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首页> 外文OA文献 >Untersuchungen zur Wirkung von östrogenwirksamen Stoffen auf die Gonadenentwicklung bei Fröschen (Rana temporaria und Xenopus laevis) sowie ultrastrukturelle und immunhistochemische Untersuchungen an Froschgonaden (Rana temporaria)
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Untersuchungen zur Wirkung von östrogenwirksamen Stoffen auf die Gonadenentwicklung bei Fröschen (Rana temporaria und Xenopus laevis) sowie ultrastrukturelle und immunhistochemische Untersuchungen an Froschgonaden (Rana temporaria)

机译：研究雌激素活性物质对青蛙（林蛙和非洲爪蟾）性腺发育的影响以及青蛙性腺（林蛙）的超微结构和免疫组化研究

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The influence of estrogen-like substances on the gonadal development in frogs (Rana temporaria and Xenopus laevis). Ultrastructural and immunohistochemical investiga-tions on the gonads of frogs (Rana temporaria).The aim of the present thesis was the investigation and assessment of the estrogen pollu-tion of a river in South-Germany above and below a sewage plant outlet and its possible effects on the gonadal development in amphibians. In an exposure experiment with differ-ent sewage dilutions the effect on the indigenous gras frog (Rana temporaria) were com-pared with the effects on the African claw frog (Xenopus laevis). Because of the lack of relevant data in current literature it was necessary to first collect some basic data on light microscopy, ultrastructure and immunohistochemistry with reference to frogs.Furthermore the sexual differentiation of two populations of different origin were compared histologically. Characterisation of the gonadsThe ovaries of Xenopus laevis displayed a garland-like structure in contrast to the compact ovaries of Rana temporaria. The ovaries of both frog species were surrounded by an epi-thelium and filled with germ cells which enclosed an ovarian cavity. The oogonies were mainly found in the peripheral parts of the organ. The oocytes were characterised by a big and irregularly shaped cell nucleolus with peripherally located nucleoli. They were sur-rounded by a thin one-layered follicular epithelium. Electron microscopical examination of the ovaries of Rana temporaria showed elongated tubular mitochondria in the cytoplasm which were found exclusively in oocytes. Some of the mitochondria from subadult frogs contained yolk crystals. Additionally an accumulation of electron-tight Granule was found just below the plasma membrane, which could be a preliminary stage of the cortical granule. The testis of the frogs were enveloped by an epithelium and a tunica albuginea which al-ready showed a tubular structure. The testis from Rana temporaria and the Xenopus laevis differed in their developmental stages. The testis of juvenile Rana temporaria contained only germ cells whereas the testis of some Xenopus laevis already contained spermato-cytes and spermatozoa. The spermatocysts, characteristic of frog testis, could be seen in conjunction with the appearance of spermatocytes only. By using light microscopy two different types of germ cells in both frog species could be distinguished which probably were primary and secondary germ cells. By electron microscopical investigation of the testis of Rana temporaria the somatic cells could, on the basis of location and morphology, be differentiated in two types. The somatic cells of the first type were located inside the Tubuli seminiferi and have probably the same function as the Sertoli cells of mammals. The cells of the second type, the Leydig cells or interstitial cells, were located outside the Tubuli and were characterized by granular vesicle in the cytoplasm.With view to the sexual differentiation of Rana temporaria a comparative histological inves-tigation of the gonads of juvenile and subadult frogs of a native midland population and a high alpine population was performed. The midland population proved to be a sexually semi-differentiated species since apart from clearly male or female animals it also com-prises intersex individuals in different stages of the transformation process. The genotypi-cally male animals developed female gonads in the first place which secondly converted into testis during a hermaphrodite stage. The transformation process from female into male gonads was, on the basis of morphological criteria, classified into three stages. The highal-pin population on the other hand proved to be a sexually differentiated race.By using the Avidin-Biotin-Complex-Technique five different antibodies were tested on the gonadal tissue of juvenile and subadult Rana temporaria. The test for Laminin, a non-collagen glycoprotein, which is part of the basal membrane in mammals, resulted in a posi-tive reaction. It seems therefore that similar to Laminin in mammals a glycoprotein plays an important part in the basal membrane of frogs. α-Actin, a fibrous protein of the smooth muscles, which was detected in the Theca externa of the layer of follicle cells in different species, could be established in the blood vessel wall only and not in the layer of follicle cells. Furthermore the occurrence of a Zona pellucida by using antibodies against the por-cine glycoprotein ZP3 was investigated. The ovum including the surrounding layer of follicle cells showed a negative reaction which however did not implicitly exclude the occurrence of a glycoprotein layer with a different antigenicity. Until now the existence of a glycoprotein layer in anurans, in connection with microscopical anatomy of ovaries in Rana temporaria or other species of frogs has not been mentioned in references. The examination of the go-nadal tissue as to the occurrence of ACE (Angiotensin Converting Enzymes) turned out negative as well.Example of applicationWith the exception of a temporarily higher concentration of alkylphenol in the beginning of the exposure, the chemical analyses revealed a relatively low degree of pollution with alcyl-phenols and steroids. The results were comparable to other results of German rivers and stayed well below other European comparative data. According to the results of this study the current level of pollution of the experimental water with estrogens does not endanger the amphibian population. The examination did not reveal any influence of the sewage on the embryonal and larval development. Furthermore, the histological investigation of the gonads in exposed and unexposed frogs with reference to the gonadal sexual differentia-tion as well as the sex ratios did not reveal significant changes. There was no correlation as to the frequency of the occurrence of intersex in the groups of exposed and unexposed frogs, neither of Xenopus laevis nor of Rana temporaria. Only the transformation process from ovaries to testis of the sexually semi-differentiated species of Rana temporaria was slowed down in the group of exposed animals in contrast to unexposed animals. The rea-son for this phenomenon could be the inhibitive influence of the low-level but more continu-ous estrogen pollution in sewage than in river water. Conversely, a link between the accel-eration of the transformation process within the group of unexposed animals and the tem-porarily higher alkylphenol level in the beginning of the exposure cannot completely be ruled out. According to references alkylphenol can cause an increase of testosterone. The semiquantitative RT-PCR detecting Vitellogenin-mRNA carried out by the Institute of Freshwater Ecology and Inland Fisheries in Berlin showed a minor increase in females of Xenopus laevis which were exposed to sewage in the ration of a 2:1 dilution in contrast to the unexposed animals. Taking into account the synergistic effects of estrogens, the in-crease could be attributed to the higher estrogen pollution of the sewage. The histopa-thological analysis for the detection of toxical effects of the sewage as well as other poten-tial influential factors provided no hints as to a possibly toxical influence of the sewage.
机译：类雌激素物质对青蛙性腺发育的影响（蛙蛙和非洲爪蟾）。蛙性腺（蛙蛙）的超微结构和免疫组化研究。本论文的目的是调查和评估德国南部一条河流中污水厂出口上方和下方的雌激素污染及其可能的发生。对两栖动物性腺发育的影响。在使用不同污水稀释度的暴露实验中，将其对本地鹅蛙（Rana temporaria）的影响与对非洲爪蛙（Xenopus laevis）的影响进行了比较。由于当前文献中缺乏相关数据，因此有必要首先收集有关青蛙的光学显微镜，超微结构和免疫组化方面的一些基本数据，然后对两个不同来源的种群的性别分化进行组织学比较。性腺的特征非洲爪蟾的卵巢表现出与花环蛙紧凑的卵巢相反的花环状结构。两种蛙类的卵巢都被上皮包围，并充满了封闭卵巢腔的生殖细胞。卵菌主要存在于器官的外围部分。卵母细胞的特征是大而形状不规则的细胞核仁，核仁位于外围。它们被薄的一层滤泡上皮所包围。电子显微镜检查的林蛙卵巢显示在细胞质中拉长的管状线粒体，仅在卵母细胞中发现。来自成年蛙的一些线粒体含有卵黄晶体。另外，在质膜的下方发现了电子密封颗粒的堆积，这可能是皮质颗粒的初级阶段。青蛙的睾丸被上皮和白膜包裹，它们已经显示出管状结构。来自Rana temporaria和Xenopus laevis的睾丸在发育阶段有所不同。幼年蛙的睾丸只含有生殖细胞，而某些非洲爪蟾的睾丸已经含有精子细胞和精子。青蛙睾丸所特有的精囊仅与精母细胞的外观结合可见。通过使用光学显微镜，可以区分两种青蛙物种中两种不同类型的生殖细胞，它们可能是原代和次生生殖细胞。通过电子显微镜研究的林蛙的睾丸，可以根据位置和形态将体细胞分为两种类型。第一类体细胞位于细小管藻中，其功能可能与哺乳动物的Sertoli细胞相同。第二类细胞是Leydig细胞或间质细胞，位于Tubuli外部，其特征是胞质中有小泡。考虑到Rana temporaria的性分化，比较了少年和性腺性腺的组织学研究。进行了本地中部种群和高高山种群的亚成体蛙的研究。中地种群被证明是有性的半分化物种，因为除了明显的雄性或雌性动物外，它还包括转换过程中不同阶段的双性恋个体。基因型雄性动物首先发育成雌性腺，然后在雌雄同体阶段再转变成睾丸。从雌性到雄性腺的转化过程根据形态学标准分为三个阶段。另一方面，高针种群被证明是有性别差异的种族。通过使用抗生物素蛋白-生物素-复杂技术，在少年和亚成年蛙的性腺组织上测试了五种不同的抗体。层粘连蛋白（一种非胶原蛋白糖蛋白）的测试导致了阳性反应，后者是哺乳动物基底膜的一部分。因此，与哺乳动物的层粘连蛋白相似，糖蛋白在青蛙的基底膜中起着重要的作用。 α-肌动蛋白是一种平滑肌纤维蛋白，仅在血管壁而不是在卵泡细胞层中建立，它在不同物种的卵泡细胞层的Theca externa中被检测到。此外，还研究了通过使用针对猪糖蛋白ZP3的抗体产生的透明带。包括卵泡细胞周围层的卵子显示出阴性反应，但是并未隐含排除具有不同抗原性的糖蛋白层的出现。到现在为止，无核糖中存在糖蛋白层在参考文献中未提及与蛙蛙或其他种类的青蛙的卵巢的显微解剖相关的。对性腺组织的ACE（血管紧张素转化酶）的检查也呈阴性反应。应用示例除了接触开始时暂时升高浓度的烷基酚外，化学分析显示烷基酚和类固醇污染程度低。该结果与德国河流的其他结果可比，并且远低于其他欧洲比较数据。根据这项研究的结果，目前的实验用水被雌激素污染的程度不会危害两栖动物。检查没有发现污水对胚胎和幼体发育的任何影响。此外，参照性腺性别差异以及性别比例对裸露和未暴露青蛙的性腺进行的组织学研究均未发现明显变化。在非洲的非洲爪蟾和印度蛙中，暴露和未暴露的青蛙群中的性交发生频率没有相关性。与未暴露的动物相比，在暴露的动物组中，只有蛙的性半分化种从卵巢到睾丸的转化过程减慢了。造成这种现象的原因可能是污水中的低水平但持续不断的雌激素污染比河水中的抑制作用更大。相反，不能完全排除在未暴露动物组中转化过程的加速与暴露开始时暂时较高的烷基酚水平之间的联系。根据参考文献，烷基酚可引起睾丸激素增加。柏林淡水生态和内陆渔业研究所进行的半定量RT-PCR检测卵黄蛋白原-mRNA显示，非洲非洲爪蟾的雌性略有增加，以2：1的稀释比暴露于污水，而未暴露动物。考虑到雌激素的协同作用，这种增加可以归因于污水中较高的雌激素污染。用于检测污水的毒性作用以及其他潜在影响因素的组织形态学分析没有提供关于污水可能的毒性影响的暗示。

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    Steineck Cornelia;

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  • 年度 2003
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  • 正文语种 {"code":"de","name":"German","id":7}
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