| In recent years, along with the advance of survival rate of tumor patients , the chemotherapy medicine with the result that lead to the medical damage is getting more attention gradually. The chemotherapy medicine has the cell toxicity to the ovary organization, damaging the ovary function, making the young female sufferer appear menopause syndrome and infertility, also, influencing their living quality and self-confident seriously. The GnRH (Gonadotropin-releasing-Hormone) repress female sex gland stalk, can reduce a chemotherapy-induced damage to the original follicle, have the protection function to ovary.The GnRHant(Gonadotropin-releasing-Hormone Antagonist) as one of the GnRHa(Gonadotropin-releasing-hormone analogues), is stronger to the female sex gland stalk, and side effect, the dose and the medicine time all low relativly. Several researches have proved that the GnRHant have an obvious protected function to the ovary, but the domestic lack the related foundation and clinical research still. This experiment aim to set up the animal model of POF to inquiry into the GnRHant protected function and possible mechanism of the ovary function, looking for a certain theory basis for the clinic.Part 1 The model Construction of chemotherapy induced POF in rats50 femal SD rats of 2-3 months were randomLy divided into four groups:the blank matched control, CTX, low,medium and high dose leve(l1,3,5 mg/kg.d) of the VP16(Etoposide).The blank matched control adopts Sodium Chloride 0.9%1mL/d. 5 groups of animals all continue belly injection for the medicine 10d.Comparing the difference of the 5 groups about the wet heavy of ovary and uterine, organizing the variety of the structure of ovary. Adopting the ELISA(Enzyme linked immunosorbant assay)method to measurese 5 groups of big rat FSH/E2 (folliclest imulating hormone /Estradiol)level.Result:The Vp16 has cell toxicity for big rat ovary. Ovary is disorganized and the reduction of follicle has something to do with the medicine amount. The mount of follicle in the high dose and medium dose VP16 groups reduced obviously, mainly the big and medium follicle, and the ovary organization has fibrosis to different degree, has follicle atresied .The grain cell can't secrete more E and P, furtherly interference HPOA (Hypothalamic-pituitary-ovarian axis), thus interference synthesis and secretion of sex hormone, expressing the high level LH/FSH, low lever E2 .This result agreed with human body ovary toxicity of the former report. The 3mg/kg.d experiment amount maches the the POF (premature ovary failure), a main clinical characteristic is low estrogen blood disease and appear the high GnRH blood disease, but the normal metabolism function of animal was basically normal. The animal model make short time, and the ovary histology, endocrine variation agree with with mankind POF extremely.Part 2 Research on the protecting function and mechanism of GnRHant to big rats ovaries under chemotherapyExperiment160 femal SD rat is randomLy divided into six groups: the blank matched control, the cetrorelix group, the CTX group, VP16 group, cetrorelix+CTX group, the cetrorelix+VP16 group, each 10s. Comparing the difference of the 6 groups about the wet heavy of ovary and uterine, and organizing the variety of the structure of ovary. Adopting the ELISA method to measures FSH/E2 evel.Result:1. The CTX and VP16 all cause big rat weight,the ovary and womb wet heavy light, and the ovary organization bleed, bad dead, highly fibrosis, the total follicle amount shows the sharply decrease, particularly is the original follicle and mature follicle .That hints that the ovary toxicity of the chemotherapy medicine may lie in the cell of fission, making the mature follicle reduce, and the follicle placed in a static period don't damage. The cetrorelix protected ovary organization compared with the groups under chemotherapy, have no obviously bleed, dead and the fibrosis ,the number of big follicle reduce and the original follicle grow up comparedly and the total follicle number have no obvious difference compared with matched control. Thus the GnRHant keep the greatly parts of original follicle, which the follicle placed in a static period escape destruction from the chemotherapy medicine, and low down the chemotherapy medicine's damage for the big rat ovary function, protect the storage function of the ovary2. The FSH and E2 can reflect the ovary storage function indirectly, the chemotherapy medicine makes the ovary has the fiber with different degree and the follicle atresied. Thus the grain cell can't secrete E/P, the further interference descends the HPOA, and the sex hormone synthesized and secreted, which expressing the FSH level increase, the E2 levels descend. The result of this research show that, in the CTX and the VP16 groups of the big rat, the FSH lever increased obviously, the E2 levels descend obviously, but the consociation treatment group, the serum E2/FSH lever and the blank matched control didn't show great difference. This list uses the chemotherapy medicine to have obvious harm to the ovary function. The consociation treatment can increase ovary storage of big rat obviously, and obstructed the destruction after using the chemotherapy medicine.As a result, the function of ovary was storaged effectively.Experiment2The chemotherapy promotes the cell apoptosis. In recent years, hot study in the chemotherapy with the result that the signal path of the ovary cell apoptosis.Result:1. The TdT lies to lead of the dUTP the indentation bitter end marking (TUNEL) technique examination is used to detect the grain cell apoptosis. The calculation of the index number of Apoptotic index (AI): Count the TUNEL positive percentage of apoptosis cell in 1000 cells. The result expresses the AI of cetrorelix group and consociation treatment groups are far and far small to the chemotherapy groups.2. SP immunohistochemistry is used to analyze the express of BCL-2 protein and caspase-3 protein in ovaries. The levels of the Bcl-2 in the two cetrorelix groups and control group are significantly higher than that in the CTX group, but the levels of the caspases -3 were on the contrary. Regulating the expression of the Bcl- 2 and caspases -3 in the ovary.may be the one of the member mechanisms that protects the ovary in the chemotherapy.3. SP immunohistochemistry was used to analyze the express of ER protein and PR protein in ovaries. The GnRHant do not change the expression of ER/PR in the rat ovary organization under this medicine level and periodicity.On the all, the GnRHant can light the destruction of chemotherapy to the big rat ovary, thus keep the ovary storage function. The protected mechanism possibility activates for depressing the apoptosis factor of caspase-3, and up-expressing the anti- apoptosis factor Bcl-2. |
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