DataSheet_2_Proteome profiling of whole plasma and plasm\a-derived extracellular vesicles facilitates the detection of tissue biomarkers in the non-obese diabetic mouse.xlsx

The mechanism by which pancreatic beta cells are destroyed in type 1 diabetes (T1D) remains to be fully understood. Recent observations indicate that the disease may arise because of different pathobiological mechanisms (endotypes). The discovery of one or several protein biomarkers measurable in readily available liquid biopsies (e.g. blood plasma) during the pre-diabetic period may enable personalized disease interventions. Recent studies have shown that extracellular vesicles (EVs) are a source of tissue proteins in liquid biopsies. Using plasma samples collected from pre-diabetic non-obese diabetic (NOD) mice (an experimental model of T1D) we addressed if combined analysis of whole plasma samples and plasma-derived EV fractions increases the number of unique proteins identified by mass spectrometry (MS) compared to the analysis of whole plasma samples alone. LC-MS/MS analysis of plasma samples depleted of abundant proteins and subjected to peptide fractionation identified more than 2300 proteins, while the analysis of EV-enriched plasma samples identified more than 600 proteins. Of the proteins detected in EV-enriched samples, more than a third were not identified in whole plasma samples and many were classified as either tissue-enriched or of tissue-specific origin. In conclusion, parallel profiling of EV-enriched plasma fractions and whole plasma samples increases the overall proteome depth and facilitates the discovery of tissue-enriched proteins in plasma. If applied to plasma samples collected longitudinally from the NOD mouse or from models with other pathobiological mechanisms, the integrated proteome profiling scheme described herein may be useful for the discovery of new and potentially endotype specific biomarkers in T1D.

1型糖尿病（type 1 diabetes, T1D）中胰腺β细胞（pancreatic beta cells）的破坏机制尚未完全阐明。近期研究表明，该疾病的发生可能源于不同的病理生物学机制（内型，endotypes）。在糖尿病前期阶段，于易获取的液体活检（liquid biopsies，如血浆）中可检测到一种或多种蛋白质生物标志物，这一发现有望实现个体化的疾病干预。

近期研究证实，细胞外囊泡（extracellular vesicles, EVs）是液体活检中组织蛋白质的来源之一。本研究利用从糖尿病前期非肥胖糖尿病（non-obese diabetic, NOD）小鼠（T1D的实验模型）中采集的血浆样本，旨在探究相较于单独分析全血浆样本，联合分析全血浆样本与血浆来源的EV组分是否能提升质谱（mass spectrometry, MS）鉴定得到的独特蛋白质数量。

对去除高丰度蛋白并经肽段分级分离的血浆样本进行液相色谱-串联质谱（LC-MS/MS）分析，共鉴定到超过2300种蛋白质；而对富集EV的血浆样本进行分析，则鉴定到超过600种蛋白质。在富集EV的样本中检测到的蛋白质中，超过三分之一未在全血浆样本中被鉴定出，且其中多数被归类为组织富集型或组织特异性起源蛋白。

综上，同时分析富集EV的血浆组分与全血浆样本，可提升整体蛋白质组深度，助力发现血浆中的组织富集型蛋白质。若将本研究提出的整合蛋白质组分析方案应用于NOD小鼠或其他具有不同病理生物学机制的模型的纵向采集血浆样本中，或可用于发现T1D中新型且可能具有内型特异性的生物标志物。