Cristhiane Favero Aguiar

Objective: Liver X receptors (LXRs) play essential roles in cholesterol homeostasis and immune response. In obesity, elevated cholesterol levels trigger proinflammatory responses; however, the specific contributions of LXRs to adipose tissue (AT) macrophage (ATM) phenotype and metabolic programming are not fully understood. In this study, we determine the role of LXR isoforms in diet‐induced obesity AT inflammation and insulin resistance.Methods: For in vivo studies, to evaluate the effects of LXR activation on AT inflammation, obese and insulin‐resistant wild‐type mice were treated with 10 mg/kg of the LXR modulator naringenin (NAR) for 4 weeks, and systemic insulin sensitivity and AT inflammation were assessed. To evaluate the effects of LXR deficiency on AT inflammation, we used LXRα, LXRβ, and LXRαβ knockout (KO) mice. For in vitro studies, to assess the role of LXRs specifically in macrophages, bone marrow‐derived macrophages from wild‐type, LXRαKO, LXRβKO, and LXRαβKO mice were treated with 0.5μM NAR 1 h prior to lipopolysaccharide (LPS) stimulation (100 ng/mL), and the effects on macrophage function and metabolism were evaluated 24 h after LPS stimulation.Results: We found that LXR deletion intensifies AT inflammation in an LXRβ‐dependent manner. LXR deficiency in immune cells exacerbates obesity‐induced AT inflammation, increasing the numbers of CD11c⁺, TNF‐α⁺, and IL‐1β⁺ ATMs. We also identified NAR as a novel LXR agonist in macrophages that reduces proinflammatory cytokine secretion by mitigating glycolysis and mitochondrial dysfunction in LPS ‐ and LPS + IFNγ‐activated macrophages. Furthermore, NAR‐treated obese mice display reduced AT inflammation, characterized by decreased CD11c⁺, IL‐1β⁺, and TNF‐α⁺ ATM numbers and monocyte infiltration compared with vehicle‐treated obese mice.Conclusions: Our study highlights distinct roles for each LXR isoform in AT inflammation regulation, with LXRβ being crucial for maintaining the anti‐ and proinflammatory balance in ATMs. Thus, LXRβ holds therapeutic potential as a target to treat AT inflammation and insulin resistance.

Tissue-resident innate immune cells have important functions in both homeostasis and pathological states. Despite advances in the field, analyzing the metabolism of tissue-resident innate lymphocytes is still challenging. The small number of tissue-resident innate lymphocytes such as ILC, NK, iNKT and γδ T cells poses additional obstacles in their metabolic studies. In this review, we summarize the current understanding of innate lymphocyte metabolism and discuss potential pitfalls associated with the current methodology relying predominantly on in vitro cultured cells or bulk-level comparison. Meanwhile, we also summarize and advocate for the development and adoption of single-cell metabolic assays to accurately profile the metabolism of tissue-resident immune cells directly ex vivo.

Natural killer T (NKT) cells are an innate-like T cell subset that recognize lipid antigens presented by CD1d-expressing antigen presenting cells (APCs), such as dendritic cells, macrophages, and B cells. They can be subdivided into two different subsets according to the variation in αβ TCR chains: type I and type II NKT cells. Type I, also called invariant NKT cells (iNKT), express restricted TCRs with an invariant α-chain (Vα24-Jα18 in humans and Vα14-Jα18 in mice) and limited β-chains. Here we have established a protocol in which iNKT cells are isolated from a donor wild-type mouse and transferred into iNKT KO (Jα18^−/−) mouse. Below we will explore the methods for cell sorting of splenic iNKTs, iNKT cells transfer, and detection of transferred cells into the liver using flow cytometry technique.

The gut microbiota is a key element to support host homeostasis and the development of the immune system. The relationship between the microbiota and immunity is a 2-way road, in which the microbiota contributes to the development/function of immune cells and immunity can affect the composition of microbes. In this context, natural killer T cells (NKT cells) are distinct T lymphocytes that play a role in gut immunity and are influenced by gut microbes. In our work, we investigated the involvement of invariant NKT cells (iNKT) in intestinal homeostasis. We found that iNKT-deficient mice (iNKT-KO) had reduced levels of fecal IgA and an altered composition of the gut microbiota, with increased Bacteroidetes. The absence of iNKT cells also affected TGF-β1 levels and plasma cells, which were significantly reduced in knockout (KO) mice. In addition, when submitted to dextran sodium sulfate colitis, iNKT-KO mice had worsening of colitis when compared with wild-type (WT) mice. To further address iNKT cell contribution to intestinal homeostasis, we adoptively transferred iNKT cells to KO mice, and they were submitted to colitis. Transfer of iNKT cells improved colitis and restored fecal IgA levels and gut microbiota. Our results indicate that intestinal NKT cells are important modulators of intestinal homeostasis and that gut microbiota composition may be a potential target in the management of inflammatory bowel diseases.