L-arginine and tetrahydrobiopterin reduce brain damage after ischaemic stroke in diabetic rats

Abstract

Introduction: This study aimed to determine whether administration of L-arginine and/or tetrahydrobiopterin (BH4) would be neuroprotective against ischaemic stroke in rat models of diabetes mellitus. Methods: Type 1 diabetes mellitus (T1DM) was induced in rats by a single intraperitoneal (i.p.) injection of streptozotocin at a high dose (60 mg/kg). Type 2 diabetes mellitus (T2DM) was induced using a combination of high-fat diet for 4 weeks and a single injection of streptozotocin at a low dose (35 mg/kg) after 2 weeks of highfat diet. Middle cerebral artery occlusion (MCAO) was induced in T1DM, T2DM, and non-diabetic (non-DM) rats. The duration of ischaemia was 75 or 90 minutes in non-diabetic rats and 75 minutes in diabetic rats. Rats were either treated with BH4 (10 mg/kg i.p.) or L-arginine (300 mg/kg i.p.) alone or in combination at 30 minutes before MCAO. After reperfusion for 24 hours, rats were subjected to a behavioural test of neurological deficit before they were sacrificed. The brains were collected for the assessment of infarct volume and oedema. Results: In non-DM rats, the percentage of infarct volume over the total volume of right brain hemisphere was 48.1% after 90 minutes of ischaemia in the vehicle group. Compared with the vehicle group, injection of BH4 or L-arginine alone prior to ischaemia reduced the percentage of infarct volume by 36.2% or 33.4%, respectively (P<0.01). The combination did not achieve any better outcome, with relative infarct volume reduced by 27.4% (P<0.01). All three treatments reduced oedema ratio and severity of neurological deficit. Mortality was 44.4% in T1DM rats and 11.1% in non-DM rats after 90 minutes of MCAO. Mortality was 12.5% in T1DM rats and 0 in non-DM rats after 75 minutes of MCAO. The percentage of infarct volume in vehicle groups after 75 minutes of ischaemia in non-DM, T1DM, and T2DM rats were 40.2%, 44.5%, and 37.6%, respectively. Treatment with tetrahydrobiopterin alone significantly reduced the relative infarct volume by 16.9% and 32.6% (P<0.05) in T1DM and T2DM rats when compared with the vehicle groups, respectively. Different treatments did not affect oedema ratio in diabetic rats. The behavioural performance improved after injection of BH4 and L-arginine in T1DM rats. Conclusion: Administration of L-arginine, BH4, or the combination was neuroprotective against stroke in nondiabetic rat. Tetrahydrobiopterin but not L-arginine reduced the brain injury after stroke in diabetic rats.