重组雌激素受体基因酵母细胞检测牛尿液中β-雌二醇残留的方法

为探索畜禽产品中雌激素残留的检验方法,加强对畜禽生产过程中激素添加的监控,本研究选择了在我国几种常见影响比较大的合成雌激素-β-雌二醇(E2)为研究对象,建立用于测定动物尿液中雌激素的重组基因酵母检测法。结果表明本方法简单、有效,能够作为雌激素残留检测体系的有益补充。     

鸡粪中雌激素的检测及微生物降解

为检测鸡饲料和鸡粪中雌激素,优化其微生物降解条件,应用高效液相色谱（HPLC）法检测蛋鸡饲料和鸡粪中的雌激素,利用11种高效降解雌激素的单一菌株和等体积混合得到的菌群开展雌激素的降解实验。结果表明：只有蛋鸡鸡粪中含有雌二醇（E2）,含量为230 ng·g^-1。微生物菌群的降解率高于单一菌株,在27℃下,10 d内能够降解76%的E2。研究表明,蛋鸡鸡粪中有雌激素存在,微生物菌群对雌激素的降解率高于单一菌株。     

固相萃取-高效液相色谱法同时测定饲料中的3种雌激素

【目的】建立饲料中3种雌激素(17β-雌二醇、苯甲酸雌二醇和戊酸雌二醇)的固相萃取-高效液相色谱(SPEHPLC)测定方法。【方法】饲料样品经甲醇+水提取,固相萃取柱净化,采用Dionex C18色谱柱,以乙腈和水为流动相进行梯度洗脱,流速1.0mL/min,检测波长230nm,柱温30℃。【结果】3种雌激素的线性关系较好;检出限为0.5~1.0mg/kg;平均回收率为89.3%~108.0%。【结论】SPE-HPLC法操作简便、线性范围宽、检出限低、准确度高,可同时测定饲料中3种雌激素的含量。     

雌二醇及其受体在北方山溪鲵精巢中的周期性分布

用免疫组织化学方法检测雌二醇(estradiol)及其受体(estrogen receptor)在北方山溪鲵(Batrachuperus tibetanus)精巢发育周期中的表达变化。结果表明,在4~5月,雌二醇及受体在精原细胞的胞质中强表达;6月,雌二醇在精母细胞的胞质中表达较弱,其受体在精母细胞的核质中有较强表达;7月,雌二醇及受体在精子细胞的胞质中表达;8月,雌二醇及受体在精原细胞的胞质中和变态后的精子头部有弱表达;9-11月,两者均在精原细胞的胞质中及精子头部有较强表达。说明雌二醇及其受体存在于整个精巢周期中,二者在生精细胞中的强弱变化基本一致,其变化规律与精子发育周期密切相关;雌激素受体多存在于生精细胞的胞质中,雌激素可能以非基因组效应调控细胞的增殖,并与精子的发育成熟密切相关。     

甘肃省市售牛羊肉中雌激素残留状况分析

以甘肃省兰州市、靖远县、临夏州、张掖地区牛羊肉为试验材料,分析了甘肃省市售牛羊肉中己烯雌酚、雌二醇、戊酸雌二醇和苯甲酸雌二醇等4种雌激素残留状况。结果表明,80份样品中(4个地方牛羊肉各10份)所测定的4种雌激素均未检出,甘肃省市售牛羊肉中无论是天然的雌二醇还是己烯雌酚、戊酸雌二醇和苯甲酸雌二醇3种人工合成雌激素均没有残留。     

益母草总黄酮对去卵巢大鼠下丘脑和海马雌激素受体α及β mRNA表达的影响

目的：观察益母草总黄酮对去卵巢大鼠下丘脑和海马雌激素受体α及β mRNA表达的影响,探讨其对绝经后骨质疏松的作用机制。方法选取10~11月龄SD雌性大鼠48只,随机分为对照组、去势组、益母草总黄酮组和雌二醇组,以双侧卵巢切除法建立大鼠骨质疏松模型。益母草总黄酮组、雌二醇组分别用益母草总黄酮、雌二醇给药4个月,采用RT-PCR法检测各组下丘脑和海马雌激素受体α及β mRNA的表达。结果大鼠卵巢切除后,血清中雌二醇水平、椎骨骨密度、子宫湿重、下丘脑和海马雌激素受体α及βmRNA的表达均明显下降。雌二醇治疗后血清中雌二醇水平、椎骨骨密度、子宫湿重升高,下丘脑和海马β mRNA的表达均明显下降。益母草总黄酮治疗后,血清中雌二醇水平、椎骨骨密度、下丘脑和海马雌激素受体α及β mRNA的表达均明显升高,子宫湿重无明显改变。结论益母草总黄酮对切除卵巢所致的骨质疏松大鼠有防治作用,对子宫无不良影响。     

基于SPE-HPLC的芹菜中环境雌激素检测

[目的]建立芹菜茎叶中6种环境雌激素（雌酮、17β-雌二醇、雌三醇1、7α-乙炔基雌二醇、双酚A和4-壬基酚）的高效液相色谱检测方法。[方法]样品切碎榨汁后,用超声波振荡提取,固相萃取小柱浓缩净化,对硝基苯甲酰氯柱前衍生,高效液相色谱法荧光测定。[结果]6种环境雌激素的线性范围为8.3～1 000.0μg/L,相关系数r=0.998 8～0.999 9,最低检出限量为3.7～8.3μg/L。样品加标平均回收率在89.60%～97.13%,相对标准偏差RSD均小于2.9%。[结论]该研究建立的方法同时测定芹菜茎叶中的6种环境雌激素,结果满意。     

牛乳中三种雌激素残留的HPLC检测法

建立了牛奶中3种雌激素残留量(雌二醇、己烯雌酚、雌酮)的HPLC(高效液相色谱)检测方法.得到色谱条件为流动相:乙腈-水(体积比为1∶1);检测波长:230 nm;流速1.0 mL.min-1.采用该方法测定了市售鲜牛奶中3种雌激素的残留量,结果表明:其分离效果良好,平均回收率范围82.2%~95.5%,相对标准偏差为2.0%~4.7%,最低检出限达1.0~2.0 ng.mL-1.该方法具有灵敏、准确、精密、无杂质干扰等优点.     

猪粪厌氧消化进程中雌激素的去除效能及机制

为探究猪粪厌氧消化进程中雌激素的去除效能及其作用机制，对厌氧消化液中的4种雌激素（雌酮E1、雌二醇E2、雌三醇E3、炔雌醇EE2）进行阶段性检测，并采用傅里叶变换红外光谱、三维荧光光谱结合荧光区域积分和16S rRNA基因扩增子高通量测序的手段对猪粪中红外官能团振动特征峰、溶解性有机物（DOM）和微生物群落结构进行分析。结果表明：在温度为35℃、周期为30 d、固体浓度为6%的条件下，厌氧消化对4种雌激素的降解速率依次为E2>E3>E1>EE2，去除效率分别为28.62%、25.83%、19.14%、11.81%，降解规律均符合有机物一级动力学降解模型。在投加雌激素之后，猪粪中微生物群落结构发生了一定改变，但仍能进行正常的厌氧消化，并促使与雌激素相结合的腐植酸类溶解性有机物含量增加，同时其中还存在酰胺、羟基、羧基等雌激素吸附位点以及雌激素高效降解菌。研究推测沼液中雌激素的去除机制包含猪粪颗粒的吸附、溶解性有机物的吸附和雌激素降解菌属的高效代谢3个方面。     

血清雌二醇水平对更年期小鼠大脑凋亡诱导因子的影响

目的评价外源性雌二醇对更年期小鼠大脑神经元凋亡的保护作用。方法以C57/BL6小鼠为实验对象,通过双侧卵巢切除术制作更年期小鼠模型。实验动物随机分为对照组、假手术组、手术组、手术+雌二醇组,其中手术+雌二醇组自卵巢去势术后第6周开始隔日给予皮下注射雌激素0.5 mg·kg-1持续2周。ELISA法测定血清雌二醇水平;免疫组化法、Western blot法检测大脑组织中凋亡诱导因子的表达。结果手术组血清雌二醇水平低于假手术组,手术组凋亡诱导因子(apotosis inducing factor,AIF)表达量明显高于假手术组,手术+雌二醇组AIF表达量明显低于手术组。结论体内雌激素的下降能够促进小鼠大脑神经元的凋亡,补充雌二醇能够减缓更年期小鼠大脑神经元的凋亡。     

Lipocalin核糖体展示文库的构建及雌二醇特异性模拟抗体的筛选

[目的]构建Lipocalin核糖体展示文库,并筛选雌二醇特异性模拟抗体。[方法]以Lipocalin蛋白家族中的BBP(胆汁三烯结合蛋白)为模板,设计包含16个随机突变位点的引物,合成BBP基因文库;引入核糖体展示所必需的全部组件,采用重叠PCR技术将BBP库与核糖体展示组件进行拼接,构建核糖体展示Anticalin模拟抗体库;再将构建的Anticalin文库进行体外转录与翻译,产生模拟抗体-核糖体-mRNA三联复合体文库。以小分子抗原雌二醇为靶标,采用固相亲和方法筛选抗雌二醇抗体,通过改变Mg2+浓度将模拟抗体-核糖体-mRNA三联复合体解离,得到与模拟抗体对应的mRNA,经过RT-PCR得到筛选后的DNA文库并重新引入核糖体展示元件进行下一轮淘筛。[结果]通过5轮生物淘筛,模拟抗体得到富集,获得高特异性抗雌二醇抗体。其中有一株克隆的亲和力为54 mmol/L(Kon=4.975 6×104个/M·S;Koff=0.002 7个/S)。IC50与LOD值分别为50和0.071 ng/ml。[结论]获得的雌二醇模拟抗体可以作为抗体并用于检测动物体内雌二醇的残留。     

Estradiol: specific binding by pituitary nuclear fraction in vitro

The nuclear fraction of a homogenate of anterior pituitary has a marked binding affinity for estradiol but not for other hormonal steroids. Characteristics of the uptake of estradiol by pituitary nuclear fraction are like those reported for the uterus. Study in vitro will be useful in elucidating how direct estrogen feedback control of anterior pituitary function is mediated.     

Transfection of v-rasH DNA into MCF-7 human breast cancer cells bypasses dependence on estrogen for tumorigenicity

The natural history of estrogen-responsive breast cancers often involves a phenotypic change to an estrogen-unresponsive, more aggressive tumor. The human breast cancer cell line, MCF-7, which requires estradiol for tumor formation in vivo and shows growth stimulation in response to estradiol in vitro, is a model for hormone-responsive tumors. The v-rasH onc gene was transfected into MCF-7 cells. The cloned MCF-7ras transfectants, which expressed the v-rasH messenger RNA and v-rasH p21 protein (21,000 daltons), were characterized. In contrast to the parental cell line, MCF-7ras cells no longer responded to exogenous estrogen in culture and their growth was minimally inhibited by exogenous antiestrogens. When tested in the nude mouse, the MCF-7ras cells were fully tumorigenic in the absence of estrogen supplementation. Thus, cells acquiring an activated onc gene can bypass the hormonal regulatory signals that trigger the neoplastic growth of a human breast cancer cell line.     

炔雌醇和菲2种环境内分泌干扰物对斑马鱼的影响研究进展

炔雌醇和菲在各种水环境中广泛分布,其对水生动物的繁殖、发育和生长等各方面有显著的影响.斑马鱼容易饲育,遗传学工具成熟,是研究脊椎动物的一种模式动物.综述了炔雌醇和菲2种内分泌干扰物对斑马鱼发育的影响及其机制的研究进展.     

Estrogen-induced factors of breast cancer cells partially replace estrogen to promote tumor growth

The hormone 17 beta-estradiol acts through its receptor system to induce MCF-7 human breast cancer cells to form tumors in athymic mice. In vitro studies have identified the production of estrogen-induced growth factors from MCF-7 cells that may have a role in growth control. These induced growth factors were sufficient to stimulate MCF-7 tumor growth in ovariectomized athymic mice, thus partially replacing estradiol. Growth factors may act as estrogen-induced "second messengers" in estrogen-responsive growth of human breast cancer.     

钒的生物学作用研究进展

综述了近年来钒的生物学作用的研究进展。大量的研究表明，钒在动物和人体中起着重要的生理作用，另外，钒与细胞凋亡、癌症和营养代谢紧密相关，并作用于免疫系统。所有的这些事实说明钒是一种动物必需的元素。     

癌症的病原物是类病毒或缺陷性病毒假说--癌症病因、机理探讨

根据现有的一些证据，结合有关生物学理论进行综合分析，提出了一个“生物导弹”致癌的新假说，即在某些生物如病毒或其他生物中存在一种比病毒还要小的致病微生物——类病毒或缺陷性病毒。病毒蛋白质壳内的类病毒或缺陷性病毒随病毒传播，并随病毒基因组一同进入动物细胞中．发生作用和复制。它们在细胞中，在某些致癌因子的帮助下，与寄主细胞染色体发生作用，实现细胞遗传上的癌化。在细胞中新形成的病毒蛋白质外壳将新复制的病毒染色体和新复制的类病毒或缺陷性病毒一同包于其中，形成新的“生物导弹”，将来可以借机转染新的对象。根据这一假说，既可以解释现有癌症理论可以解释的问题，也可以解释现有理论难以解释的问题，并指出动物对肿瘤存在抵抗机制。     

Sequence and expression of human estrogen receptor complementary DNA

The mechanism by which the estrogen receptor and other steroid hormone receptors regulate gene expression in eukaryotic cells is not well understood. In this study, a complementary DNA clone containing the entire translated portion of the messenger RNA for the estrogen receptor from MCF-7 human breast cancer cells was sequenced and then expressed in Chinese hamster ovary (CHO-K1) cells to give a functional protein. An open reading frame of 1785 nucleotides in the complementary DNA corresponded to a polypeptide of 595 amino acids and a molecular weight of 66,200, which is in good agreement with published molecular weight values of 65,000 to 70,000 for the estrogen receptor. Homogenates of transformed Chinese hamster ovary cells containing a protein that bound [3H]estradiol and sedimented as a 4S complex in salt-containing sucrose gradients and as an 8 to 9S complex in the absence of salt. Interaction of this receptor-[3H]estradiol complex with a monoclonal antibody that is specific for primate ER confirms the identity of the expressed complementary DNA as human estrogen receptor. Amino acid sequence comparisons revealed significant regional homology among the human estrogen receptor, the human glucocorticoid receptor, and the putative v-erbA oncogene product. This suggests that steroid receptor genes and the avian erythroblastosis viral oncogene are derived from a common primordial gene. The homologous region, which is rich in cysteine, lysine, and arginine, may represent the DNA-binding domain of these proteins.     

布鲁氏菌病检测技术及检测用靶蛋白抗原筛选的概况

布鲁氏菌病是一种重要的人畜共患病,也是引起动物繁殖障碍的主要疾病之一。近年来,我国人、畜间布鲁氏菌病呈高发态势,严重威胁畜牧业发展和人群身体健康。及时准确诊断对控制和净化布鲁氏菌病具有重要意义。本研究从病原学和血清学两方面对布鲁氏菌病的现有诊断技术进行概述,同时指出布鲁氏菌免疫原性蛋白抗原的筛选和研究,对布病的控制和净化意义重大。     

Oncogene-induced transformation of C3H 10T1/2 cells is enhanced by tumor promoters

The tumor promoters 12-O-tetradecanoyl-phorbol-13-acetate and teleocidin markedly enhanced the transformation of C3H 10T1/2 mouse fibroblasts when these cells were transfected with the cloned human bladder cancer c-rasH oncogene. Transfection studies with the drug resistance marker gpt and time course studies indicate that this enhancement is not simply an effect on the process of DNA transfection. These findings, together with parallel studies with NIH 3T3 fibroblasts, also indicate that the competence of animal cells for DNA transfection is a function of the recipient cell line, the transfected marker, and the growth conditions. Our findings suggest that during multistage carcinogenesis tumor promoters may complement the function of activated cellular oncogenes.