In recent years, there has been a worldwide increase in obesity, which parallels a rise in pathologies, including type 2 diabetes, collectively termed the metabolic syndrome. Chronic, low-grade inflammation has been implicated as a major link between these diseases. Recent work showed the presence of a unique subset of CD4+Foxp3+ regulatory T cells residing in visceral adipose tissue (VAT Treg) with PPAR-g being the key transcription factor responsible for their phenotype and function in controlling adipose tissue inflammation and, thereby, insulin sensitivity. VAT Tregs inversely correlated with insulin resistance. In contrast, there was a dramatic age-associated increase in frequency of VAT Tregs in lean animals, correlating with continued insulin sensitivity, despite significant increases in body and adipose tissue weights. This increase in Treg frequencies was not observed in other lymphoid and non-lymphoid tissues, including the subcutaneous fat depot.

We characterized this unique age-associated increase in VAT Tregs through the use of adoptive transfer models, in vivo labeling and tracking systems, parabiosis, and analysis of the T cell receptor (TCR) repertoire used by VAT Tregs. Our findings indicate that the progressive increase in VAT Tregs is not due to conversion of conventional CD4+ T cells nor to substantial infiltration of Tregs from the circulation and secondary lymphoid organs. However, by analyzing the TCR repertoire on a single-cell level we uncovered a striking oligo-clonal expansion of VAT Tregs, suggesting their accumulation results from in situ proliferation. We further showed that this accumulation is dependent on major histocompatibility complex (MHC) class II, but not on CD1d. Finally, we showed that IL-33 was able to induce proliferation of VAT Tregs. In parallel, we extended our analysis of TCR repertoire to the Treg population residing in skeletal muscle.

In acute and chronic models of muscle injury, muscle-resident Tregs underwent a substantial clonal expansion, with a particular clone being detected in multiple individuals. Taken together these studies highlight the importance of proliferation as a mechanism of Treg accumulation in tissues in response to acute and chronic inflammation.