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L-Citrulline restores nitric oxide level and cellular uptake at the brain capillary endothelial cell line (TR-BBB cells) with glutamate cytotoxicity

Authors
Lee, Kyeong-EunKang, Young-Sook
Issue Date
Nov-2018
Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
Keywords
L-Citrulline; Glutamate; Blood-brain barrier (BBB); Large neutral amino acid transporter 1 (LAT1); Endothelial nitric oxide synthase (eNOS); Inducible nitric oxide synthase (iNOS); Nitric oxide (NO)
Citation
MICROVASCULAR RESEARCH, v.120, pp.29 - 35
Journal Title
MICROVASCULAR RESEARCH
Volume
120
Start Page
29
End Page
35
URI
https://scholarworks.sookmyung.ac.kr/handle/2020.sw.sookmyung/4188
DOI
10.1016/j.mvr.2018.05.010
ISSN
0026-2862
Abstract
Objective: Glutamate excitotoxicity provokes neuronal cell damage and death, leading to collapse of the blood brain barrier (BBB). Recently, it has been reported that L-citrulline, a neutral amino acid and a major precursor of L-arginine in the nitric oxide (NO) cycle, can prevent both neuronal cell death and cerebrovascular cell loss in brain ischemia. Therefore, the objective of this study was to investigate the effect of L-citrulline on glutamate cytotoxicity in the BBB using the conditionally immortalized rat brain capillary endothelial cell line (TR-BBB cells) as an in vitro model of the BBB. Methods: Cell viability was determined using MIT assay. Cellular uptake of [C-14] L-citrulline and expression levels of rat large neutral amino acid transporter 1 (rLAT1), endothelial nitric oxide synthase (eNOS), and inducible nitric oxide synthase (iNOS) at mRNA level were performed using quantitative real-time polymerase chain reaction (PCR) analysis. NO production from TR-BBB cells was measured using Griess reagents. All experiments were performed after treatment of TR-BBB cells with glutamate alone or co-treatment with L-citrulline, L-arginine, and/or taurine for 24 h. Results: L-Citrulline treatment increased cell viability, [C-14] L-citrulline uptake, and the mRNA levels of LAT1 and eNOS in TR-BBB cells treated with glutamate. However, iNOS mRNA expression was inhibited by L-citrulline. NO production and transcript level of iNOS were markedly increased by glutamate treatment alone. However, co-treatment with L-citrulline, taurine, or both L-citrulline and taurine decreased NO levels and mRNA levels of iNOS in TR-BBB cells treated with glutamate. In co-treatment of TR-BBB cells with L-arginine, a NO donor, and glutamate, NO levels were increased and expression levels of iNOS mRNA were similar compared to those in cells treated with glutamate alone. Conclusion: L-Citrulline can restore NO level and its cellular uptake in TR-BBB cells with glutamate cytotoxicity. Supplying L-citrulline at the BBB may provide neuroprotective effect to improve cerebrovascular dysfunction such as a brain ischemia.
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