NEUROPROTECTIVE EFFECTS OF GREWIA carpinifolia AGAINST VANADIUM-INDUCED TOXICITY IN MICE

Abstract

Nigeria is currently the second highest gas flaring country in the world with resultant environmental vanadium discharge. Vanadium toxicity is implicated in neurodegenerative changes via free radical production. The Blood-Brain Barrier permeability for synthetic antioxidants currently used in protecting the central nervous system in vanadium toxicity remains a challenge. Grewia carpinifolia (GC) possesses antioxidant property. This study was designed to investigate the protective activities of ethanol extracts of GC in vanadium-induced toxicity in mice. Safe doses of GC leaf and stem extracts (FHI-109693) were assessed in 50 mice equally divided and administered 100, 200, 400 and 800 mg/Kg of each extract orally for 28 days using standard haematological, biochemical and histopathological methods. Distilled water served as control. Protective effects of each extract at 100 and 200 mg/Kg following vanadium toxicity in 120 mice (control, vanadium, extract-treated and standard-treated with α-tocopherol) were studied using behavioural tests (open field, hanging wire and Morris water maze), hepatic enzymes, haematological and histological parameters. In vitro antioxidant and lipid peroxidation activities were measured using ABTS, DPPH and TBARS assays. Pure bioactive compounds were isolated from leaf and stem extracts by TLC, open column chromatography, HPLC and characterised by NMR and MS. Brain Uptake Index (BUI) of each compound was determined. Neuroprotective and antioxidant activities of pure compounds following vanadium-induced toxicity were evaluated in a separate cohort of 72 mice equally divided; control, vanadium-treated, vanadium alongside compounds with high BUI and standard-treated groups by behavioural tests, antioxidant enzymes‟ activities (catalase, SOD, GPx, GSH), oxidative stress markers (MDA, NO and H2O2) measurements and immunohistochemical expression of Myelin Basic Protein (MBP) in the brain. Data were analysed using one-way ANOVA at α0.05. There were no significant differences in haematological parameters at tested doses, however, significant increase in ALP (40.85±6.78 to 81.40±6.24 IU/L), congestion of hepatic sinusoids were observed at 800 mg/Kg following 28-day administration. The extracts at 200 mg/Kg increased line crossings (18.60±4.67 to 59.00±5.93), reduced rearing (73.25±7.23 to 10.00±1.82) in open field test and increased latent time on hanging wire (27.67±5.12 to 70.34±8.05 secs). It also decreased AST (81.20±10.06 to 45.00±7.07 IU/L), ALT (75.40±9.07 to 45.00±5.82 IU/L) levels, increased PCV (32.40±4.10 to 40.50±3.54%) and prevented disorganisation of Purkinje cells of the cerebral cortex caused by vanadium. The IC50 of ABTS, DPPH and TBARS by extracts were 0.32, 0.41, 0.21 mg/mL (leaf) and 1.98, 0.80, 0.30 mg/mL (stem), respectively. Bioactive compounds isolated were iii β-spinasterol, dibutyl phthalate, β-sitosterol, benzoic acid butyl ester and stigmasterol with BUI of 70.6%, 3.5%, 76.5%, 1.1% and 87.0%, respectively. Concurrent administration of β-sitosterol and stigmasterol significantly attenuated spatial learning deficits caused by vanadium than α-tocopherol by reducing escape latency (20.66±2.13 to 37.19±4.63 secs), grooming, rearing and stretch-attend posture frequency. They also significantly increased activities of catalase (28.72±1.04 to 11.04±1.79 IU/mg protein), SOD (38.63±3.17 to 12.06±1.03 U/mg tissue), decreased oxidative stress markers and increased MBP expression. β-sitosterol and stigmasterol isolated from Grewia carpinifolia crossed the Blood-Brain Barrier, exhibited potent antioxidant and neuroprotective activities. They are therefore potential candidates in treatment of vanadium toxicity. Keywords: Vanadium toxicity, Grewia carpinifolia, β-sitosterol, Stigmasterol, Neuroprotective Word Count: 498