The Mechanism Of Mitochondria Damage And The Apoptosis Of Cell In The Toxic Effects Of Methoxychlor On Mice Ovary

There are many exogenous chemicals which can interfere the synthesis, transportation, binding, function or elimination of hormones in vivo, which maitain balance , reproduction, development and behavior in human beings.These chemicals can mimic, strengthen or inhibiti the synthesis of hormones in vivo, in some conditions, they may result in abnormality of tissues and organs as well as tumor. These kinds of chemicals are defined as endocrine disrupting chemicals. More and more evidence indicate that the exposure of these chemicals through occupation and enviorment would influence and change the generation function of human beings.Methoxychlor(MXC) is an organochlorine pesticide, which is widely used to be an alternative to dichloro-diphenyl-trichloroethane (DDT) for the past few years,Its advantages over DDT are that it is more rapidly metabolized excreted in mammals, it is acutely less toxic than DDT and it is biodegradable. It has been reported methoxychlor can affect reproduction of male mice, induce the disorder of hypothalamic-pituitary-ovarian axis, decrease secretion and synthesis of hormones, prolong estruation circle of female rats. For this reason, it is necessary to investigate the toxic effects as well as the mechanisms of MXC on mice ovary. The study was designed to analyze the effects of MXC on ovary, also to make clear whether the mitochondria exert the effects.The study were divided into three parts:Part 1. Construction of methoxychlor poisoned mice model and effects of methoxychlor on mice ovary.MXC were administratede to forty female 39-day-mice at doses of 0 (control), 16, 32, 64mg/(kg·day) via intraperitoneal injection for 20d, then mice were euthanized in estrous. Body weights, organ coefficients of ovary were observed, a quantitive analysis of corpora atretica were conducted,. Serum follicle stimulating hormone(FSH), leteinizing hormone(LH), estradiol(E2) and progestereone(P) were detected with the radio immunoassays Results:①Increasing of body weights was not different between 4 groups after treatment.②Organ weights and organ coefficients of ovaries in mice treated with MXC 32 and 64 mg groups were significantly different from the control group.③Serum follicle stimulating hormone (FSH) and estradiol (E2) were significantly decreased in 32, 64 MXC group. There were no significant differences on both serum progestereone (P) and leteinizing hormone (LH) in the 4 groups. .Conclusion:①MXC could decrease ovary weights and increase number of aatresic follicle.②MXC in experimental dose may induce the disorder of hypothalamic-pituitary-ovarian axis,which induce the decrease of E2 and FSH;③No significant influence on female gonads toxicities was found in low-dose group.Part 2. Research of methoxychlor on the apoptosis of granulosa cell.We used the ovary tissue embeded with paraffin just from the first experiment.Then the apoptosis of granulosa cell were observed by TUNEL and transmission electron microscope ;Bax and Bcl-2 in ovaries were examined with the SP immunohisochemical method. Results:①The number of apoptosis cells in 32,64 mg/ ( kg·d)groups was obviously higherthan that in 16 mg/ ( kg·d)and control groups;②The transmission electron microscope showed the apoptosis body and in 32,64mg/ ( kg·d)group; The expression of bax in 32,64 mg/ ( kg·d)group were significantly increased ,while these dose does not change the expression of bcl-2. Conclusion:①MXC in 32,64 mg/ ( kg·d)group could result in apoptosis of granulosa cell.②MXC in 32,64 mg/ ( kg·d) could increase the expression of bax in ovary ,which cause follicular atrsia.Part 3.The research of methoxychlor on the mitochondria damage of ovary.We erected the model on intraperitoneal injection MXC just like the first experiment. Then Mice were euthanized, their ovaries collected. After the preparation of mitochondria, the generation of ROS and the membrane potential of mice ovary mitochondria were detected by Fluoscence spectrophotometer, the activity of complex of mitochondria was measured by UV spectrophotomete. Results: The activity of complexⅠand the membrane potential of mitochondria were significantly lower in 32 and 64 mg/ ( kg·d) groups when compared to 16 and control groups(P<0.01),While the ROS were higher in 32 and 64 mg/ ( kg·d)groups than 16 and control groups(P<0.01). Conclusion: MXC could cause oxidative stress in mitochondria of mice ovary which may induce the apoptosis of granulosa cells, cause follicular atrsia.