Diagnosing And Evaluating Cerebral Ischemic Stroke Using Multiphoton Microscopy

Cerebral ischemic stroke is a common acute cerebral vascular disease,and it is becoming a leading cause of death and permanent disability around the world.Interrupted blood supply to the brain causes abrupt oxygen and nutrient deprivation and results in ischemic injury to the brain.In addition,vascular occlusion of variable duration times could be anticipated to result in varying degrees of injury over time.Therefore,early and accurate identification of the potentially salvageable boundary region of ischemic brain tissues may enable selection of the most appropriate candidates for early stroke therapies.Neurons are the critical cell type of brain tissue,which play an important role in cerebral ischemic stroke,are the main target of our research.In recent years,multiphoton microscopy(MPM)based on two-photon excited fluorescence(TPEF)and second harmonic generation(SHG)has been used in biomedical fields widely.It has the advantages of high resolution,low photo-bleaching and enhanced penetration depth.In this study,Sprague-Dawley(SD)rats were chosen to be the experiment objects,then MPM technology was used to visualize their normal brain tissues and ischemic brain tissues,demonstrating that this technology has the potential to diagnose and evaluate ischemic stroke.Firstly,we distinguished neurons from glial cells in the normal brain tissues via the two-photon excited fluorescence signals of intracellular intrinsic sources.The results showed that this technology not only can identify neurons,but also can provide the detailed information of microstructure in normal brain tissues.And then,TPEF microscopy was employed to identify the boundary between normal brain tissue and ischemic region,demonstrating that this technology has the ability to distinguish normal brain tissues from ischemic brain tissues.At last,MPM was employed to image the ischemic core of cerebral cortex and striatum with different duration of occlusion.The results showed that the influence of occlusion duration on neuronal morphology variation was examined.Meanwhile,our approach also presented the capability of evaluating the degree of ischemic injury via calculating the ratio of injured over normal neurons and the nuclear-to-cytoplasmic ratio of normal and injured neurons.Taken together,our results demonstrated that MPM has the potential to develop into an effective diagnostic and evaluating tool for cerebral ischemic stroke.