Role of CNPase in TBI

University Of Pittsburgh At Pittsburgh, Pittsburgh PA

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Abstract

DESCRIPTION (provided by applicant): The term "cAMP" is used universally to refer to 3',5'-cyclic adenosine monophosphate (3',5'- cAMP), the famous "second messenger" discovered by Dr. Earl Southerland. Importantly, the vast majority of investigators measure 3',5'-cAMP using various commercially available assay kits, and our group is the ONLY group in the world that routinely measures 3',5'-cAMP using high performance liquid chromatography-tandem mass spectrometry(LC-MS/MS). Dr. Jackson's lab focuses mostly on the role of purines in renal physiology/pharmacology, and routinely employs LC-MS/MS, rather than assay kits, to measure 3',5'-cAMP. In experiments unrelated to the brain, Dr. Jackson's lab serendipitously discovered that the kidney produces more 2',3'-cAMP (a positional isomer of 3',5'- cAMP) than 3',5'-cAMP. Dr. Kochanek's lab focuses primarily on mechanisms of traumatic brain injury (TBI), and has discovered that purines play a major role in protecting the brain from TBI. Because of Dr. Jackson's expertise in measuring purines and Dr. Kochanek's interest in the role of purines in TBI, a natural and productive collaboration evolved between these investigators. After the serendipitous discovery of 2',3'-cAMP in kidneys, Drs. Jackson and Kochanek decided to investigate whether 2',3'-cAMP exists in the brain. In preliminary studies, LC-MS/MS analysis of 44 samples of cerebral spinal fluid (CSF) from TBI patients showed that 2',3'-cAMP is a major constituent of "cAMP" in CSF from patients with TBI. In addition to 2',3'-cAMP, the levels of 2'-AMP, adenosine and inosine (metabolite of adenosine) were also measured in these same samples. Importantly, there was an large and significant correlation between 2',3'-cAMP and 2'-AMP, 2'-AMP and adenosine and 2'-AMP and inosine in human CSF. These surprising discoveries suggested a critical need to explore the role of 2',3'-cAMP in TBI pathophysiology, particularly with respect to the role of 2',3'-cAMP as a precursor for adenosine. Accordingly, the purpose of this exploratory project is to begin to characterize the significance of 2',3'-cAMP in TBI by testing the innovative concept that 2',3'-cAMP is involved in a "CNPase Neuroprotection Mechanism" in which brain injury leads to release of 2',3'-cAMP from mRNA, and 2',3'-cAMP is then metabolized to 2'-AMP by CNPase followed by conversion of 2'-AMP to adenosine. PUBLIC HEALTH RELEVANCE: TBI is a major source of mortality, morbidity and life-long impairment in both the civilian and military populations, with limited treatment options and generally dismal outcomes. The CNPase Neuroprotective Mechanism could be extremely important in producing extracellular adenosine during TBI, thus providing this protective, "retaliatory" metabolite to mitigate cellular damage. Manipulation of this mechanism by drugs could provide novel approaches to prevent and treat TBI.