The content of DNA and RNA in microparticles released by Jurkat and HL-60 cells undergoing in vitro apoptosis
The content of DNA and RNA in microparticles released by Jurkat and HL-60 cells undergoing in vitro apoptosis
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DOI：
10.1016/j.yexcr.2008.12.014
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发表时间：
2009-03-10
影响因子：
3.7
通讯作者：
Pisetsky, David S.
中科院分区：
文献类型：
--
作者：
Reich, Charles F., III;Pisetsky, David S.
微粒是小的膜结合囊泡，其在起泡期间从凋亡细胞释放。这些颗粒含有DNA和RNA，并显示重要的功能活动，包括免疫系统激活。此外，颗粒内的核酸可以作为各种疾病状态下的生物标志物进行分析。为了阐明微粒核酸的性质，研究了在体外经历凋亡的Jurkat T和HL-60早幼粒细胞百合的微粒中释放的DNA和RNA。通过差速离心从培养基中分离微粒，并通过流式细胞术和分子方法进行表征。在这些颗粒中，DNA通过凝胶电泳显示出梯状，并且以允许单克隆抗DNA抗体直接结合的形式存在，这表明即使没有固定，抗原也是可接近的。通过凝胶电泳分析RNA显示完整的18 s和28 s核糖体RNA条带，尽管也存在与28 s核糖体RNA降解产物一致的低分子条带。颗粒还含有信使RNA，如通过β-肌动蛋白和GAPDH序列的RT-PCR扩增所示。此外，凝胶电泳显示在microRNA的大小范围内存在低分子量RNA。总之，这些结果表明，来自凋亡Jurkat和HL-60细胞的微粒含有不同的核酸种类，表明细胞核和细胞质DNA和RNA的易位作为死亡期间的微粒释放载体。(C)2008年爱思唯尔公司All rights reserved.
Microparticles are small membrane-bound vesicles that are released from apoptotic cells during blebbing. These particles contain DNA and RNA and display important functional activities, including immune system activation. Furthermore, nucleic acids inside the particle can be analyzed as biomarkers in a variety of disease states. To elucidate the nature of microparticle nucleic acids, DNA and RNA released in microparticles from the Jurkat T and HL-60 promyelocytic cell lilies undergoing apoptosis in vitro were studied. Microparticles were isolated from Culture media by differential centrifugation and characterized by now cytometry and molecular approaches. In these particles, DNA showed laddering by gel electrophoresis and was present in a form that allowed direct binding by a monoclonal anti-DNA antibody, Suggesting antigen accessibility even without fixation. Analysis of RNA by gel electrophoresis showed intact 18s and 28s ribosomal RNA bands, although lower molecular bands consistent with 28s ribosomal RNA degradation products were also present. Particles also contained messenger RNA as shown by RT-PCR amplification of sequences for beta-actin and GAPDH. In addition, gel electrophoresis showed the presence of low molecular weight RNA in the size range of microRNA. Together, these results indicate that microparticles from apoptotic Jurkat and HL-60 cells contain diverse nucleic acid species, indicating translocation of both nuclear and cytoplasmic DNA and RNA as particle release Occurs during death. (C) 2008 Elsevier Inc. All rights reserved.