Faculty of Medicine and Life Sciences, FICAM, University of Tampere, Tampere, Finland.
Cell Biology, Faculty of Medicine and Life Sciences, University of Tampere, Tampere, Finland.
Basic Clin Pharmacol Toxicol. 2018 Sep;123 Suppl 5:62-71. doi: 10.1111/bcpt.12987. Epub 2018 Apr 27.
Many adipose tissue-related diseases, such as obesity and type 2 diabetes, are worldwide epidemics. For studying these diseases, relevant human cell models are needed. In this study, we developed a vascularized adipose tissue model where human adipose stromal cells and human umbilical cord vein endothelial cells were cocultured with natural adipogenic and defined serum-free angiogenic media for 14 days. Several different protocols were compared to each other. The protocols varied in cell numbers and plating sequences. Lipid accumulation was studied with AdipoRed reagent, relative cell number with WST-1 reagent, gene expression of glut4, leptin, aP2, adiponectin, PPARγ and PPARγ2 with RT-qPCR. Secretion of adiponectin, leptin and aP2 was analysed with ELISA. The immunostained vascular network was imaged with Cell-IQ and area quantified using ImageJ. In this study, both angiogenesis and adipogenesis were successfully induced. Protocols produced strong lipid accumulation, good vascular network formation and induced adipocyte-specific protein secretion and expression of studied adipocyte genes. Results showed that cell numbers and cell plating sequences are important factors when aiming at in vitro standardized tissue model. Presence of mature vasculature appeared leads to faster the maturation of adipocytes judged by the lipid accumulation and gene expression results. The developed vascularized adipose tissue model is simple to use, easily modifiable to suit various applications and as such, a promising new tool for adipose tissue research when, for example, studying the effect of different cell types on adipose tissue function or for mechanistic studies.
许多与脂肪组织相关的疾病，如肥胖和 2 型糖尿病，在全球范围内流行。为了研究这些疾病，需要相关的人类细胞模型。在这项研究中，我们开发了一种血管化脂肪组织模型，其中将人脂肪基质细胞和人脐静脉内皮细胞与天然脂肪生成和无血清定义的血管生成培养基共培养 14 天。将几种不同的方案进行了比较。方案在细胞数量和接种顺序上有所不同。用 AdipoRed 试剂研究脂质积累，用 WST-1 试剂研究相对细胞数量，用 RT-qPCR 研究 glut4、瘦素、aP2、脂联素、PPARγ 和 PPARγ2 的基因表达。用 ELISA 分析脂联素、瘦素和 aP2 的分泌。用 Cell-IQ 对免疫染色的血管网络进行成像，并使用 ImageJ 对其面积进行量化。在这项研究中，成功地诱导了血管生成和脂肪生成。方案产生了强烈的脂质积累、良好的血管网络形成，并诱导了脂肪细胞特异性蛋白的分泌和研究的脂肪细胞基因的表达。结果表明，细胞数量和细胞接种顺序是体外标准化组织模型的重要因素。成熟血管的存在似乎会导致脂肪细胞更快成熟，这可以从脂质积累和基因表达结果来判断。所开发的血管化脂肪组织模型使用简单，易于修改以适应各种应用，因此，当研究不同细胞类型对脂肪组织功能的影响或进行机制研究时，它是一种有前途的脂肪组织研究新工具。
