Peics Julia, Vishvanath Lavanya, Zhang Qianbin, Shan Bo, Pedersen Thomas Å, Gupta Rana K
Diabetes Pharmacology, Novo Nordisk A/S.
Touchstone Diabetes Center, Department of Internal Medicine, University of Texas Southwestern Medical Center.
J Vis Exp. 2020 Aug 16(162). doi: 10.3791/61610.
The stromal-vascular fraction (SVF) of white adipose tissue (WAT) is remarkably heterogeneous and consists of numerous cell types that contribute functionally to the expansion and remodeling of WAT in adulthood. A tremendous barrier to studying the implications of this cellular heterogeneity is the inability to readily isolate functionally distinct cell subpopulations from WAT SVF for in vitro and in vivo analyses. Single-cell sequencing technology has recently identified functionally distinct fibro-inflammatory and adipogenic PDGFRβ+ perivascular cell subpopulations in intra-abdominal WAT depots of adult mice. Fibro-inflammatory progenitors (termed, "FIPs") are non-adipogenic collagen producing cells that can exert a pro-inflammatory phenotype. PDGFRβ+ adipocyte precursor cells (APCs) are highly adipogenic both in vitro and in vivo upon cell transplantation. Here, we describe multiple methods for the isolation of these stromal cell subpopulations from murine intra-abdominal WAT depots. FIPs and APCs can be isolated by fluorescence-activated cell sorting (FACS) or by taking advantage of biotinylated antibody-based immunomagnetic bead technology. Isolated cells can be used for molecular and functional analysis. Studying the functional properties of stromal cell subpopulation in isolation will expand our current knowledge of adipose tissue remodeling under physiological or pathological conditions on the cellular level.
白色脂肪组织（WAT）的基质血管部分（SVF）具有显著的异质性，由多种细胞类型组成，这些细胞类型在成年期对WAT的扩张和重塑发挥着功能性作用。研究这种细胞异质性影响的一个巨大障碍是无法轻易地从WAT SVF中分离出功能不同的细胞亚群用于体外和体内分析。单细胞测序技术最近在成年小鼠的腹内WAT库中鉴定出了功能不同的成纤维细胞炎症性和脂肪生成性PDGFRβ + 血管周围细胞亚群。成纤维细胞炎症祖细胞（称为“FIPs”）是不产生脂肪的胶原蛋白产生细胞，可表现出促炎表型。PDGFRβ + 脂肪细胞前体细胞（APCs）在细胞移植后在体外和体内都具有高度的脂肪生成能力。在这里，我们描述了从鼠腹内WAT库中分离这些基质细胞亚群的多种方法。FIPs和APCs可以通过荧光激活细胞分选（FACS）或利用基于生物素化抗体的免疫磁珠技术进行分离。分离出的细胞可用于分子和功能分析。单独研究基质细胞亚群的功能特性将在细胞水平上扩展我们目前对生理或病理条件下脂肪组织重塑的认识。
