| ObjectivesIn 1990s, fatty acid synthase (FAS) was firstly founded in the breast cancer as cancer antigen. It correlated with cell proliferation, low-grade and poor prognosis. Since then, scientists found that FAS was highly expressed in ovarian cancer, endometrial cancer, prostate cancer and other hormone-sensitive tumors. Numerous studies have proved that sex hormones, such as estrogen, progesterone, and androgen can up-regulate the expression of FAS in sex hormone-sensitive tumors. FAS are a major energy source for tumor cells, and it provides membrane phospholipids for the rapid growth of cells. Therefore, FAS may involve in the promotion of growth, even canceration of cells. However, few studies related with the regulation mechanism of FAS in normal uterus and breast existed. The biological behavior of normal and tumor cells is substantially different. More research is needed to validate wether the theoretical basis from tumor cells applicable to normal cells. Uterus is not only the most sensitive tissue to estrogen but also periodical proliferation. Long-term continuous estrogen can promote the occurrence of endometrial hyperplasia or cancer. Therefore, the protection of endometrial is very important during hormone therapy. Meanwhile, breast often be taken into account.The study mainly focused on uterus and breast. The purpose of the study as follows: First, to observe the expression of FAS in uterus and breast through long-term exogenous estrogen and to explore the regulation mechanism of estrogen on FAS. Secondly, by detecting FAS spectrum in uterus and breast during the different growth stages of rats, investigate the influence of FAS on the growth and aging. Thirdly, to observe the expression of FAS in uterus and breast of rats during the different phase of estrous cycle, and the expression of FAS among rats with irregular estrous cycle, and the expression of FAS among rats given estrogen receptor antagonist, and to explore the effect of fluctuation in endogenous estrogen on FAS. Finally, testify the expression of FAS in women's endometrial tissue in different stage of menstrual cycle and abnormal proliferation status and try to make clinical recommendations.Methods1. Estrogen stimulating group:60 SD female rats aged 3 months randomly divided into three groups:sham operation group (Sham,6), ovariectomized group (OVX,8) and given estradiol valerate after ovariectomy one time per day for consecutive two months. The dose of estradiol valerate as follows:0.05mg/kg (7), 0.1mg/kg (10), 0.2mg/kg (10),0.4mg/kg (10) and 0.8mg/kg (10). Given 0.2mg/kg estradiol valerate for rats is equivalent to 2mg for women 60kg weight. Meanwhile, Sham group and ovariectomized group received the same volume of normal saline.2. Development and growth study groups:selected female SD rats 4 weeks,6 weeks,8 weeks,3 months,6 months,9 months and 12 months,16 months after their birth respectively. Every group has 7 rats. Rats with regular estrous cycles, samples were collected in diestrus. Otherwise, the specimens were collected at any time.3. Estrous cycle in middle-aged rats study groups:To selected middle-aged female SD rats (13 months) with regular estrous cycle verified by vaginal smear, and randomly divided them into proestrus, estrus, metestrus and diestrus groups, each have 7 rats. The rats were classified as "irregular cycle-a" if they had continuous performance of estrus or classified as "irregular cycle-b" if they had continuous performance of diestrus.4. Estrogen antagonist study group:selected 12 female SD rats (10-month) underwent bilateral ovariectomy, then, divide them into 2 groups randomly. After 2 months, one group was orally fed with estradiol valerate (1.0mg/kg) 1 time per day for 5 days. Another group was both orally fed with estradiol valerate (1.0mg/kg) and subcutaneous injected with ICI182,780 (500ug/d) 1 time per day for 5 days.5. To selected women aged 35-60 conducted diagnostic curettage for irregular uterine bleeding, and to divided them into four groups:women with proliferative endometrium, women with secretory endometrium, women with endometrial hyperplasia and women with atypical endometrial hyperplasia. Every group has 5 cases.Measurements included:(1) For rats:body fat mass, uterine wet weight, serum estradiol (E2), total cholesterol (TC), triglyceride (TG), high density lipoprotein cholesterol (HDL-C), free fatty acid (FFA), non-high density lipoprotein cholesterol (nonHDL-C), protein levels of FAS and Ki67 in uterus and breast, and gene levels of FAS, ER-a, and SREBP-lc in uterus and breast. (2) For women:protein levels of FAS, Ki67 and ER expression in endometrium of clinical specimens.Results1. Rats with different dose of exogenous estrogen(1) Body fat mass:total mass and percentage of fat mass is highest in OVX group, lowest in rats fed in 0.8mg/kg estradiol valerate.(2) Uterine wet weight/body weight ratio:uterine wet weight/body weight ratio is highest in sham operation group, followed by rats fed in 0.8mg/kg estradiol valerate. Uterine wet weight/body weight ratio was lowest in OVX group and increased dose-dependently.(3) Serum E2 and lipid profile:E2 levels increase dose-dependently. Serum TC, nonHDL-C and FFA are lowest in the rats fed in 0.8mg/kg estradiol valerate, and are highest in OVX group, and decrease with the increasing dose of estradiol valerate.(4) Expression of FAS in uterus and breast tissue:The expression of FAS in tissue of uterus and breast exists in rats according to the result of immunohistochemistry. The positive intensity of FAS is related with the positive cell ratio of nuclear antigen Ki67. The genetic testing results for tissue of uterus and breast suggested that the expression of FAS, ER-a and SREBP-lc are highest in rats fed in 0.8mg/kg estradiol valerate, and increase dose-dependently.2. Study groups during different development and growth phases(1) Body fat mass:total mass and percentage of total fat mass increased with ageing.(2) Uterine wet weight/body weight ratio:uterine wet weight/body weight ratio was lowest in rats after birth 4-weeks and significantly higher in rats after birth 42 days. There are no significant changes for rats after birth 8 weeks until 9 months. Then uterine wet weight/body weight ratio decrease after birth 9 months.(3) Serum E2 and lipid profile:serum E2 levels were lower in sexual immature status, changes in TC and nonHDL-C share the similar trends. There are three high points, respectively,4 weeks,9 months and 16 months after birth, among them 9 months occupy the highest point. Levels of TG and FFA increased with ageing.(4) Expression of FAS in uterus and breast:â‘ FAS expression in tissue of uterus increase linearly during 4-8 weeks after birth, and there are no significant changes between 8 weeks-16 months after birth. ER-a in tissue of uterus is significantly higher in group after birth of 3,6,9 months than younger and old group. There is no change of SREBP-lc during different developmental stage.â‘¡mRNA level of FAS in breast increase through ageing, and highest in 9 month after birth, then gradually decreased; ER-a is highest in 3 and 6 month after birth; SREBP-lc is highest in 9 month after birth.3. Middle-aged rat estrous cycle and estrogen antagonist study group(1) Body fat mass:there are no significant differences in total mass and percentage body fat mass in each group of rats.(2) Uterine wet weight/body weight ratio:uterine wet weight/body weight ratio is highest in "irregular cycle-a" group, and is lower in rats treated with 1.0mg/kg estradiol valerate+ICI182780 with comparison of rats treated with only 1.0mg/kg estradiol valerate.(3) Serum E2 and lipid profile:E2 levels were higher in proestrus and estrus than metestrus and diestrus. There is no significant difference in different stages of estrous cycle for lipid profile. TC, nonHDL-C is lower in rats fed in 1.0mg/kg estradiol valerate and treated with estradiol valerate+ICI 182,780 groups than others.(4) Expression of FAS in uterus and breast:there is no significant difference in expression of FAS in tissue of uterus and breast during different estrous cycle. With comparison of rat treated with 1.0mg/kg estradiol valerate and ICI182780, The expression of FAS and ER-a in uterine tissue is significantly higher in rats fed in l.Omg/kg estradiol valerate. But there is no significant difference for the gene expression in breast tissue.4. Endometrium of women from clinical sample The expressions of FAS and Ki67 in proliferative endometrium are higher than secretory endometrium. The higher expression of FAS is accompanied with the higher expression of Ki67 and ER. The expression of FAS and Ki67 in simple hyperplasia endometrium and atypical hyperplasia endometrium seem higher than proliferative endometrium.Conclusions1. Long-term exogenous estrogen can up-regulate the level of FAS expression in both rats'uterus and breast on the transcriptional level by ER-a and SREBP-lc. The expression level of FAS was stable when estrogen is lower than the treatment dose. FAS expression level was higher than the physiological state when estrogen is significantly higher than the treatment dose. The results suggest that there may be a suitable dose range for estrogen upregulating FAS.2. The level of FAS expression in uterus and breast of rats increases gradually from sexual immature status to sexual mature status. FAS may relate with the growth and development of uterus and breast.3. Physiological conditions, the cyclical fluctuations of endogenous E2 middle-aged rats had no effect on FAS expression. The effect of short-term estrogen can promote the proliferation of uterus tissue and up-regulate the level of FAS expression, which can be intercepted by estrogen receptor antagonist. But this effect does not work on breast, which implies that more time will be needed when it is regulated.4. In women, the expression of FAS is significantly higher in proliferative stage than secretory stage. The expressions of FAS seem higher in simple hyperplasy status and atypical hyperplas status than proliferative stage. |
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