Study Of The Changes In NF-κ Bp65, I-κ B_α And COX-2 Following Focal Cerebral Ischemic Reperfusion In Rats

Ischemic Cerebral Vascular Disease (ICVD) is a major cause of death and disability, especially in China. The process of thrombolysis has not been as successful as expected, because of detrimental cascades following ischemic reperfusion. Therefore it is necessary to clarify the theoretical basis of focal cerebral ischemic reperfusion. In this study using rats, we focused on the mechanism of transcription factor NF- K Bp65 activation, and I- B and cyclooxygenase-2 (COX-2) expression in both inflammatory and cell death responses following focal cerebral ischemic reperfusion. This investigation of the theoretical basis of signal transfer may lead to development of new therapeutic strategies.Part I Establishment of a rat model of cerebral ischemic reperfusion and characteristic findings in pathomorphologyObjective: To establish a model of focal cerebral ischemic reperfusion (FCIR) in rats, and to investigate the characteristics of pathomorphism.Method: Adult Wistar male rats were subjected to 2h of focal ischemia induced by middle cerebral artery occlusion (MCA0) with a filament insert, followed by 3h, 15h, 24h, and 48h of reperfusion. Sham operated rats not subjected to MCA0 served as controls. Hematoxylin and Eeosin (H&E) staining and transmission electron microscopic technology were used to characterize the pathomorphology..Results:1. All focal cerebral ischemic reperfusion rats showed focal neurological functional deficits after full recovery from anesthesia. Deficits such as hemiplegia, circling to the right, falling to the right while walking, or displaying Horner's Sign were observed. After 3h of reperfusion, the deficits partly recovered. These manifestations are similar to patients with focal cerebral ischemic reperfusion. So this model closely mimics ischemic cerebral vascular disease in humans and therefore is useful for the study of pathophysiologic mechanisms and for evaluation of the effects of stroke therapy.2. Using H&E staining, we found eosinophilic neurons with homogeneous cytoplasm and pyknotic nuclei or lacking cellular structures, features which were treated as the hallmarks of neuron death. The dead neurons (<24%) were observed in ischemic striatum at 3h of reperfusion after 2h MCA0 At 48h of reperfusion, the number of dead neurons in the cortex and striatum increased to 50-74%.3. Using Transmission Electron Microscopy we observedhippocampal damage at 24h of reperfusion following MCAo. We found in addition to the changes in neurons, there were changes in the blood-brain-barrier. In microvessels the following were observed: edema in the endothelium and pericytes, the nuclei of endothelial cells shrank, the space connected between endothelial cells and permeability were increased, the integrin-matrix attachments of endothelial cells and the basement membranes were lost, and lymphocytes were observed inthe vessels. The end-feet of astrocytes around the micro-vessel swelled and displayed regressive degeneration, and the microvescular canal became narrow. After ischemic reperfusion, there were many processes from the blood capillary endothelial cells extending into the lumen. This compensation increases the contact area and may improve the nutritional transport between blood and neurons. Perhaps this is one of the mechanisms of compensation, reparability and new-vessel regeneration. Conclusion: In conclusion, our results have demonstrated the focal neurological functional deficits in an established model of cerebral ischemic reperfusion in Wistar rats. This model can closely mimic ischemic cerebral vascular disease in humans, and it is a useful model for the study of pathophysiologic mechanisms.