Investigation of the protective effects of polyphenols on polychlorinated biphenyl-induced inflammation and oxidative stress: impact of biotransformation

多酚对多氯联苯诱导的炎症和氧化应激的保护作用的研究：生物转化的影响

• 批准号: RGPIN-2014-06291
• 负责人: Kubow, Stan
• 金额: $ 3.79万
• 依托单位: McGill University
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2016
• 资助国家: 加拿大
• 起止时间: 2016-01-01 至 2017-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=612143
• 关键词: Investigation; protective; effects; polyphenols; polychlorinated

Background: PCB exposure is linked with neurologic damage in offspring of PCB-exposed women, cognitive disorders in children and poorer memory in adults. Despite food as the main source of PCBs, little is known about direct effects of PCBs on the gut cells. The role of gut microflora in PCB biotransformation is sparsely studied. PCB toxicity mediated via oxidative stress might be protected by dietary polyphenols via an antioxidant response induced via the nuclear factor erythroid 2-related factor 2 (Nrf2). Polyphenols, however, are poorly absorbed until they undergo gut microbial biotransformation that leads to more bioavailable secondary metabolites. Polyphenol metabolites may alter metabolism of PCBs and subsequent conjugation pathways that can modulate PCB toxicity. A limitation of existing mechanistic studies using human cell culture studies is the direct use of PCBs or polyphenols on human intestinal, hepatic and neuronal cells without exposure of such compounds to intermediary digestive or metabolic processes. Research Progress: PCBs and polyphenols separately underwent human simulated in vitro digestion using a Computer Controlled Dynamic Human Gastrointestinal Model (SHGI model) that mimics the different stages of digestion throughout the human GI tract. This model includes the use of colonic vessels containing human microbiota obtained from human fecal samples. Using LC-MS and HPLC, we showed that the common dietary polyphenols led to differing polyphenol metabolite profiles and antioxidant capacities in the ascending, transverse and descending colon vessels of the GI model. We developed Caco-2/HepG2 cell co-cultures to mimic human intestinal and hepatic first pass metabolism. After the GI model colonic digests of the polyphenols were subjected to the biotransformation and absorptive processes of the co-cultures, we detected microbial polyphenol by-products previously noted in plasma of humans ingesting polyphenols. Two structurally different PCB congeners, dioxin-like (PCB-126) and non-dioxin-like PCB (PCB-153), were separately subjected to the GI model as they are dominant congeners in the human diet and in human plasma. The LC-MS profiles of the PCB metabolites generated from the GI model and co-cultures and the human microbiota profiles seen in the GI model after the polyphenol and PCB exposures are being studied. Objectives: The overall objectives are to combine the GI model with the simulated first pass metabolism co-culture system to: (1) identify biotransformed metabolites of PCBs in the study of mechanisms of PCB-induced toxicity on human intestinal, hepatic and neural cells; and (2) assess the effects of polyphenol microbial metabolites on PCB metabolism and toxicity with a focus on PCB metabolites, oxidative stress, inflammation and the Nrf2 signalling pathway. Experimental Design: Human intestinal, hepatic and neuronal cells will be exposed to parent PCBs and polyphenols and their metabolites generated by GI model and first pass metabolism system. PCB and polyphenol metabolites will be assessed via LC-MS and HPLC. Cells will be pre-treated with the identified polyphenol metabolites to test for their effects on PCB-mediated metabolism and toxicity. Neural effects will be tested on human neural progenitor cells co-cultured onto a Multiwell insert system with the human brain endothelial capillary cell line hCMEC/D3 as a model for the human blood brain barrier. Cell viability, intracellular reactive oxygen species accumulation and inflammatory cytokines will be measured. Nrf2 levels will be assessed via Western blot. Significance: This study will provide new insights of PCB-induced inflammation and oxidative-stress as mediated by gut metabolism and interactions with with dietary polyphenols.

背景资料：多氯联苯接触与多氯联苯接触妇女的后代的神经系统损伤、儿童的认知障碍和成人的记忆力下降有关。尽管食物是多氯联苯的主要来源，但对多氯联苯对肠道细胞的直接影响知之甚少。肠道微生物区系在PCB生物转化中的作用研究很少。通过氧化应激介导的PCB毒性可能受到膳食多酚的保护，通过核因子红细胞2相关因子2（Nrf 2）诱导的抗氧化反应。然而，多酚类化合物的吸收很差，直到它们经历肠道微生物生物转化，产生更多的生物可利用的次级代谢产物。多酚代谢物可能会改变PCB的代谢和随后的共轭途径，可以调节PCB的毒性。使用人类细胞培养研究的现有机制研究的局限性是在人类肠、肝和神经元细胞上直接使用多氯联苯或多酚，而不将此类化合物暴露于中间消化或代谢过程。 研究进展：使用计算机控制的动态人类胃肠道模型（SHGI模型）分别对多氯联苯和多酚进行人体模拟体外消化，该模型模拟整个人类胃肠道消化的不同阶段。该模型包括使用含有从人类粪便样品获得的人类微生物群的结肠血管。使用LC-MS和HPLC，我们表明，常见的膳食多酚导致不同的多酚代谢物的配置文件和抗氧化能力的升，横和降结肠血管的GI模型。我们开发了Caco-2/HepG 2细胞共培养物以模拟人肠和肝首过代谢。在对多酚的GI模型结肠消化物进行共培养物的生物转化和吸收过程之后，我们检测到先前在人类摄入多酚的血浆中注意到的微生物多酚副产物。两种结构不同的多氯联苯同系物，二恶英类（PCB-126）和非二恶英类多氯联苯（PCB-153），分别受到GI模型，因为它们是人类饮食和人类血浆中的主要同系物。正在研究GI模型和共培养物产生的PCB代谢物的LC-MS谱以及多酚和PCB暴露后GI模型中观察到的人体微生物群谱。 目的：本研究的总体目标是将GI模型与模拟首过代谢共培养系统相结合，以：（1）在研究多氯联苯对人体肠道、肝脏和神经细胞毒性机制中，鉴定多氯联苯的生物转化代谢物;以及（2）评估多酚微生物代谢物对PCB代谢和毒性的影响，重点是PCB代谢物，氧化应激，炎症和Nrf 2信号通路。 实验设计：人体肠道、肝脏和神经细胞将暴露于母体多氯联苯和多酚及其通过GI模型和首过代谢系统产生的代谢产物中。PCB和多酚代谢物将通过LC-MS和HPLC进行评估。将用鉴定的多酚代谢物预处理细胞，以测试它们对PCB介导的代谢和毒性的影响。将在与人脑内皮毛细血管细胞系hCMEC/D3（作为人血脑屏障模型）共培养到多孔插入系统上的人神经祖细胞上测试神经效应。将测量细胞活力、细胞内活性氧积聚和炎性细胞因子。Nrf 2水平将通过蛋白质印迹法评估。 重要性：本研究将提供新的见解多氯联苯诱导的炎症和氧化应激介导的肠道代谢和与膳食多酚的相互作用。