PLoS ONE：对精液mRNA及MicroRNA的研究

无细胞（cell-free）的mRNA及microRNA (miRNA)能够作为临床应用的分子标记物以及胞内通讯的信号分子，都是关于它们的物理性质的研究还很缺乏。近日，华中科技大学同济医学院的研究人员研究对人类无细胞的精液mRNA(cfs-mRNA)及miRNA(cfs-miRNA)开展了一项有特别意义的研究。

在该实验中，精液特异的mRNAs、miRNAs及piRNAs在所有的实验中被实时定量PCR检测。

研究人员发现，虽然通过time-course分析表明，cfs-miRNA的稳定性与cfs-mRNA相似，但是对精浆采用过滤及Triton X-100处理后，cfs-miRNA及cfs-mRNA总量的变化特征却是不同的，这暗示了它们可能有不同的物理性质。随后，精液的超微小泡(SMVs)被恢复，SMVs中的cfs-mRNA及cfs-miRNA的含量也被定量。

SMVs内的cfs-mRNAs的含量几乎与其重量一致，并且在不同大小的SMVs中高度可变。而大多数的cfs-miRNA则独立于SMVs存在于上层清液。

通过对上层清液过滤及蛋白酶K的消化，进一步研究cfs-miRNA。实验中，cfs-miRNA通过了0.10μm大小的孔，但是在强烈的蛋白酶K的消化后被显著降解。

结果表明，cfs-mRNA主要存在于SMVs内，而大部分的cfs-miRNA结合于蛋白复合体。

该研究阐明了人类精液中细胞外RNAs的稳定性，表明了它们的来源及在男性繁殖中潜在的功能，也为发展它们作为男性生殖系统生物标记物提供了新的策略。相关论文发表在4月10日的PLoS ONE。（生物谷Deepblue编译）

doi: 10.1371/journal.pone.0034566
PMC:
PMID:

Cell-Free Seminal mRNA and MicroRNA Exist in Different Forms

Honggang Li, Shiyun Huang, Cuicui Guo, Huangtao Guan, Chengliang Xiong.

The great interest in cell-free mRNA, microRNA (miRNA) as molecular biomarkers for clinical applications, and as ‘signaling’ molecules for intercellular communication highlights the need to reveal their physical nature. Here this issue was explored in human cell-free seminal mRNA (cfs-mRNA) and miRNA (cfs-miRNA).Selected male reproductive organ-specific mRNAs, miRNAs, and piRNAs were quantified by quantitative real-time PCR in all experiments.While the stability of cfs-miRNA assessed by time-course analysis (up to 24 h at room temperature) was similar with cfs-mRNA, the reductive changes between cfs-miRNA and cfs-mRNA after filtration and Triton X-100 treatment on seminal plasma were very different, implying their different physical nature. Seminal microvesicles (SMVs) were then recovered and proportions of cfs-mRNA and cfs-miRNA within SMVs were quantified.The amounts of SMVs- sequestered cfs-mRNAs almost were the same as total cfs-mRNA, and were highly variable depending on the different sizes of SMVs. But most of cfs-miRNA was independent of SMVs and existed in the supernatant.The possible form of cfs-miRNA in the supernatant was further explored by filtration and protease K digestion. It passed through the 0.10-μm pore, but was degraded dramatically after intense protease K digestion.The predominant cfs-mRNA is contained in SMVs, while most cfs-miRNA is bound with protein complexes.Our data explained the stability of extracellular RNAs in human semen, and shed light on their origins and potential functions in male reproduction, and strategy of developing them as biomarkers of male reproductive system.