Lipoproteins act as vehicles for lipid antigen delivery and activation of invariant natural killer T cells.

Invariant natural killer T (iNKT) cells act at the interface between lipid metabolism and immunity because of their restriction to lipid antigens presented on CD1d by antigen-presenting cells (APCs). How foreign lipid antigens are delivered to APCs remains elusive. Since lipoproteins routinely bind glycosylceramides structurally similar to lipid antigens, we hypothesized that circulating lipoproteins form complexes with foreign lipid antigens. In this study, we used 2-color fluorescence correlation spectroscopy to show, for the first time to our knowledge, stable complex formation of lipid antigens α-galactosylceramide (αGalCer), isoglobotrihexosylceramide, and OCH, a sphingosine-truncated analog of αGalCer, with VLDL and/or LDL in vitro and in vivo. We demonstrate LDL receptor-mediated (LDLR-mediated) uptake of lipoprotein-αGalCer complexes by APCs, leading to potent complex-mediated activation of iNKT cells in vitro and in vivo. Finally, LDLR-mutant PBMCs of patients with familial hypercholesterolemia showed impaired activation and proliferation of iNKT cells upon stimulation, underscoring the relevance of lipoproteins as a lipid antigen delivery system in humans. Taken together, circulating lipoproteins form complexes with lipid antigens to facilitate their transport and uptake by APCs, leading to enhanced iNKT cell activation. This study thereby reveals a potentially novel mechanism of lipid antigen delivery to APCs and provides further insight into the immunological capacities of circulating lipoproteins.

Cytokine Output of Adipocyte-iNKT Cell Interplay Is Skewed by a Lipid-Rich Microenvironment.

The complex direct and indirect interplay between adipocytes and various adipose tissue (AT)-resident immune cells plays an important role in maintaining local and whole-body insulin sensitivity. Adipocytes can directly interact with and activate AT-resident invariant natural killer T (iNKT) cells through CD1d-dependent presentation of lipid antigens, which is associated with anti-inflammatory cytokine production in lean AT (IL-4, IL-10). Whether alterations in the microenvironment, i.e., increased free fatty acids concentrations or altered cytokine/adipokine profiles as observed in obesity, directly affect adipocyte-iNKT cell communication and subsequent cytokine output is currently unknown. Here we show that the cytokine output of adipocyte-iNKT cell interplay is skewed by a lipid-rich microenvironment. Incubation of mature 3T3-L1 adipocytes with a mixture of saturated and unsaturated fatty acids specifically reduced insulin sensitivity and increased lipolysis. Reduced activation of the CD1d-invariant T-Cell Receptor (TCR) signaling axis was observed in Jurkat reporter cells expressing the invariant NKT TCR, while co-culture assays with a iNKT hybridoma cell line (DN32.D3) skewed the cytokine output toward reduced IL-4 secretion and increased IFNγ secretion. Importantly, co-culture assays of mature 3T3-L1 adipocytes with primary iNKT cells isolated from visceral AT showed a similar shift in cytokine output. Collectively, these data indicate that iNKT cells display considerable plasticity with respect to their cytokine output, which can be skewed toward a more pro-inflammatory profile in vitro by microenvironmental factors like fatty acids.