Scott S Spencer, Yang Xuehui, Robich Michael, Liaw Lucy, Boucher Joshua M
Maine Medical Center Research Institute.
Maine Medical Center Research Institute;
J Vis Exp. 2019 Mar 5(145). doi: 10.3791/59337.
Adipose tissue is a rich source of multi-potent mesenchymal stem cells (MSC) capable of differentiating into osteogenic, adipogenic, and chondrogenic lineages. Adipogenic differentiation of progenitor cells is a major mechanism driving adipose tissue expansion and dysfunction in response to obesity. Understanding changes to perivascular adipose tissue (PVAT) is thus clinically relevant in metabolic disease. However, previous studies have been predominately performed in the mouse and other animal models. This protocol uses human thoracic PVAT samples collected from patients undergoing coronary artery bypass graft surgery. Adipose tissue from the ascending aorta was collected and used for explantation of the stromal vascular fraction. We previously confirmed the presence of adipose progenitor cells in human PVAT with the capacity to differentiate into lipid-containing adipocytes. In this study, we further analyzed the differentiation potential of cells from the stromal vascular fraction, presumably containing multi-potent progenitor cells. We compared PVAT-derived cells to human bone marrow MSC for differentiation into adipogenic, osteogenic, and chondrogenic lineages. Following 14 days of differentiation, specific stains were utilized to detect lipid accumulation in adipocytes (Oil red O), calcific deposits in osteogenic cells (Alizarin Red), or glycosaminoglycans and collagen in chondrogenic cells (Masson's Trichrome). While bone marrow MSC efficiently differentiated into all three lineages, PVAT-derived cells had adipogenic and chondrogenic potential, but lacked robust osteogenic potential.
脂肪组织是多能间充质干细胞(MSC)的丰富来源,这些干细胞能够分化为成骨、成脂和软骨生成谱系。祖细胞的成脂分化是驱动脂肪组织扩张和肥胖相关功能障碍的主要机制。因此,了解血管周围脂肪组织(PVAT)的变化在代谢疾病中具有临床相关性。然而,以往的研究主要是在小鼠和其他动物模型中进行的。本方案使用从接受冠状动脉搭桥手术的患者身上采集的人胸部PVAT样本。收集升主动脉的脂肪组织并用于基质血管成分的外植培养。我们之前已证实在人PVAT中存在脂肪祖细胞,它们有分化为含脂脂肪细胞的能力。在本研究中,我们进一步分析了基质血管成分中细胞的分化潜能,这些细胞可能含有多能祖细胞。我们将PVAT来源的细胞与人类骨髓MSC进行比较,观察它们向成脂、成骨和软骨生成谱系的分化情况。分化14天后,使用特定染色检测脂肪细胞中的脂质积累(油红O)、成骨细胞中的钙沉积(茜素红)或软骨生成细胞中的糖胺聚糖和胶原蛋白(Masson三色染色法)。虽然骨髓MSC能有效地分化为所有这三种谱系,但PVAT来源的细胞具有成脂和软骨生成潜能,却缺乏强大的成骨潜能。