Abstract
Macrophages accumulate prominently in the visceral adipose tissue (VAT) of obese humans and high fat diet (HFD) fed mice, and this is linked to insulin resistance and type II diabetes. While the mechanisms regulating macrophage recruitment in obesity have been delineated, the signals directing macrophage persistence in VAT are poorly understood. We previously showed that the neuroimmune guidance cue netrin-1 is expressed in the VAT of obese mice and humans, where it promotes macrophage accumulation. To better understand the source of netrin-1 and its effects on adipose tissue macrophage (ATM) fate and function in obesity, we generated mice with myeloid-specific deletion of netrin-1 (Ntn1fl/fl LysMCre+/–; Ntn1Δmac). Interestingly, Ntn1Δmac mice showed a modest decrease in HFD-induced adiposity and adipocyte size, in the absence of changes in food intake or leptin, that was accompanied by an increase in markers of adipocyte beiging (Prdm16, UCP-1). Using single cell RNA-seq, combined with conventional histological and flow cytometry techniques, we show that myeloid-specific deletion of netrin-1 caused a 50% attrition of ATMs in HFD-fed mice, particularly of the resident macrophage subset, and altered the phenotype of residual ATMs to enhance lipid handling. Pseudotime analysis of single cell transcriptomes showed that in the absence of netrin-1, macrophages in the obese VAT underwent a phenotypic switch with the majority of ATMs activating a program of genes specialized in lipid handling, including fatty acid uptake and intracellular transport, lipid droplet formation and lipolysis, and regulation of lipid localization. Furthermore, Ntn1Δmac macrophages had reduced expression of genes involved in arachidonic acid metabolism, and targeted LCMS/MS metabololipidomics analysis revealed decreases in proinflammatory eicosanoids (5-HETE, 6-trans LTB4, TXB2, PGD2) in the obese VAT. Collectively, our data show that targeted deletion of netrin-1 in macrophages reprograms the ATM phenotype in obesity, leading to reduced adipose inflammation, and improved lipid handling and metabolic function.
Highlights
Obesity triggers a poorly understood systemic immune response characterized by the recruitment of macrophages and other immune cells to key metabolic tissues [1]
Our studies suggested that adipose tissue macrophages that accumulate with obesity are the source of netrin-1 that drives macrophage accrual in adipose tissue and contribute to insulin resistance
The majority of Ntn1∆mac macrophages (70%) proceeded down an alternate path to state “c” and its subtrajectories, with 18.4% of macrophages branching to a state of high migratory and immune response gene expression “d” and 42.9% of macrophages assuming a state of high expression of fatty acid transport, lipid metabolism, and negative regulation of cell adhesion genes “e”. These findings suggest that deletion of netrin-1 alters the fate of adipose tissue macrophages, with the majority of macrophages activating a program of genes specialized in lipid handling, including high expression of genes involved in fatty acid uptake and intracellular transport (Cd36, Fabp4, Fabp5), lipid droplet formation (Plin2) and lipolysis (Lipa), and regulation of lipid localization (Apoe, Pltp, Lpl)
Summary
Obesity triggers a poorly understood systemic immune response characterized by the recruitment of macrophages and other immune cells to key metabolic tissues [1]. This low-grade inflammation is causally linked to insulin resistance and type 2 diabetes, and related health complications including hepatic and cardiovascular disease [1]. Targeting of factors that regulate monocyte recruitment to tissues (e.g., the chemokine CCL2) or macrophage inflammatory gene programs (e.g., NF-kB or JNK signaling component) have been shown to reduce visceral adipose tissue inflammation and improve insulin sensitivity in high fat diet (HFD) mice [12,13,14]. The development of novel therapies targeting chronic macrophage accumulation in metabolic tissues will require a better understanding of the mechanisms regulating tissue macrophage persistence
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