27 Oct 2023

Inflammation can be a helpful response to the common cold or the bite of an insect, but dangerous if it persists. In neurodegeneration, chronic micro- and astrogliosis spell trouble for the brain. Does it have to be that way? Over the past 20 years, scientists have discovered “specialized pro-resolving mediators” that take the sting out of inflammation. Derived from long-chain fatty acids, these SPMs are potent, yet hard to come by in the body.

As described at the 2nd Symposium on Lipids in Brain Diseases, there may be ways to gin them up. Oliver Werz, Friedrich Schiller University in Jena, Germany, described small molecules that fool lipid oxygenases into producing SPMs instead of their inflammatory cousins, such as prostaglandins. Amsterdam University's Julia Konings reported ways to shift the microglial lipidome toward a “pro-resolution” phenotype. Whether these strategies could help reduce inflammatory responses in the brain needs to be studied, but some of these SPMs wane in the AD brain.   

Before we delve into the, ahem, “meat” of this fat story, a bit of background on the subtle differences between the inflammatory and pro-resolving lipids. The former mostly come from arachidonic acid, a 20-carbon fatty acid with four double bonds. Cyclooxygenases, the target of vioxx, celecoxib, and some other non-steroidal anti-inflammatories, generate a stew of pro-inflammatory prostaglandins and thromboxanes from this fatty acid by forming an aliphatic ring in the middle of the carbon chain. 5-Lipoxygenase (5-LOX) creates another group of inflammatories, the leukotrienes, by introducing a hydroxyl group on carbon number 5, hence the name 5-lipoxygenase. Here’s where it starts to get complicated. 5-LOX also makes some of the SPMs, namely the lipoxins (image below). The lipoxins can also be made from another 20-carbon fatty acid, eicosapentaenoic acid. The penta here signifies that this fatty acid has five double bonds, one more than in arachidonic acid, which is a less technical name for eicosatetraenoic acid.

There’s more. Wait till you hear about resolvins and protectins. (If you can deal with multi-omic datasets, you can handle this biochemistry.) These SPMs form when yet another enzyme, 15-LOX, oxidizes eicosapentaenoic acid or eicosatetraenoic acid. See the nomenclature pattern here—15-LOX adds a hydroxyl group to the carbon 15 of the fatty acids. Other enzymes jump in to create varieties of lipoxins, resolvins, and protectins, but the 5- and 15-LOXs are the main drivers. Yet another set of SPMs, the maresins, form when 12-LOX oxidizes docosahexaenoic acid—a 22 carbon fatty acid with, you guessed it, six double bonds.

Lives of Lipids. Long-chain fatty acids give rise to a plethora of inflammatory lipids (red) and pro-resolving mediators (blue). [Courtesy of Oliver Werz.]

Could this web of relationships be tweaked to shift the balance from pro-inflammatory to pro-resolving? Werz believes so. However, a major obstacle has stymied this line of research. 15-LOX has been notoriously difficult to activate. Scientists have a multitude of tools at hand to induce prostaglandins and leukotrienes, including lipopolysaccharide from bacteria, zymosan from fungi, complement 5a, calcium ionophores, even cell stressors such as hyperosmosis; alas, they could not figure out how to stimulate SPMs until researchers in Charles Serhan’s lab at Brigham and Women’s Hospital, Boston, discovered that bacteria do the trick (Chiang et al., 2012). While in Serhan’s lab, Werz found that E. coli and S. aureus, counterintuitively perhaps, did so by stimulating 15-LOX in pro-resolving macrophages (Werz et al., 2018). 

It now appears that some noninfectious agents mimic this effect. In Leiden, Werz reminded the audience that α-hemolysin, a toxin produced by S. aureus, bumps up production of maresins and resolvins when injected into the peritoneum of mice (Jordan et al., 2020). Better than using the bacteria, but still a toxin. What about other approaches?

Blocking cyclooxygenase to limit production of prostaglandins may seem an option, but Werz emphasized that this only tempers acute inflammation. Cox inhibitors do poorly in the long run because they prevent formation of prostaglandin E2, one of the few anti-inflammatory prostaglandins, and because with no cyclooxygenase to act on arachidonic acid, 5-LOX begins to churn out the inflammatory leukotrienes. Indeed, macrophages treated with celecoxib ramp up production of leukotriene B4 up to fourfold. The key to flipping the “lipid class switch,” i.e., producing more SPMs at the expense of inflammatory lipids, lies in 5-LOX, said Werz.

In his talk, Werz showed how he could turn 5-LOX into 15-LOX using an allosteric modulator called acetyl-keto-boswellic acid. No, not named after the Scottish author, AKBA comes from plants in the genus Boswellia. (It includes frankincense). Unlike 5-LOX inhibitors that sit in the active site of the oxygenase, AKBA sits at a distal site. From there, it breaks hydrophobic bonds between amino acid side chains and causes a conformational ripple through the enzyme right down to where arachidonic acid docks (Gilbert et al., 2020). The upshot: The fatty acid repositions in the active site in such a way that carbons 12 or 15 get attacked and oxidized instead of carbon 5.

Lipoxygenase Switch. In 5-lipoxygenase (left), AKBA (yellow) sits between the amino (gray) and catalytic domains (cyan). By disturbing that interface (right) it restructures the active site (red circle). [Courtesy of Gilbert et al., 2020.]

Werz’s group found that in cell-free systems, in HEK293 cells, and in human macrophages, monocytes and leukocytes, AKBA shifted lipid profiles toward the pro-resolving mediators (Gilbert et al., 2020; Börner et al., 2023). As a bonus, AKBA also increased 15-LOX activity, reported Werz, boosting SPMs even more. Injected into the peritonea of mice, the allosteric modulator evoked a huge increase in SPMs when the animals were injected with zymosan (image below).

Could AKBA resolve inflammation in people? A hint comes from an eight-month, open-label clinical trial of a frankincense extract in 28 people with relapsing-remitting multiple sclerosis. Within five months of starting treatment, the number of myelin lesions in the brain was reported to go down, as judged by MRI, and remain low until month eight (Stürner et al., 2018). Of 44 lipids tested in the plasma during that time, the concentrations of eight fell; seven of those were products of 5-LOX (Stürner et al., 2020). 

Participants who continued the treatment had no or few relapses up to month 36, though no further MRI scans were taken during that time. Werz said he does not know of any follow-up trial of this extract in MS, and he thinks other compounds would be worth testing. Screening a variety of natural anti-inflammatories, scientists in his lab found that celastrol and cannabidiol act similarly to AKBA, inhibiting pro-inflammatory and promoting pro-resolving mediators in mice (Pace et al., 2021; Peltner et al., 2023). 

A different strategy to tackle inflammation in MS—though still focused on 5-LOX—was presented by Konings, who works in Gijs Kooij’s lab in Amsterdam. Last June, Jelle Broos and colleagues in the same lab reported that all was not well with arachidonic acid metabolism in MS (Broos et al., 2023). Specifically, in people with the progressive form of the disease, plasma levels of lipid metabolites, including leukotrienes produced by 5-LOX, correlated with disease severity. Konings postulated that 5-LOX activating protein (FLAP), which transfers lipids to 5-LOX, might be involved, but little was known about FLAP in MS.

Why Hello, Microglia
Konings looked to see where 5-LOX and FLAP are made in the brain. While MS and control brains contained similar levels of 5-LOX, MS brains had more FLAP, mostly around active lesions. Co-staining with IBA1 and TMEM119 identified microglia as the source. To investigate, Konings used microglia derived from human induced pluripotent stem cells. When she challenged them with inflammatory molecules, such as lipopolysaccharide or interferon-γ, they churned out five times as much FLAP as did control cells.

Could this be modulated? When Konings treated microglia with experimental FLAP inhibitors such as Fiboflapon or AZX5718, the cells' lipidome shifted from a pro-inflammatory to a pro-resolving profile, suppressing leukotrienes and other 5-LOX products. How this affects microglial function needs to be worked out, but preliminary data suggest that they reduce expression of the inflammatory cytokine interleukin 1β.

At the Leiden conference, scientists were excited at the potential for shifting lipidomes in this way. “The concept is very promising,” Kooij told Alzforum. “If you are not just targeting one molecule, but shifting a whole suite of molecules from inflammation to protection, that could be extremely potent.”

Even so, they had many questions. Are SPMs perturbed in diseases besides MS? What triggers their production? How do they work?  “We don’t have the answers yet,” said Werz. Kooij told Alzforum that these lipid mediators have been extremely difficult to study because they are so hard to detect. “We believe they have direct effects only on nearby cells and, as such, they are only produced at really low concentrations,” he said.

There are signs that these molecules are important in other diseases. Previous studies reported that resolvins and lipoxins drop in the Alzheimer's brain. These same compounds tempered inflammation in mouse models of the disease (Zhu et al., 2016; Apr 2018 news; Kantarci et al., 2018). 

As for physiological triggers of SPMs, these might include certain cytokines, said Kooij, adding that this remains to be studied. Werz thinks specific receptors activate 15-Lox. Curiously, one might be ADAM10, the α-secretase that processes amyloid-precursor protein. ADAM10 binds α-hemolysin.

Downstream, SPMs mainly target immune cells, stimulating pro-resolving macrophages and blocking neutrophil infiltration and migration. How they do this needs to be understood. They appear to start out by binding G-protein coupled receptors. “At least six receptors have been proposed, but it is not entirely clear which might mediate effects of SPMs,” said Werz. It’s a buzzing area of investigation. “I think we will hear a lot more about this in the coming year,” Kooij predicted.—Tom Fagan

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References

News Citations

1. Loss of Soothing Lipids in AD Brain Furthers Microglial Mayhem 27 Apr 2018

Paper Citations

1. Chiang N, Fredman G, Bäckhed F, Oh SF, Vickery T, Schmidt BA, Serhan CN. Infection regulates pro-resolving mediators that lower antibiotic requirements. Nature. 2012 Apr 25;484(7395):524-8. PubMed.
2. Werz O, Gerstmeier J, Libreros S, De la Rosa X, Werner M, Norris PC, Chiang N, Serhan CN. Human macrophages differentially produce specific resolvin or leukotriene signals that depend on bacterial pathogenicity. Nat Commun. 2018 Jan 4;9(1):59. PubMed.
3. Jordan PM, Gerstmeier J, Pace S, Bilancia R, Rao Z, Börner F, Miek L, Gutiérrez-Gutiérrez Ó, Arakandy V, Rossi A, Ialenti A, González-Estévez C, Löffler B, Tuchscherr L, Serhan CN, Werz O. Staphylococcus aureus-Derived α-Hemolysin Evokes Generation of Specialized Pro-resolving Mediators Promoting Inflammation Resolution. Cell Rep. 2020 Oct 13;33(2):108247. PubMed.
4. Gilbert NC, Gerstmeier J, Schexnaydre EE, Börner F, Garscha U, Neau DB, Werz O, Newcomer ME. Structural and mechanistic insights into 5-lipoxygenase inhibition by natural products. Nat Chem Biol. 2020 Jul;16(7):783-790. Epub 2020 May 11 PubMed.
5. Börner F, Pace S, Jordan PM, Gerstmeier J, Gomez M, Rossi A, Gilbert NC, Newcomer ME, Werz O. Allosteric Activation of 15-Lipoxygenase-1 by Boswellic Acid Induces the Lipid Mediator Class Switch to Promote Resolution of Inflammation. Adv Sci (Weinh). 2023 Feb;10(6):e2205604. Epub 2022 Dec 25 PubMed.
6. Stürner KH, Stellmann JP, Dörr J, Paul F, Friede T, Schammler S, Reinhardt S, Gellissen S, Weissflog G, Faizy TD, Werz O, Fleischer S, Vaas LA, Herrmann F, Pless O, Martin R, Heesen C. A standardised frankincense extract reduces disease activity in relapsing-remitting multiple sclerosis (the SABA phase IIa trial). J Neurol Neurosurg Psychiatry. 2018 Apr;89(4):330-338. Epub 2017 Dec 16 PubMed.
7. Stürner KH, Werz O, Koeberle A, Otto M, Pless O, Leypoldt F, Paul F, Heesen C. Lipid Mediator Profiles Predict Response to Therapy with an Oral Frankincense Extract in Relapsing-Remitting Multiple Sclerosis. Sci Rep. 2020 May 29;10(1):8776. PubMed.
8. Pace S, Zhang K, Jordan PM, Bilancia R, Wang W, Börner F, Hofstetter RK, Potenza M, Kretzer C, Gerstmeier J, Fischer D, Lorkowski S, Gilbert NC, Newcomer ME, Rossi A, Chen X, Werz O. Anti-inflammatory celastrol promotes a switch from leukotriene biosynthesis to formation of specialized pro-resolving lipid mediators. Pharmacol Res. 2021 May;167:105556. Epub 2021 Mar 31 PubMed.
9. Peltner LK, Gluthmann L, Börner F, Pace S, Hoffstetter RK, Kretzer C, Bilancia R, Pollastro F, Koeberle A, Appendino G, Rossi A, Newcomer ME, Gilbert NC, Werz O, Jordan PM. Cannabidiol acts as molecular switch in innate immune cells to promote the biosynthesis of inflammation-resolving lipid mediators. Cell Chem Biol. 2023 Aug 21; PubMed.
10. Broos JY, Loonstra FC, de Ruiter LR, Gouda M, Fung WH, Schoonheim MM, Heijink M, Strijbis EM, Teunissen C, Killestein J, de Vries HE, Giera M, Uitdehaag BM, Kooij G. Association of Arachidonic Acid-Derived Lipid Mediators With Disease Severity in Patients With Relapsing and Progressive Multiple Sclerosis. Neurology. 2023 Aug 1;101(5):e533-e545. Epub 2023 Jun 8 PubMed.
11. Zhu M, Wang X, Hjorth E, Colas RA, Schroeder L, Granholm AC, Serhan CN, Schultzberg M. Pro-Resolving Lipid Mediators Improve Neuronal Survival and Increase Aβ42 Phagocytosis. Mol Neurobiol. 2016 May;53(4):2733-49. Epub 2015 Dec 9 PubMed.
12. Kantarci A, Aytan N, Palaska I, Stephens D, Crabtree L, Benincasa C, Jenkins BG, Carreras I, Dedeoglu A. Combined administration of resolvin E1 and lipoxin A4 resolves inflammation in a murine model of Alzheimer's disease. Exp Neurol. 2018 Feb;300:111-120. Epub 2017 Nov 7 PubMed.

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