Expunging Microglial Lipid Droplets Boosts Phagocytosis

In Alzheimer’s disease, microglia pack on fat, which impairs phagocytosis. Now, scientists led by Christiane Ruedl of Nanyang Technological University, Singapore, report that blocking lipid buildup in these cells restores their capacity to engulf Aβ, leading to fewer plaques in the brains of APP knock-in mice. Their paper ran in the February 5 Science Advances.

  • Microglia overloaded with lipids choke on Aβ in APP knock-in mice.
  • Depleting the fat-storage-associated protein FIT2 reduced lipid droplet formation.
  • Sans FIT2, microglia rein in plaques.

“This adds to the growing body of work, from mouse and fly models, implicating lipid metabolism in neurodegeneration, particularly AD,” Amita Sehgal, Perelman School of Medicine, University of Pennsylvania, told Alzforum. “The manuscript also suggests a possible therapeutic intervention to reduce lipid load in microglia.”

Previously, Tony Wyss-Coray and colleagues at Stanford University, Palo Alto, California, had identified lipid-droplet–accumulating microglia lurking in the hippocampi of aging mice. These fat-stuffed cells spelled trouble, fanning inflammation and reactive oxygen species while struggling to clear debris (Marschallinger et al., 2020; Aug 2019 news). Since then, two studies—one from the same group and one led by Gaurav Chopra, Purdue University, West Lafayette, Indiana—showed that exposure to Aβ fibrils triggered this lipid droplet buildup, as it summoned microglia to cluster near plaques (Haney et al., 2024; Prakash et al., 2023; Sep 2023 news). Critically, the lipid impaired Aβ clearance from the brain through phagocytosis.

Building on this research, Ruedl and colleagues ask which factors influence lipid buildup, and they identify the types of fats involved. First author Xiaoting Wu and colleagues cultured microglia from wild-type and APP-NL-G-F knock-in mice. As previously reported, the number of fat-laden microglia grew with age in wild-type mice, from 5 percent at 2 months to 35 percent at 6 months. This was even more pronounced in APP-knock-in mice, in which half of all isolated microglia carried fatty deposits by 6 months. Throw in a high-fat diet, and lipid-stuffed microglia grew to 60 percent.

Weighing Heavy. Microglia cultured from 2-month-old APP N-L-GF mice looked normal (left), but those cultured from 6-month-old animals (middle) accumulated lipid droplets (green). A high-fat diet made this worse (right). [Courtesy of Wu et al., Sci. Advances 2025.]

What’s in these droplets? Lipid profiling revealed elevated levels of triglycerides and cholesterol esters—the main fats that make up lipid droplets (Fujimoto and Parton, 2011). Kimberley Bruce, University of Colorado in Aurora, noted a dearth of the arachidonic acid, a precursor to prostaglandins and leukotrienes, highlighting potential deficits in the inflammatory response (comment below).

High-Profile Lipids. Microglia from 6-month-old APP KI mice accumulate (red) 64 triglycerides and five cholesterol esters, among other lipids. Sphinganine metabolites and several free fatty acids tanked (blue). [Courtesy of Wu et al., Sci. Advances 2025.]

Which genes are behind this shift? Wu isolated microglia from 6-month-old wild-type and APP knock-in mice and ran single-cell RNA sequencing. This identified 10 distinct microglial clusters and three clusters of border-associated macrophages. The latter reside in perivascular spaces in the brain and have been linked to cerebral amyloid angiopathy (Apr 2023 conference news; Aug 2024 conference news).

Reactive microglia—including Nlrp3-positive inflammasome-related, TNF-hi, and CD11c-positive “activated response” microglia (ARM)—were more prevalent in the knock-ins, the CD11c+ cluster being the most abundant. This cluster was chock-full of lipid metabolism genes typically associated with foam cells—lipid-rich macrophages that accumulate near atherosclerotic plaques in cardiovascular disease. These included some familiar genes, such as ApoE, Abca1, and TREM2 (image below).

Foaming at the Cell. Microglia and border-associated macrophage clusters boost expression of some genes (right) associated with foam cells. Cluster 1, activated response microglia—prevalent in APP-KIs—highly expressed these genes. [Courtesy of Wu et al., Sci. Advances, 2025.]

Wu and colleagues confirmed that cultured-microglia-carrying lipid droplets struggled with phagocytosis, poorly engulfing fluorescently labeled inactivated E. Coli or Aβ. What if microglia were free of excessive lipids? To find out, researchers targeted FIT2, a protein embedded in the endoplasmic reticulum that helps lipid droplets form and bud.

Because all cells express FIT2 and whole-body knockout is lethal (Goh et al., 2015), the authors created a conditional knockout. When APP-N-L-GF mice were 2 months old, Wu used a tamoxifen-inducible transgene to selectively deplete Fit2 in CX3CR1-positive brain macrophages, including microglia. By 6 months, lipid staining revealed that microglia in these mice had 30 percent smaller lipid droplet area than control microglia. The FIT2-deficient microglia were not entirely devoid of droplets, in keeping with the protein primarily unleashing triglyceride-laden droplets. Other lipids, such as sterol esters, may still contribute.

Would absence of FIT2 make microglia function more smoothly? The moment of truth came when Wu tested phagocytosis of FIT2-depleted microglia. They slurped more E. coli particles and Aβ oligomers than FIT2-positive cells. Most compellingly, FIT2 depletion in the mouse brain enhanced Aβ clearance. Immunofluorescence staining revealed significantly fewer Aβ plaques in the cortex and hippocampus of FIT2-deficient mice (image below).

“These exciting proof-of-concept data support a growing body of literature highlighting microglial lipid metabolism as a rational, and clinically relevant, target to improve AD pathology,” Bruce told Alzforum.

Back to Work. Six-month-old APP-NL-G-F mice accumulate amyloid plaques (red) in the cortex and hippocampus (left). Knocking out Fit2 in microglia reduces lipid droplet and plaque burden (right). [Courtesy of Wu et al., Sci. Advances, 2025.]

Still, whether targeting FIT2 improves memory in the mice is unclear, wrote Laura Piccio of the University of Sydney (comment below). Wu noted that APP knock-in mice have only modest memory problems, making it difficult to assess any impact of microglial lipid depletion. Nevertheless, the authors see clinical potential, writing, “Reducing microglial LD load via FIT2 depletion or other pharmacological interventions is a feasible therapeutic strategy.” Piccio was more cautious. “The study does not explore the broader effects of FIT2 depletion in vivo,” she wrote. “Interfering with its function could have systemic and long-term effects.”—George Heaton

George Heaton is a freelance writer in Durham, North Carolina.

Comments

  1. Reducing microglial lipid load enhances Aβ phagocytosis in an Alzheimer’s disease mouse model

    Wu and co-authors investigate the role of lipid-droplet-accumulating microglia (LDAM) and border-associated macrophages (BAMs) in an APP-KI model, which harbors a humanized Aβ sequence containing the Swedish, Beyreuther/Iberian, and Arctic mutations. The experiments were conducted in 6-month-old mice, an age at which memory deficits are observed in this model (Saito et al., 2014). By integrating multi-omics approaches, including transcriptomic and lipidomic profiling, the authors document how LD formation influences microglial function. Notably, genetic deletion of FIT2 which is a known regulator of lipid droplet biogenesis, restored microglial phagocytosis and enhanced Aβ clearance, suggesting a potential therapeutic avenue.

    Targeting FIT2 effectively reduced LD formation, appeared to restore microglial phagocytosis, and improved plaque clearance in APP-KI mice. Interestingly, CD11c+ microglia in FIT2-deficient APP-KI mice exhibited a downregulation of disease-associated microglial (DAM) genes, including Trem2, Apoe, and Cd9, which may be a consequence of reduced amyloid burden. However, these microglia retained high expression of MHC II genes, particularly Cd74. Notably, Cd74 and Axl were previously reported to show higher expression in LD-low microglia compared to LD-high microglia (Marschallinger et al., 2020). 

    Furthermore, ACSL1, an enzyme that catalyzes the conversion of free fatty acids (FFAs) into Acyl-CoAs, plays a crucial role in lipid droplet biogenesis and has been implicated in microglial dysfunction in AD (Haney et al., 2024). The elevated FFA levels observed in FIT2-deficient microglia warrant further investigation, particularly in the context of lipid metabolism and microglial homeostasis.

    From a therapeutic standpoint, targeting FIT2 would likely present significant challenges due to its higher expression in neurons and astrocytes than in microglia within the brain. Notably, homozygous FITM2 mutations have been associated with deafness-dystonia syndrome, motor regression, ichthyosis, and sensory neuropathy, raising concerns about potential adverse effects of systemic FIT2 inhibition (Zazo Seco et al., 2017). Another critical question is whether peripheral immune cells exhibit distinct FIT2-regulated pathways and whether FIT2 modulation could be CX3CR1-specific. Addressing these questions will be essential for the development of targeted therapeutic strategies that minimize unintended consequences while effectively modulating FIT2.

    Another key question emerging from this and other studies on LDs in mice is their relevance to human brain aging and AD, as well as the role different ApoE isoforms play. Previous research from our lab, building on Alzheimer’s original observations of LDs in AD, suggests that APOE4 carriers may be more prone to LD formation and more susceptible to their toxic effects. However, further studies are needed to determine in which cell types LDs accumulate in humans, the molecular processes driving LD formation, and how these deposits interfere with cellular functions.

    Overall, this study elegantly highlights the importance of microglial lipid droplets in amyloid pathology and provides novel insights into their role in microglial function, lipid metabolism, and disease progression.

    References:

    Saito T, Matsuba Y, Mihira N, Takano J, Nilsson P, Itohara S, Iwata N, Saido TC. Single App knock-in mouse models of Alzheimer's disease. Nat Neurosci. 2014 May;17(5):661-3. Epub 2014 Apr 13 PubMed.

    Marschallinger J, Iram T, Zardeneta M, Lee SE, Lehallier B, Haney MS, Pluvinage JV, Mathur V, Hahn O, Morgens DW, Kim J, Tevini J, Felder TK, Wolinski H, Bertozzi CR, Bassik MC, Aigner L, Wyss-Coray T. Lipid-droplet-accumulating microglia represent a dysfunctional and proinflammatory state in the aging brain. Nat Neurosci. 2020 Feb;23(2):194-208. Epub 2020 Jan 20 PubMed. Correction.

    Haney MS, Pálovics R, Munson CN, Long C, Johansson PK, Yip O, Dong W, Rawat E, West E, Schlachetzki JC, Tsai A, Guldner IH, Lamichhane BS, Smith A, Schaum N, Calcuttawala K, Shin A, Wang YH, Wang C, Koutsodendris N, Serrano GE, Beach TG, Reiman EM, Glass CK, Abu-Remaileh M, Enejder A, Huang Y, Wyss-Coray T. APOE4/4 is linked to damaging lipid droplets in Alzheimer's disease microglia. Nature. 2024 Apr;628(8006):154-161. Epub 2024 Mar 13 PubMed.

    Zazo Seco C, Castells-Nobau A, Joo SH, Schraders M, Foo JN, van der Voet M, Velan SS, Nijhof B, Oostrik J, de Vrieze E, Katana R, Mansoor A, Huynen M, Szklarczyk R, Oti M, Tranebjærg L, van Wijk E, Scheffer-de Gooyert JM, Siddique S, Baets J, de Jonghe P, Kazmi SA, Sadananthan SA, van de Warrenburg BP, Khor CC, Göpfert MC, Qamar R, Schenck A, Kremer H, Siddiqi S. A homozygous FITM2 mutation causes a deafness-dystonia syndrome with motor regression and signs of ichthyosis and sensory neuropathy. Dis Model Mech. 2017 Feb 1;10(2):105-118. Epub 2016 Dec 15 PubMed.

    View all comments by Tony Wyss-Coray View all comments by Andy Tsai

  2. Although the sequestering of lipids within intracellular lipid droplets protects against lipid oxidation, excess lipid droplet accumulation is associated with aging and disease. In AD, microglia accumulate excess lipid droplets, which leads to reduced functionality, such as phagocytosis, promoting AD neuropathogenesis. While many research groups, including our own, have been interested in understanding the mechanisms leading to lipid droplet accumulation, in this study, Wu et al. focus on strategies that reduce LDs to increase microglial phagocytosis and prevent AD pathology.

    The authors use cell sorting to confirm that lipid-droplet-containing microglia increase with aging and disease, in this case, a murine model of amyloid accumulation (APP-KI). In further support, untargeted lipidomics showed that microglia isolated from APP-KI mice contained more neutral lipids typically associated with LDs (e.g., triglycerides and cholesterol esters). I also found it interesting that the very-long-chain polyunsaturated fatty acid and eicosanoid precursor arachidonic acid was depleted in microglia from the APP-KI mice, highlighting potential deficits in the inflammatory response. It would also be interesting to probe the saturation index of lipids within the microglia, especially given recent papers highlighting increased unsaturated lipids in (pathology-associated) lipid droplets.

     To further ascertain the phenotype of microglia in response to increased amyloid load, the authors use single-cell transcriptomics, identifying lipid-accumulating disease-associated microglia (DAM)-like signatures (e.g., expressing ApoE, LPL, Trem2, and Cst7) that have also been observed in other AD models (e.g., 5xFAD). Going forward, methodological advances permitting, performing lipidomic and transcriptomic analysis on the same cells will advance our mechanistic understanding of microglial lipid metabolism and lipid droplet formation.

    Chasing a therapeutic intervention, Wu and colleagues genetically ablate FIT2 (CX3CR1; FIT2flox), a protein that binds to triglycerides and diacylglycerols in the endoplasmic reticulum to increase lipid droplet formation. Microglia-specific FIT2 depletion in APP-KI mice resulted in reduced microglial lipid droplet accumulation, triglyceride composition, expression of DAM-like genes (e.g., Plin2, ApoE, Trem2, and Cts7), and amyloid load. FIT2 depletion also results in increased microglial phagocytosis ex vivo, and in vitro, supporting the idea that reducing lipid droplet accumulation in microglia could preserve functionality and rescue AD-like pathology.

    These exciting proof-of-concept data support a growing body of literature highlighting microglial lipid metabolism as a rational, and clinically relevant, target to improve AD pathology. However, it is important to reiterate that at baseline lipid droplets serve to protect the cell. Therefore, selectively inhibiting lipid droplet formation in subsets of microglia and at specific disease stages will be an attractive strategy going forward.

    View all comments by Kimberley Bruce

  3. This study investigates the role of the protein FIT2, a key protein in lipid droplet biogenesis, in LD accumulation within microglia and BAMs in an AD mouse model. They show that LDs accumulate with age and AD progression in the two cell types in the CNS and that this is exacerbated by a high-fat diet. Other key findings include that FIT2 deficiency reduces LD formation in microglia and BAMs, leading to changes in their lipid metabolism and gene-expression profiles. Furthermore, FIT2 depletion enhances microglial phagocytosis, increasing the uptake of E. coli bioparticles and Aβ oligomers in vitro, as well as reducing Aβ accumulation in the brain tissue.

    This study is novel and has the following strengths:

    1. Identifies FIT2 as a key driver of LD accumulation in microglia and BAMs, which impaired their phagocytic function and clearance of amyloid in an AD mouse model.
    2. Suggests the potential for a therapeutic intervention by reducing FIT2 expression which resulted in enhanced phagocytic function in microglia and BAMs, suggesting this as a possible strategy to improve Aβ clearance in AD.
    3. Integrates single-cell transcriptomic and lipidomic analyses of microglia, to identify specific microglial lipid-associated phenotypes and lipid profiles in an AD mouse model setting with the potential to provide new mechanistic insights. Importantly, certain microglial clusters had similar transcriptomic profiles to foam cells, well-characterized lipid-rich macrophages found in atherosclerotic plaques in cardiovascular diseases.
    4. Another interesting aspect is related to the impact of diet (high-fat diet) in modulating LD accumulation in microglia and BAMs, which could have relevant clinical implications.

    Limitations include:

    1. Lack of in vivo functional validation of these findings: while FIT2 deficiency improved microglial functions and lowered Ab deposition, the study does not clearly demonstrate whether this leads to in vivo cognitive or behavioural improvement in the AD model.
    2. It seems the role of Ab accumulation in AD progression and cognitive decline is currently a matter of intense debate.
    3. The study does not explore broader effects of FIT2 depletion in vivo of this ubiquitously expressed protein. Interfering with its function could have systemic and long-term effects as well as impact other cell types beyond microglia; this could hinder its clinical translation.

    In conclusion, this study provides important insights into the metabolic dysfunction of microglia in AD, as a result of altered lipid-homeostasis. Furthermore, it identifies FIT2 as a promising target for intervention. However, further in vivo and human studies are needed to confirm its therapeutic potential and to understand long-term effects of interfering with its function in the brain.

    View all comments by Laura Piccio
    • User Profile Image Pinar Ayata
      CUNY Advanced Science Research Center
    • Posted:

    This study contributes to a growing body of research on the role of microglial lipid metabolism in Alzheimer’s disease. Important studies from the labs of Tony Wyss-Coray, Li-Huei Tsai, Mathew Blurton-Jones, Gaurav Chopra, Erik Musiek, and many others showed that amyloidosis induces a shift in microglial lipid metabolism and lipidomic profile, as well as an increase in lipid-droplet-accumulating microglia near sites of pathology. A common theme in these studies is that these microglia have reduced phagocytic capabilities and a neurotoxic effect.

    Christiane Ruedl’s research group shows that this phenomenon is not limited to microglia but also seen in brain macrophages of the meninges. This study goes another step further, showing us that reducing LDs in microglia/macrophages via genetic depletion of FIT2 not only restores their phagocytic activity but also reduces amyloid burden in APP knock-in. 

    View all comments by Pinar Ayata

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References

News Citations

  1. Newly Identified Microglia Contain Lipid Droplets, Harm Brain
  2. Lipid-Laden, Sluggish Microglia? Blame Aβ.
  3. Macrophages Blamed for Vascular Trouble in ApoE4 Carriers
  4. Implicated in ARIA: Perivascular Macrophages and Microglia

Research Models Citations

  1. APP NL-G-F Knock-in

Paper Citations

  1. Marschallinger J, Iram T, Zardeneta M, Lee SE, Lehallier B, Haney MS, Pluvinage JV, Mathur V, Hahn O, Morgens DW, Kim J, Tevini J, Felder TK, Wolinski H, Bertozzi CR, Bassik MC, Aigner L, Wyss-Coray T. Lipid-droplet-accumulating microglia represent a dysfunctional and proinflammatory state in the aging brain. Nat Neurosci. 2020 Feb;23(2):194-208. Epub 2020 Jan 20 PubMed. Correction.
  2. Haney MS, Pálovics R, Munson CN, Long C, Johansson PK, Yip O, Dong W, Rawat E, West E, Schlachetzki JC, Tsai A, Guldner IH, Lamichhane BS, Smith A, Schaum N, Calcuttawala K, Shin A, Wang YH, Wang C, Koutsodendris N, Serrano GE, Beach TG, Reiman EM, Glass CK, Abu-Remaileh M, Enejder A, Huang Y, Wyss-Coray T. APOE4/4 is linked to damaging lipid droplets in Alzheimer's disease microglia. Nature. 2024 Apr;628(8006):154-161. Epub 2024 Mar 13 PubMed.
  3. Prakash P, Manchanda P, Paouri E, Bisht K, Sharma K, Wijewardhane PR, Randolph CE, Clark MG, Fine J, Thayer EA, Crockett A, Gasmi N, Stanko S, Prayson RA, Zhang C, Davalos D, Chopra G. Amyloid β Induces Lipid Droplet-Mediated Microglial Dysfunction in Alzheimer's Disease. bioRxiv. 2023 Jun 6; PubMed.
  4. Fujimoto T, Parton RG. Not just fat: the structure and function of the lipid droplet. Cold Spring Harb Perspect Biol. 2011 Mar 1;3(3) PubMed.
  5. Goh VJ, Tan JS, Tan BC, Seow C, Ong WY, Lim YC, Sun L, Ghosh S, Silver DL. Postnatal Deletion of Fat Storage-inducing Transmembrane Protein 2 (FIT2/FITM2) Causes Lethal Enteropathy. J Biol Chem. 2015 Oct 16;290(42):25686-99. Epub 2015 Aug 24 PubMed.

Further Reading

Papers

  1. Marschallinger J, Iram T, Zardeneta M, Lee SE, Lehallier B, Haney MS, Pluvinage JV, Mathur V, Hahn O, Morgens DW, Kim J, Tevini J, Felder TK, Wolinski H, Bertozzi CR, Bassik MC, Aigner L, Wyss-Coray T. Lipid-droplet-accumulating microglia represent a dysfunctional and proinflammatory state in the aging brain. Nat Neurosci. 2020 Feb;23(2):194-208. Epub 2020 Jan 20 PubMed. Correction.
  2. Haney MS, Pálovics R, Munson CN, Long C, Johansson PK, Yip O, Dong W, Rawat E, West E, Schlachetzki JC, Tsai A, Guldner IH, Lamichhane BS, Smith A, Schaum N, Calcuttawala K, Shin A, Wang YH, Wang C, Koutsodendris N, Serrano GE, Beach TG, Reiman EM, Glass CK, Abu-Remaileh M, Enejder A, Huang Y, Wyss-Coray T. APOE4/4 is linked to damaging lipid droplets in Alzheimer's disease microglia. Nature. 2024 Apr;628(8006):154-161. Epub 2024 Mar 13 PubMed.
  3. Prakash P, Manchanda P, Paouri E, Bisht K, Sharma K, Wijewardhane PR, Randolph CE, Clark MG, Fine J, Thayer EA, Crockett A, Gasmi N, Stanko S, Prayson RA, Zhang C, Davalos D, Chopra G. Amyloid β Induces Lipid Droplet-Mediated Microglial Dysfunction in Alzheimer's Disease. bioRxiv. 2023 Jun 6; PubMed.
  4. Fujimoto T, Parton RG. Not just fat: the structure and function of the lipid droplet. Cold Spring Harb Perspect Biol. 2011 Mar 1;3(3) PubMed.
  5. Kadereit B, Kumar P, Wang WJ, Miranda D, Snapp EL, Severina N, Torregroza I, Evans T, Silver DL. Evolutionarily conserved gene family important for fat storage. Proc Natl Acad Sci U S A. 2008 Jan 8;105(1):94-9. Epub 2007 Dec 26 PubMed.

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