7 Dietary sulforaphane reduces peri-infarct constriction frequency in mouse brain following focal cerebral ischaemia via a Nrf2-independent mechanism

Introduction The dietary isothiocyanate sulforaphane (SFN) obtained from precursors in cruciferous vegetables such as broccoli is a potent inducer of endogenous antioxidant defences. As a strong electrophile, it oxidises cysteine residues in the redox sensor Keap1 inducing activity of the transcription factor Nrf2 and upregulation of target antioxidant and detoxifying genes. We have previously demonstrated that oral administration of doses achievable through diet for three days prior to surgically induced ischaemic stroke results in a reduced infarct volume 70 hour later in wild type but not Nrf2 deficient mice, possibly as a consequence of enhanced Nrf2-mediated redox protection in the context of ischaemic-reperfusion injury. Materials and methods To further investigate mechanisms by which dietary sulforaphane affords neuroprotection, we measured acute blood flow changes during and after focal cerebral ischaemia in mice fed vehicle (corn oil) or sulforaphane at 5 mg/kg for 3 days. Ischaemic stroke was induced via the Longa procedure, employing a surgically inserted filament to transiently occlude the middle cerebral artery under stereotaxic anaesthesia. Ischaemia followed by 60 min of reperfusion resulted in a reproducible temporal lobe stroke. Transcranial laser speckle imaging (Moor FLIPI 2) was used to assess blood flow with high temporal (20 s) and spatial (20 µm) resolution, enabling discrimination of three functional regions during stroke in the unaffected contralateral hemisphere, partially responsive per-infarct region and the ischaemic infarct core. Results While sulforaphane did not alter initial baseline blood flow or affect the depth of ischaemia at the start of the procedure, a clear effect was resolved on the number of evoked peri infarct constriction (PIC) waves. While in both vehicle and treatment groups, an initial PIC was observed in all animals within 10 min of the onset of ischaemia, sulforaphane pre-treatment in wild type mice significantly reduced the number of subsequent PICs. In Nrf2 deficient mice, an equivalent and statistically significant reduction in PIC number was seen associated with sulforaphane pre-treatment, implying that the effects of sulforaphane on PIC frequency are unlikely to be mediated by Nrf2 but rather associated with associated redox gene induction. As sulforaphane significantly improved post-reperfusion core blood flow in wild type not Nrf2 deficient mice. Conclusions Our findings suggest that peri-infarct depolarisation waves or baseline Nrf2 genetic status are not major contributors to post-reperfusion, no-reflow in ischaemic stroke. However, activation of Nrf2 target genes by dietary sulforaphane protects against no-reflow pathology with persistent benefits post stroke. Supported by the British Heart Foundation