Urine Metabolomics Signatures in Reversible Cerebral Vasoconstriction Syndrome

Background: The pathophysiology of reversible cerebral vasoconstriction syndrome (RCVS) is unclear. An unbiased systems-based approach might help to illustrate the metabolite profiling and underlying pathophysiology. Methods: Urine samples were collected from RCVS patients and matched controls recruited in Taipei Veterans General Hospital. 1H-Nuclear magnetic resonance (1H-NMR) was used to initially explore the metabolic profile, and liquid chromatography tandem mass spectrometry (LC-MS/MS) was then used to identify metabolic alterations in RCVS. Untargeted metabolite screening was randomly performed on 10 RCVS patients and 10 control subjects in the discovery phase. The selected untargeted metabolites were further validated on 47 RCVS patients during their ictal stage (with 40 of them having remission samples) and 47 controls in the replication phase. Findings: Six metabolites-hippurate, citrate, 1,3,7-trimethyluric acid, ascorbic acid, D-glucurono-6,3-lactone, and D-threo-isocitric acid-with t-test derived p-value 1, were identified as potential urine signature that can well distinguish RCVS subjects at ictal stage from controls. Among them, citrate, hippurate, ascorbic acid, and D-glucurono-6,3-lactone were significantly lower, and 1,3,7-trimethyluric acid and D-threo-isocitric acid were higher in RCVS patients. Of these, four selected metabolites, citrate, D-glucurono-6,3-lactone, ascorbic acid, and 1,3,7-trimethyluric acid, returned to normal levels in remission. These metabolites are related to pathways associated with free radical scavenging, with the hub molecules being associated with endothelial dysfunction or sympathetic overactivity. Our study demonstrated that six urine metabolites are significantly changed in RCVS patients, with the implicated networks highly relevant to the pathogenesis of RCVS. Funding: This work was supported by the grants from Ministry of Science and Technology of Taiwan MOST 105-2320-B-039-055-MY3, 105-2811-B-039-026, 106-2811-B-039-014 and 106-2320-B-039-053 (to YLL & WHH); Brain Research Center, National Yang-Ming University from The Featured Areas Research Center Program within the framework of the Higher Education Sprout Project by the Ministry of Education (MOE) in Taiwan (to SJW & SPC); Taipei Veterans General Hospital [V100E6-001, V106C-117] (to SWJ & SPC); MOST 104-2314-B-010-015-MY2, MOST 104-2314-B-075-006 -MY3, and MOST 107-2314-B-010-021 -] (to SWJ & SPC); and Ministry of Health and Welfare, Taiwan [MOHW 103-TDU-B-211-113-003, MOHW 104-TDU-B-211-113-003, MOHW 105-TDU-B-211-113-003] (to SJW). Declaration of Interest: The authors have disclosed no potential conflicts of interest. Ethical Approval: The study protocol was approved by the Institutional Review Board of TVGH (IRB No. 98-05-04A). All participants provided written informed consent before entering the study. All clinical investigations were conducted according to the principles expressed in the Declaration of Helsinki.