双酚A和β-六氯环己烷对小鼠雌激素活性的实验研究

目的　利用无内源性雌激素模型小鼠 ,探讨双酚A(BPA)和 β-六氯环己烷 (β -HCH)单独与联合投予时的雌激素活性。方法　手术切除小鼠双侧卵巢 17天后 ,给予不同剂量雌二醇 (17β -E2 )并建立其与子宫脏器系数和POD(过氧化物酶 )活性的剂量效应关系。单独或联合腹腔注射给予BPA和 β -HCH ,测定雌激素活性。 结果　 17β -E2剂量与子宫脏器系数和POD活性之间有剂量效应关系。BPA高剂量组子宫脏器系数和POD活性均升高 (P <0 0 5、P <0 0 1)。β -HCH高剂量组使子宫脏器系数升高 (P <0 0 1) ,而POD活性升高无显著性差异 (P >0 0 5 )。在联合投予实验中 ,子宫脏器系数均未见协同作用。而BPA和 β -HCH剂量为 10 0和 5 0 0 μmol/kg体重时对POD活性呈现出协同作用。结论　BPA和 β -HCH单独作用时 ,在一定剂量范围内有雌激素活性。两者联合作用时 ,在一定浓度下表现出协同作用。     

二氧化钛纳米颗粒和雌二醇对小鼠子宫脏器系数和过氧化物酶活性的影响

目的　探讨二氧化钛 (TiO2 )纳米颗粒对 1 7β 雌二醇 (1 7β E2 )生物活性的影响。方法　去卵巢小鼠连续 3天 ,每日 1次同时腹腔注射纳米TiO2 [1 0 ,50 ,2 50mg/ (kg·d) ]和 1 7β E2 [0 0 2 μmol/ (kg·d) ]后 ,测定子宫脏器系数和过氧化物酶 (POD)活性。结果　同时给予纳米材料TiO2 和 1 7β E2 使小鼠子宫脏器系数和POD活性均升高。其中同时给予 1 0mg/ (kg·d)纳米TiO2 组增加比较明显 ,而 2 50mg/ (kg·d)纳米TiO2 组增加最少。结论　纳米TiO2 具有增强 1 7β E2 生物活性的作用。     

黑升麻的雌激素活性及其对人类乳腺癌MCF-7细胞雌激素受体水平的影响

为了检测黑升麻 (CR)的雌激素活性及其对人类乳腺癌 MCF- 7细胞雌激素受体 (ERs)水平的影响 ,未成年雌性小鼠按体重以 75、15 0和 30 0 m g/ kg的 CR灌胃 14d,观察处于动情期的时间 ,第 15天处死后取子宫和卵巢称重。以 MTT实验筛选 CR的最佳细胞作用剂量 ,观察对照组、CR组 (4.75μg/ L )和 17β-雌二醇组 (0 .3nmol/ L)的细胞生长曲线 ,并在流式细胞仪上用间接免疫荧光法测定各组 ERs水平。结果发现子宫重量随 CR剂量增加而增加 ,30 0 mg/ kg的 CR可显著增加动情天数 (P<0 .0 5 )。 4.75 μg/ L CR对 MCF- 7细胞的促增殖作用最强 ,达 6 4.7% ;CR组和 17β-雌二醇组的细胞群体倍增时间 (TD)分别为 32 .1h和31.7h,明显低于对照组 (TD=35 .3h) ;4.75 μg/ L 的 CR可显著增加细胞的 ERs水平 (P<0 .0 1)。结果表明CR具有雌激素活性 ,其升高 ERs水平的作用可能是该物质可治疗更年期综合征的机制之一。     

典型天然雌激素MCF-7细胞增殖实验的条件优化研究

目的 研究不同试验条件对MCF-7增殖实验检测雌激素效应的影响.方法 选用两种来源(ATCC和某肿瘤医院)的MCF-7细胞,研究不同浓度、不同培养时间、不同溶剂、不同培养液条件下MCF-7细胞对17B-雌二醇(E2)、乙炔雌二醇(EE2)、雌三醇(E3)、雌酮(E1)的反应,以优化MCF-7增殖实验条件.结果 不同E2...     

白藜芦醇的雌激素样作用研究

为研究白藜芦醇在体内的雌激素样作用 ,给刚断乳小鼠分别皮下注射和灌胃不同剂量的白藜芦醇 ,观察该物质对子宫和阴道的影响。结果显示 ,2 0mg kg .BW的白藜芦醇可明显促进未成熟小鼠阴道开口 (P <0 0 5 )和阴道上皮角化 ,显著增加子宫重量及其系数 (P <0 0 5 ,P <0 0 1) ,并可使子宫内膜柱状上皮增厚或腺体增生。结果表明白藜芦醇在体内具有雌激素样作用。而口服白藜芦醇的雌激素活性明显弱于皮下注射     

白藜芦醇对卵巢切除大鼠骨丢失的抑制作用

利用卵巢切除大鼠 ,研究白藜芦醇对雌激素缺乏诱导的骨丢失是否具有抑制作用。大鼠卵巢切除后 ,每日灌胃给予白藜芦醇 0 7mg kgBW ,持续 1 2周 ,同时设置假手术组和卵巢切除对照组。结果发现 ,白藜芦醇能显著增加卵巢切除大鼠股骨重量、钙含量和股骨干骺端的骨密度 (P <0 0 5 ,P <0 0 1 ) ,表明白藜芦醇能有效抑制雌激素缺乏所诱导的骨丢失     

五种多环芳烃对大鼠子宫雌激素受体影响的实验研究

采用大鼠子宫湿重实验和体外雌激素受体结合实验评价了 5种多环芳烃的拟雌激素活性。结果表明苯并 (a)芘、7,12 -二甲基苯并蒽和 2 ,3,6 ,7-二苯并蒽的中、高剂量可导致初断乳大鼠子宫湿重明显增加 (P <0 0 1或P <0 0 5 )。苯并 (a)蒽和 1,2 -苯并菲各剂量均不引起大鼠子宫湿重明显增加 (P >0 0 5 )。苯并 (a)芘、7,12 -二甲基苯并蒽和 2 ,3,6 ,7-二苯并蒽能明显抑制雌二醇与雌激素受体的结合 (P <0 0 1)。苯并 (a)蒽和 1,2 -苯并菲各剂量均不能明显抑制雌二醇与雌激素受体的结合 (P >0 0 5 )。通过体内外实验表明苯并 (a)芘、7,12 -二甲基苯并蒽和 2 ,3,6 ,7-二苯并蒽具有明显的拟雌激素活性 ,而苯并 (a)蒽和 1,2 -苯并菲没有检测到拟雌激素活性     

β-胡萝卜素对香烟烟气诱导脂质过氧化的交互作用

目的　研究 β 胡萝卜素对香烟烟气诱导脂质过氧化的交互作用。 方法　建立了大鼠肺Ⅱ型细胞体外培养模型 ,加入 β 胡萝卜素或 和香烟烟气颗粒提取物染毒 2 4h后 ,检测细胞的丙二醛 (MDA)含量。并构建短期动物模型 ,使小鼠暴露于香烟烟气和 或补充 β 胡萝卜素 10天 ,检测血清MDA的含量。结果　在大鼠肺Ⅱ型细胞模型中 ,随香烟烟气颗粒提取物的含量增加 ,MDA水平明显上升 ,而 β 胡萝卜素各剂量组间MDA水平的差异无显著性。0 5 μg ml的 β 胡萝卜素可抑制香烟烟气颗粒提取物对肺细胞脂质过氧化的诱导作用 ,表现为拮抗作用。在小鼠动物模型中 ,烟气暴露组的MDA水平显著高于正常对照组 ,而 β 胡萝卜素各剂量组间MDA水平的差异无显著性。给予 β 胡萝卜素 2 5mg kgBW的烟气暴露组小鼠血清中MDA水平下降。结论　β 胡萝卜素对香烟烟气诱导的脂质过氧化具有拮抗作用 ,其有效作用剂量在大鼠肺Ⅱ型细胞实验中为 0 5 μg ml ,在小鼠实验中为 2 5mg kgBW。     

天然植物提取物抗CCl_4所致大鼠慢性肝损伤的病理研究

目的　研究天然植物提取物对CCl4 所致大鼠慢性肝损伤的病理影响。方法　采用 4 0 %CCl4 2次 周腹腔注射 (2ml kgBW)建立慢性肝损伤模型 ,干预组在造模同时给予天然植物提取物 (0 2 7g kgBW)灌胃 ,7周后取肝组织 ,常规病理切片 ,HE、Masson、James染色后 ,光镜下观察病理改变 ,电镜下观察超微结构变化。结果　天然植物提取物能有效拮抗CCl4 诱导的大鼠肝组织内网状纤维、胶原纤维的形成 ,有效减轻CCl4 所致的肝组织病理改变。结论　天然植物提取物对CCl4 所致大鼠慢性肝损伤有一定防治作用     

植物甾醇酯对实验大鼠血脂水平的影响

目的探讨植物固醇酯(PSE)对大鼠血脂水平的影响。方法(1)在高脂饲料喂养模型下,4组大鼠分别给予不含(模型组)或含有3个剂量PSE(分别为4·0、16·0、32·0mg/kgbw)的油溶液,30天后测定血脂水平。(2)摘除卵巢的去势大鼠分为模型组,E2组(22·5μg/kgbw己烯雌酚),PSE组(32·0mg/kgbwPSE)和FPE组(500mg/kgbw游离型植物固醇),60天后测定血脂、雌激素,及大鼠子宫、肝脏重量。各实验均设正常对照组以确认造模成功。结果PSE使高脂饲料喂养大鼠甘油三酯(TG)水平下降,高剂量PSE组大鼠总胆固醇(TC)、低密度脂蛋白胆固醇(LDL-C)明显降低,高密度脂蛋白胆固醇(HDL-C)及HDL-C/TC升高(P<0·05)。去势模型使大鼠体重、肝脏重量、TC、TG和LDL-C明显升高,雌激素水平和子宫重量显著下降(和对照组比,P<0·05)。PSE和FPS可显著降低去势大鼠体重和肝重,使TC和TG水平下降,子宫重量略有增加(P<0·05),但作用未及E2(P<0·05),对雌激素水平也没有显著影响。结论植物固醇酯具有明显降血脂作用。     

Estrogenic Effects of Cimicifuga racemosa (Black Cohosh) in Mice and on Estrogen Receptors in MCF-7 Cells

The estrogenic effects of Cimicifuga racemosa or Actacea racemosa (black cohosh, CR) extracts were tested in mice, and their effects on estrogen receptor (ER) levels in human breast cancer MCF-7 cells were also investigated. Four groups of weanling female Kunming mice were given 0 (control), 75, 150, or 300 mg/kg body weight CR extracts orally for 14 days. The estrus cycle and the weights of the uterus and ovary of mice, as well as serum estradiol (E(2)) were measured. The proliferation patterns of MCF-7 cells exposed to CR extracts or 17beta-estradiol were studied by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Subsequently, growth of MCF-7 cells in 0 (control) or 4.75 &mgr;g/L of CR extracts or 0.3 nmol/L of 17beta-estradiol groups were observed for 5 days. ER levels in MCF-7 cells were analyzed by indirect immunofluorescence assay using flow cytometry. The uterine weights of mice increased with the increase in dosage of CR extracts, and the estrus duration was significantly prolonged in the group receiving 300 mg/kg body weight (P <.05). However, CR extracts did not increase the serum E(2) concentration significantly. In the in vitro study, a dose-response relationship was demonstrated when cells were treated with low doses of CR extracts, and the optimal enhancement concentration of CR extracts was 4.75 &mgr;g/L on MCF-7 cells. The doubling times (T(D)) of cell growth in the CR extracts group and the 17beta-estradiol group were 32.1 and 31.7 hours, respectively, both shorter than that of the negative control group (T(D) = 35.3 hours). Additionally, 4.75 &mgr;g/L of CR extracts resulted in significantly increased ER levels compared with the control group (P <.01). In conclusion, CR extracts produced an estrogenic action. The effect of increasing ER levels by CR extracts may be one of the potential mechanisms of its phytotherapeutic effects for postmenopausal symptoms.     

The intact immature rodent uterotrophic bioassay: possible effects on assay sensitivity of vomeronasal signals from male rodents and strain differences

The vomeronasal organ in rodents is an important social and sexual signaling pathway. We have investigated whether the housing of intact immature females in close proximity to mature males would interfere with the sensitivity of the immature rodent uterotrophic bioassay as the result of vomeronasal signals transmitted by male urinary proteins. The hypothesis was that the proximity of males might induce early puberty, thereby increasing mean uterine weight and reducing the responsiveness of the assay. The hypothesis was tested in both rats and mice by housing mature males above immature females, separated only by a wire screen, for 3 days and determining possible changes in uterine weight. The results were negative. Neither the mean uterine weight nor the group mean standard deviation of the uterine weights were changed in the uterotrophic bioassay. Given that the timing of sexual maturation may vary with the strain of mouse used, we also evaluated the sensitivity of the immature mouse uterotrophic assay to diethylstilbestrol (DES) using four strains of mice. Similar sensitivity was observed for the CD-1, C57Bl6, and Alpk strains, but B6CBF(1) mice were marginally less sensitive to DES than were the other strains. These findings add to earlier data indicating the robustness of the rodent uterotrophic assay protocol.     

农药联苯菊酯对大鼠内分泌干扰作用的研究

目的通过大鼠子宫增重试验和Hershberger试验研究农药联苯菊酯(BIF)的内分泌干扰作用。方法子宫增重试验中,60只雌性SD大鼠随机分为6组,每组10只:BIF低、中、高剂量组分别经口灌胃给予1.47、4.41和13.23mg/kgBW的BIF;溶剂对照组经口灌胃给予玉米油;17α-乙炔雌二醇(EE)经口阳性组经口灌胃给予1.0μg/kg BW的EE,EE经皮阳性组经口灌胃给予玉米油并皮下注射0.6μg/kg BW的EE。连续给药3天后,测定大鼠子宫湿重、干重,计算其子宫系数,并观察病理变化。在Hershberger试验中,60只去势雄性SD大鼠随机分为6组,每组10只:BIF低、中、高剂量组分别经口灌胃给予1.4、4.2和12.6mg/kg BW的BIF,阳性对照组经口灌胃给予3.0mg/kg BW的氟他胺,溶剂对照组和丙酸睾酮对照组(TP组)经口灌胃给予玉米油,除溶剂对照组外,其余各组均皮下注射给予0.2mg/kgBW丙酸睾酮。所有剂量组连续给药10天后,称量腹侧前列腺(VP)、精囊腺和凝固腺(SVCG)、肛提肌加球海绵体肌(LABC)、阴茎头(GP)、尿道球腺(COW)、肝、肾、肾上腺的重量,计算其脏器系数,测定血清中三碘甲腺原氨酸(T3)和甲状腺素(T4)水平。结果子宫增重试验中,与溶剂对照组相比,BIF高剂量组子宫湿重系数和干重系数显著增加(P<0.05)。组织病理学检查发现,BIF中、高剂量组子宫内膜增生、增厚,子宫内膜上皮细胞高度增高(P<0.05)。Hershberger试验,与TP组相比,BIF高剂量组VP、LABC重量显著减轻,T3、T4水平显著降低(P<0.05)。结论子宫增重试验和Hershberger试验提示BIF具有潜在的内分泌干扰作用。     

Roxatidine,an H2 Receptor Blocker,is an Estrogenic Compound—Experimental Evidence

Roxatidine is an H₂ receptor blocker frequently used in the treatment of peptic ulcers. H₂ receptor blockers are reported to show antifertility activity. To examine the mechanism of antifertility, estrogenic and antiestrogenic activity was studied using an in vitro rat and rabbit uterine receptor binding assay and in vivo using the uterotrophic assay in immature Wistar rats. The results revealed that roxatidine showed mild receptor binding affinity to both rat and rabbit uterine receptors when compared to estradiol. Interstingly, in vivo roxatidine increases the wet uterine weight of immature Wistar rats significantly (P<0.001) when compared to a control group. The increase in uterine weight within the roxatidine treated group was somewhat similar to that of the estradiol treated group. Histopathological results and the structure of the roxatidine support that H₂ receptor blocker roxatidine is an estrogenic compound.     

The OECD program to validate the rat uterotrophic bioassay. Phase 2: dietary phytoestrogen analyses

Many commercial laboratory diets have detectable levels of isoflavones (e.g., phytoestrogens such as genistein [GN]) that have weak estrogenic activity both in vitro and in vivo. During validation studies of the uterotrophic bioassay, diet samples from 20 participating laboratories were collected and analyzed for three major phytoestrogens: GN, daidzein (DN), and coumestrol (CM). Soy phytoestrogens GN and DN were found at total phytoestrogen levels from 100 to 540 microg/g laboratory diet; a forage phytoestrogen, CM, ranged from nondetectable to 4 microg/g laboratory diet. The phytoestrogen levels were compared with both baseline uterine weights of the control groups and with the relative uterine weight increase of groups administered two weak estrogen agonists: bisphenol A (BPA) and nonylphenol (NP). The comparison uses a working assumption of additivity among the phytoestrogens, despite several significant qualifications to this assumption, to estimate total genistein equivalents (TGE). Some evidence was found that phytoestrogen levels in the diet > 325-350 microg/g TGE could diminish the responsiveness of the uterotrophic bioassay to weak agonists. This was especially true for the case of the intact, immature female version of the uterotrophic bioassay, where higher food consumption relative to body weight leads to higher intakes of dietary phytoestrogens versus ovariectomized adults. This dietary level is sufficient in the immature female to approach a biological lowest observable effect level for GN of 40-50 mg/kg/day. These same data, however, show that low to moderate levels of dietary phytoestrogens do not substantially affect the responsiveness of the assay with weak estrogen receptor agonists such as NP and BPA. Therefore, laboratories conducting the uterotrophic bioassay for either research or regulatory purposes may routinely use diets containing levels of phytoestrogens < 325-350 microg/g TGE without impairing the responsiveness of the bioassay.     

Confirmation of uterotrophic activity of 3-(4-methylbenzylidine)camphor in the immature rat

In this study we found that the ultraviolet sunscreen component 3-(4-methylbenzylidine)camphor (4MBC) is uterotrophic in immature rats when administered by either subcutaneous injection or oral gavage. These data confirm earlier reports of uterotrophic activity for this agent when administered to immature rats in the diet or by whole-body immersion; however, they are in contrast to negative unpublished immature rat uterotrophic assay results. Data also indicate that 4MBC binds to isolated rat uterine estrogen receptors and shows activity in a human estrogen receptor yeast transactivation assay; however, we considered both of these effects equivocal. In this study, we confirmed the original observation that 4MBC was active as a mitogen to MCF-7 breast cancer cells. We evaluated and discounted the possibility that the estrogenic activity of 4MBC is related to its bulky camphor group, which is of similar molecular dimensions to that of the weak estrogen kepone. Uncertainty remains regarding the mechanism of the uterotrophic activity of 4MBC.     

Immature rat uterotrophic assay of bisphenol A

We used the immature rat uterotrophic assay to determine the estrogenicity of bisphenol A (BPA). We administered BPA (in sesame oil) to rats subcutaneously (sc; 0, 8, 40, and 160 mg/kg/day) or orally (0, 40, 160, and 800 mg/kg/day) for 3 days beginning on postnatal day (PND) 18; rats were sacrificed 24 hr after the last administration. Uterine wet, blotted, and relative weights increased in all groups given BPA sc. After oral administration, uterine relative weight increased in 160 and 800 mg/kg BPA groups, and wet and blotted weights increased in the 800 mg/kg BPA group. Plasma concentrations of BPA at 1 hr after the last administration were detected in all groups given BPA sc and in groups given 160 and 800 mg/kg BPA orally, with a dose-response effect. The study was then reproduced under the same conditions. After sc injections, uterine wet and blotted weights increased in the 40 and 160 mg/kg BPA groups, and relative weight increased in all groups given BPA sc. By contrast, uterine wet, blotted, and relative weights increased only in the 160 and 800 mg/kg oral BPA groups. Also, to examine time-course changes in uterine weight, we administered BPA (in sesame oil) sc from PND 18 to PND 20 for 3 days at doses of 0, 8, 40, and 160 mg/kg/day; uterine weights were then measured at 6, 12, 18, and 24 hr after the last administration. Uterine wet, blotted, and relative weights increased in all BPA groups at 6 and 24 hr and in 40 and 160 mg/kg BPA groups at 12 hr. By contrast, at 18 hr, uterine wet, blotted, and relative blotted weights increased in all BPA groups and relative wet weight increased in 40 and 160 mg/kg BPA groups. The percentage increases in uterine wet and relative weights of 40 and 160 mg/kg BPA groups at 6 hr were higher than those at 24 hr relative to the controls, but the coefficient of variation in these weights in the group given 8 mg/kg BPA at 24 hr was smaller than that at 6 hr. These findings demonstrate BPA-induced uterotrophy in the immature uterotrophic assay in rats administered 8 mg/kg/day sc and in rats given 160 mg/kg/day orally, and suggest that the autopsy at 24 hr after the last administration is suitable.     

某自来水厂水源水有机提取物的雌激素样活性研究

[目的]了解某自来水厂水源水有机提取物的雌激素样活性。[方法]用固相萃取法富集水样,分低、中、高剂量染毒组（体外试验为每毫升染毒量分别相当于10、100、1000mL水源水中有机提取物的剂量;体内试验为每千克体重灌胃量相当于2、5、10、25L水源水中有机提取物的剂量）、阴性对照组和雌二醇阳性对照组,通过体外人乳腺癌（MCF-7）细胞增殖试验、报告基因试验和初断乳大鼠子宫增重试验对水源水有机提取物的雌激素样活性进行研究。[结果]体外细胞增殖试验中,高剂量组较阴性对照组出现细胞增殖效应（P〈0．05）,细胞增殖率为（130．85±13．68）％;而报告基因试验中,中剂量组检测到荧光强度增加（P〈0．05）,相对荧光强度为（207．67±12．40）％。子宫增重试验中,相当于低、中、高剂量染毒组大鼠子宫湿重和黏液重脏器系数与阴性对照组相比差异无统计学意义。[结论]体外试验表明水源水中含有雌激素样活性物质,但体内试验尚不能确定该水源水具有明确的雌激素样活性。     

Induction of hyperplasia and increased DNA content in the uterus of immature rats exposed to coumestrol.

Administration of the phytoestrogen coumestrol to ovariectomized rats leads to increases in both wet and dry uterine weights in the absence of an increase in uterine DNA content, as reported by Markaverich et al. [Effects of Coumestrol on Estrogen Receptor Function and Uterine Growth in Ovariectomized Rats. Environ Health Perspect 103:574-581 (1995)]. It was not possible to know if the observed atypical uterotrophic response of coumestrol was associated uniquely with the ovariectomized uterotrophic assay protocol. This question is answered in the present paper. Two experiments are described in which three daily oral gavage administrations of 60 mg/kg/day coumestrol to immature AP rats were followed by assessment of the reproductive tract on the fourth day. In both experiments coumestrol increased uterine fluid content and increased the weights of the uterus, cervix, and vagina. In addition, bromodeoxyuridine staining of uterine sections enabled confirmation of uterine hyperplasia for the coumestrol-treated animals. In the second experiment, total uterine DNA was determined; it doubled in the coumestrol-treated animals. Estradiol benzoate acted as the positive control agent for both of these experiments, and in each case it gave similar responses to those seen for coumestrol. We conclude that the uterotrophic activity of the phytoestrogen coumestrol in the immature intact rat is typical of the activity of the natural estrogen estradiol.     

Uterotrophic effects of benzophenone derivatives and a p-hydroxybenzoate used in ultraviolet screens

Ultraviolet (UV) sunscreen products are popular because of concerns about UV radiation and skin cancer. Unfortunately, some of these products contain agents with estrogenic activity. We used an ovariectomized rat uterotrophic assay to measure the estrogenic activities of 2,4-dihydroxybenzophenone (2,4-DHBP), 2,2',4,4'-tetrahydroxybenzophenone (2,2',4,4'-THBP), and 4-hydroxybenzoic acid isobutyl ester (isobutyl-paraben), which are agents in UV sunscreens, and ethynyl estradiol (EE) and bisphenol A (BPA), which are positive controls. All chemicals increased rat uterine weights. The 10% effective doses (ED10, mg/kg/day) of EE, BPA, 2,4-DHBP, 2,2',4,4'-THBP, and isobutyl-paraben, as determined by Hill equation analysis, where 5E-5, 41.1, 544.6, 33.0, and 230.9, respectively, and their relative potencies against EE were about 1/800,000, 1/10,000,000, 1/600,000, and 1/4,000,000, respectively. Our findings indicated that UV screens contain weak estrogenic compounds.