Promoting stroke recovery using delayed growth factor delivery: Contribution of ipsilesional versus contralesional pyramidal tract plasticity

Stroke incidence is increasing due to the rapidly aging population in developed countries. Whereas untreated acute middle cerebral artery occlusion (MCAO) causes death in 20% of patients and long-term disability in more than 70% of patients, acute stroke therapy with rapid vessel recanalization significantly reduces mortality without influencing functional recovery beyond the acute stroke phase. This lack of functional recovery suggests a need for innovative therapies that can restore function after stroke. The purpose of these studies was to examine the effects of delayed administration of the growth factors erythropoietin (Epo) and vascular endogenous growth factor (VEGF) on functional neurological recovery and pyramidal tract plasticity in mice. The first study investigated how subacute delivery of Epo, starting at 3 days after stroke onset and continuing for 30 days (1 I.U. /day or 10 I.U. /day; via mini osmotic pump), influenced neuronal survival, axonal sprouting and neurological function recovery in C57Bl6/j mice submitted to 30-min MCAO. Epo administered in a 10 I.U. /day dose, in contrast to the 1 I.U. /day administration, showed a significant increase in neuronal survival and CD31 + newly-formed capillaries. This vascular growth enabled further neuroregeneration processes. Functional behavioral tests showed a significant improvement of motor coordination (RotaRod test) and grip strength (Grips strength test) among mice with 10 I.U. /day Epo administration, with no improvement for the low dose group. To investigate the neurological changes underlying these results, two anterograde tract tracers (dextrane amines) were injected in the motor cortex ipsilateral and contralateral to the ischemic lesion, in mice treated with the higher Epo dose. Histological evaluation of the tracers, both at the level of rubral and facial nucleus, showed that functional recovery in these animals was due to an increase of contralateral projections, accompanied by a compensatory decrease of ipsilateral projections. In the second study, VEGF was investigated due to its dual actions on vessels and neurons, which have potential for promoting long distance axonal plasticity in the ischemic brain. Mice were submitted to 30 minutes MCAO, followed by the intraventricular delivery of normal saline or VEGF (0.004 or 0.02 µg/day) starting 3 days post-ischemia. The outcome parameters were functional neurological recovery, long distance axonal plasticity by anterograde tract tracing and cellular and molecular responses examined by histochemistry, RT-PCR and Western blots. VEGF promoted neurological recovery when administered at the higher dosage, by stimulating long distance axonal plasticity in the contralesional but not ipsilesional pyramidal tract system. This observation was accompanied by deactivation of matrix metalloproteinase-9 (MMP9) in the ipsilesional brain tissue and downregulation of axonal growth inhibitors and guidance molecules in the contralesional brain tissue. The results support the concept that brain plasticity is consistent with coordinated axonal growth responses both ipsilateral and contralateral to the site of stroke. Considering that Epo is well tolerated in humans, clinical studies are now conceivable in which Epo is applied in patients in the post-acute stroke phase.