2大会报告.pdf

中华医学会肾脏病学分会2014 年学术年会 大会报告 PL-01 Using 2-Photon Microscopy to Understand Kidney Injury and Repair Bruce A. Molitoris, Indiana University and the Rouderbush VA Medical Center, Indianapolis, Indiana, USA. Two-photon microscopy, along with the development of new fluorescent probes and innovative computer software, has advanced the study of intracellular and intercellular processes in organs. Researchers can now follow in dynamic fashion and determine the subcellular distribution, behavior, and interactions of labeled chemical probes and proteins in live kidney tissue in real time without fixation artifacts. Fluorescently labeled dextrans, have extended our understanding of dynamic events, including glomerular filtration characteristics and proximal tubule reabsorption at the subcellular level. To accomplish expression of specific proteins in vivo, cDNAs of fluorescently labeled proteins as plasmids or cloned into adenovirus vectors and infused by micropuncture or injected via retrograde renal vein under hydrodynamic pressure conditions to induce multiple cell protein expression with high efficacy. The localization and intensity of the expressed fluorescent proteins can be observed repeatedly at different time points allowing for enhanced quantitative analysis while limiting animal use. Optical sections of images acquired with the two-photon microscope can be 3-D reconstructed and quantified with Metamorph, Voxx and Amira software programs. This approach has opened many new avenues to study the pathophysiology and therapy of kidney disease in both acute and chronic settings. Specifically, in acute kidney injury proximal tubule injury, peritubular dysfunction and glomerular blood flow alterations have been observed