体外培养鸽嗉囊组织形态学、酶活力及基因表达动态变化规律

本试验旨在探究体外培养鸽嗉囊组织形态学、酶活力及基因表达变化规律。取60周龄美国王鸽嗉囊组织于体外培养7 d,动态检测其形态学、代谢和凋亡相关酶活力以及角蛋白和凋亡相关基因表达的变化。形态学结果表明:鸽嗉囊组织于体外培养第2~4天结构清晰,上皮层细胞排列紧密,体外培养5 d后嗉囊结构显著退化,形态学受到破坏。酶活力和基因表达结果显示:Caspase-3酶活力于培养第1、5、6、7天显著高于培养第2~4天(P0.05);琥珀酸脱氢酶、Na~+-K~+-ATP酶、Ca~(2+)-Mg~(2+)-ATP酶和总ATP酶活力以及Bcl-2和角蛋白19基因表达均于培养第1、6、7天显著下调(P0.05),于培养第2~4天显著升高(P0.05);Bak1基因表达于培养第2~5天稳定,随后于培养第6天显著上调(P0.05)。综上所述,体外培养鸽嗉囊组织形态学、酶活力和基因表达随培养时间延长而显著改变。本试验条件下,鸽嗉囊组织体外培养稳定时间为4 d,且开展后续药理、病理及生理学试验前的1 d预培养十分必要。

Dynamic Changes of Histological Structure,Enzymes Activities and Genes Expressions of Pigeon Crop in Tissue Culture in vitro

This study was conducted to investigate dynamic changes of histological structure, enzymes activities and genes expressions of pigeon crop in tissue culture in vitro. The crop from health White King meat-type pigeons(at the age of 60) were collected and cultivated for 7 days in vitro. Histological structure, activities of metabolism-and apoptosis-related enzymes and expressions of cytokertin-and apoptosis-related genes of pigeon crop were analyzed in tissue culture in vitro. The histological assessment indicated that structure of crop were integrated, cells in the epithelial layer lined up densely from 2 to 4 days after culture in vitro, then the epithelial structures of crop were severely degenerated after 5 days culture in vitro. The results of enzymeological assay and genes expressions showed that activity of Caspase-3 on the 1 st, 5 th, 6 th and 7 th day after culture was significantly higher than that on the 2 nd, 3 rd and 4 th day after culture in vitro(P<0.05). Activities of succinate dehydrogenase, Na~+-K~+-ATPase, Ca~(2+)-Mg~(2+)-ATPase and total ATP aseand genes expressions of Bcl-2 and cytokertin-19 were significantly reduced on the 1 st, 6 th and 7 th day of culture in vitro, and increased from 2 to 4 days after culture in vitro(P<0.05). Bak1 gene expression remained stable from 2 to 5 days after culture in vitro, then it was highly upregulated again on the 6 th day after culture in vitro(P<0.05). The results indicated that the histological structure, enzymes activities and genes expressions of pigeon crop changed significantly in tissue culture in vitro with the incubation going on, and the optimal culture time was 4 days.Additionally, pigeon crop tissueshould bepre-cultured in vitro for 1d before other researches of pharmacology, pathology and physiology.