The objective of this Shared Equipment Evaluation Program (ShEEP) proposal is to request Keyence BZX800E All-in-One Automated Imaging System which will offer researchers an instrument that is capable of: confocal-like multi-stack imaging and real-time focusing of thick tissue sections; rapid automatic high- magnification image acquisition and stitching of composite images; and three-dimensional profiling of serial tissue sections imaged at high resolution. The control software is simple and intuitive. Thus, with minimal training a novice user can scan a series of tissue sections from an entire organ, generate high-resolution composites of each tissue section, and merge all composites into a single three-dimensional rendering of the entire tissue. Traditional microscopes available at the Baltimore VA Medical Center (BVAMC) do not possess these capabilities. The VA projects cover an array of specialties as described below. Each will benefit from Keyence BZX800E All-in- One Automated Imaging System. The BZX800E Microscope optimizes the imaging process by designing a fully automated, easy-to-use system that captures high-resolution images and allows researchers to achieve their desired results faster. These applications are of great use for VA Biomedical Laboratory Research and Development Service (BLR&D), Clinical Science Research and Development (CSR&D), and Rehabilitation Research and Development (RR&D) funded investigators. Tibor Kristian, Ph.D. will serve as the Project Director (PD)/ Principal Investigator (PI) for this proposal. Dr. Kristian?s study focuses on understanding injury mechanisms that lead to mitochondrial and ultimately cellular bioenergetic failure in stroke victims. This imaging system is essential for imaging and quantification of intracellular organelles damage following stroke- induced ischemic brain injury. Bingren Hu, M.D., Ph.D. is testing a novel hypothesis that brain ischemia- reperfusion leads to a cascade of events resulting in inactivation; as well as study novel mechanisms underlying brain ischemia-reperfusion (IR) injury. Many antibodies will be used in Aims 1 and 2 of Dr. Hu?s studies; and the BZX800 microscope is necessary for imaging and quantification of intracellular organelles damage following IR brain injury. Bogdan Stoica, M.D. will use this new system to investigate traumatic brain injury (TBI). Dr. Stoica?s project requires significant imaging of chronic neuroinflammation markers and secondary damaging markers. Tapas Makar, Ph.D.?s project studies the effects of 7, 8- dihydroxyflavone (7,8- DHF); which has potent brain derived neurotrophic factor (BDNF) agonist activity and will use the imaging system in his specific aims 2 and 3. Dr. Makar will determine whether DHF treatment modulates inflammation and immunity in the brain and spinal cord in encephalomyelitis (EAE), a murine model of Multiple sclerosis, using monoclonal antibodies. Leonardo Tonelli, Ph.D.?s project studies the stress of deployment and exposure to traumatic events that puts soldiers at a greater risk than the general public in developing psychological disorders. Dr. Tonelli?s analysis will consist of immunohistochemistry in adjacent sections for: homing markers, chemokine receptors, microglia and macrophage activation and/or differentiation- all of which can be viewed with the Keyence BZ-X800. Guofeng Xie, M.D. studies colon cancer and his long-term goal is to develop innovative, effective, and safe therapies by targeting pivotal signal transduction molecules underlying colon cancer progression. The microscope will allow Dr. Xie to examine the expression patterns of matrix metalloproteinase 7 (MMP7) using immunofluorescent anti-MMP7 antibody in human colon cancer cells and surgical specimens. The funded projects form the basis for translational studies that will improve treatment in Veterans- potentially yielding new therapeutics.
The purpose of this ShEEP application is to request the acquisition of a Keyence BZ-X800E All-In- One Automated Imaging System. This fully-automated instrument is capable of ten times more work than conventional microscopes with superior imaging quantity. This is particularly critical for the participating VA-funded projects in this ShEEP application; since histological quantification of brain damage alone will require the use of 600 sections in a drug treatment study. Therefore, this instrument will provide a major contribution to the VA's long-term research goals; and impact a wide range of translational research at the Baltimore VA Medical Center (VAMC) by reducing overall costs for tissue processing and speeding the development of many critical therapies. The Keyence Microscope will aid researchers in revealing new mechanisms of cell death following injury, and will facilitate identification of new targets that can be potentially used to improve Veterans health.