New research links triglyceride metabolism to microglial inflammation using LEGENDplex profiling

 

In a recent Cell Reports study, researchers explored how lipid metabolism—specifically triglyceride synthesis and breakdown—regulates inflammation in microglia, the brain’s resident immune cells. This work is especially relevant in the context of APOE4, a genetic variant strongly associated with Alzheimer’s disease risk. A key highlight of the study is the use of LEGENDplex™, our bead-based multiplex immunoassay, to quantify cytokine and chemokine secretion from human iPSC-derived microglia. LEGENDplex enabled precise measurement of inflammatory markers following LPS stimulation and DGAT enzyme inhibition, revealing how lipid droplet dynamics within microglia influence immune signaling.¹

 

 

Lipid droplets and immune function

 

Lipid droplets are dynamic organelles primarily involved in lipid storage, but they also play key roles in cellular signaling and metabolism. In the context of inflammation, lipid droplets serve as platforms for the synthesis of inflammatory mediators and have been shown to modulate immune cell function, contributing to both the initiation and resolution of inflammatory responses.²

 

Using human iPSC-derived microglia, the study demonstrated that lipid droplet formation is not just a byproduct of inflammation, but a regulatory mechanism. Inhibiting DGAT1 and DGAT2, enzymes responsible for triglyceride synthesis, altered microglial responses to inflammatory stimuli like LPS. These changes were evident in both gene expression and cytokine secretion profiles.

 

 

LEGENDplex: a key tool for cytokine profiling iPSC derived microglia

 

To quantify the immune response, the researchers turned to LEGENDplex multiplex immunoassays. This workflow enabled simultaneous measurement of multiple cytokines and chemokines from small sample volumes. LEGENDplex Human Inflammation Panel 1 revealed that iPSC-derived microglia treated with DGAT inhibition using an adipose triglyceride lipase (ATGL) inhibitor, atglistatin, significantly increased the number of lipid droplets and reduced secretion of pro-inflammatory cytokines such as IL-6, TNF-α, and IL-1β, under inflammatory stimuli, highlighting the role of triglyceride metabolism in shaping immune phenotypes (Figure 1).

 

Flow-based LEGENDplex assays can be used to assess cytokine production and immune activation in in vitro and biological samples caused by a novel biologic, therapeutic, or pathogen. Our panels can simultaneously detect multiple cytokines derived from monocytes, macrophages, microglia, T cells, and NK cell activation and provide an in-depth analysis of the immune cell response following stimulation. Our multiplex assays permit a broad assessment of cytokine production, which can help characterize immune cell responses in multiple research applications.

 

 

(A) Schematic of inhibition of lipid droplet catabolism using atglistatin. 
(B) Transcript abundance of adipose triglyceride lipase (ATGL) in LPS-treated microglia relative to vehicle-treated controls (vehicle, white; LPS, blue; n = 3 independent treatments). Data are represented as mean ± SD. *p ≤ 0.05 by two-tailed unpaired t test. 
(C) Fluorescence images of APOE3/APOE3 microglia treated with vehicle or ATGL inhibitor and stained with LipidSPOT. Bottom panels show ROI in top panels. Scale bars, 50 μm. 
(D) Lipid droplet number per cell, quantified across 3 wells with at least 40 cells analyzed per well. n ≥ 6 frames per condition. Data are represented as mean ± SD. **p ≤ 0.01 by two-tailed unpaired t test. 
(E) Fold change of cytokines secreted by LPS-treated APOE3/APOE3 microglia with and without DGAT inhibitor treatment. Data represent 6 technical replicates across n = 3 microglial derivations.
Image from Stephenson et al. 2025. Cell Reports, 42(10):113456.¹ Creative Commons CC BY 4.0.

 

 

 

Using humanized APOE4 transgenic microglia (APOE4/APOE4), the authors also used LipidSpot™ Lipid Droplet Stains (Biotium) to visualize and quantify lipid droplet accumulation in microglia (Figure 1C and D) and observed elevated cytokine expression at both the transcript and protein levels, indicating a heightened inflammatory state and dysregulated immune activation in these models (Figure 2).

 

Gene expression levels of cytokines (from normalized read counts) from APOE4/APOE4 microglia relative to average vehicle-treated controls (vehicle, white; LPS, blue; DGAT inhibition, gray; DGAT inhibition + LPS, green. n = 3 treatments) (top). Data are represented as mean ± SD. ns p > 0.05, *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001 by two-way ANOVA and post-hoc Tukey test. Fold change (relative to vehicle controls) of cytokines secreted by APOE4/APOE4 microglia treated DGAT inhibitors or vehicle. Data represent 6 technical replicates across n = 3 microglial derivations (bottom). Image from Stephenson et al. 2025. Cell Reports, 42(10):113456.¹ Creative Commons CC BY 4.0.

 

 

Implications for Alzheimer’s disease

 

The findings suggest that targeting triglyceride metabolism could modulate microglial activation in APOE4 carriers, potentially offering a new therapeutic angle for neurodegenerative diseases. By combining transcriptomic analysis with LEGENDplex cytokine profiling, the study provides a comprehensive view of how metabolic pathways intersect with immune signaling in the brain.

 

 

Summary

 

Stephenson et al. highlights lipid droplets as key modulators of neuroinflammation.¹ Their work shows that inhibiting triglyceride biosynthesis in microglia also suppressed their inflammatory response. This occurred even in APOE4-expressing cells, the APOE variant that is known to have significant genetic risk factor for developing late-onset Alzheimer’s disease. LEGENDplex’s Human Inflammation Panel 1 enabled high-throughput quantification of cytokines and chemokines from iPSC-derived microglia and was pivotal in providing critical insight into the link between lipid metabolism and immune dysfunction.

 

This dual-platform approach—combining transcriptomics with LEGENDplex cytokine profiling—offered a powerful lens into microglial immunometabolism and opens new avenues for therapeutic modulation of neuroinflammation.

 

Our reagents and solutions are designed for all aspects of your immunology research. Focus on single targets with our highly-sensitive ELISAs and ELISpot kits, or analyze multiple cytokines with our FluoroSpot and LEGENDplex multiplex immunoassays.  We offer specific LEGENDplex panels for key cell types and pathways including Human Macrophage/Microglia Panel, Human Neuroinflammation Panel 1 and Human Neuroinflammation Panel 2, and Human Neurodegenerative Biomarker Panel 1. For high throughput detection of cytokines and additional biomarkers, learn more about Revvity’s no-wash immunoassay technologies.

 

 

1. Stephenson, R. et al. (2025). Triglyceride metabolism controls inflammation and microglial phenotypes associated with APOE4. Cell Reports, 42(10), 113456. https://doi.org/10.1016/j.celrep.2025.113456. PubMed.
2. Fu, L., Luo, T., Hao, Z., Pan, Y., et al. (2025). Exploring novel roles of lipid droplets and lipid metabolism in regulating inflammation and blood–brain barrier function in neurological diseases. Frontiers in Neuroscience, 19, 1603292. https://doi.org/10.3389/fnins.2025.1603292  PubMed.