In Vitro Model of Cytokine-Induced Inflammatory 3T3-L1 Adipocytes Mimicking Obesity

Obesity is a risk factor for many diseases. The 3T3-L1 cell line is often used to obtain mature adipocytes, but these lack the inflammatory phenotype observed in obesity. Using a cocktail of cytokines that mimics the secretome of macrophages found in the inflammatory adipose tissue, we developed a protocol for obtaining mature inflammatory adipocytes. This model was validated at gene (RT-qPCR) and protein levels (multiplex adipokine array) as we found a decrease of adipogenic markers (C/EBPα, PPARУ, adiponectin, and CD36) and an increase of pro-inflammatory cytokines (IL-6, IL-1β, CXCL1, CXCL10, TNF-α, ICAM-1, and lipocalin-2). We provide a relevant in vitro model for studying the impact of low-grade chronic inflammation caused by obesity and its downstream effects on metabolic disorders and tumor microenvironments. Key features • Currently available protocols of adipocyte differentiation are not relevant for studying obesity in vitro. • We developed a simple and reproducible method to generate inflammatory adipocytes in vitro using a cytokine cocktail. • Gene expression analysis (qPCR) confirms the downregulation of adipogenic and protective markers (e.g., adiponectin, CD36) and the upregulation of pro-inflammatory cytokines (e.g., IL-6, IL-1β, TNF-α). • Adipokine array reveals decreased secretion of anti-inflammatory molecules (adiponectin, IGFBPs, FGF-21, HGF) and increased release of pro-inflammatory adipokines (serpin E1, IGF-1, lipocalin-2, IL-6, ICAM-1). • This protocol provides a relevant and versatile method for investigating obesity-related inflammation and its role in disease progression.