Bassols Judit, Bonet Sergi, Belghazi Maya, Dacheux Françoise, Dacheux Jean-Louis
Biotechnology of Porcine Reproduction, Department of Biology, Faculty of Sciences, University of Girona, Campus de Montilivi s/n, 17071 Girona, Spain.
Theriogenology. 2007 Jul 1;68(1):76-86. doi: 10.1016/j.theriogenology.2007.04.009. Epub 2007 May 21.
A proteomic approach was used in this study to follow the protein expression of epididymal cells during the different phases of a cell culture protocol which was able to obtain an epididymal cell monolayer. The secretory activity of intact proximal and middle caput epididymal fragments and caput, corpus and cauda epithelial cell monolayers was examined on different days of culture. Transcriptomic activity was also followed by RT-PCR for the mRNA of several previously identified major proteins. During the establishment of epididymal cell cultures, a progressive shift was found in the pattern of protein secretion. The normal epididymal protein profile, specific for each epididymal region, was progressively replaced by a less specific profile with the secretion of new proteins. A correlation between protein secretion and the presence of the mRNA of the marker proteins was observed only in the first phase of culture. Most of the new proteins which appeared were characteristic of the secretion of cell monolayers cultivated over several weeks. Despite the significant modifications of the epididymal cell secretome, the presence of new proteins secreted only by cell cultures originating from a specific epididymal region shows the presence of remaining endogenous differentiation.
在本研究中,采用蛋白质组学方法追踪细胞培养方案不同阶段附睾细胞的蛋白质表达情况,该方案能够获得附睾细胞单层。在培养的不同天数,检测完整的附睾近端和附睾头中段片段以及附睾头、体和尾上皮细胞单层的分泌活性。还通过RT-PCR对几种先前鉴定的主要蛋白质的mRNA进行转录组活性追踪。在建立附睾细胞培养过程中,发现蛋白质分泌模式逐渐发生变化。每个附睾区域特有的正常附睾蛋白质谱逐渐被一种不太特异的谱所取代,同时分泌新的蛋白质。仅在培养的第一阶段观察到蛋白质分泌与标记蛋白mRNA的存在之间存在相关性。出现的大多数新蛋白质是培养数周的细胞单层分泌的特征。尽管附睾细胞分泌组有显著改变,但仅由特定附睾区域来源的细胞培养物分泌的新蛋白质的存在表明仍存在内源性分化。