An ex vivo Mouse Model for High-Fat Diet-Like Adipose Tissue Inflammation

This is the authors' abstract. We don't add key points for this paper.

This protocol describes an ex vivo mouse model that recapitulates selected cellular and inflammatory features observed in visceral adipose tissue (VAT) following high-fat diet (HFD) feeding, without dietary intervention in vivo. Small VAT explants from lean mice are cultured for 7-10 days, during which they spontaneously develop an inflammatory phenotype that closely resembles that of VAT from HFD-fed animals. Importantly, the model preserves adipose tissue (AT) architecture, enabling the investigation of inflammatory processes in a physiologically relevant context. During culture, key hallmarks of obesity-related inflammation emerge, including the formation of crown-like structures (CLS), the accumulation of metabolically activated macrophages (MMes), and increased inflammatory cytokine production. Despite adipocyte death, other drivers of obesity-related inflammation, including metabolic stressors such as saturated fatty acids, altered lipid handling, glucose excess, oxidative stress, hypoxia, adipokine imbalance, and paracrine adipocyte-immune cell signaling, can be partially represented in the model. The system supports a wide range of downstream applications, including live imaging, flow cytometry, quantitative PCR, ELISA, immunostaining, and bulk or single-cell RNA sequencing. In addition, pharmacological and cytokine-based treatments can be applied to investigate signaling pathways and cell type-specific responses. This easily customizable model aligns with the 3R principles by reducing animal use for in vivo dietary studies. The model is reproducible, cost-effective, and straightforward to implement, providing a versatile platform for studying adipocyte death, immune cell dynamics, and therapeutic strategies in AT inflammation.