Postsurgical treatments for human malignant gliomas prove ineffective as there are high rates of tumor reoccurrence due to their elevated DNA-repair mechanisms. There is a group of human gliomas, oligodendrogliomas, that present longer survival periods of patients receiving chemotherapy and/or radiation. Certain genetic aberrations, specifically deletions of chromosome arms 1p and 19q, are associated with oligodendrogliomas that may lead to better responses to postsurgical treatment. My work addresses the hypothesis that this observed trend is due to decreased O6-alkylguanine DNA methyltransferase(MGMT) activity, which may be a result of the 1p and 19q deletions. MGMT is a known repair enzyme of DNA that contributes to tumor reoccurrence following alkylating agent based therapies. The MGMT activity is determined by estimating the number of MGMT molecules present in a homogenate through an assay developed in the Silber lab. The highly-sensitive assay exploits the reaction of an alkyl group irreversibly binding to MGMT. By incubating [3H]-labeled O6-methylguanine contained in double-stranded DNA with tumor homogenate, each MGMT molecule is covalently bound to a methyl group. The radioactivity is then counted by a liquid scintillation spectrometer. Preliminary data collected indicates that oligodendrogliomas with the deletions typically have lower amounts of MGMT activity than oligodendrogliomas without the deletion. When analyzing the clinical outcome information available, patients whose tumors have lower MGMT activity tend to follow the trend of higher responsiveness to treatment and longer periods of survival. By better understanding the DNA repair mechanisms of human malignant gliomas, new therapies can be developed that can improve the prognosis of patients.

Approximately half a million mortalities occur in the United States due to cardiac arrest alone; only a few survive from the initial shock. To improve these chances, first responders must resuscitate patients before their arrival to the hospital for proper medical treatment. Unfortunately, none of the devices on the market can be deployed to guide resuscitation by generating useful hemodynamic information to the responder. Animal studies further simulate that obtaining hemodynamic data during medical procedures such as CPR provides optimum patient outcomes. We have created a non-invasive ultrasound-controlled system that measures flow patterns of oxygenated blood within the ascending carotid artery, as a means to guide resuscitation during cardiac arrest. We designed and tested our ultrasound transducers on a water-submerged string phantom that mimics the flow properties of blood. The transducer propagates a pressure wave towards the moving string, and then records the reflected and frequency shifted wave resulting from the Doppler effect. Once the data has been collected in MATLAB, we produce a graphical user interface (GUI) containing a real-time pulsed Doppler spectrogram displaying the blood flow speeds, which are proportional to the Doppler-shifted frequencies. Our team collected data from multiple swine in-vivo to capture the flow rate in the carotid artery. We analyzed and compared the baseline, trauma, and resuscitation measurements to significant parameters such as heart rate and mean arterial pressure. Successful analysis of those pig studies will motivate retrospective human trials that, in turn, should provide sufficient results to motivate commercialization of this technology.

 Glioblastoma multiforme (GBM) is the most common and deadliest cancer of the central nervous system. The existence of stem-like cells within GBM tumors - Glioblastoma stem cells (GSCs) - may explain some of the malignant features of Glioblastoma multiforme (GBM) owing to their rapid self renewal and diffuse migratory behavior. Increasingly apparent is the role ion channels play in many cancer cell properties, including regulation of cell cycle progression and proliferation, tumor cell motility and invasion, and regulation of metabolic rate. We hypothesize that ion channel dysregulation contributes to GSC malignancy, including increased viability, proliferation, and migration. To test this, multiple GSC isolates derived from human GBM tumors were maintained in culture over many passages while retaining stem cell properties. RNA-seq expression profiles revealed ion channel families that were highly enriched in GSC samples. Real-time qPCR was next used to confirm increased expression of voltage gated calcium, sodium, and potassium channels, ionotropic glutamate channels and inwardly-rectifying potassium channels. Identifying ion channels uniquely enriched in GSCs led to functional analyses to determine how ion channel blockers affected their proliferative and migratory properties. We performed MTT cell viability assays to assess how ion channel blockers altered the viability of GSCs compared to a control neural stem cell line. We were able to show a significant decrease in cell viability across GSC lines compared to control at a variety of drug concentrations. Utilizing live-image cell tracking, we determined decreased migratory behavior and rates of these GSC lines with the same blockers. Finally, to elucidate the role of the extrinsic tumour microenvironment on the proliferation and migration of GSCs, unique coculture models involving GSCs cultured on mouse organotypic brain slices were utilized, and showed the affects of stimulating and inhibitory DREADD receptors to modulate local neuronal activity.

Previous work in my lab has shown that extracranial applications of high intensity focal ultrasound can affect the activity of specific regions in the brain, even going as far as producing specific movements in the extremities of rodents if applied to the motor cortex. With the recent discovery that activation using light stimulation of neurons whose conductive coating (myelin) has been damaged results in higher-than-normal remyelination processes, we proposed that the activation of similarly damaged neurons using high frequency focused ultrasound will also result in increased remyelination and restoration of the targeted neurons. Such a treatment could be used for various neurodegenerative diseases, including multiple sclerosis. Since multiple sclerosis is a disease which damages the myelin coating of neurons and impairs their function, this treatment could revolutionazie multiple sclerosis therapy. Earlier this year, I began work on developing such a treatment using a mouse model which induces brain lesions similar to those found in human multiple sclerosis patients. These experiments include therapies using ultrasound transducers at three different center frequencies, and effectiveness of the treatment is assessed by EEG recordings of the mouse brains during application of ultrasound, as well as MRI scans and stained brain slices of treated and control mice. This work will run through May, and by then I will be able to determine whether high frequency focused ultrasound therapy can be effective in either slowing down demyelination or speeding up remyelination in a well-known animal model of multiple sclerosis. So far we have collected about half of our mouse data, and the post-therapy MRI scans look promising but brain slice staining is currently underway, which will provide much more concrete data on the extent of remyelination.