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寒冷刺激下鱼类细胞驯化适应机制研究
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摘要
在常年寒冷的环境下,南极鱼类逐步适应并形成了一系列抗寒抗冻的机制。为了研究南极鱼的抗寒抗冻机制,我们将斑马鱼胚胎成纤维样细胞(ZF4细胞)于28℃野生型(对照组)或18℃培养驯化。两种细胞传代15代以上后放入10℃进行处理,我们发现18℃驯化后经过驯化的细胞凋亡程度明显降低。核型分析结果显示18℃驯化后的细胞更趋向于成为四倍体,细胞经PI染色也证实驯化后细胞内的DNA含量较未驯化的高。这表明在持续性冷刺激下细胞内的基因组可能会出现扩增。此外,驯化后细胞内线功能性线粒体的含量较未驯化细胞显著上调。为了探究这一现象所涉及的通路,我们又将正常培养的细胞分别在18℃、13℃、10℃短期处理7天并检测线粒体含量。结果显示:28℃野生型ZF4细胞在不同温度处理7d时,功能性线粒体的含量显著提升,但是不同温度处理的细胞线粒体数量并未出现显著性差异。由此而知,鱼类为了适应寒冷环境需要更多基因和蛋白的表达,以维持正常的生理代谢,而其中涉及的具体通路及调控与进化的机制还有待进一步探究。
Perennially in cold,Antarctic fish gradually formed a series of cold resistance mechanism of antifreeze.To identify the antifreeze mechanism of Antarctic fish,ZF4 cells were cultured under 18℃ for cold acclimation,and 28℃ as control.Cells which cultured more than 15 generations were treated under severe cold stress(10 ℃). Our data showed that cells underwent cold acclimation had a lower level of apoptosis. The results of karyotype analysis illustrated that cells underwent cold acclimation were tend to be tetraploid. And PI staining also confirmed that the DNA content in the cells after acclimation was higher than that of non-acclimated cells. It indicated that the genome doubling may occur when cells under persistent cold stress. The level of functional mitochondria of acclimated cells was significantly up-regulated. To explore the pathways involved in this phenomenon, we treated normal cultured cells at 18℃, 13℃, and 10℃for 7days and assayed for mitochondrial content.
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