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

肾脏 2,3-cAMP-腺苷途径

• 批准号: 8479345
• 负责人: EDWIN Kerry JACKSON
• 金额: $ 31.8万
• 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/8479345
• 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.

描述（由申请人提供）：在2008年，我们发现完整的肾脏产生了大量的2'，3'-camp（位置异构体3'，5'-camp），2'，3'-camp源自mRNA降解，源于能量耗尽，大量增加了肾脏2'，3'-tamp Biosynthessess。这是任何器官，组织或细胞中内源性2'，3'-训练的第一个文献。这一令人兴奋的发现引发了尤里卡时刻：2'，3'-cAMP可能是一个未被发现但至关重要的细胞外腺苷来源！现在，我们已经证实了这一假设，我们指的是2'，3'-camp的生产，然后通过2'-Amp和3'-Amp转换为腺苷，为2'，3'- 3'-营地 - 腺苷途径。该应用的重点是2'，3'-循环核苷酸-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转换为2'-AMP的酶，并且是唯一已知促进该反应的酶。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'-训练室打开了线粒体通透性转变孔（MPTPS），众所周知，MPTPS的开放会导致细胞凋亡和坏死。此外，许多研究表明，腺苷通过特定的受体保护肾脏免受缺血/再灌注损伤。因此，CNPase应该是重新保护的，因为它可能会降低细胞内TOXN（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2'，3'-cAMP和腺苷的肾脏水平中的重要性。我们将使用13C标记的内标进行对2'，3'-汤匙代谢组的LC-MS/MS分析来检查：1）由CNPase-// - /cnpase cnpase cnpase-casse cnpase-classe cnpase-+和cnpase的外源性2'，3'-camp至2'-amp，3'-amp，3'-amp，3'-amp，3'-amp，腺苷，腺苷和inosine的代谢； 2）内源性（能量耗尽）的代谢2'，3'-训练到2'-Amp，3'-Amp，3'-Amp，腺苷，腺苷和肌苷，由CNPase - / - ，CNPase-cnpase-，/+和CNPase-/+和CNPase+/cnpase+/+/+小鼠由完整的肾脏和肾细胞。如果CNPase参与2'，3'-cAMP的肾脏代谢，则应反映在外源性和内源性2'，3'-cAMP的肾脏代谢中。最后，我们将量化来自CNPase -/ - ，CNPase-/+和CNPase+/+小鼠的肾脏缺血/再灌注损伤引起的肾脏损伤。如果CNPase参与了2'训练，3'-cAMP或增加细胞外腺苷的脱落细胞，则应在CNPase缺乏的肾脏中加剧肾脏损伤。我们的假设具有几种至关重要的含义：1）肾脏CNPase可能是保护肾脏免受侮辱的关键酶； 2）这种酶（由于疾病或药物）的故障可以解释为什么某些患者会因肾脏损伤而患上急性或慢性肾衰竭； 3）2'，3'-营地腺苷途径的药理或分子操作可能会提供一种有效的方法来降低急性和慢性肾衰竭的风险，昂贵，普遍和生活的疾病。