Diabetogenic Mine Tailings: Mechanistic Link Between Arsenic, NRF2, Autophagy, and Diabetes

总结(项目1：Donna Zhang) 含金属有害物质对土壤和水的污染，特别是在矿场附近 尾矿和冶炼厂，导致附近社区长期暴露在有毒金属混合物中，构成了 严重的健康问题。根据有毒物质疾病登记局的数据， 与这些地点的尾矿有关的一种污染物是有毒的类金属砷(As)。流行病学 研究表明，无论是通过饮用水还是通过饮用水，慢性砷暴露都与 或食物，糖尿病发病率增加。因此，暴露在含砷尾矿中，这可能 导致吸入或摄入砷，可能是导致接触砷者患病风险增加的重要因素 社区。重要的是，尽管已知对健康的严重影响，但其分子机制 含砷尾矿对糖尿病表型的促进作用尚未阐明。此前，我们 报道称，低剂量、与环境相关的砷阻止自噬，是细胞降解的关键 对维持蛋白质平衡至关重要的途径。此外，我们已经证明，自噬功能障碍会导致 在长时间激活关键的抗氧化剂转录因子NRF2。通常维持在较低水平 通过Keap1介导的泛素化和26S蛋白酶体的降解，NRF2在 通过氧化修饰Keap1(Keap1-C151依赖，规范)或隔离 在AS诱导的自噬功能障碍期间，Keap1进入自噬小体(p62依赖，非规范)。而当 通过Keap1-C151依赖的规范机制控制的Nrf2激活是保护性的，延长了 AS暴露中依赖p62的NRF2的非规范激活导致细胞功能障碍和组织 损坏，表明NRF2的“阴暗面”。我们假设含砷的尾矿能促进 糖尿病通过依赖p62的、长时间激活的Nrf2。这一假设得到了我们的 初步数据显示，暴露于AS的野生型(WT)小鼠表现出糖耐量和 增强胰岛素抵抗，这在Nrf2-/-、p62-/-或Nrf2-/-p62-/-小鼠中没有观察到。我们最近的RNAseq 暴露于砷20周的小鼠的肝脏产生的数据也显示出显著的变化 葡萄糖、胰岛素、胆固醇和脂类代谢相关基因的表达。在此应用程序中，我们将 通过以下方法验证我们的假设：1)表征慢性前列腺癌的时间和剂量依赖性糖尿病的潜在致病作用 WT小鼠暴露于饮用水或尾矿中的砷颗粒物(PM10)(目标1)；2)测定 延长NRF2激活在推动AS诱导的糖尿病相关细胞代谢重编程中的作用 LINES(目标2)；和3)体内确认AS和延长的NRF2诱导的重要分子改变 促进糖尿病的活动(目标3)。对砷引起的改变的机械性理解 事实证明，糖尿病在诊断、预防和治疗策略的产生中具有极其重要的价值 适用于接触含砷尾矿的人群和有砷暴露风险的人群。

SUMMARY (Project 1: Donna Zhang) Contamination of soil and water by metal-containing hazardous substances, particularly at sites near mine tailings and smelters, has led to chronic exposure of nearby communities to toxic metal mixtures, posing a serious health problem. Based on data from the Agency for Toxic Substances Disease Registry, the number one contaminant associated with mine tailings at these sites is the toxic metalloid arsenic (As). Epidemiological studies have demonstrated a positive correlation between chronic As exposure, either through drinking water or food, with an increased incidence of diabetes. Thus, exposure to As-containing mine tailings, which could result in inhalation or ingestion of As, may be a significant contributor to enhanced risk of disease in exposed communities. Importantly, despite the known severity of the health effects, the molecular mechanisms by which As-containing mine tailings enhance diabetic phenotypes have not yet been elucidated. Previously, we reported that low, environmentally relevant doses of arsenic block autophagy, a key cellular degradation pathway critical to maintaining proteostasis. Furthermore, we have shown that autophagic dysfunction results in prolonged activation of the key antioxidant transcription factor NRF2. Normally maintained at low levels through KEAP1-mediated ubiquitination and degradation by the 26S proteasome, NRF2 is upregulated at the protein level via oxidative modification of KEAP1 (KEAP1-C151 dependent, canonical) or sequestration of Keap1 into autophagosomes during As-induced autophagy dysfunction (p62-dependent, non-canonical). While controlled Nrf2 activation through the Keap1-C151 dependent canonical mechanism is protective, prolonged p62-dependent non-canonical activation of NRF2 during As exposure causes cellular dysfunction and tissue damage, indicative of a “dark side” to NRF2. We hypothesize that As-containing mine tailings promote diabetes through p62-dependent, prolonged activation of Nrf2. This hypothesis is supported by our preliminary data indicating that wild type (WT) mice exposed to As showed impaired glucose tolerance and enhanced insulin resistance, which was not observed in Nrf2-/-, p62-/-, or Nrf2-/-p62-/- mice. Our recent RNAseq data generated from the liver of mice exposed to As for 20 weeks also showed significant changes in the expression of genes involved in glucose, insulin, cholesterol, and lipid metabolism. In this application, we will test our hypothesis by: 1) characterizing the time and dose-dependent diabetogenic potential of chronic exposure to As in drinking water or mine tailing As-particles (PM10) in WT mice (Aim 1); 2) determining the role of prolonged NRF2 activation in driving As-induced metabolic reprogramming in diabetes-relevant cell lines (Aim 2); and 3) in vivo confirmation of important molecular alterations induced by As and prolonged NRF2 activity in promoting diabetes (Aim 3). A mechanistic understanding of arsenic-mediated alterations that lead to diabetes will prove extremely valuable in the generation of diagnostic, preventive, and therapeutic strategies for populations exposed to As-containing mine tailings and populations at risk of arsenic exposure.