Seo DO, O'Donnell D, Jain N, Ulrich JD, Herz J, Li Y, Lemieux M, Cheng J, Hu H, Serrano JR, Bao X, Franke E, Karlsson M, Meier M, Deng S, Desai C, Dodiya H, Lelwala-Guruge J, Handley SA, Kipnis J, Sisodia SS, Gordon JI, Holtzman DM. ApoE isoform- and microbiota-dependent progression of neurodegeneration in a mouse model of tauopathy. Science. 2023 Jan 13;379(6628):eadd1236. PubMed.

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  1. In this heroic work, Seo et al. provide evidence for an ApoE-dependent and ApoE-isoform-dependent role of the gut microbiome in neuropathology in the P301S tau model, expressing human ApoE3 (TE3) or ApoE4 (TE4). Neuropathology in conventionally reared (Conv-R) TE4 mice, germ-free (GF) TE4 mice, and GF TE4 mice that received fecal implants at 12 weeks of age from sex-matched 40-week-old TR4 mice (Ex-GF), was analyzed at 40 weeks of age, when TE4 mice showed greater neuropathology than TE3 mice in earlier studies. Compared to E4 mice without P301S tau, ConV-R female and male TE4 mice showed hippocampal volume loss and enlargement of the lateral ventricle (LV) volume. GF TE4 female and male mice showed less hippocampal volume loss and less LV volume than ConV-R female and male TE4 mice, but less hippocampal volume and more LV volume than ConV-R and GF E4 mice.

    These effects showed anatomical specificity, because no difference in neuropathology in the entorhinal-piriform cortex was seen. The protective effects seen for hippocampal volume and hippocampal tau phosphorylation in GF TE4 mice were not seen in Ex-GF TE4 mice, while the protective effects seen for LV volume seemed partially retained, especially in males. The protective effects in GF TE4 mice were associated with reduced immunoreactivity for markers of astrocytes and activated microglia and with large and more branched astrocytes, an astrocytic phenotype already seen at 12 weeks of age and preceding severe tau pathology. Consistent with these changes, differential gene expression at 40 weeks of age was seen comparing ConV-R and GF-E4 mice but not comparing ConV-R and Ex-GF TE4 mice.

    Antibiotic treatment from postnatal 16 through 22 increased hippocampal volume at 40 weeks in TE3 male, but not female, mice. There was a trend toward a treatment effect in TE4 male, but not female, mice but it did not reach significance. Antibiotic treatment improved nest building in TE3 and TE4 male, but not female, mice, and in both genotypes nest building was positively correlated with hippocampal volume. Effects of antibiotic treatment on LV volume were also seen only in TE3 and TE4 males but not females. Consistent with this pattern, hippocampal tau phosphorylation was significantly reduced only in TE3 male mice following antibiotic treatment.

    Single-nucleus RNA sequencing revealed that specific neuronal populations were reduced and the microglial population increased in the presence of tau pathology in the hippocampi of ConV-R TE3, TE4, and TE mice without ApoE (TEKO) male mice and these effects were reduced with antibiotic treatment. The antibiotic treatment was associated with an increased cecal/body weight ratio and an altered gut microbiome, as analyzed by 16S ribosomal RNA. The effects of the gut microbiome on glial function might involve microbiota-generated short-chain fatty acids (SCFAs) and a peripheral cytokine response. Bacteria generating SCFAs acetate, propionate, and butyrate were reduced by antibiotic treatment in TE3 and TE4 males. In addition, administration of SCFAs in the drinking water to 10- or 31-week-old TE4 GF mice for five or four weeks, respectively, altered alveolar macrophage gene expression and increased gliosis and tau phosphorylation in the hippocampi of the older TE4 GF mice.

    The results of the current study are consistent with the fact that the human gut microbiome diversifies with age, reflects healthy versus unhealthy aging, associates with a healthy lipid profile, predicts survival (Wilmanski et al., 2021), and that alterations in microbiome composition link to Alzheimer’s disease (AD) and impact AD-associated behaviors and brain pathologies (Kundu et al., 2022; Kundu et al., 2021; Marizzoni et al., 2020). A relationship between the gut microbiome and the brain is not limited to AD but is also seen in other neurodegenerative conditions. Taxa that comprise the gut microbiome link to Parkinson’s disease (PD) as well (Fonseca et al., 2019; Elfil et al., 2020; Keshvarzian et al., 2020; Koutzoumis et al., 2020). Consistent with these human data, gut microbiota regulate motor impairments and neuroinflammation in a PD mouse model overexpressing α-synuclein (Sampson et al., 2016) and we showed that in wild-type mice and mice lacking the metabotropic receptor 8 (mGlu8), the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and genotype affected the diversity of the gut microbiome and that there were significant associations between microbiome α-diversity and sensorimotor performance and between microbiome composition and fear learning (Torres et al., 2018). Interestingly, MPTP affected the swim speeds of E4, but not E3, mice in the water maze (Torres et al., 2020), consistent with an ApoE isoform-dependent response using treatments that affect the gut microbiome.

    In the current study, based on the age difference between the donor and recipient mice, and that only one genotype was used as donor, the role of age and/or the genotype of the donor in the effects of the fecal implant are unclear. Our prior work supports the notion that genotype impacts the microbiome’s contribution to cognition: Using 6-month-old knock-in (KI) mice expressing human amyloid precursor protein with dominant mutations (AppNL-F or AppNL-G-F), we showed that the microbiome’s composition as well as its association with mouse cognition differs based on the APP genotype and DNA methylation of the APOE gene in the hippocampus (Kundu et al., 2021). In addition, transplanting microbiomes from AppNL-G-F and AppNL-G-F mice also expressing E4 (AppNL-G-F/E4) into germ-free WT mice revealed donor genotype-dependent differences in recipient mouse behavioral and cognitive performance Insoluble cortical Ab40 levels were detected in AppNL-G-F and AppNL-G-F/E4 recipient mice. Although tau pathology is more associated with cognitive impairments than Aβ pathology, recipients of AppNL-G-F donor mice had cortical insoluble Aβ40 levels that positively correlated with activity levels on the first and second day of open field testing. This relationship was genotype-dependent and not seen in recipients of AppNL-G-F/E4 donor mice. In addition to genotype, the relationship between the gut microbiome and behavioral and cognitive measures are also modulated by sex (Raber et al., 2020). This is consistent with the sex-dependent effects of antibiotic treatments on neuropathology seen in the current study.

    Especially as beneficial interventions in AD will likely need to start relatively early and be safe and affordable, manipulation of the gut microbiome is an attractive therapeutic strategy to consider and future studies are warranted to further assess this strategy. However, as the current and earlier studies indicate, genotype and sex of the recipient, and in case of fecal transplants of the donor as well, need to be carefully considered.

    References:

    Wilmanski T, Diener C, Rappaport N, Patwardhan S, Wiedrick J, Lapidus J, Earls JC, Zimmer A, Glusman G, Robinson M, Yurkovich JT, Kado DM, Cauley JA, Zmuda J, Lane NE, Magis AT, Lovejoy JC, Hood L, Gibbons SM, Orwoll ES, Price ND. Gut microbiome pattern reflects healthy ageing and predicts survival in humans. Nat Metab. 2021 Feb;3(2):274-286. Epub 2021 Feb 18 PubMed.

    Kundu P, Stagaman K, Kasschau K, Holden S, Shulzhenko N, Sharpton TJ, Raber J. Fecal Implants From App NL-G-F and App NL-G-F/E4 Donor Mice Sufficient to Induce Behavioral Phenotypes in Germ-Free Mice. Front Behav Neurosci. 2022;16:791128. Epub 2022 Feb 8 PubMed.

    Kundu P, Torres ER, Stagaman K, Kasschau K, Okhovat M, Holden S, Ward S, Nevonen KA, Davis BA, Saito T, Saido TC, Carbone L, Sharpton TJ, Raber J. Integrated analysis of behavioral, epigenetic, and gut microbiome analyses in AppNL-G-F, AppNL-F, and wild type mice. Sci Rep. 2021 Feb 25;11(1):4678. PubMed.

    Marizzoni M, Cattaneo A, Mirabelli P, Festari C, Lopizzo N, Nicolosi V, Mombelli E, Mazzelli M, Luongo D, Naviglio D, Coppola L, Salvatore M, Frisoni GB. Short-Chain Fatty Acids and Lipopolysaccharide as Mediators Between Gut Dysbiosis and Amyloid Pathology in Alzheimer's Disease. J Alzheimers Dis. 2020;78(2):683-697. PubMed.

    Santos SF, de Oliveira HL, Yamada ES, Neves BC, Pereira A Jr. The Gut and Parkinson's Disease-A Bidirectional Pathway. Front Neurol. 2019;10:574. Epub 2019 Jun 4 PubMed.

    Elfil M, Kamel S, Kandil M, Koo BB, Schaefer SM. Implications of the Gut Microbiome in Parkinson's Disease. Mov Disord. 2020 Jun;35(6):921-933. Epub 2020 Feb 24 PubMed.

    Keshavarzian A, Engen P, Bonvegna S, Cilia R. The gut microbiome in Parkinson's disease: A culprit or a bystander?. Prog Brain Res. 2020;252:357-450. Epub 2020 Mar 5 PubMed.

    Koutzoumis DN, Vergara M, Pino J, Buddendorff J, Khoshbouei H, Mandel RJ, Torres GE. Alterations of the gut microbiota with antibiotics protects dopamine neuron loss and improve motor deficits in a pharmacological rodent model of Parkinson's disease. Exp Neurol. 2020 Mar;325:113159. Epub 2019 Dec 13 PubMed.

    Sampson TR, Debelius JW, Thron T, Janssen S, Shastri GG, Ilhan ZE, Challis C, Schretter CE, Rocha S, Gradinaru V, Chesselet MF, Keshavarzian A, Shannon KM, Krajmalnik-Brown R, Wittung-Stafshede P, Knight R, Mazmanian SK. Gut Microbiota Regulate Motor Deficits and Neuroinflammation in a Model of Parkinson's Disease. Cell. 2016 Dec 1;167(6):1469-1480.e12. PubMed.

    Torres ER, Akinyeke T, Stagaman K, Duvoisin RM, Meshul CK, Sharpton TJ, Raber J. Effects of Sub-Chronic MPTP Exposure on Behavioral and Cognitive Performance and the Microbiome of Wild-Type and mGlu8 Knockout Female and Male Mice. Front Behav Neurosci. 2018;12:140. Epub 2018 Jul 18 PubMed.

    Torres ER, Weber Boutros S, Meshul CK, Raber J. ApoE isoform-specific differences in behavior and cognition associated with subchronic MPTP exposure. Learn Mem. 2020 Sep;27(9):372-379. Print 2020 Sep PubMed.

    Raber J, Fuentes Anaya A, Torres ER, Lee J, Boutros S, Grygoryev D, Hammer A, Kasschau KD, Sharpton TJ, Turker MS, Kronenberg A. Effects of Six Sequential Charged Particle Beams on Behavioral and Cognitive Performance in B6D2F1 Female and Male Mice. Front Physiol. 2020;11:959. Epub 2020 Aug 28 PubMed.

    View all comments by Jacob Raber

  2. Seo et al. add to our understanding of the microbiome’s impact on murine AD models by showing that neurodegeneration is reduced in gnotobiotic P301S mice. Several lines of evidence suggest microglia are key to this neuroprotection because (i) neurodegeneration is reduced in P301S mice lacking TREM2, a microglial activator (Leyns et al., 2017), (ii) microglial function is reduced with gnotobiotic conditions, an effect reversed by short-chain fatty acid (SCFA) supplementation (Erny et al., 2015), and (iii) Seo et al. report that SCFA supplementation reversed the gnotobiotic neuroprotection in the P301S model.

    Germ-free conditions have also been reported to decrease amyloid burden in APPPS1 mice (Harach et al., 2017). Although this effect was also reversed by SCFA treatment (Colombo et al., 2021), the mechanism for gnotobiotic reduction of amyloid burden is less clear. In particular, Harach et al. noted that germ-free conditions resulted in an increase in amyloid clearing enzymes, and that this increase was blunted by SCFA. Hence, SCFA may act in both microglial-dependent and -independent pathways to alter disease pathology. The effects of SCFA supplementation appear most robust in gnotobiotic conditions, noting that inconsistent results have been obtained in SCFA-treated APP mice with a conventional microbiome (Colombo et al., 2021; Fernando et al., 2020; Zajac et al., 2022).

    References:

    Leyns CE, Ulrich JD, Finn MB, Stewart FR, Koscal LJ, Remolina Serrano J, Robinson GO, Anderson E, Colonna M, Holtzman DM. TREM2 deficiency attenuates neuroinflammation and protects against neurodegeneration in a mouse model of tauopathy. Proc Natl Acad Sci U S A. 2017 Oct 24;114(43):11524-11529. Epub 2017 Oct 9 PubMed.

    Erny D, Hrabě de Angelis AL, Jaitin D, Wieghofer P, Staszewski O, David E, Keren-Shaul H, Mahlakoiv T, Jakobshagen K, Buch T, Schwierzeck V, Utermöhlen O, Chun E, Garrett WS, McCoy KD, Diefenbach A, Staeheli P, Stecher B, Amit I, Prinz M. Host microbiota constantly control maturation and function of microglia in the CNS. Nat Neurosci. 2015 Jun 1; PubMed.

    Harach T, Marungruang N, Duthilleul N, Cheatham V, Mc Coy KD, Frisoni G, Neher JJ, Fåk F, Jucker M, Lasser T, Bolmont T. Reduction of Abeta amyloid pathology in APPPS1 transgenic mice in the absence of gut microbiota. Sci Rep. 2017 Feb 8;7:41802. PubMed.

    Colombo AV, Sadler RK, Llovera G, Singh V, Roth S, Heindl S, Sebastian Monasor L, Verhoeven A, Peters F, Parhizkar S, Kamp F, Gomez de Aguero M, MacPherson AJ, Winkler E, Herms J, Benakis C, Dichgans M, Steiner H, Giera M, Haass C, Tahirovic S, Liesz A. Microbiota-derived short chain fatty acids modulate microglia and promote Aβ plaque deposition. Elife. 2021 Apr 13;10 PubMed.

    Fernando WM, Martins IJ, Morici M, Bharadwaj P, Rainey-Smith SR, Lim WL, Martins RN. Sodium Butyrate Reduces Brain Amyloid-β Levels and Improves Cognitive Memory Performance in an Alzheimer's Disease Transgenic Mouse Model at an Early Disease Stage. J Alzheimers Dis. 2020;74(1):91-99. PubMed.

    Zajac DJ, Shaw BC, Braun DJ, Green SJ, Morganti JM, Estus S. Exogenous Short Chain Fatty Acid Effects in APP/PS1 Mice. Front Neurosci. 2022;16:873549. Epub 2022 Jul 4 PubMed.

    View all comments by Diana Zajac

  3. In this paper Dong-oh Seo and colleagues have investigated the impact of ApoE isoforms on tau-mediated neurodegeneration. The authors show that mice raised in germ-free conditions, or treated with antibiotics, are protected against tau pathology and neurodegeneration, and that these benefits were more pronounced in ApoE-3-expressing male mice. Intriguingly, short-chain fatty acids produced by the gut microbes may have played a role in promoting this harmful phenotype.

    This is a very exciting and thought-provoking work from the Holtzman group which adds further support to the role of the APOE genotype on gut-brain axis modulation, and suggests that the gut microbiome is worth further investigation as a potential target to mitigate the deleterious impact of the APOE genotype on cognitive decline and the prevention of AD.

    View all comments by David Vauzour

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Mutations

  1. APOE C130R (ApoE4)