20 Oct 2023

You might well think that myelin, the fatty insulation that speeds action potentials between neurons, falls solely under the purview of oligodendrocytes. After all, these specialized cells wrap cholesterol-laden myelin around axons in the brain's white matter beginning early in development, and throughout life they sustain and repair this insulation. Well, it turns out they need microglia to help them do that. At the 2nd Symposium on Lipids in Brain Diseases, held September 13-15 in Leiden, The Netherlands, scientists reported that microglia are essential for regular maintenance of myelin.

They showed that in demyelinating conditions, microglia can scupper remyelination. In diseases such as multiple sclerosis, Charcot-Marie-Tooth disease, and adult-onset leukoencephalopathy with axonal spheroids and pigmented glia, microglia are summoned to mop up debris. In so doing, they become both phagocytic and inflammatory. If they get stuck in that state, persistent inflammation hinders oligodendrocytes' repairing the insulation. The findings could have implications not only for MS, but for other neurodegenerative diseases that degrade myelin, including Alzheimer’s.

Niamh McNamara, Netherlands Institute for Neuroscience, Amsterdam, explained how microglia are crucial for the normal growth and maintenance of myelin. Without them, the myelin begins to unravel and develops bulges, also called outfolds, McNamara found. In the absence of microglia, oligodendrocyte lipid and cholesterol metabolism falters, and the cells soon struggle to sustain the insulation. McNamara carried out most of this research while in Veronique Miron’s lab at the University of Edinburgh (McNamara et al., 2023). 

The effect is subtle and required sleuthing to figure out. McNamara and colleagues knew that when they ablated microglia, or macrophages, myelination crumbled in mice. But which of these immune cells were to blame? To find out, she turned to mice lacking the fms-intronic regulatory element (FIRE) in the gene for macrophage colony stimulating factor 1 receptor. CSF1R is essential for microglial/macrophage proliferation and survival. FIRE is a super enhancer for transcription; without it, macrophages and monocytes cannot make CSF1R, and die. Curiously, only certain cells depend on FIRE to produce this receptor. These include microglia, but not border-associated and perivascular macrophages in the brain (Rojo et al., 2019). Would FIREΔ/Δ mice have normal myelin, then?

They did—at first. In 1-month-old mice, it was business as usual for oligodendrocytes. They insulated axons of all diameters in the white matter; myelin basic protein levels seemed normal.

Even so, on closer inspection, McNamara found that all was not well with the myelin around FIREΔ/Δ axons. Under the electron microscope, it appeared to be coming apart in places and beginning to blister. Myelin's “inner tongue”—the area where oligodendrocytes start laying it down—was abnormally thick.

Deformities persisted as the mice aged, suggesting it was not a developmental phenomenon. By 3 to 4 months of age, axons became hypermyelinated; by 6 months the myelin had begun to break down, endangering axon insulation. This waning of myelin also happened in wild-type mice fed PLX-5622, the CSF1R inhibitor that stymies both microglial and macrophage proliferation.

FIRE Damage. Myelin around axons in FIREΔ/Δ mice blebs (left), comes undone (center), and its inner tongue (brown, right) thickens. [Courtesy McNamara et al., 2022.]

These structural changes had consequences. In the Barnes circular maze, FIREΔ/Δ  mice made more errors than wild-type when McNamara moved their escape hole 180 degrees. This switcheroo forces mice to adapt, hence the FIREΔ/Δ mice's shortcomings imply loss of cognitive flexibility. There may be parallels in a rare disease that goes by the mouthful adult-onset leukoencephalopathy with axonal spheroids and pigmented glia. In ALSP, heterozygous mutations in CSF1R make microglia scarce, and neurons in white matter degenerate (Oosterhof et al., 2018). People with ALSP have trouble with memory and executive function.

Why do FIREΔ/Δ mice fail at myelin upkeep? They do have normal numbers of oligodendrocytes. To see if these cells might be compromised in other ways, McNamara teamed up with the labs of Josef Priller at Charité-Universitätsmedizin Berlin and U Edinburgh and Anna Williams at U Edinburgh. Profiling transcriptomes via single-nucleus RNA-Seq, the scientists found a preponderance of oligodendrocytes expressing Serpina3n.

This serine protease inhibitor marks an inflammatory type of oligodendrocyte found in mouse amyloidosis models (Jan 2020 news). Serpina3n also ticks up in plasma in early stages of AD, and in the cerebrospinal fluid of people with multiple sclerosis (Aug 2023 news; Fissolo et al., 2021). 

To figure out what these Serpina3n cells do, the scientists surveyed their transcriptomes for hints of altered biological pathways. This showed that these oligodendrocytes poorly regulate their own cholesterol and lipid metabolism. Lipidomic analysis of white matter backed this up. In collaboration with Jerome Hendricks at Hasselt University, Hasselt, Belgium, McNamara found that white-matter triglyceride levels were lower, and cholesteryl ester levels higher in FIREΔ/Δ mice than in wild-type.

What might connect the lipid metabolism of oligodendrocytes to a dearth of microglia? McNamara suspects TGF-β. Gene pathway analysis of the Serpina3n oligodendrocytes placed this growth factor upstream of the dysregulated lipid metabolism genes. In FIRED/D mice, twice as many oligodendrocytes lacked the TGF-β receptor. Conditionally knocking out this receptor in mouse oligodendrocytes had the same effect as ablating microglia, i.e., myelin started unraveling and the myelin tongue swelled. The plot thickened further when McNamara treated FIREΔ/Δ mice with SRI-011381, a TGF-β agonist that bypasses the receptor to trigger SMAD2–SMAD3 transcription factors downstream. Injected into the peritoneum three times a week beginning when mice were 2 months old, this agonist rescued myelination deficits in FIREΔ/Δ mice a month later.

All told, the findings suggest that microglia, by releasing TGF-β, help oligodendrocytes keep their lipid metabolism on an even keel.

In Leiden, Edorardo Marcora, Icahn School of Medicine at Mount Sinai, New York, asked if TREM2 loss-of-function variants linked to Alzheimer’s affect myelination. McNamara considers this question important but doesn’t think anyone has investigated.

Anil Cashikar, Washington University, St. Louis, wondered what happens to the excess cholesteryl esters made by the Serpina3n oligodendrocytes. “Do they form cholesterol crystals?” he asked. Such crystals have been linked to faulty myelin repair in old mice (Cantuti-Castelvetri et al., 2018). McNamara saw no such crystals in the FIRE mice, but noted that low ApoE expression by oligodendrocytes might mean the cells do not export cholesterol. “We have not yet nailed down the link between the pathology and lipid metabolism,” she said.

Pointers toward that link may come from work on myelin repair done in Gesine Saher’s lab at the Max-Planck-Institute of Experimental Medicine, Gottingen, Germany. In Leiden, Saher talked about how cholesterol helps microglia corral amyloid and how sterols made by microglia can help repair myelin lesions in mouse models of multiple sclerosis.

Myelin holds 70 percent of the cholesterol in the brain. When axons lose this insulation, oligodendrocytes trying to repair it churn out the enzymes needed to make cholesterol and other sterols. They include squalene synthase (SQS) and HMG-CoA reductase, the target of statins. Remyelination typically follows in two steps—an acute phase and a chronic phase. To their surprise, Stefan Berghoff and colleagues in Saher’s lab found that oligodendrocytes boosted sterol synthesis only in the chronic phase of remyelination. In the acute phase, it was microglia that churned out these enzymes. Furthermore, conditionally knocking out SQS in microglia severely limited acute remyelination (Berghoff et al., 2021). 

How do microglia support remyelination? Once again, their transcriptomes might point to an answer. In MS, and related animal models, degenerating myelin nudges microglia and macrophages into a phagocytic, inflammatory state so they can clear myelin debris. This state must resolve before remyelination can occur, but microglia unable to make sterols never reverted to their “normal” selves, reports Saher (image below). Instead, they kept making nitric oxide, interleukin-1β, Cxcl10, and other inflammatory mediators.

These mediators had something in common. Most are activated by liver X receptor signaling. Originally found in the liver, LXR receptors are expressed throughout the body. They regulate cholesterol, lipid, and glucose metabolism. Could microglial LXR signaling matter to remyelination?

Pursuing this idea, Berghoff found that phagocytic cells in three different models of MS upregulated LXR-dependent genes, including ApoE and ABCA1. But what set off this transcriptional change? Given that sterols activate LXR, and that remyelination requires microglial sterol synthesis, he looked for a sterol that might do the trick. He found that 24-dehydrocholesterol reductase (Dhcr24) was downregulated in the three models of MS—an important clue. If this enzyme were to stop working, then the LXR agonist desmosterol would accumulate provided the rest of the sterol synthesis pathway remained operational (image below).

Indeed, Saher said that’s exactly what happens. Using mass spectrometry, Berghoff found that desmosterol levels rose in mouse models of MS, but only when microglia synthesized squalene, a desmosterol precursor. In fact, adding squalene to mouse chow calmed microglial inflammation in MS mice, as did the Dhcr24 inhibitor, SH42. Squalene also reduced paralysis scores in the MS mice. So did the LXR agonist N,N-dimethyl-3β-hydroxycholenamide. It and squalene worked even better when given together.

All told, Saher explained, it is sterols, the very thing myelin is made of, that ultimately regulate microglial response to myelin injury. Curiously, even though microglia are packed with cholesterol they have phagocytosed from damaged myelin, the cells cannot simply recycle sterols from it. If they are to revert to homeostasis, they also need to boost sterol synthesis, because desmosterol cannot be made from cholesterol.

This work was done in mice and in cells. That said, it could be relevant to MS, and Alzheimer’s and other neurodegenerative disorders that feature demyelination (Roher et al., 2002; Aug 2021 news). If microglia in MS lesions cannot switch back to a non-inflammatory state, a vicious cycle of inflammation and demyelination may ensue, said Saher. Indeed, studies have found that Dhcr24 is down- and LXR target genes are upregulated in MS lesions. Might the same anti-inflammatory mechanisms be at play (Boven et al., 2006; Hendrickx et al., 2017; Mailleux et al., 2018)?—Tom Fagan

Comments

1. • Charles Stromeyer
     
   • Posted: 22 Oct 2023

   In AD and in vascular dementia, microglia also mop up myelin debris. Since this debris is high in iron content, the microglia are killed via ferroptosis (Adeniyi et al., 2023).

   References:

   Adeniyi PA, Gong X, MacGregor E, Degener-O'Brien K, McClendon E, Garcia M, Romero O, Russell J, Srivastava T, Miller J, Keene CD, Back SA. Ferroptosis of Microglia in Aging Human White Matter Injury. Ann Neurol. 2023 Dec;94(6):1048-1066. Epub 2023 Sep 14 PubMed.

   View all comments by Charles Stromeyer

Make a Comment

To make a comment you must login or register.

References

News Citations

1. Human and Mouse Microglia React Differently to Amyloid 17 Jan 2020
2. Proteins in Biofluids Foreshadow Dementia by 30 Years 29 Aug 2023
3. Flipping the Script: Could Myelin Degeneration Drive Amyloidosis? 27 Aug 2021

Paper Citations

1. McNamara NB, Munro DA, Bestard-Cuche N, Uyeda A, Bogie JF, Hoffmann A, Holloway RK, Molina-Gonzalez I, Askew KE, Mitchell S, Mungall W, Dodds M, Dittmayer C, Moss J, Rose J, Szymkowiak S, Amann L, McColl BW, Prinz M, Spires-Jones TL, Stenzel W, Horsburgh K, Hendriks JJ, Pridans C, Muramatsu R, Williams A, Priller J, Miron VE. Microglia regulate central nervous system myelin growth and integrity. Nature. 2023 Jan;613(7942):120-129. Epub 2022 Dec 14 PubMed.
2. Rojo R, Raper A, Ozdemir DD, Lefevre L, Grabert K, Wollscheid-Lengeling E, Bradford B, Caruso M, Gazova I, Sánchez A, Lisowski ZM, Alves J, Molina-Gonzalez I, Davtyan H, Lodge RJ, Glover JD, Wallace R, Munro DA, David E, Amit I, Miron VE, Priller J, Jenkins SJ, Hardingham GE, Blurton-Jones M, Mabbott NA, Summers KM, Hohenstein P, Hume DA, Pridans C. Deletion of a Csf1r enhancer selectively impacts CSF1R expression and development of tissue macrophage populations. Nat Commun. 2019 Jul 19;10(1):3215. PubMed.
3. Oosterhof N, Kuil LE, van der Linde HC, Burm SM, Berdowski W, van Ijcken WF, van Swieten JC, Hol EM, Verheijen MH, van Ham TJ. Colony-Stimulating Factor 1 Receptor (CSF1R) Regulates Microglia Density and Distribution, but Not Microglia Differentiation In Vivo. Cell Rep. 2018 Jul 31;24(5):1203-1217.e6. PubMed.
4. Fissolo N, Matute-Blanch C, Osman M, Costa C, Pinteac R, Miró B, Sanchez A, Brito V, Dujmovic I, Voortman M, Khalil M, Borràs E, Sabidó E, Issazadeh-Navikas S, Montalban X, Comabella Lopez M. CSF SERPINA3 Levels Are Elevated in Patients With Progressive MS. Neurol Neuroimmunol Neuroinflamm. 2021 Mar;8(2) Print 2021 Mar PubMed.
5. Cantuti-Castelvetri L, Fitzner D, Bosch-Queralt M, Weil MT, Su M, Sen P, Ruhwedel T, Mitkovski M, Trendelenburg G, Lütjohann D, Möbius W, Simons M. Defective cholesterol clearance limits remyelination in the aged central nervous system. Science. 2018 Feb 9;359(6376):684-688. Epub 2018 Jan 4 PubMed.
6. Berghoff SA, Spieth L, Sun T, Hosang L, Schlaphoff L, Depp C, Düking T, Winchenbach J, Neuber J, Ewers D, Scholz P, van der Meer F, Cantuti-Castelvetri L, Sasmita AO, Meschkat M, Ruhwedel T, Möbius W, Sankowski R, Prinz M, Huitinga I, Sereda MW, Odoardi F, Ischebeck T, Simons M, Stadelmann-Nessler C, Edgar JM, Nave KA, Saher G. Microglia facilitate repair of demyelinated lesions via post-squalene sterol synthesis. Nat Neurosci. 2021 Jan;24(1):47-60. Epub 2020 Dec 21 PubMed.
7. Roher AE, Weiss N, Kokjohn TA, Kuo YM, Kalback W, Anthony J, Watson D, Luehrs DC, Sue L, Walker D, Emmerling M, Goux W, Beach T. Increased A beta peptides and reduced cholesterol and myelin proteins characterize white matter degeneration in Alzheimer's disease. Biochemistry. 2002 Sep 17;41(37):11080-90. PubMed.
8. Boven LA, Van Meurs M, Van Zwam M, Wierenga-Wolf A, Hintzen RQ, Boot RG, Aerts JM, Amor S, Nieuwenhuis EE, Laman JD. Myelin-laden macrophages are anti-inflammatory, consistent with foam cells in multiple sclerosis. Brain. 2006 Feb;129(Pt 2):517-26. Epub 2005 Dec 19 PubMed.
9. Hendrickx DA, van Scheppingen J, van der Poel M, Bossers K, Schuurman KG, van Eden CG, Hol EM, Hamann J, Huitinga I. Gene Expression Profiling of Multiple Sclerosis Pathology Identifies Early Patterns of Demyelination Surrounding Chronic Active Lesions. Front Immunol. 2017;8:1810. Epub 2017 Dec 21 PubMed.
10. Mailleux J, Vanmierlo T, Bogie JF, Wouters E, Lütjohann D, Hendriks JJ, van Horssen J. Active liver X receptor signaling in phagocytes in multiple sclerosis lesions. Mult Scler. 2018 Mar;24(3):279-289. Epub 2017 Feb 1 PubMed.

Further Reading

No Available Further Reading