Imagine boosting the plaque-degrading prowess of astrocytes, the most abundant brain cell. Researchers led by Kalipada Pahan, Rush University Medical Center, Chicago, suggest this may be possible, at least in mice, by using a widely prescribed drug and vitamin A. In the October 26 Science Signaling, they reported that adding gemfibrozil and retinoic acid to mouse astrocyte cultures spurred clearance of Aβ. The drugs increased expression of LDL receptors and prodded lysosome activity in the cells. In a mouse model of amyloidosis, the combo slowed plaque accumulation and preserved memory and spatial learning. The findings highlight the potential of stimulating astrocytes and suggest the drug duo may be worth testing in clinical trials.
- Gemfibrozil and retinoic acid made astrocytes clear more Aβ.
- The cells' LDL receptors multiplied, lysosomes went into overdrive.
- Peroxisome proliferator-activated receptor α may drive the changes.
“Astrocytes have an underappreciated capability to clear extracellular amyloid,” Ralph Nixon, New York University, told Alzforum. He noted that while microglia carry up to five times as much autophagic machinery as astrocytes, the latter are more plentiful. “If you add up all the astrocyte autophagic capability in the brain, it may be very significant,” he said.
Gemfibrozil is a cholesterol-lowering drug that can cross the blood-brain barrier. It activates the peroxisome proliferator-activated receptor α. Retinoic acid receptors aid PPARa function, which may explain why the two work better in combination (van Neerven et al., 2008).
Gary Landreth, Indiana University School of Medicine, Indianapolis, noted that the effects of these drugs are not limited to PPARα. “The drug cocktail not only stimulates PPARα, but also all other type II receptors and RARs, which have a broad range of target genes," he wrote to Alzforum (Feb 2012 news).
Previously, Pahan's and other groups reported that these drugs improved plaque clearance and memory in mouse models of amyloidosis (Chandra and Pahan, 2019; Luo et al., 2020). Gemfibrozil and retinoic acid also upregulate the transcription factor TFEB, a driver of lysosomal activity in astrocytes (Ghosh et al., 2015). Astrocyte-specific expression of TFEB in amyloidosis mice revved lysosomes to clear plaques (Xiao et al., 2014). Could the two drugs have cleared plaques in the present study by similarly ramping up astrocyte autophagy?
To find out, co-first authors Sumita Raha and Arunava Ghosh added the drugs to mouse astrocytes in culture, then “fed” them fluorescent Aβ. After four hours, the treated cells took up 80 percent more Aβ than did untreated astrocytes. The fluorescent Aβ co-localized with LysoTracker Red, a lysosome marker (see image below).
How did the drugs work? First, the scientists focused on how the cells took up Aβ. Micropinocytosis—the pinching of cell-membrane invaginations—accounted for some but not all of the drug’s effects. Raha et al. wondered if low-density lipoprotein receptors might explain the rest. LDLRs boost plaque clearance in the mouse brain, and their expression in the liver has been linked to PPARα (Dec 2009 news; Mar 2012 conference news; Nov 2002 news; Huang et al., 2008). Indeed, the drug combo raised LDLR levels in the cultured astrocytes, and knocking down the LDLR attenuated Aβ uptake.
Next, the authors assessed degradation within the astrocytes. To determine how fast the cells broke down Aβ, they let them imbibe the fluorescently labelled peptide, then watched the fluorescence fade. Compared to controls, astrocytes “on” the drugs cleared Aβ twice as fast. They also produced twice as much cathepsin B and D, two lysosomal proteases that degrade Aβ. On the other hand, neither Aβ clearance nor cathepsin levels budged if the scientists knocked down TFEB, or when they blocked acidification of the lysosomes, which is essential for proteolysis in these organelles. All told, the findings suggested that gemfibrozil and retinoic acid together ramped up TFEB expression, which boosted lysosomal function and accelerated Aβ degradation.
Supporting this interpretation was the finding that PPARα was essential for these drug-induced changes. In cultured astrocytes lacking the PPARα receptor, the drug combination left LDLR expression, Aβ uptake and clearance, and lysosome production unchanged.
How about in vivo? The scientists fed 6-month-old 5xFAD mice both drugs for two months. Treated mice expressed more astrocyte LDLR than did untreated mice. They also had more TFEB and lysosome membrane proteins in their astrocytes and neurons, indicating ramped-up lysosome production.
Plaques are abundant in 5xFAD mice by 8 months of age, but compared to controls, treated mice had fewer plaques in their hippocampi at this age (see image below). They navigated a T maze more easily, identified novel objects placed in their cages faster and more accurately, and chose the tunnel containing food in a Barnes maze more often than did untreated mice.
Treatment had no benefit if the scientists knocked out the PPARα gene in mouse astrocytes, though Jungsu Kim and Hande Karahan, Indiana University, Indianapolis, offered a cautionary note about the mice used for this experiment. "It would strengthen this study if astrocyte-specific PPARα deletion could be corroborated by co-staining other cell types," they wrote (full comment below).
Nixon noted that astrocytes' ability to clear plaques seems powerful but latent. “If astrocytes are capable of clearing Aβ from 6-month-old 5xFAD mice after treatment, why doesn’t that happen without drugs?” he asked.
Moreover, Nixon and others questioned if the memory improvement in 5xFAD mice is due to Aβ clearance by astrocytes, or something else. “The drugs seem to target astrocytes, but whether they mediate behavior through clearance of Aβ or through some other biological role in other brain cells is unclear,” Nixon said. Takahisa Kanekiyo, Mayo Clinic, Jacksonville, thinks astrocytes may communicate with microglia and neurons, perhaps by altering lipid metabolism. “The therapeutic effects of gemfibrozil and retinoic acid might be achieved through multiple mechanisms beyond astrocytic Aβ degradation,” he wrote (full comment below).
What could these findings mean for treatment? Other research groups have previously, and separately, tested gemfibrozil and the retinoic acid receptor agonist acitretin in pilot AD trials, to no avail (Dec 2019 conference news; Endres et al., 2014).
“Preclinical experiments suggest retinoic acid might be useful to treat AD, but we still need larger clinical trials with cognitive measurements,” Kristina Endres, Johannes Gutenberg University, Mainz, Germany, wrote to Alzforum. She noted that the current paper did not compare the effects of the drug combination to those of each individually. “One cannot judge the superiority of the combined drugs versus the already multifunctional effects of retinoic acid alone,” Endres added.
Peter McCaffery, University of Aberdeen, Scotland, wondered if retinoid X receptor agonists, such as bexarotene, could be combined with gemfibrozil, rather than retinoic acid itself, to achieve similar effects (full comment below). Bexarotene, which stimulates retinoic receptors to bind PPARα’s cousin PPARγ and boost formation of ApoE, failed in early stage AD trials, including one that assessed its effect on Aβ levels (Aug 2019 news; Feb 2016 news).
Kelly Dineley, University of Texas Medical Branch at Galveston, wondered which species of Aβ, or rather its removal, was behind the pathology and cognition improvement reported in this paper (full comment below).
Pahan has licensed the gemfibrozil/retinoic acid combination to a company that is working on an IND application to test it in an AD trial.—Chelsea Weidman Burke
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Research Models Citations
- Chandra S, Pahan K. Gemfibrozil, a Lipid-Lowering Drug, Lowers Amyloid Plaque Pathology and Enhances Memory in a Mouse Model of Alzheimer's Disease via Peroxisome Proliferator-Activated Receptor α. J Alzheimers Dis Rep. 2019 May 18;3(1):149-168. PubMed.
- Luo R, Su LY, Li G, Yang J, Liu Q, Yang LX, Zhang DF, Zhou H, Xu M, Fan Y, Li J, Yao YG. Activation of PPARA-mediated autophagy reduces Alzheimer disease-like pathology and cognitive decline in a murine model. Autophagy. 2019 Mar 22;:1-18. PubMed.
- Ghosh A, Jana M, Modi K, Gonzalez FJ, Sims KB, Berry-Kravis E, Pahan K. Activation of peroxisome proliferator-activated receptor α induces lysosomal biogenesis in brain cells: implications for lysosomal storage disorders. J Biol Chem. 2015 Apr 17;290(16):10309-24. Epub 2015 Mar 6 PubMed.
- Xiao Q, Yan P, Ma X, Liu H, Perez R, Zhu A, Gonzales E, Burchett JM, Schuler DR, Cirrito JR, Diwan A, Lee JM. Enhancing astrocytic lysosome biogenesis facilitates Aβ clearance and attenuates amyloid plaque pathogenesis. J Neurosci. 2014 Jul 16;34(29):9607-20. PubMed.
- Huang Z, Zhou X, Nicholson AC, Gotto AM Jr, Hajjar DP, Han J. Activation of peroxisome proliferator-activated receptor-alpha in mice induces expression of the hepatic low-density lipoprotein receptor. Br J Pharmacol. 2008 Oct;155(4):596-605. Epub 2008 Aug 18 PubMed.
- Endres K, Fahrenholz F, Lotz J, Hiemke C, Teipel S, Lieb K, Tüscher O, Fellgiebel A. Increased CSF APPs-α levels in patients with Alzheimer disease treated with acitretin. Neurology. 2014 Nov 18;83(21):1930-5. Epub 2014 Oct 24 PubMed.
- Lauer AA, Janitschke D, Dos Santos Guilherme M, Nguyen VT, Bachmann CM, Qiao S, Schrul B, Boehm U, Grimm HS, Hartmann T, Endres K, Grimm MO. Shotgun lipidomics of liver and brain tissue of Alzheimer's disease model mice treated with acitretin. Sci Rep. 2021 Jul 27;11(1):15301. PubMed.
- Wójtowicz S, Strosznajder AK, Jeżyna M, Strosznajder JB. The Novel Role of PPAR Alpha in the Brain: Promising Target in Therapy of Alzheimer's Disease and Other Neurodegenerative Disorders. Neurochem Res. 2020 May;45(5):972-988. Epub 2020 Mar 13 PubMed.
- Raha S, Ghosh A, Dutta D, Patel DR, Pahan K. Activation of PPARα enhances astroglial uptake and degradation of β-amyloid. Sci Signal. 2021 Oct 26;14(706):eabg4747. PubMed.