Identifying And Evaluating Sources Of Variability In Rodent Studies

识别和评估啮齿动物研究中变异的来源

基本信息

批准号：
7734436
负责人：
grace e kissling
金额：
$ 7.22万
依托单位：
NATIONAL INSTITUTE OF ENVIRONMENTAL HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/7734436
关键词：
Accounting Adopted Affect Animals Bacteria Biological Biological Assay Blood Body Weight Bone Marrow Cancer Etiology Carcinoma Cell Count Cell Nucleus Cells Chemical Exposure Chemicals Chromosomes Collection Computer Simulation Conflict (Psychology)Consumption Count Data Data Analyses Data Quality Databases Diagnostic Diet End Point Ensure Environmental Exposure Erythrocytes Estrogens Exhibits Female Flow Cytometry Future Genetic Goals Hepatitis Infection Kidney Kidney Neoplasms Laboratories Laboratory Study Liver Liver neoplasms Measurement Metabolic Methodology Methods Micronucleus Tests Microscope Microscopic Microscopy Morphology Mouse Strains Mus Mutagenicity Tests Noise Numbers Organ Outcome Pathologist Phytoestrogens Play Population Predisposition Range Rate Rattus Research Design Rodent Role Sampling Studies Signal Transduction Source Sprague-Dawley Rats Statistical Models Statistically Significant Techniques Time Use of New Techniques Vagina Variant adenoma base carcinogenicity feeding improved male micronucleus research study response triethanolamine tumor

项目摘要

Short-term rodent bioassays, such as the uterotrophic bioassay, have been developed to assess estrogenic effects of chemicals. The outcomes of these assays may be affected by factors other than the chemicals under study. For example, many laboratory diets contain phytoestrogens that have estrogenic effects. Also, diets vary in the amount of metabolic energy that is available. High energy consumption may interfere with estrogenic responses. Furthermore, different rodent strains exhibit differing sensitivities to estrogens. Because bioassays for estrogenic effects are conducted at many laboratories around the world, using different diets and different rodent strains, conflicting results sometimes occur. For the endpoint of timing of vaginal opening, we showed that the commonly used Sprague-Dawley rat is less sensitive to estrogenic chemicals than the Fischer 344 rat or the CD-1 mouse. We also found that phytoestrogen level in the diet is more highly correlated with some estrogenic responses than is metabolic energy. We recommend that rodents in bioassays for assessing estrogenic effects should be fed a low metabolic energy diet that is free of phytoestrogens. Furthermore, sensitivity to estrogenic effects varies among strains of mice and rats as well as among endpoints, so strain selection should be made carefully. The NTP conducts rodent micronucleus tests as part of its battery of genetic toxicity testing. Normally, as red blood cells mature, they shed their nucleus. If chromosome damage has occurred in the red blood cells, small parts of the nucleus (micronuclei) may remain in the cell. In the micronucleus test, blood or bone marrow of mice or rats exposed to a chemical is examined under the microscope and numbers of mature red blood cells containing micronuclei are counted. Recently, a new technique using flow cytometry has been proposed for counting micronuclei in a less mature population of red blood cells. An advantage of this technique is that many more cells can be examined per animal than is feasible with microscopy. We contributed to two studies comparing microscopic and flow cytometry enumerations of micronuclei. 1) Blood and bone marrow samples from studies of nine chemicals in mice and rats were evaluated using both microscopic and flow cytometry techniques. We found that the two techniques produced very similar micronucleus counts and they resulted in the same conclusion about whether a chemical is genotoxic. 2) Using a range of typical values of micronucleus occurrences, we conducted computer simulations to study the effects of the number of cells examined per animal on the power and sensitivity of the micronucleus test. Based on both of these studies, the NTP has decided to adopt flow cytometry for the micronucleus test. In the mid-1990s, several NTP studies of chemicals were partially compromised by an infection of Heliobacter hepaticus. This bacterium causes hepatitis in mice which often leads to liver tumors. Because liver tumors were observed in these studies, it was not clear whether H. hepaticus or the chemical exposure was responsible. For most of these studies, the chemical caused cancer in other organs unaffected by the infection, so the carcinogenic effect of the chemical could be determined. For triethanolamine, however, tumors were observed only in the liver. Several years later, the NTP conducted a second, identical study of triethanolamine in mice that were free of H. hepaticus infection. However, at that time, the diet that the mice were fed had been changed and animals were larger in the second study. Because liver tumor rates increase with body weight, among other factors, our analyses included statistical modeling of liver tumor rates using historical control data from the each study. We analyzed data from both studies and determined that liver tumor rates in males were only slightly increased by triethanolamine exposure and that the H. hepaticus infection accounted for the statistically significant increase seen in the initial study. Liver tumors in female mice were significantly increased by triethanolamine exposure, independently of the H. hepaticus infection. Several pathologists examining kidneys of NTP rats have noticed a distinctive tumor morphology that they believe occurs spontaneously. The current diagnostic rules, however, do not distinguish this morphology from chemically-induced adenomas and carcinomas. We analyzed the occurrences of this tumor morphology from among all NTP studies and determined that the occurrences are, indeed, random. Based on this study, the diagnostic rules that the NTP uses for kidney tumors may be refined to separate these tumors from chemically-induced tumors, potentially removing noise from the signal of chemical induction of kidney tumors. This would improve the accuracy of NTPs determinations regarding carcinogenicity. Because study design is critical to the collection of high quality data and may control factors involved in extraneous variability of data, we contributed to NTP study design teams for 6 chemicals. We are also providing advice to the new NTP Host Susceptibility Branch as they seek to understand the role that genetic differences play in the variation of responses to environmental exposures.

已经开发出短期啮齿动物生物测定（例如子宫营养生物测定法）来评估化学物质的雌激素作用。这些测定的结果可能会受到研究化学品以外的其他因素的影响。例如，许多实验室饮食含有具有雌激素作用的植物雌激素。同样，饮食的可用代谢能量量也有所不同。高能量消耗可能会干扰雌激素反应。此外，不同的啮齿动物菌株对雌激素表现出不同的敏感性。由于使用不同的饮食和不同的啮齿动物菌株在世界各地的许多实验室都进行了雌激素作用的生物测定，因此有时会发生冲突的结果。对于阴道开口的时间安排的终点，我们表明常用的sprague-dawley大鼠对雌激素化学物质的敏感性不如Fischer 344大鼠或CD-1小鼠。我们还发现，饮食中的植物雌激素水平与某些雌激素反应高于代谢能量。我们建议在生物测定中评估雌激素作用的啮齿动物应喂养不含植物雌激素的低代谢能量。此外，对雌激素作用的敏感性在小鼠和大鼠菌株之间以及端点之间各不相同，因此应仔细选择应变。 NTP作为其遗传毒性测试的一部分进行啮齿动物的微核测试。通常，随着红细胞的成熟，它们脱落了核。如果在红细胞中发生染色体损伤，则细胞核的一小部分可能保留在细胞中。在微核测试中，在显微镜下检查了暴露于化学物质的小鼠或大鼠的血液或骨髓，并计算含有微核的成熟红细胞的数量。最近，已经提出了一种使用流式细胞仪的新技术来计算微核中的微核中，在不太成熟的红细胞种群中。该技术的一个优点是，每只动物可以检查许多细胞比显微镜可行的细胞多。我们有助于两项研究，比较了微核的显微镜和流式细胞仪列表。 1）使用显微镜和流式细胞仪技术评估了从小鼠和大鼠研究的九种化学物质研究中的血液和骨髓样品。我们发现，这两种技术产生了非常相似的微核计数，并且对化学物质是否具有遗传毒性得出了相同的结论。 2）使用一系列典型的微核发生值，我们进行了计算机模拟，以研究每只动物检查的细胞数量对微核测试的功率和灵敏度的影响。基于这两项研究，NTP决定采用流式细胞仪进行微核测试。在1990年代中期，几项对化学物质的NTP研究因肝杆菌的感染而部分损害。这种细菌会导致小鼠肝炎，这通常会导致肝肿瘤。由于在这些研究中观察到肝肿瘤，因此尚不清楚肝甲基肝炎或化学暴露是造成的。对于大多数研究，化学物质在不受感染影响的其他器官中引起癌症，因此可以确定化学物质的致癌作用。然而，对于三乙醇胺，仅在肝脏中观察到肿瘤。几年后，NTP在没有肝螺旋杆菌感染的小鼠中对三乙醇胺进行了第二项相同的研究。但是，当时，在第二项研究中，喂养小鼠的饮食已经改变，动物也更大。由于肝肿瘤率随体重的增加，除其他因素外，我们的分析包括使用每项研究的历史控制数据对肝肿瘤率进行统计建模。我们分析了这两项研究的数据，并确定只有三乙醇胺暴露仅通过暴露于男性的肝脏肿瘤率略有增加，而肝硬化感染则是最初研究中统计学上显着的升高。雌性小鼠的肝肿瘤通过三乙醇胺暴露显着增加，而与肝癌感染无关。几位检查NTP大鼠肾脏的病理学家注意到一种独特的肿瘤形态，他们认为这是自发发生的。但是，目前的诊断规则并未将这种形态与化学诱导的腺瘤和癌区分开。我们从所有NTP研究中分析了这种肿瘤形态的发生，并确定发生的确是随机的。基于这项研究，可以将NTP用于肾脏肿瘤使用的诊断规则将这些肿瘤与化学诱导的肿瘤分开，从而从化学诱导的肿瘤中分离出来，从而从肾脏肿瘤的化学诱导信号中消除了噪声。这将提高NTPS确定有关致癌性的准确性。由于研究设计对于收集高质量数据至关重要，并且可能控制与数据无关可变性有关的因素，因此我们为6种化学物质的NTP研究设计团队做出了贡献。我们还为新的NTP宿主易感分支提供了建议，因为他们试图了解遗传差异在对环境暴露的反应变化中的作用。