Establishment of human mini-Testis for reproductive toxicity testing

总结 在制造和使用这些化学品过程中的职业接触最多 可能对健康造成潜在不良影响的暴露水平。仍然存在相当大的知识差距 关于工作场所接触工业化学品与可能的有害健康之间的关系 方面的影响.监管机构制定的当前测试方法主要依赖于动物测试。的成本和 为进行风险评估而进行动物试验的时间严重限制了广泛的化学品筛选工作。这些 数据缺口需要实施新的毒性测试愿景，如国家 研究理事会的报告“21世纪的毒性测试”。在NIOSH R21 OH 010473的支持下 grant，我们建立了啮齿动物睾丸细胞共培养模型，发现这种体外共培养模型可以 鉴定生殖毒物虽然来自这些体内或体外啮齿动物模型的结果是有益的， 这些模型不能像啮齿类动物那样再现人类精子发生的全过程 与人类相比，以一种明显不同的方式进行。因此，开发体外人类模型， 生殖毒性测试对于推进该领域和理解细胞和 这些分子机制可能是人类精子发生中断的基础。在本提案中，我们 将开发人类睾丸细胞共培养模型，并证明其可靠性和敏感性。 工作场所化学品的高通量和多参数高含量分析筛选模型 生殖和发育系统。具体目的是：（1）建立睾丸细胞共培养体系 人精原干细胞（hSSCs）、支持细胞和间质细胞的体外培养模型;（2） 基于不良结果途径的测定;（3）应用多参数高通量和高- 含量分析（HT/HCA），检查和比较生殖毒性和无毒化合物，以验证 体外睾丸细胞共培养模型。通过这项假设驱动的研究，我们将测试 体外3D人类睾丸细胞共培养模型创建适当微环境的假设 其模拟人类睾丸以促进生殖细胞的体外生长，并产生可重复的结果， 预测生殖毒性。拟议的研究将导致出版物，并产生毒性数据， 工作场所的化学品，并提供数据进行风险评估。因此，该提案符合 癌症、生殖和心血管研究计划（CRC）的使命。的识别 生殖毒物将减少有害接触，降低生殖风险， 故障因此，我们提出的项目是在NIOSH CRC的研究实践（r2 p） 促进将研究成果转化为预防措施的战略。

Summary Occupational exposures during manufacture and application of these chemicals represent the highest level of exposure that could cause potential adverse health effects. There remains a considerable knowledge gap regarding the relationship between workplace exposure of industrial chemicals and possible adverse health effects. Current test methods set by regulatory agencies predominately rely on animal testing. The cost and time with the animal testing for risk assessment seriously limit efforts for extensive chemical screening. These data gaps necessitate the implementation of a new vision for toxicity testing as reflected in the National Research Council’s report “Toxicity Testing in the 21st Century”. Under the support of NIOSH R21 OH010473 grant, we established a rodent testicular cell co-culture model and found this in vitro co-culture model could identify the reproductive toxicants. While results from these in vivo or in vitro rodent models are informative, these models could not recapitulate the whole process of human spermatogenesis as rodent spermatogenesis proceeds in a distinctly different fashion compared to humans. Thus, developing in vitro human models for reproductive toxicity testing is paramount for advancing the field and for understanding the cellular and molecular mechanisms that may underlie these disruptions in human spermatogenesis. In this proposal, we will develop a human testicular cell co-culture model, and demonstrate the reliability and sensitivity of this model in high-throughput and multi-parametric high-content analytical screening for workplace chemicals on the reproductive and developmental systems. The specific aims are (1) to establish testicular cell co-culture model from human spermatogonial stem cells (hSSCs), Sertoli cell and Leydig cells; (2) to develop integrated assays based on the adverse outcome pathways; (3) to apply the multi-parametric high-throughput and high- content analysis (HT/HCA) to examine and compare reproductive toxic and non-toxic compounds to validate the in vitro testicular cell co-culture model. Through this hypothesis-driven research, we will test the hypothesis that an in vitro 3D human testicular cell co-culture model creates a proper microenvironment that mimics a human testis to promote the growth of germ cells in vitro and generate reproducible results for predicting reproductive toxicity. The research proposed will result in publications and generate toxicity data of the chemicals in the workplace, and provide data for risk assessment. Therefore, this proposal meets the mission of Cancer, Reproductive, and Cardiovascular Research Program (CRC). The identification of reproductive toxicants will reduce hazardous exposures and lower the risk of reproductive malfunction. Therefore, our proposed project is within the NIOSH CRC’s Research to Practice (r2p) strategy to promote the transfer and translation of research findings into prevention practices.