I am a neurologist with specialty board certification in Vascular Neurology. I have run large, multi-center clinical trials for industry and NIH. I have a laboratory effort that has been funded for over 30 years by NIH, VA and AHA focused on translational pre-clinical stroke modeling, pharmacology, and vascular biology. Our lab has recently focused on the role of thrombin mediating cytotoxicity and cytoprotection in the brain. Also, we have a long-standing interest in pre-clinical models of therapeutic hypothermia, motivated by my role as PI of the largest clinical trial of therapeutic hypothermia for stroke, the ICTuS program. We have considerable experience with a variety of experimental models; behavioral testing; histology; and cell biology. My team and I were recently selected by NINDS to develop the Stroke Preclinical Assessment Network (SPAN) as the Coordinating Center. The SPAN will test 6 novel stroke treatments in a state-of-the-art, rigorous network of expert pre-clinical stroke testing centers. We have considerable experience with a variety of experimental stroke models; behavioral testing; histology; and cell biology. Details follow:
I joined the Justin Zivin lab as a post-doctoral Fellow at the time he published his seminal demonstration that rt-PA powerfully protected neurological function after cerebral embolism (Zivin et al, 1985; Science 230:1289). During a 5-year R29 (NS267883) under Zivin, I created a model of post-embolic hemorrhage and contributed to the development of rt-PA as a clinical therapeutic. I was honored to be selected as the San Diego site PI of the NINDS for rt-PA Acute Stroke Trial (N01-N292332 and N01-NS02377), during which I helped draft the protocol and main results for that landmark trial. I wrote, produced and directed the NIH Stroke Scale training and certification video that has now been viewed by over 2M individuals around the world—I oversaw the clinimetric validation of these video tools. The rt-PA for stroke trial, published in the New England Journal of Medicine in 1995, is one of the most influential trials in the history of modern neurology.
My earliest research focused on agonists of the GABA-A receptor as neuroprotectants, which I demonstrated were at least as powerful in vivo as glutamate channel antagonists. The combination of both classes of drug exhibited powerful synergism. These studies led to several clinical trials for which I was the San Diego site PI. Then, I was chosen to serve as the global principal investigator of the GABA-A agonist clomethiazole trial known as CLASS, the global PI of the CHANT trial, and the North American coordinating investigator for SAINT 1 and SAINT 2.
In later studies, my lab studied angiogeneis, and the potential for angiogenic growth factors to serve as neuroprotectants. We noted a blood-brain barrier effect of VEGF, used in these studies, which led to several studies of angiogenesis (R01 NS43300) as we explored the role of putative neuroprotective growth factors. We eventually discovered that ischemic brain produces angiogenic growth factors to open capillaries and to promote entry of phagocytic macrophages, a phenomenon we labeled “the Clean-up hypothesis”. We showed convincingly that the angiogenesis documented around infarcts serves ONLY the clean-up of necrosis and is not a protective or neo-restorative effect.
During the vascular leakage studies in the lab, we became aware of work showing that thrombin powerfully opens the BBB by a direct effect on endothelial cells and kills neurons and astrocytes. Given our interest in leakage, we began working with thrombin. We also were aware of the seemingly contradictory effect of very-low-dose thrombin to protect brain (pre-conditioning). One of my PhD students noted that neurons expressed prothrombin message, an effect we dismissed at first as lab error. After confirming his results and finding traces of a similar finding in the literature, we began to ask what the purpose could be of neuronal-expressed thrombin. We showed (R01 NS075930) that thrombin powerfully kills cells in a dose-dependent manner via the PAR-1 receptor. Thrombin activity is associated with direct neuron killing during stroke. Thrombin antagonists, such as argatroban, are powerful neuroprotectants, and a clinical trial studied PAR-1 agent 3K3A-APC for which I served as the national PI (U01 NS088312). Argatroban itself is under study as stroke therapy, and I served as the Los Angeles site PI (P50 NS044227, Grotta).
Although these translational results were gratifying, we were left with the question, why does brain make the serum clotting factor prothrombin? We have used oxygen-glucose deprivation (OGD) in cell culture to model cellular responses to stroke or cardiac arrest. In a series of elegant in vitro experiments, my colleague Padmesh Rajput and I have demonstrated the OGD stressed neurons secrete thrombin into the culture media; OGD neuronal conditioned-media causes astrocyte activation; such astrocyte activation can be reproduced with low doses of thrombin; and is blocked by thrombin inhibitors or by knocking down or knocking out PAR-1 from the astrocytes. We then created an inducible, neuron-targeted prothrombin knock out experimental. After inducing the neuronal prothrombin knock out, the region of induced infarction in these experimentals is significantly enlarged and there is greatly reduced evidence of astrocyte activation in the ischemic bed. Taken together, these results suggest the novel hypothesis that neurons produce prothrombin as a distress signal to adjacent astrocytes, which respond with the newly described astrocyte protective response. We are actively engaged in multiple studies to support/refute this hypothesis; extend it to other elements in the NVU; and identify the active agent(s) comprising the astrocyte protective response.
Another area of interest in our laboratory is brain cooling, or therapeutic hypothermia, the most powerful neuroprotectant ever documented in stroke models. Therapeutic hypothermia is of proven benefit for victims of cardiac arrest or neonatal hypoxic-ischemic injury. For over a decade I worked on the ICTuS program (P50 NS044148) to deliver therapeutic hypothermia to patients. In the first trial, ICTuS, we showed that therapeutic hypothermia could be safely delivered with an endovascular cooling catheter. In the ICTuS-L program, we safely combined endovascular cooling with rt-PA. In the ICTuS 2/3 trial we attempted to document efficacy of therapeutic hypothermia and thrombolysis. The trial was stopped early, however, after several other trials of intra-arterial neurothrombectomy were overwhelmingly positive. We revised the ICTuS 3 protocol to accommodate neurothrombectomy, but during analysis of the small ICTuS 2 dataset we noted a troubling trend (not statistically significant) toward pneumonia risk during cooling. Also, other trials were published showing lack of efficacy for therapeutic hypothermia in head trauma, and no discernible difference between 33ºC vs 36ºC target temperatures for hypothermia after cardiac arrest. We asked if we were using therapeutic hypothermia correctly in treating these illnesses. Based on our observations of astrocyte protection of neurons, we asked whether cooling could disrupt the astrocyte mediated protection of neurons. In fact, we showed that hypothermia interferes with the astrocytes, in a graded, temperature-dependent manner. We recently showed that ultra-fast cooling to 33ºC for a short time was more powerful in the MCAo model than longer cooling periods. These exciting and novel data, if confirmed, suggest a reason for the failures of therapeutic hypothermia in some clinical trials. Moreover, our data suggests testable hypotheses about the effects of temperature in modulating NVU protection during ischemia.