O'Connor J C, Cook J C, Craven S C, Van Pelt C S, Obourn J D
Haskell Laboratory for Toxicology and Industrial Medicine, E.I.du Pont de Nemours & Company, Newark, Delaware 19714, USA.
Fundam Appl Toxicol. 1996 Oct;33(2):182-95.
We have combined several endpoints into a single 5-day in vivo screening procedure to identify estrogenic compounds and dopaminergic modulators, both of which play important roles in enhancing mammary tumorigenesis in rodents. The endpoints evaluated as markers of estrogenicity included increases in uterine fluid and vaginal cornification incidence, serum prolactin levels, uterine weight, uterine epithelial cell height, uterine stromal cell proliferation, and uterine progesterone receptor (PR) number and decreases in uterine estrogen receptor (ER) number. The endpoints evaluated for changes in dopamine regulation included increases in prolactin and decreases in growth hormone levels. The estrogen agonist estradiol (E2) and estriol (E3), the mixed estrogen agonist/ antagonist tamoxifen (TAM), the full antiestrogen ICI-182, 780 (ICI),and the dopamine modulators haloperidol (HAL) and reserpine (RES) were tested using a three-time/day (8-hr intervals) intraperitoneal dosing regimen in sexually mature ovariectomized female Crl:CD BR rats. All compounds were evaluated over a range of concentrations. This in vivo battery was used to evaluate the effects of different classes of endocrine modulators on the selected endpoints. For example, the estrogen receptor agonists E2 and E3 display a unique profile based on changes in the uterotrophic endpoints (estrus conversion, uterine fluid imbibition, increases in uterine weight, and uterine endometrial cell proliferation) where full and partial agonists can be distinguished by the magnitude of these responses. Both the estrogen receptor antagonist ICI and the dopamine modulators HAL and RES lack these uterotrophic responses. Dopamine modulators can be distinguished from estrogen receptor agonists by the profile of increased prolactin levels with no uterotrophic changes. Estrogen receptor antagonists can be distinguished from agonists by comparing their effects on ER, PR, and uterotrophic responses. For instance, the full estrogen receptor antagonist ICI decreased ER (to almost 0) and PR levels, but has no uterotrophic effects, while TAM decreases ER (to almost 0) and increases PR with uterotrophic effects. The most useful endpoints for distinguishing estrogen agonists and dopamine modulators were uterine fluid imbibition, uterine weight, uterine stromal cell proliferation, and serum prolactin levels. In order to distinguish an estrogen agonist from an antagonist, other endpoints, such as receptor levels, are necessary. The advantage of an in vivo screen is that it utilizes a metabolically and physiologically defined system which is especially important with highly integrative system such as the endocrine system. This battery can be used as a screening tool to identify potential endocrine modulators as well as to identify mode of action following adverse findings in toxicology studies. Last, additional endpoints may be added to identify other classes of endocrine modulators.
我们将多个终点指标整合到一个为期5天的体内筛选程序中，以识别雌激素化合物和多巴胺能调节剂，这两类物质在增强啮齿动物乳腺肿瘤发生过程中均发挥重要作用。评估雌激素活性的终点指标包括子宫液增加、阴道角化发生率、血清催乳素水平、子宫重量、子宫上皮细胞高度、子宫基质细胞增殖以及子宫孕酮受体（PR）数量增加，子宫雌激素受体（ER）数量减少。评估多巴胺调节变化的终点指标包括催乳素增加和生长激素水平降低。使用每天三次（间隔8小时）腹腔给药方案，对性成熟的去卵巢雌性Crl:CD BR大鼠进行雌激素激动剂雌二醇（E2）和雌三醇（E3）、雌激素激动剂/拮抗剂他莫昔芬（TAM）、完全抗雌激素ICI-182,780（ICI）以及多巴胺调节剂氟哌啶醇（HAL）和利血平（RES）的测试。所有化合物均在一系列浓度范围内进行评估。这个体内检测组合用于评估不同类别的内分泌调节剂对选定终点指标的影响。例如，雌激素受体激动剂E2和E3基于子宫营养终点指标（发情转换、子宫液吸收、子宫重量增加和子宫内膜细胞增殖）的变化呈现出独特的特征，其中完全激动剂和部分激动剂可通过这些反应的程度来区分。雌激素受体拮抗剂ICI以及多巴胺调节剂HAL和RES均缺乏这些子宫营养反应。多巴胺调节剂可通过催乳素水平升高且无子宫营养变化的特征与雌激素受体激动剂区分开来。雌激素受体拮抗剂可通过比较其对ER、PR和子宫营养反应的影响与激动剂区分开来。例如，完全雌激素受体拮抗剂ICI可使ER（降至几乎为0）和PR水平降低，但无子宫营养作用，而TAM可使ER（降至几乎为0）并增加PR，同时具有子宫营养作用。区分雌激素激动剂和多巴胺调节剂最有用的终点指标是子宫液吸收、子宫重量、子宫基质细胞增殖和血清催乳素水平。为了区分雌激素激动剂和拮抗剂，还需要其他终点指标，如受体水平。体内筛选的优势在于它利用了一个代谢和生理特征明确的系统，这对于像内分泌系统这样高度整合的系统尤为重要。这个检测组合可作为一种筛选工具，用于识别潜在的内分泌调节剂以及在毒理学研究中出现不良发现后确定作用模式。最后，可以添加其他终点指标以识别其他类别的内分泌调节剂。
