White adipose tissue (WAT) fibrosis, characterized by an excess of extracellular (ECM) matrix components, is strongly associated with WAT inflammation and dysfunction due to obesity. Interleukin (IL)-13 and IL-4 were recently identified as critical mediators in the pathogenesis of fibrotic diseases. However, their role in WAT fibrosis is still ill-defined. We therefore established an ex vivo WAT organotypic culture system and demonstrated an upregulation of fibrosis-related genes and an increase of α-smooth muscle actin (αSMA) and fibronectin abundance upon dose-dependent stimulation with IL-13/IL-4. These fibrotic effects were lost in WAT lacking il4ra, which encodes for the underlying receptor controlling this process. Adipose tissue macrophages were found to play a key role in mediating IL-13/IL-4 effects in WAT fibrosis as their depletion through clodronate dramatically decreased the fibrotic phenotype. IL-4-induced WAT fibrosis was partly confirmed in mice injected intraperitoneally with IL-4. Furthermore, gene correlation analyses of human WAT samples revealed a strong positive correlation of fibrosis markers with IL-13/IL-4 receptors, whereas IL13 and IL4 correlations failed to confirm this association. In conclusion, IL-13 and IL-4 can induce WAT fibrosis ex vivo and partly in vivo, but their role in human WAT remains to be further elucidated.

Obesity is concurrent with immunological dysregulation, resulting in chronic low-grade inflammation and cellular dysfunction. In pancreatic islets, this loss of function has been correlated with mature β-cells dedifferentiating into a precursor-like state through constant exposure to inflammatory stressors. As mature adipocytes likewise have the capability to dedifferentiate in vitro and in vivo, we wanted to analyze this cellular change in relation to adipose tissue (AT) inflammation and adipose tissue macrophage (ATM) activity. Using our organotypic AT explant culture method combined with a double-reporter mouse model for labeling ATMs and mature adipocytes, we were able to visualize and quantify dedifferentiated fat (DFAT) cells in AT explants. Preliminary testing showed increased dedifferentiation after tamoxifen (TAM) stimulation, making TAM-dependent lineage-tracing models unsuitable for quantification of naturally occurring DFAT cells. The regulatory role of ATMs in adipocyte dedifferentiation was shown through macrophage depletion using Plexxicon 5622 or clodronate liposomes, which significantly increased DFAT cell levels. Subsequent bulk RNA sequencing of macrophage-depleted explants revealed enrichment of the tumor necrosis factor α (TNFα) signaling pathway as well as downregulation of associated genes. Direct stimulation with TNFα decreased adipocyte dedifferentiation, while application of a TNFα-neutralizing antibody did not significantly alter DFAT cell levels. Our findings suggest a regulatory role of resident ATMs in maintaining the mature adipocyte phenotype and preventing excessive adipocyte dedifferentiation. The specific regulatory pathways as well as the impact that DFAT cells might have on ATMs, and vice versa, are subject to further investigation.