【關鍵詞】 高溫；神經元；骨架蛋白；熱休克蛋白質70
【Abstract】 AIM: To observe the effects of hyperthermia (39℃) on the expressions of skeleton protein (βtubulin) and heat shock protein 70 (HSP70) of cortical neurons in vitro cultured and explore the neuroprotective effect of HSP70. METHODS: The cerebral cortical neurons of embryonic mice were cultured. The cultured neurons were divided as control group which was cultured at 37℃ and hyperthermia group which was put in high temperature (39℃) for 30 min. The immunofluorescence and laser scanning confocal imaging were used to detect HSP70 and βtubulin expressions at different time points (0, 2, 6, 24, 72 h) after hyperthermia. RESULTS: The fluorescence intensity of βtubulin in hyperthermia group was lower than that in control group, and it increased as recovery time went along. The fluorescence intensity of HSP70 in former group was higher than that in the latter, and it increased firstly and then decreased. The level of HSP70 reached the highest level at 2 h and remained a high level for 24 h. The expression of HSP70 was in nucleus after hyperthermia and then in cytoplasm as time went on. CONCLUSION: Hyperthermia may lead to the structural change of the cerebral cortical neuron probably through influencing skeleton protein structure. HSP70 may be involved in the protection of neurons in hyperthermia.
【Keywords】 hyperthermia; neurons; skeleton protein; Heat shock proteins 70
【摘要】 目的： 觀察39℃環境對體外培養的小鼠皮層神經元骨架蛋白(βtubulin)和熱休克蛋白70（HSP70）表達的影響，探討HSP70的神經保護作用. 方法： 取小鼠胚胎大腦皮層進行神經元原代培養，將培養好的神經元分為37℃培養的對照組和39℃刺激30 min後0, 2, 6, 24和72 h的高溫組. 分別進行熒光免疫染色，應用鐳射共聚焦掃描觀察神經元βtubulin和HSP70的表達. 結果： 39℃刺激後，高溫組神經元βtubulin熒光強度均低於37℃對照組，且隨著恢復時間的'延長其熒光強度值逐漸升高. 高溫組神經元HSP70熒光強度均高於37℃對照組，且隨著恢復時間的延長其熒光強度值先升高後降低，於2 h時達高峰，24 h仍維持在較高的水平. 高溫可導致HSP70熒光分佈轉移至細胞核，並隨著恢復時間的延長逐漸轉移至細胞漿. 結論： 高溫可以導致神經元形態變化，骨架蛋白結構紊亂可能是神經元形態變化的重要原因之一，HSP70可能參與並起到保護作用.
【關鍵詞】 高溫；神經元；骨架蛋白；熱休克蛋白質70
0引言
高溫環境下，當人體內溫度上升到40℃以上時，就會出現中樞神經系統功能障礙如：譫妄、驚厥或昏迷，甚至死亡. 但其複雜的詳細機制目前仍不清楚. 機體在高溫環境下受到熱應激時，正常的蛋白質合成會受到抑制，但一組高度保守的熱休克蛋白（heat shock proteins, HSPs）會迅速合成［1］. 熱休克蛋白70（heat shock protein 70, HSP70）是熱休克蛋白家族中的主要成員之一，是重要的分子伴侶. 在各種細胞器中，核仁和細胞膜對熱刺激特別敏感，HSP70廣泛存在於細胞骨架和核骨架內. 為了探討高溫環境下神經元HSP70和βtubulin表達的變化，我們採用體外培養的神經元，模擬高溫環境，應用鐳射共聚焦掃描方法觀察了高溫刺激對神經元HSP70和βtubulin表達的影響，並初步探討其可能的生物學意義.
1材料和方法
