(P>0.05).(7) There were no significant differences in the m RNA relative expression levels on P450 arom, P450 scc, PPARγ, St AR, 3β-HSD, 17β-HSDI and 17β-HSDII of ovaries among treatment and control groups(P>0.05).Conclusion(1) Prenatal DEHP exposure could disturb the development of embryo in female offspring rats, showing that the body weight of DEHP treatment groups on born day were lower than the control.(2) Prenatal DEHP exposure could disorder the normal estrous cycle and serum steroid hormone levels in female rats.(3) The effects of prenatal DEHP exposure indicated the low-dose effects on the development of reproductive systems in female rats.(4) In this study, there were no significant differences in the m RNA relative expression levels on the synthesis pathway of steroid hormone in ovary. The further study of the related mechanisms should be continued. |
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