小柯机器人

瑞士洛桑联邦理工学院Bart Deplancke小组发现，确定性单细胞RNA测序（scRNA-seq）可捕捉肠隐窝和类器官组成的变化。2022年2月14日，《自然—方法学》杂志在线发表了这项成果。

研究人员开发了一个确定性的mRNA捕获珠和细胞共包裹的液滴系统，DisCo，旨在处理低投入的样品（<500个细胞）。研究人员证明DisCo通过结合机器视觉和多层微流控技术实现了精确的粒子和细胞定位以及液滴分选控制，能够以高捕获效率（>70%）和每小时350个细胞的速度连续处理低投入的单细胞悬浮液。为了突出Disco的独特能力，研究人员分析了31个处于不同发育阶段的单个肠道类器官。这显示了广泛的类器官异质性，并确定了不同的亚型，包括一个再生的胚胎样Ly6a⁺干细胞群，即使在分化条件下也能保持对称的隐窝或球状体，以及一个未定性的"球状体"亚型，主要由前体和成熟（Muc2⁺）球状体细胞组成。为了补充这个数据集，并证明DisCo处理低输入、体内衍生组织的能力，研究人员还分析了单个小鼠肠道隐窝。这表明存在着与球状体成分相似的隐窝，它们主要由再生干细胞组成，表明在平衡肠道中存在着再生隐窝。这些发现证明了DisCo在提供小型个体组织中细胞异质性高分辨率捕获方面的独特能力。

据悉，scRNA-seq方法已经改变了人们解析跨系统细胞特性的能力，但目前是针对大细胞输入（>1,000个细胞）的。这使得它们在处理小的、单独的组织样品时效率低下且成本高，这个问题往往通过加载大宗样品来解决，产生混杂的细胞群读数。

附：英文原文

Title: Deterministic scRNA-seq captures variation in intestinal crypt and organoid composition

Author: Bues, Johannes, Bioanin, Marjan, Pezoldt, Joern, Dainese, Riccardo, Chrisnandy, Antonius, Rezakhani, Saba, Saelens, Wouter, Gardeux, Vincent, Gupta, Revant, Sarkis, Rita, Russeil, Julie, Saeys, Yvan, Amstad, Esther, Claassen, Manfred, Lutolf, Matthias P., Deplancke, Bart

Issue&Volume: 2022-02-14

Abstract: Single-cell RNA sequencing (scRNA-seq) approaches have transformed our ability to resolve cellular properties across systems, but are currently tailored toward large cell inputs (>1,000 cells). This renders them inefficient and costly when processing small, individual tissue samples, a problem that tends to be resolved by loading bulk samples, yielding confounded mosaic cell population read-outs. Here, we developed a deterministic, mRNA-capture bead and cell co-encapsulation dropleting system, DisCo, aimed at processing low-input samples (<500 cells). We demonstrate that DisCo enables precise particle and cell positioning and droplet sorting control through combined machine-vision and multilayer microfluidics, enabling continuous processing of low-input single-cell suspensions at high capture efficiency (>70%) and at speeds up to 350 cells per hour. To underscore DisCo’s unique capabilities, we analyzed 31 individual intestinal organoids at varying developmental stages. This revealed extensive organoid heterogeneity, identifying distinct subtypes including a regenerative fetal-like Ly6a+ stem cell population that persists as symmetrical cysts, or spheroids, even under differentiation conditions, and an uncharacterized ‘gobloid’ subtype consisting predominantly of precursor and mature (Muc2+) goblet cells. To complement this dataset and to demonstrate DisCo’s capacity to process low-input, in vivo-derived tissues, we also analyzed individual mouse intestinal crypts. This revealed the existence of crypts with a compositional similarity to spheroids, which consisted predominantly of regenerative stem cells, suggesting the existence of regenerating crypts in the homeostatic intestine. These findings demonstrate the unique power of DisCo in providing high-resolution snapshots of cellular heterogeneity in small, individual tissues.

DOI: 10.1038/s41592-021-01391-1

Source: https://www.nature.com/articles/s41592-021-01391-1

Nature Methods：《自然—方法学》，创刊于2004年。隶属于施普林格·自然出版集团，最新IF：47.99
官方网址：https://www.nature.com/nmeth/
投稿链接：https://mts-nmeth.nature.com/cgi-bin/main.plex