Protective Role of Arginase II in Cerebral Ischemia and Excitotoxicity

Abstract

Background: Arginase (Arg), one of the enzymes involved in the urea cycle, provides an essential route for the disposal of excess nitrogen resulting from amino acid and nucleotide metabolism. Two reported subtypes of Arg (ArgI and II) compete with nitric oxide synthase (NOS) to use L-arginine as a substrate, and subsequently regulate NOS activity. It has been reported that Arg has significant effects on circulation that suggest the potential role of this enzyme in regulating vascular function. However, the role of Arg following brain damage has not been elucidated. In this study, we hypothesize that the deletion of ArgII will lead to aggravated brain injury following cerebral ischemia and excitotoxicity.

Methods and findings: To test our hypothesis, male C57BL/6 wildtype (WT) and ArgII^-/- mice were subjected to permanent distal middle cerebral artery occlusion and survived for 7 d. Cerebral blood flow (CBF) data revealed a statistically non-significant decrease in CBF in ArgII^-/- mice. However, ArgII^-/- mice had significantly higher neurologic deficit scores and brain infarctions. The hypothesis was further tested in a more specific N-methyl-D-aspartate (NMDA)-induced acute excitotoxic model. WT and ArgII^-/- mice were given a single intrastriatal injection of 15 nmol NMDA. Forty-eight hours later, the excitotoxic brain damage was significantly worse in ArgII^-/- mice. The data from both models confirm the neuroprotective effect of ArgII.

Conclusion: Targeting ArgII could be considered an integrative part of a multi-modal approach to fight acute brain damage excitotoxicity, ischemic brain injury, and other forms of brain trauma.

Keywords: Cerebral blood flow; Mice; NMDA; Neuroprotection; Nitric oxide synthase; Stroke; Vasodilation.

Figure 1

Deletion of ArgII leads to increased neurological deficit score (NDS) and infarct volume. WT (n=8) and ArgII^-/- (n=10) mice were subjected to permanent distal middle cerebral artery occlusion (pMCAO) and were allowed to survive for 7 days. On day 7, mice were sacrificed immediately After being tested for NDS, and brain infarction was analyzed After triphenyl tetrazolium chloride staining. (A) As an indicator of neurological dysfunction, assessment on day 7 After ischemia shows significantly larger NDS in ArgII^-/- mice than in WT mice. (B) Representative macrographs showing the slices of brain infarction from WT (left) and ArgII^-/- (right) mice. (C) Histogram shows the percent corrected cortex infarct volume of WT and ArgII^-/- mice. The infarct size was significantly larger in ArgII^-/- than in WT mice. Data represented as mean ± S.E.M; *p=0.0019, **p=0.0079.

Figure 2

ArgII deletion attenuates relative cerebral blood flow rCBF. rCBF was recorded at baseline, at induction of ischemia, and at 15-min intervals during ischemia until 120 min in WT and ArgII^-/- mice (n=5, each group). ArgII^-/- exhibited a non-significant decrease in CBF. Changes in CBF were recorded as a percent of baseline. Data are represented as mean ± S.E.M.

Figure 3

ArgII deletion increases NMDA-induced neurotoxicity. WT and ArgII^-/- mice (n=7, each group) were given a single stereotaxic injection of 15 nmol NMDA in the striatum and sacrificed 48 h later. Brain sections were stained with Cresyl violet to analyze the brain lesions. (A) Representative macrographs of coronal sections of the WT (left panel) and ArgII^-/- (right panel) mice brain After intrastriatal injection with 15 nmol NMDA. The brain sections from the ArgII^-/- mise appears to show greater lesion volume. (B) Analysis of the brain sections show that the ArgII^-/- mice were more vulnerable to the NMDA-induced neurotoxicity than were the WT mice. Values are represented as means ± S.E.M.; *p=0.035, when compared with WT group.