Utilizing a human stem cell model of the esophagus to understand racial disparities during injury repair

项目摘要/摘要 非裔美国人(AA)和欧洲裔美国人(EA)胃食道反流的患病率相似 疾病(GERD)。尽管如此，与EA相比，AA显示出较低的食道损伤发生率， 化生和食管腺癌。种群遗传学和分子研究已经牵连到 人类组织中这些差异的特定基因；然而，缺乏种族多样性的人类食道 模型阻碍了对机制和潜在治疗选择的进一步研究。我们开发了一种 人类食道来源多样的干细胞/有机生物库和高含量的基于图像的 胆汁酸损伤反应的筛选试验。结果显示，AA食道细胞对 与电针来源的细胞显著不同，反映了组织特征和临床发现。此外，我们 曾报道过一种关键酶，谷胱甘肽转移酶-2(GSTT2)负责 使活性氧失活，从而减少DNA损伤，并在AA中高表达 食管。利用祖先多样化的干细胞模型，我们展示了GSTT2低水平的关键关联 具有较高的损伤程度，与人体组织对损伤的原始反应一致。然而，GSTT2在以下方面的直接作用 维持上皮细胞动态平衡和对食道细胞适应性的这种反应和机制/药物 仍有待阐明。假设：非裔美国人的食道组织对胃的反应不同 由于解毒酶GSTT2和可稳定的化合物的高表达而造成的酸/胆汁损伤 GSTT2将保护细胞免受损伤。这项提案中要调查的三个具体目标将涉及 原代组织和干细胞来源的体外培养以验证损伤的分子特征和差异 EA和AA细胞在单细胞水平上的反应(目标1)，GSTT2的基因操作 确定直接调解对伤害的保护(目标2)，以及高通量、无偏见 损伤反应的特征与药物筛选相结合以确定将抑制的化合物 胆汁/酸损伤(目标3)。费雷尔-托雷斯博士在K99.R00计划中的主要研究目标是开发高 吞吐量技术，这将使她能够研究祖先不同的种群及其对伤害的反应。 因此，K99阶段已经计划在干细胞遗传修饰和高含量方面进行培训 基于表型的药物发现。在这个指导阶段，费雷尔-托雷斯博士将与杰森·斯宾塞博士合作 乔纳森·塞克斯顿博士的共同导师。指导K99计划是为费雷尔-托雷斯博士设计的 获得这些领域的专业知识。此外，朱尔斯·林博士和马西娅·克鲁兹-科雷亚博士将担任顾问 博士后委员会成员和临床沉浸顾问。这将使用 密歇根大学提供的特殊资源。这将推动费雷尔-托雷斯博士的目标和 帮助她建立了独立的研究计划，专注于#年食道疾病的种族差异 她的R00阶段。

PROJECT SUMMARY/ABSTRACT African Americans (AA) and European Americans (EA) have a similar prevalence of gastro-esophageal reflux disease (GERD). Nonetheless, when compared to EA, AA show a lower incidence of esophagus damage, metaplasia, and esophageal adenocarcinoma. Population genetics and molecular studies have implicated specific genes for these differences in human tissue; however, a lack of racially diverse human esophagus models hinders further investigation into the mechanisms and potential treatment options. We developed an ancestrally diverse stem cell/organoid biobank of human esophagus and a high-content, image-based screening assay to interrogate bile-acid injury response. Results showed that AA esophageal cells responded significantly differently than EA-derived cells, mirroring tissue profiling and clinical findings. Furthermore, we have previously reported that a key enzyme, glutathione-transferase theta-2 (GSTT2), is responsible for inactivating reactive oxygen species, thus reducing DNA damage, and is highly expressed in the AA esophagus. Utilizing the ancestrally diverse stem cell model, we show key associations of GSTT2 low levels with higher injury, consistent with primary human tissue response to injury. However, a direct role of GSTT2 in this response and mechanism/drugs to maintain epithelial homeostasis and fitness to esophageal cells remains to be elucidated. Hypothesis: Esophageal tissue from African Americans respond differently to gastric acid/bile injury due to higher expression of detoxifying enzyme GSTT2, and compounds that can stabilize GSTT2 will protect cells against injury. The three specific aims to be investigated in this proposal will involve primary tissue and stem cell-derived in vitro cultures to validate the molecular profiles and differences in injury response between EA and AA cells at the single-cell level (Aim 1), with genetic manipulation of GSTT2 to determine direct mediation of protection against injury (Aim 2), and a high-throughput unbiased characterization of injury response coupled with a drug screen to determine compounds that will inhibit bile/acids injury (Aim 3). Dr. Ferrer-Torres’ primary research goals in the K99.R00 program is to develop high throughput techniques that will allow her to study ancestrally diverse populations and their response to injury. Therefore, the K99 phase has been planned to train in stem cell genetic modifications and high-content phenotypic-based drug discovery. For this mentored phase, Dr. Ferrer-Torres will work with Dr. Jason Spence and co-mentor by Dr. Jonathan Sexton. The mentored K99 program has been designed for Dr. Ferrer-Torres’ gain expertise in these areas. In addition, Dr. Jules Lin and Dr. Marcia Cruz-Correa will serve as advisory postdoctoral committee members and advisors for clinical immersions. This will be carried out utilizing the exceptional resources available at the University of Michigan. This will impulse Dr. Ferrer-Torres’ goals and help her establish her independent research program focusing on racial disparities in esophageal diseases in her R00 phase.