Pelisek Jaroslav, Yundung Yankey, Reutersberg Benedikt, Meuli Lorenz, Rössler Fabian, Rabin Laetitia, Kopp Reinhard, Zimmermann Alexander
Department of Vascular Surgery, University Hospital Zurich, 8091 Zurich, Switzerland.
Department of Surgery and Transplantation, University Hospital Zurich, 8091 Zurich, Switzerland.
J Clin Med. 2023 Aug 3;12(15):5109. doi: 10.3390/jcm12155109.
Proper biobanking is essential for obtaining reliable data, particularly for next-generation sequencing approaches. Diseased vascular tissues, having extended atherosclerotic pathologies, represent a particular challenge due to low RNA quality. In order to address this issue, we isolated RNA from vascular samples collected in our Swiss Vascular Biobank (SVB); these included abdominal aortic aneurysm (AAA), peripheral arterial disease (PAD), healthy aorta (HA), and muscle samples. We used different methods, investigated various admission solutions, determined RNA integrity numbers (RINs), and performed expression analyses of housekeeping genes (), ribosomal genes (, ), and long non-coding RNAs (, ). Our results show that RINs from diseased vascular tissue are low (2-4). If the isolation of primary cells is intended, as in our SVB, a cryoprotective solution is a better option for tissue preservation than RNAlater. Because RNA degradation proceeds randomly, controls with similar RINs are recommended. Otherwise, the data might convey differences in RNA degradation rather than the expressions of the corresponding genes. Moreover, since the 18S and 28S genes in the diseased vascular samples were degraded and corresponded with the low RINs, we believe that DV200, which represents the total RNA's disintegration state, is a better decision-making aid in choosing samples for omics analyses.
恰当的生物样本库建设对于获取可靠数据至关重要，特别是对于下一代测序方法而言。由于RNA质量较低，患有晚期动脉粥样硬化病变的患病血管组织带来了特殊挑战。为了解决这个问题，我们从我们的瑞士血管生物样本库（SVB）收集的血管样本中分离RNA；这些样本包括腹主动脉瘤（AAA）、外周动脉疾病（PAD）、健康主动脉（HA）和肌肉样本。我们使用了不同方法，研究了各种保存溶液，测定了RNA完整性数值（RIN），并对管家基因（ ）、核糖体基因（ ， ）和长链非编码RNA（ ， ）进行了表达分析。我们的结果表明，患病血管组织的RIN较低（2 - 4）。如果像我们的SVB那样打算分离原代细胞，对于组织保存而言，冷冻保护溶液比RNAlater是更好的选择。由于RNA降解是随机进行的，建议使用具有相似RIN的对照。否则，数据可能传达的是RNA降解的差异而非相应基因的表达差异。此外，由于患病血管样本中的18S和28S基因发生了降解且与低RIN值相对应，我们认为代表总RNA降解状态的DV200在选择用于组学分析的样本时是更好的决策辅助指标。
