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

通过干扰激素的功能和作用，内分泌干扰物(EDCs)，如增塑剂邻苯二甲酸二(2-乙基己基)酯(DEHP)，可对发育和生殖产生负面影响。在研究促进奖(SC2)的支持下，Traore博士在伊丽莎白城市州立大学担任教员时发现，怀孕期间体内暴露于DEHP对成年人的睾酮(T)产生负面影响，早期暴露会加剧后续暴露的影响。现在，在坎贝尔大学医学院，特劳雷博士提议对急性和慢性DEHP对间质细胞产生T的影响进行体外机制研究。直到最近，通过现有方法分离和培养的间质细胞只能产生并维持高水平的T细胞3-5天。因此，使用体外培养的原代细胞来检测环境相关浓度的EDCs的直接、长期影响或对其作用于间质细胞的机制进行批判性评估是不可行的。然而，齐尔金博士在约翰·霍普金斯大学的实验室 彭博公共卫生学院(JHSPH)最近开发出一种方法，通过这种方法，从LeyDig干细胞分化而来的Leydig细胞能够产生并维持较长时间，甚至几个月的高水平T细胞产生。曾担任SC2奖导师的齐尔金博士将继续与特拉雷博士和他的学生们合作。能够维持高水平T产生的原代间质细胞的可用性将对我们拟议的高度控制条件下环境相关邻苯二甲酸盐浓度的慢性影响及其作用机制的研究具有重要价值。随着这些细胞的获得，我们将在体外进行MEHP对干细胞形成间质细胞的直接影响，MEHP暴露对成年间质细胞的影响，以及早期暴露对后续暴露后果的影响。我们还将对邻苯二甲酸盐诱导的氧化应激对T细胞的急性和长期影响进行机制研究。 制作。坎贝尔大学拥有拟议研究所需的设备和设施，以及 将参与本研究项目的优秀医学生、研究生和本科生。有了这笔REAP赠款的资金，特拉雷博士将能够支持和扩大他在坎贝尔大学实验室正在进行的研究，坎贝尔大学是一所位于北卡罗来纳州农村地区、服务不足的机构。约翰霍普金斯大学公共卫生学院和坎贝尔大学之间的广泛合作不仅有助于激发坎贝尔大学的学生，还将增加他们考虑从事基础科学或临床研究的兴趣。此外，该项目无疑将吸引坎贝尔大学的其他教职员工寻求资金来开展他们的研究，并有助于加强校园内的研究环境。

By interfering with hormone function and action, endocrine-disrupting chemicals (EDCs) such as the plasticizer di-(2-ethylhexyl) phthalate (DEHP) can have negative effects on development and reproduction. With the support of a Research Enhancement (SC2) Award, when he was a faculty member at Elizabeth City State University, Dr. Traore found that in vivo exposure to DEHP during gestation had negative effects on testosterone (T) production in the adult, and that early exposure exacerbated the effects of subsequent exposures. Now at Campbell University School of Medicine, Dr. Traore is proposing to conduct in vitro mechanistic studies of both acute and chronic DEHP effects on T production by Leydig cells. Until recently, Leydig cells isolated and cultured by available methods have been able to produce and maintain high levels of T production for only 3-5 days. Therefore, it has not yet been feasible to use primary cells in vitro to examine the direct, long-term effects of environmentally relevant concentrations of EDCs or to critically evaluate mechanisms involved in their actions on Leydig cells. However, Dr. Zirkin’s lab at the Johns Hopkins Bloomberg School of Public Health (JHSPH) has recently developed a method by which Leydig cells derived from the differentiation of Leydig stem cells are able to produce and sustain high levels of T production for longer periods, several months. Dr. Zirkin, who served in a mentorship position for the SC2 Award, is continuing to collaborate with Dr. Traore and his students. The availability of primary Leydig cells able to sustain high levels of T production will be of great value for our proposed studies of the chronic effects of environmentally relevant phthalate concentrations under highly controlled conditions, and the mechanisms by which they function. With the availability of these cells, we will conduct in vitro studies of the direct effects of MEHP on Leydig cell formation from stem cells, the effects of MEHP exposure on the adult Leydig cells, and the effects of early exposures on the consequences of subsequent exposures. We also will conduct mechanistic studies of acute and long-term effects of phthalate-induced oxidative stress on T production. Campbell University has the equipment and facilities necessary for the proposed studies, as well as outstanding medical students, graduate students and undergraduates who will be involved in this research project. With funds of this REAP grant, Dr. Traore will be able to support and expand ongoing research in his lab at Campbell University, an institution located in a rural, underserved area of North Carolina. The extensive collaboration between the Johns Hopkins School of Public Health and Campbell University will not only help excite Campbell University students but also increase their interest in considering careers in basic science or clinical research. Moreover, the project undoubtedly will attract other Campbell University faculty to seek funding to develop their research, and help strengthen the research environment on campus.