| Objective: Cerebrovascular disease (CVD) is commonly and frequently encountered , usually with acute onset, which is an emergent disease endangering the people's life and health because of its high mortality and serious disability. Cerebral ischemia accounts for seventy percents of CVD. Temperature is an important factor affecting the prognosis of cerebral ischemia. Pyrexia may be directly or indirectly correlated with stroke via several mechanisms. A vast amount of clinical and experimental researches have demonstrated that mild hypothermia therapy has a definite effect on cerebrovascular diseases, However, its specific mechanism are not very clear. Hypothermia may inhibit various mechanism of ischemic injury in neuron. Survivin is a new member of the inhibitor of apoptosis protein and has a significant effect on anti-apoptosis. The recent investigations of survivin were all located in the tumor therapy because of its unique disposition. There were few reports about survivin expression in brain tissue after ischemia. Bcl-2 was firstly separated from follicular lymphoma, it has been confirmed as a gene to inhibit cell's apoptosis. These cells include tumor cell, myocardial cell, vascular endothelial cell, nerve cell, et al. In this study, we practiced hypothermia on cerebral ischemic animal model to observe the Bcl-2 and survivin expression in brain tissue, and try to explain the mechanism of hypothermia.Methods: Fifty-four male SD rats were randomly divided into a sham-operated group (eighteen rats, n=18) and a ischemia group(thirty-six rats, n=36), the latter also was divided into a normothermia group and a mild hypothermia group. Every group was observed at three time points(12,24,48h), and there were six rats in every time point(n=6). Model building: Every SD rat was treated with modified artery occlusion by a clew. The brain temperature was lowered to 32-34°C in focal cerebral ischemia rodent models by cooling therapy, started at once after the ischemia onset and maintained 4-6h, then rewarming slowly. The temperature of normothermia group maintained as incipient temperature. The sham-operated group was only anesthetized to expose blood vessel, without clew. At every time point after operated, all the rats were examined by neurological deficit scoring with three methods. Then in every group, the rats(n=6) were overdose anesthetized and the brains were taken out. Six coronal brain sections prepared at 2-mm intervals. The section 4 was formalin fixation with 4% paraform, then was imbedded in paraffin after 24h. The brain slices were stained with hematoxylin and eosin (HE) for histopathological evaluation and with routine immunohistochemical methods for the expression of Survivin and Bcl-2 in and around ischemia and other brain area. The infarct volume was examined with 2,3,5-triphenyl-tetrazolium chloride staining.Results1 The neurological deficit scores of mild hypothermia group(2.17±0.75) resulted in significant difference as compared with the normothermia group(3.17±0.75). P<0.05.2 Microscopic examination of HE stained sections revealed that brain infarct area is poorly stained involving the ipsilateral side of the middle cerebral artery in groups of the ischemia. In this brain region, endothelial cells damage and thrombus were observed at the middle artery and perforating arteries. In cortex and hippocampus, the cellular nucleus were crenulated, the endochylema vacuolization were observed. No brain infarct was observed in sham-operated group. In this group, the endochylema is satiated and stained evenly.3 A significant reduction in infarct volume was found in the hypothermia group(0.12±0.02) compared with the normothermia group(0.15±0.01). P<0.05.4 The results of immunohistochemistry reveal that Bcl-2 expression is significant shortly after ischemia. The masculine cells of Bcl-2 were mainly localized in neuronal nucleolemma and cell membrane, not in endochylema. Compared to the normothermia group(57.30±3.39), the number of masculine cells in hypothermia group(72.83±4.58) increased markedly. There were slight expression of Bcl-2 in the sham-operated group(6.83±4.31). The difference has statistical significance. P<0.05. The masculine cells of surviving were mainly localized in neuronal endochylema and vascular endothelial cell. Compared to the normothermia group(35.83±4.58), the number of masculine cells in hypothermia group(69.33±8.69) increased markedly. There were slight expression of survivin in the sham-operated group(5.32±1.02). The difference also has statistical significance. P<0.05.Conclusions1 Mild hypothermia is a effective neuroprotective measure to date. Mild hypothermia may promote Bcl-2 and surviving expression and affect neuron-related gene expression, and reduce neuronal apoptosis caused by cerebral ischemia and thus protect the brain.2 Survivin, as a new member of anti-apoptosis family, can express in the brain tissue after ischemia and hypoxia.3 The expression of Bcl-2 has the same tendency with survivin. These two genes may protect the brain tissue damage synergistically through different anti-apoptosis pathway.
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