Abstract W P216: Inducible Glutamate Oxaloacetate Transaminase: A New Target for Stroke Therapy

Introduction: Elevated extracellular glutamate plays a central role in mediating ischemic brain damage. The current work stems from our key observation demonstrating that induction of glutamate oxaloacetate transaminase (GOT), a glutamate metabolizing enzyme in the brain, can attenuate stroke-induced injury. At the stroke site, inducible GOT (iGOT) catalyzes the metabolism of extracellular glutamate via a truncated TCA cycle to generate energy and limit concentrations. Hypotheses: Phytoestrogen Biochanin induces GOT. iGOT inducer Biochanin protects against stroke in vivo. Methods: In vitro. Primary cortical neuronal cells were isolated from the cerebral cortex of C57/BL6 mice feti and seeded in 12-well plates maintained for 24h under standard media and culture conditions prior to experimental treatment. After 24h, cells were incubated with vehicle control (DMSO) or one of several screened phytoestrogen isoflavones for 72h. After 72h sample were collected for mRNA and protein expression analyses of GOT. In vivo. Mice (C57/BL6) received daily IP injection of vehicle control (75% DMSO in water, n=4) or Biochanin A (10mg/kg body weight, n=6) for 3 weeks prior to 90min middle cerebral artery occlusion (MCAO). Following 48h, mice were subjected to 11.7T MRI for determination of stroke induced lesion volume and brain tissue were collected for downstream mRNA and protein expression analyses of GOT. Results: The following phytoestrogen isoflavones were screened for their ability to induce GOT: biochanin A, flavaprenin, diadzein, formononetin, glycitin, and genistein. Biochanin A was identified as a potent (3-fold) inducer of GOT. Biochanin A significantly induced GOT mRNA expression in neural cells. In an in vivo setting of acute ischemic stroke, Biochanin A treated mice had significantly (n=6; p<0.05) smaller stroke-induced lesions as compared to vehicle controls. Conclusion: In this work we have identified a natural small molecule inducer of GOT - Biochanin A. Biochanin A is a safe, naturally occurring phytoestrogen isoflavone that represents a therapeutic target that lends itself to be translated to clinical study.