Autophagy in Polycystic Kidney Disease (PKD)

多囊肾病 (PKD) 中的自噬

• 批准号: 10266045
• 负责人: CHARLES Louis EDELSTEIN
• 金额: --
• 依托单位: VA EASTERN COLORADO HEALTH CARE SYSTEM
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-07-01 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10266045
• 关键词: 3-methyladenine; Apoptosis; Apoptotic; Autophagocytosis; Autophagosome; Autosomal Dominant Polycystic Kidney; Caloric Restriction; Cells; Chemicals; Chloroquine; Chronic Kidney Failure; Clinical Research; Cyst; Data; Defect; Diagnosis; Dominant-Negative Mutation; Epithelial Cells; Eukaryotic Cell; Future; Genetic; Genetic Techniques; Glucose; Growth; Healthcare; Hereditary Disease; Homeostasis; Human; Hypertension; Intervention; Kidney; Kidney Transplantation; Knock-out; Knockout Mice; Lead; Life; Lysosomes; Membrane; Methods; Mitochondria; Mus; Organelles; Pathway interactions; Patients; Peptides; Pharmacology; Play; Polycystic Kidney Diseases; Process; Proprotein Convertase 1; Proteins; Protocols documentation; Publishing; Recycling; Renal function; Role; Techniques; Testing; Therapeutic; Time; Trehalose; Tubular formation; United States Department of Veterans Affairs; Veterans; care burden; deprivation; detection method; experimental study; improved; in vivo; inhibition of autophagy; insight; new therapeutic target; novel; prevent; protective effect

Autophagy is a process that takes place in all eukaryotic cells that keeps cells alive under stressful conditions. In autophagy there is the sequestration of damaged organelles into double-membraned autophagosomes that subsequently fuse with lysosomes where their cargoes are delivered for degradation and recycling. In the healthy kidney, autophagy plays an important role in the homeostasis and viability of renal tubular epithelial cells. There is indirect evidence that PKD is a case of suppressed autophagy: 1) Many of the agents that protect against PKD are autophagy inducers. 2) Preliminary/published data demonstrate decreased autophagic flux in Pkd1 -/- mouse kidneys and PC1 -/- cells. 3) There is cross-talk between autophagy and apoptosis, a key process in cyst growth. The overall hypothesis is that autophagic flux (autophagosome-lysosome fusion and degradation) is decreased early in the polycystic kidney and that interventions that block autophagy will worsen PKD and conversely autophagy inducers like caloric restriction, 2-deoxy-glucose or trehalose will increase autophagy, decrease apoptosis and improve PKD in an autophagy–dependent manner. The presence of autophagy (autophagic flux, presence of autophagosomes and autolysosomes), mitophagy (autophagosomes containing mitochondria) and the effect of autophagy inhibition (using genetic techniques), autophagy induction (using autophagy-inducers that are known to protect against PKD) on cyst growth and apoptosis will be determined in Pkd1 -/- mice and PKD1 -/- cells. An improved understanding of the mechanism by which induction of autophagy can prevent PKD may lead to the identification of new targets for both diagnosis and therapy of PKD. In Aim 1, the effect of autophagy inhibition or induction in will be tested Pkd1 -/- mouse kidneys. Autophagic flux and apoptosis will be determined during a time course in Pkd1 -/- mouse kidneys. It will be determined whether autophagy knockout in kidney- specific double knockout ATG7, Pkd1 -/- mice worsens apoptosis, proliferation, PKD and kidney function. The effect of autophagy inducers (caloric restriction, 2-deoxy-glucose or trehalose) on autophagy, apoptosis and proliferation, PKD and renal function will be determined in Pkd1 -/- mice and ATG7, Pkd1 double knockout mice. The effect of a novel autophagy inducer (trehalose) on PKD will be determined. In Aim 2, pathways of autophagy in PKD1 -/- tubular cells will be determined. These experiments will use more sophisticated techniques of detection of autophagic flux and mitophagy and pharmacological and genetic techniques of autophagy induction and inhibition that cannot be performed in vivo. The effect of autophagy induction by mechanistically distinct methods (glucose deprivation, trehalose, Beclin1 peptide) and autophagy inhibition (3 methyladenine, chloroquine, expression of dominant negative ATG4) on autophagy pathways and flux, mitophagy, apoptosis, proliferation, cyst size/number will be determined. Our results will provide a definitive test of the hypothesis that PKD represents a case of decreased autophagy. Our results will offer mechanistic insights into the mechanism of action of caloric restriction or chemical caloric restriction that protect against PKD and that are also autophagy inducers. Our results will provide therapeutic insights into the potential future use of autophagy inducers in PKD.

自噬是发生在所有真核细胞中的一个过程，它使细胞在压力条件下保持存活。在 自噬是将受损的细胞器隔离成双膜自噬体， 随后与溶酶体融合，在那里它们的货物被递送用于降解和再循环。在健康 在肾脏中，自噬在肾小管上皮细胞的稳态和活力中起重要作用。有 间接证据表明PKD是一种抑制自噬的情况：1）许多防止PKD的药物是 自噬诱导剂2)初步/已发表的数据表明Pkd 1-/-小鼠自噬通量降低 肾脏和PC 1-/-细胞。3)自噬和细胞凋亡之间存在相互作用，这是囊肿生长的关键过程。 总体假设是自噬通量（自噬体-溶酶体融合和降解）减少 在多囊肾早期，阻断自噬的干预措施会使PKD恶化，反之亦然。 自噬诱导剂如热量限制、2-脱氧葡萄糖或海藻糖将增加自噬，减少自噬。 细胞凋亡并以自噬依赖的方式改善PKD。自噬（autophagic flux， 自噬体和自溶酶体的存在）、线粒体自噬（含有线粒体的自噬体）和 自噬抑制（使用遗传技术）、自噬诱导（使用 已知保护免受PKD）对囊肿生长和细胞凋亡的影响将在Pkd 1-/-小鼠和PKD 1-/-小鼠中测定。 细胞对诱导自噬预防PKD的机制的进一步理解可能导致 为PKD的诊断和治疗确定新的靶点。在目标1中，自噬的作用 将在Pkd 1-/-小鼠肾脏中测试抑制或诱导。将测定自噬通量和凋亡 在Pkd 1-/-小鼠肾脏中的时间过程中。它将确定是否自噬敲除在肾脏- 特异性双基因敲除ATG 7、Pkd 1-/-小鼠的凋亡、增殖、PKD和肾功能。的 自噬诱导剂（热量限制、2-脱氧葡萄糖或海藻糖）对自噬、细胞凋亡和 将在Pkd 1-/-小鼠和ATG 7、Pkd 1双敲除小鼠中测定增殖、PKD和肾功能。 将确定新型自噬诱导剂（海藻糖）对PKD的影响。在目标2中，自噬途径 在PKD 1-/-肾小管细胞中的表达将被确定。这些实验将使用更复杂的探测技术 自噬通量和线粒体自噬的药理学和遗传技术以及自噬诱导和 不能在体内进行的抑制。不同机制诱导自噬的效果 （葡萄糖剥夺，海藻糖，Beclin 1肽）和自噬抑制（3-甲基腺嘌呤，氯喹， 显性负性ATG 4）在自噬途径和流量、线粒体自噬、凋亡、增殖、囊肿 数量/数量将被确定。我们的研究结果将提供一个明确的测试的假设，PKD代表一个 自噬减少的病例。我们的研究结果将为热量的作用机制提供机械的见解 限制或化学热量限制，防止PKD，也是自噬诱导剂。我们 这些结果将为自噬诱导剂在PKD中的潜在未来应用提供治疗见解。