Defining the lineage of thermogenic perivascular adipose tissue [Pup Aorta]

Brown adipose tissue can expend large amounts of energy and thus increasing its amount or activity is a promising therapeutic approach to combat metabolic disease. In humans, major deposits of brown fat cells are found intimately associated with large blood vessels, corresponding to perivascular adipose tissue (PVAT). However, the cellular origins of PVAT are poorly understood. We applied single cell transcriptomic analyses, ex vivo adipogenesis assays, and genetic fate mapping to determine the identity of perivascular adipocyte progenitors. In mice, we found that thoracic PVAT develops from a fibroblastic lineage, consisting of progenitor cells (Pdgfra+; Ly6a+; Pparg-) and preadipocytes (Pdgfra+; Ly6a+; Pparg+). Progenitor and preadipocyte cells in PVAT shared transcriptional similarity with analogous cell types in white adipose tissue, pointing towards a conserved hierarchical structure of adipose lineage cells. Interestingly, the aortic adventitia of adult animals contained a novel population of adipogenic smooth muscle cells (SMCs) (Myh11+; Pdgfra-; Pparg+) that contributed to perivascular adipocyte formation. Similarly, human PVAT contained presumptive fibroblastic and SMC-like adipocyte progenitors, as revealed by single nucleus RNAseq. Taken together, these studies define distinct populations of progenitor cells for thermogenic PVAT, providing a foundation for developing strategies to augment brown fat activity Transcriptome (mRNA) profiles of FACS isolated cell populations from the aorta and perivascular fat (intermediate cells, progenitor cells, preadipocytes, and smooth muscle cells (SMC)) from wild type (WT) 3 day old (P3) mixed male and female mouse pups were generated by deep sequencing, in triplicate, followed by DESeq2 analysis for differential expression of genes between the various groups.棕色脂肪组织（Brown adipose tissue）可消耗大量能量，因此提升其数量或活性是对抗代谢性疾病的极具潜力的治疗策略。在人类体内，大量棕色脂肪细胞沉积于大血管旁，对应血管周脂肪组织（perivascular adipose tissue, PVAT）。然而，目前对PVAT的细胞起源尚不清楚。本研究采用单细胞转录组分析、体外脂肪生成实验及遗传命运图谱技术，以明确血管周脂肪细胞祖细胞的身份。在小鼠中，我们发现胸腔PVAT起源于成纤维细胞谱系，该谱系由祖细胞（Pdgfra+; Ly6a+; Pparg-）和前脂肪细胞（Pdgfra+; Ly6a+; Pparg+）组成。PVAT中的祖细胞与前脂肪细胞与白色脂肪组织中的同类细胞具有转录相似性，提示脂肪谱系细胞存在保守的层级结构。值得注意的是，成年动物的主动脉外膜中存在一类新型成脂性平滑肌细胞（smooth muscle cells, SMC）（Myh11+; Pdgfra-; Pparg+），这类细胞可参与血管周脂肪细胞的生成。同样，通过单细胞核RNA测序（single nucleus RNAseq）分析发现，人类PVAT中存在推测的成纤维细胞样及平滑肌细胞样脂肪祖细胞。综上，本研究明确了产热型PVAT的不同祖细胞群，为开发增强棕色脂肪活性的策略奠定了基础。本研究对野生型（wild type, WT）3日龄（postnatal day 3, P3）雌雄混合小鼠幼崽的主动脉及血管周脂肪中经荧光激活细胞分选（fluorescence-activated cell sorting, FACS）分离得到的细胞群（包括中间细胞、祖细胞、前脂肪细胞及平滑肌细胞（SMC））的转录组（mRNA）谱进行了深度测序，每组设置三次生物学重复，随后采用DESeq2分析各组间的基因差异表达情况。