Pathogenesis of Diabetic Nephropathy

摘要 糖尿病肾病 (DN) 的特点是代谢和细胞信号传导事件紊乱，导致 增加ECM的合成。它们包括葡萄糖中间体和多元醇的增强通量、畸变 燃料传感分子，例如 AMPK；并增加 PKC 活性、有害 AGE 的产生、 MAP/ERK 激酶和 Smad 蛋白、促纤维细胞因子和活性氧 (ROS) 的产生。 后者被认为是 DN 发病机制的核心。这些信号事件已被研究 肾小球细胞和肾小管或间质细胞的信息有限。有趣的是，肾小管间质的变化 与肾功能参数紊乱更好地相关；因此，有合理的需要来定义 DN 生物学与“糖尿病肾小管病”有关。除了影响的大量信号事件之外 肾小球的病理学，最近发现的葡萄糖醛酸-木酮糖 (G-X) 途径，显然与 特别是在肾小管室中发生的糖尿病肾病很少受到关注。 G-X 途径的事件由肌醇加氧酶 (MIOX) 启动，MIOX 是一种管状酶，具有增加的 DN 中的表达。在 G-X 事件期间，NADPH:NADP 和 NAD:NADH 比率存在严重扰动， 因此，存在极大程度的“氧化还原失衡”和“NAD 缺乏”，从而导致相应的后果 肾小管稳态不良（JASN 2015，JBC 2016-1）。由于 MIOX 促进剂包括碳水化合物、氧化剂/ 抗氧化、渗透和甾醇反应元素可预期 MIOX 的循环刺激如下 高血糖、血脂和化学氧化应激（JBC-2011、JBC 2016-2、JASN-2017）以及持续的 MIOX 的上调和 ROS 的产生。它的调节也受到表观遗传修饰的调节 （AJP 2017）。考虑到上述考虑，我们希望探讨其中涉及的机制 使用各种转基因 MIOX 小鼠模型研究肾小管间质的生物学。目标 I 是描绘 使用高血糖小鼠模型调节 MIOX 表达的各种表观遗传机制 高脂血症。 MIOX 启动子的 DNA 甲基化/去甲基化、组蛋白甲基化/去甲基化和 将研究 MIOX-TG 和 MIOX-KO 小鼠中的乙酰化/去乙酰化，并将其与乙酰化程度相关联。 肾小管间质损伤。 AIM II 旨在描述 MIOX-TG 中加重肾小管间质损伤的机制 与高血糖和 AGE 超载期间的 WT 或 -KO 小鼠相比。肾功能、细胞氧化还原的干扰， 将研究线粒体动力学。救援实验将包括秋田犬与 MIOX-/- 的杂交 小鼠和评价的反映损伤改善的各种参数。目标 III 是描述机制 与 WT 和 KO 小鼠相比，高脂血症中 MIOX-TG 的肾损伤加重。 MIOX 结束后的事件 表达，即 NAD 缺乏、谷胱甘肽消耗和 ROS 生成增加、扰动 Sirtuins 的活性、p-AMPK、PGC-1α、线粒体动力学、ER 应激和肾小管间质纤维化将被 评估。救援实验将包括 MIOX-/- 与 ob/ob 和 PPARαΔob/ob 小鼠的杂交。

ABSTRACT Diabetic nephropathy (DN) is characterized by disturbances in metabolic & cellular signaling events leading to increased synthesis of ECM. They include accentuated flux of glucose intermediaries & polyols, aberrations in fuel sensing molecules, e.g., AMPK; and increased PKC activity, generation of nocuous AGEs, expression of MAP/ERK kinases and Smad proteins, profibrogenic cytokines & production of reactive oxygen species (ROS). The latter are regarded as central to the pathogenesis of DN. These signaling events have been studied in glomerular cells and information for tubular or interstitial cells is limited. Interestingly, tubulointerstitial changes correlate better with derangement in renal functional parameters; thus there is a legitimate need to define the biology of DN with respect to “diabetic tubulopathy”. In addition to a multitude of signaling events that affect the pathology of the glomerulus, a recently discovered glucuronate-xylulose (G-X) pathway, apparently relevant to diabetic nephropathy that is operative specifically in the tubular compartment, has received very little attention. Events of G-X pathway are initiated by myo-inositol oxygenase (MIOX), a tubular enzyme that has an increased expression in DN. During G-X events there are severe perturbations in NADPH:NADP+ & NAD+:NADH ratios, as a result there is a tremendous degree of "redox imbalance" and "NAD+ deficiency" leading to consequential adverse tubular homeostasis (JASN 2015, JBC 2016-1). Since MIOX promoter includes carbohydrate, oxidant/ antioxidant, osmotic and sterol response elements a cyclic stimulation of MIOX would be anticipated following hyperglycemia, lipidemia and chemical oxidant stress (JBC-2011, JBC 2016-2, JASN-2017) along with sustained up-regulation of MIOX and generation of ROS. Its regulation is also modulated by epigenetic modifications (AJP 2017). Keeping in perspective the above considerations we wish to explore the mechanisms involved in the biology of tubulo-interstitium using various genetically modified MIOX mice models. AIM I is to delineate various epigenetic mechanisms that modulate MIOX expression, using mice models of hyperglycemia and hyperlipidemia. DNA methylation/demethylation of MIOX promoter, histone methylation/demethylation and acetylation/decetylation in MIOX-TG and MIOX-KO mice will be investigated and correlated with the extent of tubulointerstitial injury. AIM II is to delineate mechanisms that accentuate tubulointerstitial injury in MIOX-TG vs WT or -KO mice during hyperglycemia and AGEs' overload. Perturbations in renal functions, cellular redox, mitochondrial dynamics will be investigated. Rescue experiments will include cross-breeding Akita with MIOX-/- mice and various parameters reflecting amelioration of injury appraised. AIM III is to delineate mechanisms that augment renal injury in MIOX-TG vs WT & KO mice in hyperlipidemia. The events following MIOX over- expression, i.e., NAD+ deficiency, glutathione depletion and augmented generation of ROS, perturbations in sirtuins' activity, p-AMPK, PGC-1α, mitochondrial dynamics, ER stress and tubulo-interstitial fibrosis will be assessed. Rescue experiments will include cross-breeding of MIOX-/- with ob/ob and PPARαΔob/ob mice.