Want more evidence of the importance of microglia for Alzheimer’s disease? A study in the February 27 Science Advances suggests that spicing up the metabolism of these cells can tamp down Alzheimer’s pathology. Researchers led by Zengqiang Yuan at the Beijing Institute of Basic Medical Sciences investigated the effects of a flavonoid salt, sodium rutin, in two AD models. After consuming the salt for several months, the transgenic mice accumulated fewer plaques than untreated mice, while maintaining synapses and memory at wild-type levels. The authors attributed these improvements to healthier microglia and less inflammation. They report that cultured microglia exposed to sodium rutin expressed more phagocytic receptors, produced fewer pro-inflammatory cytokines, and made more ATP. A larger number of microglia surrounded plaques in treated mice than controls, and the cells internalized more Aβ.

  • The flavonoid sodium rutin stimulates microglial metabolism.
  • It boosts microglial phagocytosis of plaques, and lowers amyloid pathology in AD mouse models.

“This is a very interesting and important study that proposes sodium rutin as a potential candidate for AD treatment, through its amelioration of neuroinflammation,” Radosveta Koldamova at the University of Pittsburgh wrote to Alzforum. However, she cautioned that the authors examined but a handful of mice per group, highlighting the importance of replicating these findings in other studies. In the AD field, many treatments have worked in mice but failed to translate to people.

Rutin occurs naturally in plants such as apples, citrus fruits, green tea, and buckwheat. It acts as an antioxidant and anti-inflammatory, and has been found to inhibit Aβ aggregation in vitro (Jiménez-Aliaga et al., 2011; Wang et al., 2012). A previous study reported less neuroinflammation, fewer Aβ oligomers, and better memory in transgenic mice that consumed rutin (Xu et al., 2014). Other groups noted a beneficial effect of rutin on memory in injection models of neurotoxicity (Javed et al., 2012; Moghbelinejad et al., 2013; Choi et al., 2015). 

Microglial Invasion. More microglia (green) crowd around amyloid plaques (red) in APP/PS1 mice fed sodium rutin (right) than in untreated controls (left). Nuclei are blue. [Courtesy of Science/AAAS.]

However, the compound dissolves poorly in water, limiting its bioavailability. To get around this limitation, first author Rui-Yuan Pan dissolved rutin in sodium hydroxide, then purified the salt. Sodium rutin was 200-fold more soluble than its precursor, and achieved fivefold higher concentration in mouse serum. The compound crosses the blood-brain barrier, appearing in brain tissues two hours after ingestion. The authors did not report the concentration in brain.

Pan administered 0.25 mg/ml sodium rutin in water to six-month-old APP/PS1 mice for seven months. At 13 months, treated animals had half as many amyloid plaques in the prefrontal cortex as untreated controls did. They had less microgliosis, astrogliosis, and pro-inflammatory cytokines, but higher levels of anti-inflammatory cytokines such as IL-10. Treatment maintained dendritic spine density at wild-type levels, and rescued performance in the Morris water maze. In a second mouse model, 5xFAD, sodium rutin treatment lowered plaque burden by two-thirds and rescued long-term potentiation in slices. For behavioral studies, the authors analyzed about a dozen mice per group, while pathology studies used three to six mice per group.

How might sodium rutin produce these effects in the brain? The authors report that expression of several microglial phagocytic receptors, including TREM2, CR3, GPR34, MerTK, and P2Y6, bumped up by 50 percent or more in prefrontal cortex of treated APP/PS1 mice. Sodium rutin doubled the amount of TREM2 in microglia and enhanced its recycling to the cell surface. The authors did not report expression data for the 5xFAD mice.

TREM2 helps maintain mitochondrial energy production (Aug 2017 news). This led the authors to measure metabolism in cultured microglia exposed to sodium rutin. The treatment increased ATP production and oxygen consumption, they report. Sodium rutin localized to mitochondria in these cells. When the authors tested sodium rutin on microglia from TREM2 knockout mice, which have impaired oxidative phosphorylation, the treatment rescued ATP production. Thus, the authors concluded, the effects of sodium rutin do not depend on TREM2.

Would fitter mitochondria facilitate phagocytosis, which makes high energy demands on the cell? In support of this, the authors measured twice as many CD68-positive phagosomes and four times as much internalized Aβ in the prefrontal cortices of treated than untreated APP/PS1 mice. In addition, more microglia clustered around plaques in treated mice (see image above).

Overall, the data so far suggest that sodium rutin exerts its effects mainly through microglia, Yuan told Alzforum. He said that tracing experiments with biotinylated sodium rutin showed that it appeared almost exclusively in microglia in mouse cortex. In addition, the flavonoid had no effect on oxidative phosphorylation in cultured neurons or astrocytes. In future work, Yuan is searching for the cellular targets of sodium rutin to determine how it upregulates receptors and revs up metabolism. He hopes to test the compound in clinical trials.—Madolyn Bowman Rogers

Comments

  1. This study tests the effect of Rutin (quercetin-3-rutinoside), a natural flavonoid with different biological functions that has been used in traditional medicines. The team processed rutin into sodium rutin using alkaline aqueous solution, significantly improving its water solubility and bioavailability. This makes it a much more promising drug than the natural compound.

    The authors found a remarkable reduction of Aβ plaques in two mouse models of AD, namely APP/PSdE9 and 5xFAD. In both, the treatment started at a stage of already developed amyloid deposition so the effect could be characterized as therapeutic that reduces already formed amyloid. Pan et al. found that sodium rutin treatment did not affect APP processing and microglia proliferation but enhanced microglial recruitment around the plaques, which potentially can promote Aβ phagocytosis and clearance.

    Furthermore, they found increased expression and recycling of phagocytic receptors such as GPR34, P2Y6, and others. However, in vitro experiments demonstrated that the phagocytic effect of sodium rutin was not affected by knockdown of phagocytic receptors such as Trem2, CR3, and MerTK.

    To investigate the effect of sodium rutin on the phagocytic receptor Trem2 further, the authors used Trem2-deficient mice. Interestingly, the amyloid pathology in Trem2-deficient AD mice was ameliorated by sodium rutin treatment similarly as in Trem2-WT mice. Together, the in vitro and in vivo data suggest that at least these phagocytic receptors are not essential for the sodium rutin-mediated enhancement of Aβ phagocytosis and clearance.

    Finally, the authors demonstrated that sodium rutin promotes a metabolic switch from anaerobic glycolysis to mitochondrial OXPHOS (oxidative phosphorylation), which could provide microglia with sufficient energy (ATP) for Aβ clearance.

    This is a very interesting and important study that proposes sodium rutin as a potential candidate for AD treatment through amelioration of neuroinflammation.

    The number of mice per group is low. For example, the effect on amyloid deposition is analyzed on 14 to 18 slices from three mice per group (Fig.3). For some of the experiments (Fig.4) the data are presented as number of plaques, but it is not clear from how many mice are these plaques were collected.

  2. Pan et al. report that oral administration of the flavonoid rutin to mouse models of Alzheimer’s disease results in amelioration of cognition deficits as well as a broad array of AD-related phenotypes. They report a dramatic reduction in plaque burden, an increase in plaque-associated microglia, enhanced phagocytic activity, and reduction in neuroinflammation. Remarkably, rutin elicits a robust increase in microglial (but not neuronal) oxidative metabolism. Indeed, it appears to be an elixir.

    The novelty of this study arises principally from the reformulation of the compound as a sodium salt, improving its aqueous solubility. Previous studies reported in 2014 found salutary effects in an AD model on a more restricted range of endpoints, thus conceptually the study doesn’t break much new ground. There are >6,000 papers investigating the actions of rutin. There are myriad mechanisms of action that have been described, most prominently anti-oxidative and anti-inflammatory. The present study does not explore the underlying mechanism for any of rutin’s positive effects and it is difficult to understand how they are achieved and unlikely they can be ascribed to any single mechanism. It is notable that despite the very extensive study of this natural product there are few, if any, credible examples of its actions in humans. This raises the question of whether the robust effects of rutin (or maybe other flavonoids) observed in rodents in a large number of experimental settings are at all relevant in humans.

    One of the most provocative findings is that rutin treatment results in a shift in the relative contribution of mitochondrial oxidative phosphorylation to microglial metabolism. This finding is of considerable significance and it is not clear how rutin might effect this change. I am not persuaded that overall microglial energetics is rate-limiting for phagocytosis. The authors state that the ability of rutin to enhance phagocytosis is due to induction of phagocytic receptors. This is an overstatement. They only have a correlation.

    There are a few technical issues that need to be considered, most prominently the use of mice over a period of four to eight months of age in 5XFAD mice and six to 13 month APP/PS1 for behavioral experiments and synapse analysis. This is problematic as this spans the period in which rapid plaque deposition is occurring, and unless the cohorts were tightly matched for age and sex it is hard to understand how they achieve statistical significance. In our experience, it is hard to detect a genotype effect in the 5XFAD mice at four months. The change in synapse number and morphology is very small and of questionable biological significance. The ages of the animals were not stated for the other experiments (I didn’t have access to the supplemental material) and if there is a similar spread in ages it colors the interpretation of the data significantly.

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References

Research Models Citations

  1. APPswe/PSEN1dE9
  2. 5xFAD (B6SJL)

News Citations

  1. Without TREM2, Microglia Run Out of Gas

Paper Citations

  1. . Quercetin and rutin exhibit antiamyloidogenic and fibril-disaggregating effects in vitro and potent antioxidant activity in APPswe cells. Life Sci. 2011 Dec 19;89(25-26):939-45. PubMed.
  2. . Rutin inhibits β-amyloid aggregation and cytotoxicity, attenuates oxidative stress, and decreases the production of nitric oxide and proinflammatory cytokines. Neurotoxicology. 2012 Jun;33(3):482-90. PubMed.
  3. . Rutin improves spatial memory in Alzheimer's disease transgenic mice by reducing Aβ oligomer level and attenuating oxidative stress and neuroinflammation. Behav Brain Res. 2014 May 1;264:173-80. Epub 2014 Feb 7 PubMed.
  4. . Rutin prevents cognitive impairments by ameliorating oxidative stress and neuroinflammation in rat model of sporadic dementia of Alzheimer type. Neuroscience. 2012 May 17;210:340-52. PubMed.
  5. . Rutin activates the MAPK pathway and BDNF gene expression on beta-amyloid induced neurotoxicity in rats. Toxicol Lett. 2013 Oct 20;224(1):108-113. PubMed.
  6. . The n-Butanol Fraction and Rutin from Tartary Buckwheat Improve Cognition and Memory in an In Vivo Model of Amyloid-β-Induced Alzheimer's Disease. J Med Food. 2015 Jun;18(6):631-41. Epub 2015 Mar 18 PubMed.

Further Reading

Primary Papers

  1. . Sodium rutin ameliorates Alzheimer's disease-like pathology by enhancing microglial amyloid-β clearance. Sci Adv. 2019 Feb;5(2):eaau6328. Epub 2019 Feb 27 PubMed.