Title: In Vitro Analysis of the Effects of Acute and Chronic Phthalate Exposures on Leydig Cell Testosterone Production, and the Molecular Mechanisms Involved

项目摘要 通过干扰正常的激素功能和作用，内分泌干扰化学品（EDCs），如 增塑剂邻苯二甲酸二（2-乙基己基）酯（DEHP）可能对男性生殖系统产生负面影响。在 Traore博士先前的研究发现，子宫内接触DEHP可导致血清睾酮水平降低 成人的水平。在体外进行的研究中，他发现急性（短期）接触单-2-乙基己基 邻苯二甲酸酯（MEHP）是DEHP的代谢产物，可抑制分离的睾丸间质细胞产生睾酮， 通过MA-10小鼠肿瘤Leydig细胞的孕酮产生。此外，他发现MEHP可以消极地 影响MA-10细胞中的线粒体功能和胆固醇移位。直到最近， 使用从睾丸中分离的Leydig细胞进行长期研究，因为尽管这些细胞能够 产生高水平的睾丸激素，他们不这样做超过1-2天。因此， 使用分离的Leydig细胞来检查环境相关浓度的直接，长期影响 或严格评估邻苯二甲酸酯作用的机制。兹金博士在约翰霍普金斯的实验室 彭博公共卫生学院（JHSPH）最近开发了一种方法， 从与生精小管相关的干细胞的体外分化中， 细胞能够产生并维持高水平的睾酮产生数个月。的可用性 这些细胞的数量将对我们提出的研究睾丸激素产生的影响具有重要价值 Leydig细胞急性、慢性和重复暴露于环境相关的邻苯二甲酸酯 浓度，特别是用于解决邻苯二甲酸酯- 导致睾丸激素减少。我们在R15更新申请中提出的研究将： (1)增加我们对环境压力因素介导抑制的机制的理解， 睾酮激素的产生;（2）提供了关于间质细胞多次暴露于 邻苯二甲酸盐，我们一生中接触的化学品;以及3）解决早期接触是否 在以后的生活中易受随后的无毒剂量暴露的影响。坎贝尔大学 优秀的医学生，研究生和本科生谁将参与拟议的 research.随着资金从REAP赠款，目前的应用程序寻求更新，特劳雷博士支持和 他在坎贝尔大学的实验室里训练了许多学生，坎贝尔大学是一所位于农村的机构， 北卡罗来纳州。赠款的更新将允许Traore博士继续并大大扩展正在进行的研究 同时增加参与学生对基础科学和/或临床研究的兴趣。而且他 研究的成功无疑会影响吸引其他坎贝尔大学的教师寻求资金来发展 在他们的实验室里进行研究。

Project Summary By interfering with normal hormonal function and action, endocrine-disrupting chemicals (EDCs) such as the plasticizer di-(2-ethylhexyl) phthalate (DEHP) can have negative effects on the male reproductive system. In previous studies, Dr. Traore found that in utero exposure to DEHP can result in reduced serum testosterone levels in the adult. In studies conducted in vitro, he found that acute (short-term) exposure to mono-2-ethylhexyl phthalate (MEHP), a metabolite of DEHP, can inhibit testosterone production by isolated Leydig cells and progesterone production by MA-10 mouse tumor Leydig cells. Additionally, he found that MEHP can negatively affect mitochondrial function and cholesterol translocation in MA-10 cells. Until recently, it has not been feasible to conduct long-term studies using Leydig cells isolated from the testis because although these cells are able to produce high levels of testosterone, they do not do so for longer than 1-2 days. Therefore, it has not been feasible to use isolated Leydig cells to examine the direct, long-term effects of environmentally relevant concentrations of EDCs, or to critically evaluate mechanisms involved in phthalate actions. Dr. Zirkin’s lab at the Johns Hopkins Bloomberg School of Public Health (JHSPH) recently developed a method by which Leydig cells can be derived from the in vitro differentiation of stem cells associated with the seminiferous tubules, and that the derived Leydig cells are able to produce and sustain high levels of testosterone production for several months. The availability of these cells will be of great value for our proposed studies of the effects on testosterone production of acute, chronic, and repeat exposures of Leydig cells to environmentally relevant phthalate concentrations, and particularly so for addressing the molecular mechanisms involved in phthalate- induced reductions in testosterone. The studies that we propose in this R15 renewal application will: (1) increase our understanding of the mechanism by which environmental stressors mediate the inhibition of testosterone hormone production; and (2) provide new insights regarding multiple exposure of Leydig cells to phthalates, chemicals to which we are exposed throughout life; and 3) address whether early exposures predispose susceptibility to subsequent non-toxic dose exposure later in life. Campbell University has outstanding medical students, graduate students and undergraduates who will be involved in the proposed research. With funds from the REAP grant that the current application seeks to renew, Dr. Traore supported and trained many students in his lab at Campbell University, an institution located in a rural, underserved area of North Carolina. Renewal of the grant will allow Dr. Traore to continue and greatly extend ongoing research while increasing the interest in basic science and/or clinical research of the involved students. Moreover, his research success undoubtedly will influence attract other Campbell University faculty to seek funding to develop research programs in their labs.