The Renal 2',3'-cAMP-Adenosine Pathway

肾脏 2,3-cAMP-腺苷途径

• 批准号: 8282204
• 负责人: EDWIN Kerry JACKSON
• 金额: $ 32.95万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-06-15 至 2017-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8282204
• 关键词: 2' -adenylic acid; Acute; Adenosine; Anabolism; Apoptosis; Biological Assay; Cells; Chronic Kidney Failure; Cyclic AMP; Cyclic Nucleotides; Disease; Documentation; Electrolytes; Enzymes; Excretory function; Glomerular Filtration Rate; Goals; Histopathology; Inflammation; Injury; Inosine; Ischemia; Isomerism; Kidney; Knockout Mice; Label; Life; Measures; Metabolism; Molecular; Mus; Necrosis; Organ; Pathway interactions; Patients; Pharmaceutical Preparations; Production; Proteinuria; Reaction; Renal Blood Flow; Renal function; Reperfusion Injury; Reporting; Risk; Role; Sampling; Source; Structure; Time; Tissues; Toxin; United States; Urine; Venous; Wild Type Mouse; extracellular; in vivo; indexing; kidney cell; mRNA Transcript Degradation; metabolic poison; mitochondrial permeability transition pore; phosphoric diester hydrolase; receptor; renal ischemia; response

DESCRIPTION (provided by applicant): In 2008, we discovered that intact kidneys produce large quantities of 2',3'-cAMP (a positional isomer of 3',5'-cAMP), that 2',3'-cAMP derives from mRNA degradation and that energy depletion massively increases renal 2',3'-cAMP biosynthesis. This was the FIRST documentation of endogenous 2',3'-cAMP in any organ, tissue or cell. This exciting discovery triggered an eureka moment: 2',3'-cAMP may be an unrecognized, but critically important, source of extracellular adenosine! We have now confirmed this hypothesis, and we refer to the production of 2',3'-cAMP, followed by its conversion to adenosine, via 2'-AMP and 3'-AMP, as the 2',3'- cAMP-Adenosine Pathway. The focus of this application is the role of 2',3'-cyclic nucleotide-3'- phosphodiesterase (CNPase) in the renal 2',3'-cAMP-adenosine pathway. CNPase is an enzyme that can convert 2',3'-cAMP to 2'-AMP and is the ONLY enzyme known to promote this reaction. Why do we think that renal CNPase is important? A recent report demonstrates that intracellular 2',3'-cAMP opens mitochondrial permeability transition pores (mPTPs), and it is well known that opening of mPTPs leads to apoptosis and necrosis. Moreover, many studies demonstrate that adenosine, via specific receptors, protects the kidney from ischemia/reperfusion injury. Therefore, CNPase should be renoprotective because it likely reduces the intracellular levels of an intracellular toxn (2',3'-cAMP) and concomitantly increases the extracellular levels of a protective metabolite (adenosine). The main goal of the present proposal is to quantify the importance of renal CNPase in regulating renal levels of 2',3'-cAMP and adenosine. We will employ a LC-MS/MS assay for the 2',3'-cAMP metabolome using a 13C-labeled internal standard to examine: 1) the metabolism of exogenous 2',3'-cAMP to 2'-AMP, 3'-AMP, adenosine and inosine by intact kidneys and renal cells from CNPase -/-, CNPase -/+ and CNPase +/+ mice; and 2) the metabolism of endogenous (triggered by energy depletion) 2',3'-cAMP to 2'-AMP, 3'-AMP, adenosine and inosine by intact kidneys and renal cells from CNPase -/-, CNPase -/+ and CNPase +/+ mice. If CNPase is involved in the renal metabolism of 2',3'-cAMP, then this should be reflected in the renal metabolome of exogenous and endogenous 2',3'-cAMP. Finally, we will quantify renal damage induced by ischemia/reperfusion injury in kidneys from CNPase -/-, CNPase -/+ and CNPase +/+ mice. If CNPase is involved in ridding cells of 2',3'-cAMP or increasing extracellular adenosine, then renal injury should be exacerbated in kidneys deficient in CNPase. Our hypothesis has several critically important implications: 1) Renal CNPase may be a key enzyme in protecting the kidneys from insults; 2) Malfunction of this enzyme (because of disease or drugs) could explain why some patients develop acute or chronic renal failure in response to renal injury; and 3) Pharmacological or molecular manipulation of the 2',3'- cAMP-adenosine pathway may offer an effective approach to reducing the risk of acute and chronic renal failure, diseases which are costly, prevalent and life altering. PUBLIC HEALTH RELEVANCE: We hypothesize that 2',3'-cyclic nucleotide-3'-phosphodiesterase (CNPase) protects the kidneys from injury. Our hypothesis, if correct, has several critically important implications: 1) renal CNPase should be viewed as a key enzyme in protecting the kidneys from insults; 2) malfunction of this enzyme (because of disease or drugs) could explain why some patients develop acute or chronic renal failure in response to renal injury; and 3) pharmacological or molecular manipulation of the 2',3'-cAMP-adenosine pathway may offer an effective approach to reducing the risk of acute and chronic renal failure, diseases which are costly, prevalent and life altering.

描述（申请人提供）：2008年，我们发现完整的肾脏产生大量的2',3'-cAMP（3',5'-cAMP的位置异构体），2',3'-cAMP来源于mRNA 降解和能量消耗会大量增加肾脏 2',3'-cAMP 生物合成。这是任何器官、组织或细胞中内源 2',3'-cAMP 的第一个记录。这一激动人心的发现引发了一个灵光一现的时刻：2',3'-cAMP 可能是一种未被认识但极其重要的细胞外腺苷来源！我们现在已经证实了这一假设，我们将 2',3'-cAMP 的产生，然后通过 2'-AMP 和 3'-AMP 转化为腺苷，称为 2',3'-cAMP-腺苷途径。该应用的重点是 2',3'-环核苷酸-3'-磷酸二酯酶 (CNPase) 在肾脏 2',3'-cAMP-腺苷途径中的作用。 CNPase 是一种可以将 2',3'-cAMP 转化为 2'-AMP 的酶，也是已知唯一能促进该反应的酶。为什么我们认为肾脏 CNPase 很重要？最近的一份报告表明，细胞内2',3'-cAMP会打开线粒体通透性转换孔（mPTP），众所周知，mPTP的打开会导致细胞凋亡和坏死。此外，许多研究表明，腺苷通过特定受体可以保护肾脏免受缺血/再灌注损伤。因此，CNPase 应该具有肾脏保护作用，因为它可能会降低细胞内毒素 (2',3'-cAMP) 的细胞内水平，并同时增加保护性代谢物（腺苷）的细胞外水平。本提案的主要目标是量化肾脏 CNPase 在调节肾脏 2',3'-cAMP 和腺苷水平中的重要性。我们将使用 13C 标记的内标对 2',3'-cAMP 代谢组进行 LC-MS/MS 测定，以检查：1) 外源 2',3'-cAMP 到 2'-AMP 的代谢，3来自 CNPase -/-、CNPase -/+ 和 CNPase +/+ 小鼠的完整肾脏和肾细胞的'-AMP、腺苷和肌苷； 2) 完整肾脏和肾细胞从 CNPase -/-、CNPase -/+ 代谢内源性（由能量消耗触发）2',3'-cAMP 为 2'-AMP、3'-AMP、腺苷和肌苷和 CNPase +/+ 小鼠。如果 CNPase 参与 2',3'-cAMP 的肾脏代谢，那么这应该反映在外源性和内源性 2',3'-cAMP 的肾脏代谢组中。最后，我们将量化 CNPase -/-、CNPase -/+ 和 CNPase +/+ 小鼠肾脏缺血/再灌注损伤引起的肾损伤。如果 CNPase 参与清除细胞中的 2',3'-cAMP 或增加细胞外腺苷，则缺乏 CNPase 的肾脏会加剧肾损伤。我们的假设有几个至关重要的意义：1）肾 CNP 酶可能是保护肾脏免受损伤的关键酶； 2）这种酶的功能障碍（由于疾病或药物）可以解释为什么一些患者因肾损伤而出现急性或慢性肾功能衰竭； 3) 2',3'-cAMP-腺苷途径的药理学或分子操作可以提供一种有效的方法来降低急性和慢性肾衰竭的风险，这些疾病是昂贵的、普遍的和改变生活的。 公共健康相关性：我们假设 2',3'-环核苷酸-3'-磷酸二酯酶 (CNPase) 可以保护肾脏免受损伤。我们的假设如果正确，将具有几个至关重要的意义：1）肾 CNPase 应被视为保护肾脏免受损伤的关键酶； 2）这种酶的功能障碍（由于疾病或药物）可以解释为什么一些患者因肾损伤而出现急性或慢性肾功能衰竭； 3) 2',3'-cAMP-腺苷途径的药理学或分子操作可以提供一种有效的方法来降低急性和慢性肾衰竭的风险，这些疾病是昂贵的、普遍的和改变生活的。