Spheroid Culture System Confers Differentiated Transcriptome Profile and Functional Advantage to 3T3-L1 Adipocytes

This study highlights functional differences between 2-D monolayer and 3-D spheroid 3T3-L1 adipocyte culture models and explores the underlying genomic mechanisms responsible for the different phenotypes present. The spheroids showed higher triglyceride accumulation than the monolayer culture and further increase with larger spheroid size. Whole transcriptome analysis indicated significant differential expression of genes related to adipogenesis, including adipocytokine signaling, fatty acid metabolism, and PPAR-γ signaling. Spheroids also showed downregulation of matrix metalloproteinases (MMPs), integrin, actin-cytoskeleton associated genes, and Rho/GTPase3 expression relative to 2-D monolayer, indicating suppression of the Rho-ROCK pathway and thereby promoting adipogenic differentiation. When exposed to linoleic acid (500 μM) and TNF-α (125 ng/mL) to promote chronic adiposity, linoleic acid treatment resulted in increased intracellular triglycerides and subsequent TNF-α treatment resulted in significantly altered adipocytokine signaling, fatty acid metabolism, and PPAR signaling, in addition to upregulation of multiple MMPs in spheroids vs. monolayer. Overall, 3-D spheroids showed enhanced adipogenic phenotype as indicated by triglyceride synthesis and transcriptome changes while retaining sensitivity to a pro-inflammatory stimulus. The 3-D spheroid culture thus may provide a simple, convenient, and sensitive in vitro model to study adipocyte response to metabolic stresses relevant to clinical pathologies.