Development of non-invasive system for the evaluation of free radicals in brain with encephalopathy.

近年来的研究表明，自由基（包括活性氧（ROS）和活性氮（RNS））与体内氧化还原平衡的破坏有关，导致多种疾病的发生。相反，由于缺乏无创地估计脑中氧化还原平衡的技术，很少有关于个体受试者中自由基与脑病相关性的研究。本研究将自旋探针法与活体ESR技术相结合，建立了一种新的无创检测脑内自由基反应的方法，并合成了一种新的硝酰基自旋探针--羧基-丙氧基乙酰氧基甲酯（CxP-AM），用于活体动物脑内自由基反应的检测和成像。CxP-AM被设计成被酯酶水解而不被脂肪酶水解，使得其在静脉注射后穿过血脑屏障并保留在脑实质细胞的胞质相中。通过电子自旋共振计算机断层成像技术证明了探针在脑中的滞留性。此外，我们将此探针应用于脑缺血再灌注自由基反应的评价。本研究的目的是建立一种新的无创评价脑氧化还原平衡的技术，并将一种新的氧化还原代谢生物学参数应用于脑组织。

Recently, some studies indicated that free radicals including reactive oxygen species (ROS) and reactive nitrogen species (RNS) associates with a collapse of redox balance in vivo, resulted in induction of several types of diseases. On the contrary, there were few studies on the association of free radicals with encephalopathy in an individual subject, because of a lack of technique, by which redox balance in brain was estimated noninvasively. In the present study, we established a novel technique to noninvasively estimate free radical reactions in brain by using the combination of in vivo ESR and a spin-probe method.In order to estimate free radical reactions and image them in the brain of living animals, a nitroxyl spin-probe, carboxy-PROXYL acetoxymethyl ester (CxP-AM) was newly synthesized. CxP-AM was designed to be hydrolyzed by esterase, but not by lipase, so that it would pass through the blood-brain barrier and be retained in the cytosolic phase of parenchymal cells in the brain after intravenous injection. The retentivity of the probe in brain was proved through an electron spin resonance computed tomographic imaging technique. Furthermore, we applied this probe to the evaluation of free radical reactions in brain with ischemia-reperfusion. Through these studies, it is elucidated that free radical reactions in the brain with ischemia-reperfusion are significantly enhanced.The goal of our study is to establish a novel technique for the non-invasive evaluation of redox balance in brain, and apply a new biological parameter on redox metabolism to brain.