Abstract
Regulatory T cells (Tregs) mediate anti-inflammatory functions and have been most associated with self-
immunity and autoimmune diseases. However, given the underlying role of subclinical or persistent low-level
inflammation in metabolic and cardiovascular diseases, attention has recently focused on the potential role of
Tregs in controlling the proinflammatory milieu in these pathologies. A subset of Tregs, the effector memory
Tregs, are the most suppressive, but are also the most transient. We made the striking observation that high-
density lipoproteins (HDL), but not other lipoproteins, significantly improved Treg survival. These findings offer
an explanation for the positive correlation we found between HDL cholesterol and Treg abundance. Our
preliminary data show that HDL preferentially bind and are internalized by Treg memory subsets, particularly by
effector memory Tregs, promoting their survival by reducing effector caspase activation. Mechanistically, our
new data also suggest that this HDL pro-survival effect may occur via activation of the AKT signaling pathway,
which results in enhanced de novo fatty acid synthesis and reduced mitochondrial oxidative stress. Since HDL
is a family of related particles with complex mixtures of lipids and proteins, we started exploring which HDL
subspecies or components stimulate Treg survival. We found that the pro-survival effect was mediated by the
HDL protein component, and particularly the relatively minor HDL protein constituent, apolipoprotein (APO)E,
and/or other proteins that co-reside on APOE-containing HDL. In contrast, the major HDL scaffold proteins from
the APOA family did not play a major role. Our overarching hypothesis is that APOE-rich HDL specifically interact
with Tregs to trigger pathways that limit caspase-dependent apoptosis. Aim 1 will identify the intracellular
signaling pathways involved in HDL-mediated survival of Treg, notably the receptor(s) involved in HDL binding/
internalization by memory Tregs and the mechanisms by which HDL promote Treg survival. We will also
determine the effect of HDL on Treg suppression. Aim 2 will identify specific HDL subspecies and protein
components that promote Treg survival. We will also perform deletional and site-directed mutagenesis
experiments to identify the critical region(s) of APOE, to inform explorations of synthetic peptides that can mimic
APOE’s pro-survival effects on memory Tregs. Aim 3 will assess the involvement of APOE-rich HDL in Treg
survival in vivo. We will leverage the Cincinnati Pediatric Diabetes and Obesity Center cohort and probe the
association between absolute numbers of Treg subsets, their functional characteristics, and HDL subspecies,
before and after weight reduction surgery. The ability of HDL subspecies collected before and after surgery to
promote memory and effector memory Treg survival will be studied. By identifying specific cellular pathways and
specific lipoprotein species responsible for the Treg survival effects, we will open new avenues for potential
therapeutic intervention aimed at modulating Treg function in a host of auto-immune and chronic metabolic
diseases.