Dose- and time-dependent neuroprotective effects of Pycnogenol following traumatic brain injury

Abstract

After traumatic brain injury (TBI), both primary and secondary injury cascades are initiated, leading to neuronal death and cognitive dysfunction. We have previously shown that the combinational bioflavonoid, Pycnogenol (PYC), alters some secondary injury cascades and protects synaptic proteins when administered immediately following trauma. The purpose of the present study was to explore further the beneficial effects of PYC and to test whether it can be used in a more clinically relevant fashion. Young adult male Sprague-Dawley rats were subjected to a unilateral moderate/severe cortical contusion. Subjects received a single intravenous (i.v.) injection of PYC (1, 5, or 10 mg/kg) or vehicle, with treatment initiated at 15 min, 2 h, or 4 h post injury. All rats were killed at 96 h post TBI. Both the cortex and hippocampus ipsilateral and contralateral to the injury were evaluated for possible changes in oxidative stress (thiobarbituric acid reactive species; TBARS) and both pre- and post-synaptic proteins (synapsin-I, synaptophysin, drebrin, post synaptic density protein-95, and synapse associated protein-97). Following TBI, TBARS were significantly increased in both the injured cortex and ipsilateral hippocampus. Regardless of the dose and delay in treatment, PYC treatment significantly lowered TBARS. PYC treatment significantly protected both the cortex and hippocampus from injury-related declines in pre- and post-synaptic proteins. These results demonstrate that a single i.v. treatment of PYC is neuroprotective after TBI with a therapeutic window of at least 4 h post trauma. The natural bioflavonoid PYC may provide a possible therapeutic intervention in neurotrauma.

FIG. 1.

Changes in the level of oxidative stress (TBARS) in both the cortex (A) and hippocampus (B) at 96 h after a moderate unilateral cortical contusion. Levels of TBARS were significantly increased in both brain regions following the brain injury. Vehicle-treated animals showed the greatest increases (200%). Animals treated with PYC showed significantly lower levels of oxidative stress whether treatment was delayed for 15 min, 2 h, or 4 h. Each bar represents the group mean±SD. *p<0.001 compared to vehicle; #p<0.05 compared to the lower 1 mg/kg dose.

FIG. 2

Changes in the levels of synaptic proteins in the ipsilateral cortex following a cortical contusion injury at 96 h. Both pre (synapsin-I (B); synaptophysin (C)) and post (drebrin (D), SAP-97 (E), PSD-95 (F)) synaptic proteins were significantly decreased in vehicle-treated animals, while PYC treatment afforded significant protection. (A) Typical immunoblot showing densities of the bands used in analysis. Time indicates the delay following injury when treatment was initiated. Each bar represents the group mean±SD. *p<0.001 compared to vehicle; #p<0.05 compared to the lower 1 mg/kg dose.

FIG. 3.

Changes in the levels of synaptic proteins in the ipsilateral hippocampus after a cortical contusion injury at 96 h. Both pre (synapsin I (B); synaptophysin (C)) and post (drebrin (D), SAP-97 (E), PSD-95 (F)) synaptic proteins were significantly decreased in vehicle-treated animals. Subjects treated with PYC were afforded significant neuroprotection. Panel (A) shows typical immunoblotting bands used in the analysis. Time indicates the delay following injury when treatment was initiated. Each bar represents the group mean±SD. *p<0.001 compared to vehicle; #p<0.05 compared to the lower 1 mg/kg dose.