CAJ | 학술논문

组织构建；组织工程；热打击；神经元；细胞凋亡；核因子κB；活性氧；caspase-3；抗-核因子κB65蛋白；国家自然科学基金 　　背景：大量研究显示高热可诱导机体细胞发生广泛凋亡，但对于高热如何介导神经元细胞凋亡并没有深入研究报道。 　　目的：检测热打击细胞模型中核因子κB信号通路对活性氧诱导细胞凋亡的影响。方法：使用细胞培养箱建立细胞热打击模型，热打击组分别将细胞置于39，41，43℃培养箱中进行热打击2 h，对照组(37℃组)将细胞置于标准37℃、体积分数5%CO 2细胞培养箱。使用Annexin V-FITC/PI双染色方法检测不同温度热打击下神经元细胞凋亡率，Westen blot 检测caspase-3及抗-核因子КB65蛋白表达，DCFH法检测细胞内活性氧含量，同时检测活性氧抑制剂MnTMPyP及核因子κB抑制剂PDTC对热打击细胞凋亡的影响。 　　结果与结论：39℃热打击对细胞凋亡无影响，41℃热打击诱导细胞少量凋亡(10.19%)，43℃热打击诱导诱导细胞大量凋亡(43.02%)。caspase-3和抗-核因子κB65蛋白的表达于热打击温度依赖的方式增加。MnTMPyP及PDTC均可有效阻断热打击引起的caspase-3、抗-核因子κB65蛋白的表达和细胞凋亡。结果证实热打击后细胞内活性氧增加诱导神经元细胞凋亡，提示核因子κB可能作为中间信号通路参与了热打击引起的细胞凋亡。
조직구건；조직공정；열타격；신경원；세포조망；핵인자κB；활성양；caspase-3；항-핵인자κB65단백；국가자연과학기금 　　배경：대량연구현시고열가유도궤체세포발생엄범조망，단대우고열여하개도신경원세포조망병몰유심입연구보도。 　　목적：검측열타격세포모형중핵인자κB신호통로대활성양유도세포조망적영향。방법：사용세포배양상건립세포열타격모형，열타격조분별장세포치우39，41，43℃배양상중진행열타격2 h，대조조(37℃조)장세포치우표준37℃、체적분수5%CO 2세포배양상。사용Annexin V-FITC/PI쌍염색방법검측불동온도열타격하신경원세포조망솔，Westen blot 검측caspase-3급항-핵인자КB65단백표체，DCFH법검측세포내활성양함량，동시검측활성양억제제MnTMPyP급핵인자κB억제제PDTC대열타격세포조망적영향。 　　결과여결론：39℃열타격대세포조망무영향，41℃열타격유도세포소량조망(10.19%)，43℃열타격유도유도세포대량조망(43.02%)。caspase-3화항-핵인자κB65단백적표체우열타격온도의뢰적방식증가。MnTMPyP급PDTC균가유효조단열타격인기적caspase-3、항-핵인자κB65단백적표체화세포조망。결과증실열타격후세포내활성양증가유도신경원세포조망，제시핵인자κB가능작위중간신호통로삼여료열타격인기적세포조망。
BACKGROUND:Hyperpyrexia can induce a wide range of cel apoptosis in organisms, but no study has introduced the mechanism of heat stress-induced neuronal apoptosis. OBJECTIVE:To observe the effect of nuclear factor kappa B (NF-κB) signal pathway on heat stress-induced neuronal apoptosis through reactive oxygen species. METHODS:Heat stress model was established in the cel incubator. Heat stress group of cel s were incubated at 39,41,43℃for2hours,whilecontrolgroupofcelswereincubatedat37 ℃in5% CO2 for 2 hours. Apoptosis was analyzed by flow cytometry using Annexin V-FITC/PI staining. The expression levels of caspase-3 and p-NF-κB65 were determined by western blot analysis. The amounts of intracel ular reactive oxygen species were assayed by DCFH staining. In addition, the effect of MnTMPyP and PTDC on heat stress-induced apoptosis was also studied. RESULTS AND CONCLUSION:39 ℃ heat stress had no impact on the apoptosis, 41 ℃ heat stress induced a smal amount of apoptosis (10.19%), and 43 ℃ heat stress triggered a large amount of apoptosis (43.02%). The expression of caspase-3 and p-NF-κB65 was increased, in a temperature-dependent manner. In addition, both MnTMPyP and PTDC significantly decreased the heat stress-induced apoptosis and expression of caspase-3 and p-NF-κB65. Experimental findings indicate that, the increase of intracel ular reactive oxygen species may induce neuronal apoptosis, and NF-κB participates in the heat stress-induced neuronal apoptosis as the intermedial signal pathway.