The Anti-apoptotic Mechanism Of ASA In Focal Cerebral Ischemia /reperfusion In Rats

Background and purpose: Neuronal cell death following cerebral ischemia is an extremely complex pathophysiologic process, and the cell death has two forms necrosis and apoptosis. Necrosis located in the core of the lesion, and apoptotic cell mainly occurs in the penumbra. With the therapy of thrombolysis, the reperfusion after cerebral ischemia may reduce the infarct volume obviously, while at the same time the existence of ischemia-reperfusion injury may induce apoptosis and enlarge the infarction size. In brief, the final infarct size is determined by apoptotic cells. To suppress apoptosis is the main target of clinical treatment. Fas ligand (FasL) is a type II membrane protein, which binding to fas, can initiates a chain of intracellular events that ultimately leads to activation of caspases and death of the target cell. Caspase-3 is one of the most important proteinases in caspase family, and amplifies the proteinase cascade, causes the cells move to death by and by. AIF is an apoptosis factor delivered by mitochondria, which can induce apoptosis through a caspase-independent pathway. Aspirin (ASA) is a classical non-steroidal anti-inflammatory drug. Recently researchs discovered that ASA also had the direct neuroprotective effects, such as increase of the ATP output, suppression of the glutamic acid release; decrease of iNOS expression, and increase nNOS expression; inhibition of NF-kB activation; elimination of the free radical; activation of cdk5/P35; prevention of Zn²⁺ inflow.However the reports about the anti-apoptotic mechanism of ASA are few. In the former experiment of this study showed that ASA could reduce the infarct volume, up-regulated the proportion of Bcl-2/Bax in rat, indicating the anti-apoptotic action of ASA. At present, the anti-apoptotic drugs mainly interfere with apoptotic stimulation, such as clearing free radical, stabilizing the Ca²⁺ imbalance, while the drugs which interfere with the apoptotic pathway are few. So the aim of this study is to clarify the anti-apoptotic mechanism of ASA and how ASA effect the apoptotic approaches by observing the expression of FasL, caspase-3 and AIF in rat MCA-CI/RP model.Methods: The rat model of MCA- CI/RP were established with suture occlusion technique according to modified Zea Longa method. TTC (Triphenyl tetrazolium chloride) staining was used to observe the infarction. The rats were divided into five groups according to the time of reperfusion in experimental groups (ASA groups): A (6h), B (24h), C (48h), D (4d) and E (7d), so did the control groups (A'^E'). ASA were given to the experimental groups with the dose of 80 mg · kg^-1 via an i.p. route, while the same dose of diaminocaproic acid were given to the control groups right after the CI/RP and the next few days till killed at assigned time. The neurologic detect scale were used by Bederson scores. The FasL, caspase-3 and AIF expression were detected by immunohistochemistry technique.Results: The rats presented neurologic defect in various degree after CI/RP. TTC stain may display the infarct size clearly, the infarct zone is pallor, while the normal tissue showed red in colour. The neurologic impairment of the rats was evaluated according to Bederson score. The scores of ASA group were lower than that of the control group during 4⁷ days p < 0.05. The expression of FasL could be detected six hours after CI/RP, reached to peak twenty-four hours, and disappeared in one week. The positive FasL cells were mainly seen in the neuron and the neuroglia, and there were some brown particles on the membrane or the cytoplasm.