Analysis of Protein Expression in Human Cells Cocultured with Porcine Peripheral Blood Mononuclear Cells
Analysis of Protein Expression in Human Cells Cocultured with Porcine Peripheral Blood Mononuclear Cells
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DOI：
10.1159/000495179
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发表时间：
2019-03
期刊：
影响因子：
4.6
通讯作者：
Yuyuan Ma;Xiong Zhao;J. Jia;Yong-xin Yang;Rui Fan;M. Lv;F. Ding;Jianmin Wu;Jingang Zhang
中科院分区：
文献类型：
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作者：
Yuyuan Ma;Xiong Zhao;J. Jia;Yong-xin Yang;Rui Fan;M. Lv;F. Ding;Jianmin Wu;Jingang Zhang
目的：猪内源性逆转录病毒（Porcine endogenous retroviruses，PERV）在猪体内普遍存在，并能在体外感染人细胞，因此在猪-人异种移植中引起了人们的极大关注。虽然PERV感染的人胚肾293（HEK 293）细胞上没有明显的细胞病变效应，但我们进行了蛋白质组学分析，以研究蛋白质谱的差异，以进一步表征PERV感染的效应。方法：将HEK 293细胞与猪外周血单个核细胞（PBMCs）共培养。PERV感染和非感染HEK 293细胞的蛋白质谱进行了分析，通过二维凝胶电泳（2-DE）。通过高清晰度质谱（HDMS）分析鉴定具有至少1.5倍改变的蛋白质点。采用实时荧光定量RT-PCR和Western blotting方法对蛋白质组学结果进行验证。结果：PERV感染和未感染HEK 293细胞的差异分析，通过2-DE显示10个差异调节蛋白。通过HDMS鉴定的蛋白质参与多种细胞途径，包括信号转导、细胞凋亡和蛋白质合成。结论：本研究的结果显示，差异表达的蛋白质在HEK 293细胞与猪PBMC共培养，并暗示这些变化可能是由PERV感染诱导。这些结果为了解嗜人PERV感染的分子效应提供了线索和潜在的联系。
Objective: Porcine endogenous retroviruses (PERV) involved in pig to human xenotransplantation have raised great concerns because of their ubiquitous nature in pigs and their ability of infecting human cells in vitro. Although no significant cytopathic effect attributed to PERV was evident on PERV-infected human embryonic kidney 293 (HEK293) cells, we did proteomic analysis to investigate the differences of protein profile in order to further characterize the effect of PERV infection. Methods: HEK293 cells were cocultured with porcine peripheral blood mononuclear cells (PBMCs). Protein profiles of PERV-infected and -noninfected HEK293 cells were analyzed by two-dimensional gel electrophoresis (2-DE). Protein spots with at least 1.5-fold alteration were identified by high-definition mass spectrometry (HDMS) analysis. Then real-time RT-PCR and Western blotting were performed to validate the proteomic results. Results: Differential analysis of PERV-infected and -noninfected HEK293 cells by 2-DE revealed ten differentially regulated proteins. The proteins identified by HDMS were involved in various cellular pathways including signal transduction, cell apoptosis, and protein synthesis. Conclusion: The results of this study revealed differentially expressed proteins in HEK293 cells cocultured with porcine PBMCs and implied that these changes were probably induced by PERV infection. These results provide clues and potential links to understanding the molecular effect of the infection by human-tropic PERV.