Circulating extracellular RNA markers of liver regeneration

Background and Aims: Although the liver can regenerate after sustaining injury, the tools to detect ongoing regeneration are lacking. The restoration of the liver after hepatectomy involves rapid, sequential, and coordinated changes in gene expression that result in systemic and local changes, such as the activation of progenitor cell populations and proliferation of quiescent hepatocytes. We postulated that changes in gene expression and regenerative programs could be activated within target cells through the transfer of extracellular RNA across cells or within the circulation, coordinating tissue response, and these extracellular RNA could represent biomarkers of the hepatic regenerative response. Methods: We identified temporal changes in circulating extracellular RNA at selected time periods up to 24 hours after partial hepatectomy in mice. (CD-1 males) Results: A peak increase in extracellular RNA content occurred 6 hours after hepatectomy. RNA sequencing was performed to identify the circulating extracellular non-coding RNA released after partial hepatectomy. To identify candidate markers, we used bioinformatics analyses to find the enriched RNA in the circulation after hepatectomy. We then performed tissue expression analysis for selected non-coding RNA and correlated it to these findings. Finally, a digital PCR assay was used to detect and quantify the expression of selected extracellular RNA in serum samples. Conclusions: Extracellular RNA that were selectively enriched during acute regeneration were detected within the circulation and represent biomarkers of ongoing liver regeneration in mice. The ability to detect ongoing active regeneration would benefit the evaluation of hepatic recovery after liver injury. Compare small RNA expression in time points after partial hepatectomy in CD-1 mice.背景与目的：尽管肝脏在遭受损伤后具备再生能力，但目前仍缺乏能够检测其正在进行的再生过程的工具。肝部分切除术（partial hepatectomy）后肝脏的修复过程涉及基因表达层面快速、有序且协同的变化，进而引发全身与局部的改变，例如祖细胞群体的激活以及静息肝细胞（hepatocyte）的增殖。我们提出假说：靶细胞内的基因表达与再生程序的改变，可通过细胞间或循环系统中的细胞外RNA（extracellular RNA）传递得以激活，以此协调组织应答反应；而这类细胞外RNA或可作为肝脏再生应答的生物标志物（biomarker）。 方法：我们针对CD-1品系雄性小鼠，在肝部分切除术后至多24小时的选定时间节点，检测了循环中外源性细胞外RNA的时序变化。 结果：肝部分切除术后6小时，细胞外RNA含量出现峰值升高。我们通过RNA测序（RNA sequencing）鉴定了肝部分切除术后释放至循环系统中的细胞外非编码RNA（non-coding RNA）。为筛选候选标志物，我们借助生物信息学分析找出了肝部分切除术后循环系统中富集的RNA。随后针对选定的非编码RNA开展组织表达分析，并将其与上述结果进行关联验证。最终采用数字PCR（digital PCR）检测与定量血清样本中选定的细胞外RNA的表达水平。 结论：在急性再生阶段选择性富集的细胞外RNA可在循环系统中被检测到，可作为小鼠体内正在进行的肝脏再生的生物标志物。能够检测当前活跃的再生过程，将有助于评估肝损伤后的肝脏恢复情况。本研究比较了CD-1小鼠肝部分切除术后不同时间点的小RNA表达情况。