Astrocytic interleukin-3 programs microglia and limits Alzheimer's disease

doi:10.1038/s41586-021-03734-6

. 2021 Jul;595(7869):701-706.

doi: 10.1038/s41586-021-03734-6. Epub 2021 Jul 14.

Astrocytic interleukin-3 programs microglia and limits Alzheimer's disease

Cameron S McAlpine^{1

2

3}, Joseph Park⁴, Ana Griciuc⁴, Eunhee Kim⁴, Se Hoon Choi⁴, Yoshiko Iwamoto¹, Máté G Kiss¹, Kathleen A Christie⁵, Claudio Vinegoni¹, Wolfram C Poller^{1

2}, John E Mindur¹, Christopher T Chan¹, Shun He¹, Henrike Janssen¹, Lai Ping Wong^{6

7}, Jeffrey Downey¹, Sumnima Singh¹, Atsushi Anzai¹, Florian Kahles¹, Mehdi Jorfi⁴, Paolo Fumene Feruglio⁸, Ruslan I Sadreyev^{6

9}, Ralph Weissleder¹, Benjamin P Kleinstiver⁵, Matthias Nahrendorf¹, Rudolph E Tanzi¹⁰, Filip K Swirski^{11

12}

Affiliations

¹ Center for Systems Biology and Department of Radiology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA.
² Cardiovascular Research Institute and Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
³ Friedman Brain Institute and Nash Family Department of Neuroscience, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
⁴ Genetics and Aging Research Unit, McCance Center for Brain Health, Mass General Institute for Neurodegenerative Disease, Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA, USA.
⁵ Center for Genomic Medicine, Department of Pathology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA.
⁶ Department of Molecular Biology, Massachusetts General Hospital, Boston, MA, USA.
⁷ Department of Genetics, Harvard Medical School, Boston, MA, USA.
⁸ Department of Neuroscience, Biomedicine and Movement Sciences, University of Verona, Verona, Italy.
⁹ Department of Pathology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA.
¹⁰ Genetics and Aging Research Unit, McCance Center for Brain Health, Mass General Institute for Neurodegenerative Disease, Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA, USA. rtanzi@mgh.harvard.edu.
¹¹ Center for Systems Biology and Department of Radiology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA. filip.swirski@mssm.edu.
¹² Cardiovascular Research Institute and Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY, USA. filip.swirski@mssm.edu.

PMID: 34262178
PMCID: PMC8934148
DOI: 10.1038/s41586-021-03734-6

Astrocytic interleukin-3 programs microglia and limits Alzheimer's disease

Cameron S McAlpine et al. Nature. 2021 Jul.

. 2021 Jul;595(7869):701-706.

doi: 10.1038/s41586-021-03734-6. Epub 2021 Jul 14.

Authors

Affiliations

¹ Center for Systems Biology and Department of Radiology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA.
² Cardiovascular Research Institute and Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
³ Friedman Brain Institute and Nash Family Department of Neuroscience, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
⁴ Genetics and Aging Research Unit, McCance Center for Brain Health, Mass General Institute for Neurodegenerative Disease, Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA, USA.
⁵ Center for Genomic Medicine, Department of Pathology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA.
⁶ Department of Molecular Biology, Massachusetts General Hospital, Boston, MA, USA.
⁷ Department of Genetics, Harvard Medical School, Boston, MA, USA.
⁸ Department of Neuroscience, Biomedicine and Movement Sciences, University of Verona, Verona, Italy.
⁹ Department of Pathology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA.
¹⁰ Genetics and Aging Research Unit, McCance Center for Brain Health, Mass General Institute for Neurodegenerative Disease, Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA, USA. rtanzi@mgh.harvard.edu.
¹¹ Center for Systems Biology and Department of Radiology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA. filip.swirski@mssm.edu.
¹² Cardiovascular Research Institute and Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY, USA. filip.swirski@mssm.edu.

PMID: 34262178
PMCID: PMC8934148
DOI: 10.1038/s41586-021-03734-6

Abstract

Communication within the glial cell ecosystem is essential for neuronal and brain health^1-3. The influence of glial cells on the accumulation and clearance of β-amyloid (Aβ) and neurofibrillary tau in the brains of individuals with Alzheimer's disease (AD) is poorly understood, despite growing awareness that these are therapeutically important interactions^4,5. Here we show, in humans and mice, that astrocyte-sourced interleukin-3 (IL-3) programs microglia to ameliorate the pathology of AD. Upon recognition of Aβ deposits, microglia increase their expression of IL-3Rα-the specific receptor for IL-3 (also known as CD123)-making them responsive to IL-3. Astrocytes constitutively produce IL-3, which elicits transcriptional, morphological, and functional programming of microglia to endow them with an acute immune response program, enhanced motility, and the capacity to cluster and clear aggregates of Aβ and tau. These changes restrict AD pathology and cognitive decline. Our findings identify IL-3 as a key mediator of astrocyte-microglia cross-talk and a node for therapeutic intervention in AD.

PubMed Disclaimer

Conflict of interest statement

Competing interests

C.S.M., F.K.S., and R.E.T. are inventors on a patent application filed by Mass General Brigham that describes targeting IL-3 signaling in Alzheimer’s disease (invention record no. 2020-568). B.P.K. is an inventor on patent applications filed by Mass General Brigham that describe genome engineering technologies and methods, is an advisor to Acrigen Biosciences, and consults for Avectas Inc. and ElevateBio.

Figures

**Extended Data Fig. 1.. Analysis of *Il3^−/−* mice.**
a. FITC-dextran (mol. wt. 4000) was i.v. injected into WT and *Il3^−/−* mice prior to sacrifice. Blood brain barrier integrity was determined by measuring FITC signal in brain homogenate (n=4-6). b. Prior to sacrifice WT and *Il3^−/−* mice were i.v. injected with an anti-GR1 antibody conjugated to PE to label all circulating monocytes and neutrophils. PE signal among CD45⁺ cells was assessed in the brain by flow cytometry (n=4-7). c. Doublecortin staining and quantification in the hippocampus of WT and *Il3^−/−* mice at 4 months of age (n=3). d. Absence of Caspase 3 staining in the hippocampus of WT and *Il3^−/−* mice along with a representative image of rare positively stained cells from the thalamus (n=3). e. Assessment of MHCII⁺ microglia in the brain of WT and *Il3^−/−* mice (n=4-7). f. Analysis of Ki67⁺ proliferating microglia (n=5). g. Time in new arm during Y-maze testing (n=6-7). Groups of mice are of evenly mixed sex. Error bars indicate mean ± SEM.

**Extended Data Fig. 2.. Hematopoiesis and peripheral immune cell dynamics in WT, *5xFAD*, and *Il3^−/−5XFAD* mice.**
a. Average swim speed during acquisition days of Morris water maze (n=9-10). b. Flow cytometry assessment of blood leukocytes. c. Flow cytometry analysis of Lin⁻Sca1⁺cKit⁺ (LSKs), multi-potent progenitors (MPP)-4 and -3, short-term hematopoietic stem cells (StHSCs), long-term HSCs (LtHSCs), common myeloid progenitors (CMPs), granulocyte macrophage progenitors (GMPs), and monocyte dendritic progenitors (MDPs) in the bone marrow of 5-month-old WT, *5xFAD*, and *Il3^−/−5xFAD* mice (n=7-12). d. FITC-dextran (mol. wt. 4000) was i.v. injected into *5xFAD* and *Il3^−/−5xFAD* mice prior to sacrifice. Blood brain barrier integrity was determined by measuring FITC signal in brain homogenates (n=4). e. WT, *5xFAD, Il3^−/−5xFAD* mice were joined by parabiosis with UbiGFP mice from the age of 2 to 6 months (4 months total). f. GFP chimerism in blood Ly6C^hi monocytes and brain CD45⁺ cells was assessed by flow cytometry in the WT, *5xFAD*, and *Il3^−/−5xFAD* parabiont (n=4-5). Groups of mice are of evenly mixed sex. *p<0.05, **p<0.01, ***p<0.001. Error bars indicate mean ± SEM.

**Extended Data Fig 3.. Generation and validation of *Il3^GFPfl/flAldh1/1Cre^ERT25XFAD* and *Il3rɑ^fl/flCx3cr1Cre^ERT25XFAD* mice.**
Schematics of the endogenous loci and editing strategies to generate conditional/reporter models for *Il3* (a) and *Il3rɑ* (b). Mice were generated by excising large fragments of the endogenous loci by co-delivery of SpCas9 and two gRNAs, and in the presence of a long single stranded DNA donor encoding a loxP-cDNA-P2A-EGFP-loxP cassette. Representative Sanger sequencing traces validating insertion of the loxP-cDNA-P2A-EGFP-loxP cassettes at the endogenous loci of *Il3* (c) and *Il3rɑ* (d). Missense mutation quench the GFP signal in *Il3rɑ* targeted mice. e. GFP signal in *ex-vivo* stimulated splenic and lymph node T-cells, known IL-3 sources, from WT and *Il3^GFPfl/fl* mice. f. Flow cytometry analysis of astrocyte IL-3 production in 5-month-old *Il3^GFPfl/fl5xFAD* and *Il3^GFPfl/flAldh1/1Cre*^ERT25xFAD mice injected with tamoxifen. g. qPCR analysis of *Il3* mRNA expression in sorted astrocytes. h. CSF IL-3 levels. Filled circles represent male mice, open circles represent female mice. i. STAT5 phosphorylation in *ex-vivo* heart macrophages from *Il3rɑ*^fl/fl mice simulated with recombinant IL-3. j. Flow cytometry analysis of microglia IL-3Rα production in 5-month-old *Il3rɑ*^fl/fl5xFAD and *Il3rɑ*^fl/flCx3cr1Cre^ERT25xFAD mice injected with tamoxifen. k. qPCR analysis of *Il3rɑ* mRNA expression in sorted microglia. All *Il3^GFPfl/fl5xFAD, *Il3*^GFPfl/flAldh1/1Cre*^ERT25xFAD, Il3ra^fl/fl5xFAD, and *Il3rɑ*^fl/flCx3cr1Cre^ERT25xFAD mice were injected with tamoxifen beginning at 2 months of age. **p<0.01, ***p<0.001. Error bars indicate mean ± SEM.

**Extended data Fig. 4.. Flow cytometry gating strategy and histology controls.**
a. Gating strategy used to identify cell populations in the brain of all mice except *Aldh1/1*^GFP mice. b. Backgating of GFP⁺ astrocytes in *Il3^GFPfl/fl* mice. c. Gating Strategy Used to identify cell populations in the brain of *Aldh1/1*^GFP mice. d. Gating and isotype control plots for microglia IL-3Rɑ staining. e. Representative images of IgG control antibody staining and IL-3 staining in the brain.

**Extended Data Fig. 5.. Astrocyte IL-3 and microglia IL-3Rɑ production.**
a. *Il3* expression in astrocytes sorted from WT and *5xFAD* mice at 5 and 12 months of age. b. Tissue IL-3 levels in various brain regions in WT mice at 4 months of age (n=4). c. Scheme of daily i.p. LPS injection into WT mice over 4 days and qPCR analysis of *complement C3* and *gfap* in sorted astrocytes after LPS injection (n=6). d. Cerebrospinal fluid IL-3 levels (n=6-8). e. Representative images of GFAP⁺ astrocytes in the cortex of WT, *Il3^−/−*, *5xFAD*, and *Il3^−/−5xFAD* mice. f. Representative images of Aβ deposits (6E10) and astrocytes (GFAP) in *5xFAD* and *Il3^−/−5XFAD* mice. g. Proportion of IL-3Rɑ⁺ microglia in the brain of WT mice at various ages (n=4). h. *Il3rα* transcript expression in brain homogenate of WT mice at various ages (n=4). i. Proportion of IL-3Rɑ⁺ macrophages in heart, liver, lung (interstitial and alveolar), and brain of WT and *5xFAD* mice at 8 months of age (n=4). *p<0.05, **p<0.001. Error bars indicate mean ± SEM.

**Extended Data Fig. 6.. *Il3rɑ* expression in homeostatic microglia, TREM2-independent type 1 DAMs, and TREM2-dependent type 2 DAMs.**
Analysis of *Il3rɑ* and other cytokine receptors expressed in resting microglia, DAM stage 1, and DAM stage 2 microglia. Data are published in Keren-Shaul *et al. Cell*, 2017.

**Extended Data Fig 7.. Characterization of IL-3Rɑ^hi and IL-3Rɑ^lo microglia.**
a. Gating strategy for IL-3Rɑ^hi and IL-3Rɑ^lo microglia in *5xFAD* mice. b. Flow cytometry analysis of IL-3Rɑ^hi and IL-3Rɑ^lo microglia (n=3-6). c. mRNA transcript expression in sorted IL-3Rɑ^hi and IL-3Rɑ^lo microglia (n=5-7). *p<0.05, **p<0.01. Error bars indicate mean ± SEM.

**Extended Data Fig. 8.. IL-3 does not influence microglia proliferation, the recognition and phagocytosis of Aβ, or the production of inflammatory cytokines.**
a. BrdU incorporation into microglia (n=6-7). b. Heatmap of microglia RNAseq expression data of genes important to cell cycle and proliferation. c. Assessment of *ex vivo* phagocytosis of Aβ42 conjugated to a pH-sensitive dye (PHrodo Red) in sorted microglia lacking *Il3* or stimulated with recombinant IL-3 (n=4). d. Heatmap of microglia RNAseq expression data of genes important to Aβ recognition and phagocytosis. e. Heatmap of microglia RNAseq expression data of inflammatory cytokines. Groups of mice are of evenly mixed sex. Error bars indicate mean ± SEM. ns = not significant

**Extended Data Fig. 9.. Human AD iPS triculture system.**
a. *Il3rɑ* expression in human iPS microglia exposed to Aβ (n=2). b. chemokine and cytokine levels in the media of the human AD iPS triculture system. (n=2-3). Error bars indicate mean ± SEM.

**Extended Data Fig. 10.. rIL-3 delivery to the cortex or periphery of *5xFAD* mice and summary figure.**
a. Recombinant IL-3 or PBS was delivered into the cortex of *5xFAD* mice. Three days later microglia localization to Aβ aggregates was assessed (n=6-7). b. Scheme of recombinant IL3 delivery intraperitoneally twice a week for 10 weeks to *5xFAD* mice. c. Prior to sacrifice Ymaze behavioral testing was performed and time in the new arm was quantified (n=7-8). d. The amount of Aβ in the cortex of mice was quantified by analyzing histological sections (n=6). Groups of mice are of evenly mixed sex. **p<0.01. Error bars indicate mean ± SEM. e. Model of IL-3’s role in AD. Astrocytes produce IL-3. In response to Aβ, TREM2 signaling increases microglia IL-3Rɑ, rendering microglia responsive to astrocyte-derived IL-3. IL-3 signaling instigates microglia transcriptional and functional reprogramming leading to a signature of immune regulation, motility, and migration. IL-3-dependent reprogramming promotes clustering of microglia around Aβ enabling Aβ clearance and mitigation of AD pathology.

**Fig. 1.. IL-3 protects against β-amyloid accumulation and cognitive impairment in *5xFAD* mice.**
a. Representative immunofluorescence images of β-amyloid (Aβ) plaques (6E10) in 5-month-old *5xFAD* and *Il3^−/−5XFAD* mice. b. Quantification of Aβ area and Aβ plaque size in the cortex (n=5 *5xFAD* mice; n=7 *Il3^−/−5xFAD* mice). c. Levels of tris-buffered saline (TBS)- and formic acid (FA)-soluble Aβ40 and Aβ42 in cortex homogenates of *5XFAD* and *Il3^−/−5XFAD* mice (n=6-7 *5xFAD* mice; n=7–9 *Il3^−/−5xFAD* mice). d. Quantification of time in new arm and entries made into the new arm during Y-maze testing (n=8 *5xFAD* mice; n= 12 *Il3^−/−5xFAD* mice). Morris water maze testing. e. Time for male mice needed to reach hidden platform plotted across training days (two-way ANOVA for groups p=0.0074) and area under the curve (AUC) of escape latency (n=10 *5xFAD* mice; n=9 *Il3^−/−5xFAD* mice). f. Time in target zone on probe day (8th day) (n=10 *5xFAD* mice; n=9 *Il3^−/−5xFAD* mice). Filled circles represent male mice, open circles represent female mice. *p<0.05, **p<0.01, ***p<0.001. Error bars indicate mean ± SEM.

**Fig. 2.. Microglia become responsive to astrocyte-derived IL-3 in Alzheimer’s disease.**
a. IL-3 levels in the plasma (PL) and cerebrospinal fluid (CSF) of WT and *5xFAD* mice at 5 and 12 months (m.) of age (n=4 WT and *5xFAD* 5m.plasma; n=14 WT 5m. CSF; n=6 *5xFAD* 5m. CSF; n=5 WT 12m. CSF; n=4 *5xFAD* 12m. CSF). b. Flow cytometry analysis of the brain of WT, *Il3^GFPfl/fl*, and *Il3^GFPfl/fl5xFAD* reporter mice. c. Quantification of IL3⁺ (GFP⁺) astrocytes in the brain of *Il3^GFPfl/fl* and *l3^GFPfl/fl5xFAD* mice (n=4 mice per group). d. *Il3* mRNA transcript expression in Aldh1l1-GFP⁺ astrocytes, CD11b⁺CD45^mid microglia, and CD45⁻ cells sorted from *Aldh1/1*^GFP mice (n=7 mice per group). e. Representative immunofluorescence images showing co-localization of IL-3-GFP⁺ with GFAP⁺ astrocytes in *Il3^GFPfl/fl* mice. f. Quantification of IL3⁺ astrocytes (Aldh1/1-GFP⁺) in various brain regions and representative immunofluorescence images from numerous regions showing co-localization of IL-3 with GFP⁺ astrocytes in *Aldh1/1*^GFP mice (n=4 per group). g. Flow cytometry and **(h)** analysis of IL-3Rɑ⁺ microglia in the brain of WT and *5xFAD* mice (n=5 5m. WT mice; n=11 5m. *5XFAD* mice; n=3 8m. WT mice; n=5 8m. *5XFAD* mice). i. *Il3ra* mRNA expression in sorted microglia (n=5 5m. WT mice; n=4 5m. and 8m. *5xFAD* mice; n=3 8m. WT mice). j. STAT5 phosphorylation in *ex-vivo* microglia simulated with recombinant IL-3. k. Representative immunofluorescence images showing co-localization of IL-3Rɑ with Iba1 in the cortex of *5xFAD* mice. l. *Il3rɑ* transcript expression in sorted microglia from WT, *Trem2^−/−, 5xFAD*, and *Trem2^−/−5xFAD* mice displayed as Log₂ fold change (FC) for indicated comparisons (for 4 month old mice p=3.45e⁻²⁷ for *5xFAD* vs WT, p=3.4e⁻⁰⁶ for *Trem2^−/−5xFAD* vs WT, p=1.82e⁻⁰⁸ for *Trem2^−/−5xFAD* vs *5xFAD*; for 8 month old mice p=1.12e⁻⁹³ for *5xFAD* vs WT, p=1.4e⁻¹³ for *Trem2^−/−5xFAD* vs WT, p=1.6e⁻⁴⁰ for *Trem2^−/−5xFAD* vs *5xFAD*; FDR<0.0002; for WT n=13M/14F at 4 m., n=8M/8F at 8m.; for *Trem2^−/−* n=11M/12F at 4m., 2M/2F at 8m.; for *5xFAD* n=14M/14F at 4 m., 10M/9F at 8m.; and for *Trem2^−/−5xFAD* n=11M/11F at 4m., 9M/8F at 8m.). ns=not significant. m. Flow cytometry analysis of IL3Rɑ⁺ on microglia from *5xFAD* and *Trem2^−/−5xFAD* mice (n=4 mice per group). Groups are of evenly mixed sex. *p<0.05, **p<0.01, ***p<0.001. Error bars indicate mean ± SEM.

**Fig. 3.. IL-3 signaling correlates with disease pathology in the brain of humans with Alzheimer’s disease.**
a. Representative immunofluorescence images of the cortex of control subjects (C) or Alzheimer’s disease (AD) patients stained for IL-3 and GFAR b. Measurement of IL-3 levels in cortex tissue homogenates from control and AD patients (n=15 controls, n=23 AD patients, see Extended Data Table S6 for characteristics). c. Representative immunofluorescence images of the cortex of humans with or without Alzheimer’s disease (AD) stained for IL-3Rɑ and Iba1. d. qPCR analysis of fold change in *IL3Rɑ* expression in the frontal cortex of control non-demented individuals and AD patients (n=28 controls, n=30 AD patients, see Extended Data Table S7 for characteristics). e. Assessment of brain *IL3Rɑ* expression with *APOE* genotype (n=30 AD patients). f. Correlation of mean *IL3Rɑ* expression with years (yrs) of disease duration (n=30 AD patients). Correlation of *IL3Rɑ* expression with formic acid (FA)-soluble Aβ40 (g) and Aβ42 (h) in the frontal cortex of AD patients (n=23 AD patients). *p<0.05, **p<0.01, ***p<0.001. Open circles represent control subjects and red circles represent AD patients. Error bars indicate mean ± SEM.

**Fig. 4.. IL-3 reprograms microglia promoting motility and clustering of β-amyloid in the murine brain and in a 3D human AD iPS triculture system.**
a. Enumeration, by flow cytometry, of microglia in WT, *5XFAD, Il3^−/−5XFAD* mice (n=10 *5xFAD* mice; n=8 *Il3^−/−5XFAD* mice). b. Multidimentional scaling (MDS) plot of RNAseq data from 5 month old *5XFAD* and *Il3^−/−5XFAD* mice (n=12, each data point represents 4 pooled mice of 2M/2F). c. Volcano plot indicating differentially regulated genes (FC>1.6, FDR<0.1, p<0.005) in microglia of *5XFAD* and *II3^−/−5XFAD* mice. d. Heatmap of key differentially regulated genes (FC>1.6, FDR<0.1, p<0.005), except *Il3rɑ* which is not significantly different between *5xFAD* and *Il3^−/− 5xFAD* microglia (n=12, each data point represents 4 pooled mice of 2M/2F). e. Pathway analysis of significantly regulated genes. f. Representative immunofluorescence images of microglia from *5XFAD* and *Il3^−/−5XFAD* mice along with skeletal analysis of microglia morphology (n=5 *5xFAD* mice; n=4 *Il3^−/−5XFAD* mice). g. Representative immunofluorescence images of cortex sections from *5XFAD* and *Il3^−/−5XFAD* mice stained for Aβ (6E10), Iba1 and DAPI and quantification of the number of microglia within 25μm of Aβ plaques (n=5 *5xFAD* mice; n=7 *Il3^−/−5XFAD* mice). h. Schematic depicting segmentation and spatial analysis along with computed density gradient and diffusion rate of microglia surrounding Aβ (n=5 *5xFAD* mice; n=7 *Il3^−/−5XFAD* mice). i. Three dimensional confocal images of mouse cortex (634x250x634μm) stained for Aβ (6E10) and microglia (Iba1). All mouse data are of groups of 5 month old animals of evenly mixed sex. j. Scheme of 3D human AD triculture microfluidic system where human neuron ReNcell VM progenitor derived astrocytes and neuronal cells, with or without the K670N/M671L (Swedish) and V717I (London) familial AD mutations resulting overproduction of Aβ and neurofibrillary tangle (NFT) p-tau (AD), are plated in the central chamber while human iPS-derived microglia labeled with CellTracker are plated in side chambers. k. Confocal images demonstrated p-tau (PHF1) localization with neurons (Tuj1) and the presence of astrocytes (GFAP) in the central chamber and microglia (P2RY12) in the side chamber. l. ELISA quantification of Aβ40, Aβ38, and Aβ42 in media of central chamber in 3D microfluidic system plated with control or AD cells (n=3 per group). m. Confocal imaging of IL-3 localization to GFAP⁺ human astrocytes and IL-3Rɑ localization to CellTracker labeled human iPS-derived microglia in 3D microfluidics system. n. Flow cytometry quantification of iPS-derived microglia that have migrated to the central chamber of the 3D microfluidics system with or without addition of human recombinant IL-3 (n=3 per group). o. Representative confocal images and image quantification of iPS microglia migration to central chamber (n=3 per group for astrocytes+neurons and AD astrocytes+neurons; n=4 AD astrocytes+neurons+rlL3). p. CCL2 and CCL4 levels in media of 3D microfluidics system (n=3 per group). *p<0.05, **p<0.01, ***p<0.001. Error bars indicate mean ± SEM.

**Fig. 5.. Astrocyte IL-3 or microglia IL-3Rɑ deletion instigate while IL-3 infusion resolves β-amyloid burden and cognitive decline.**
a. Representative immunofluorescent images and Aβ quantification of brain sections probed for Aβ (6E10) and DAPI of 5-month-old *Il3GFP^fl/fl5xFAD* and *Il3^GFPfl/flAldh1/1Cre*^Ert25xFAD mice injected with tamoxifen (n=6 *Il3^GFPfl/fl5xFAD* mice; n=9 *Il3^GFPfl/flAldh1/1Cre*^Ert25xFAD mice). b. Levels of TBS- and FA-soluble Aβ40 and Aβ42 in cortex homogenates (n=6 *Il3^GFPfl/fl5xFAD* mice; n=9 *Il3^GFPfl/flAldh1/1Cre*^Ert25xFAD mice). c. Gene expression in sorted microglia (n=6 *Il3^GFPfl/fl5xFAD* mice; n=9 *Il3^GFPfl/flAldh1/1Cre*^Ert25xFAD mice). d. Representative immunofluorescent images and quantification of microglia-Aβ co-localization (n=6 *Il3^GFPfl/fl5xFAD* mice; n=9 *Il3^GFPfl/flAldh1/1Cre*^Ert25xFAD mice). e.Y-maze testing time in new arm (n=6 *Il3^GFPfl/fl5xFAD* mice; n=9 *Il3^GFPfl/flAldh1/1Cre*^Ert25xFAD mice). f. Representative immunofluorescent images and Aβ quantification of brain sections probed for Aβ (6E10) and DAPI of 5-month-old *Il3rɑ*^fl/fl5xFAD and *Il3rɑ*^fl/flCx3cr1Cre^Ert25xFAD mice injected with tamoxifen. (n=7 per group). g. Levels of TBS- and FA-soluble Aβ40 and Aβ42 in cortex homogenates (n=7 per group). h. Representative immunofluorescent images and quantification of microglia-Aβ co-localization (n= per group). i. Y-maze testing time in new arm (n=8 *Il3rɑ*^fl/fl5xFAD mice; n=6 *Il3rɑ^fl/flCx3cr1Cre^Ert25xFAD*). j. Experimental approach and representative immunofluorescent images of brain sections probed for Aβ (6E10) and DAPI from mice receiving brain infusion of PBS or recombinant IL-3. k. Quantification of cortex Aβ (n=6 *Il3^−/−5xFAD*+PBS; n=5 *Il3^−/−5xFAD+rIL*-3). l. Levels of TBS- and FA-soluble Aβ40 and Aβ42 in cortex homogenates (n=7 per group). m. Representative Immunofluorescence images and co-localization quantification of microglia within 25μm of Aβ in the cortex of mice receiving PBS or rIL-3 (n=6 *Il3^−/−5xFAD*+PBS; n=5 *Il3^−/−5xFAD*+rIL-3). n. Y-maze testing time in new arm (n=13 *Il3^−/−5xFAD*+PBS; n=10 *Il3^−/−5xFAD*+rIL-3). Filled circles represent male mice, open circles represent female mice. *p<0.05, **p<0.01, ***p<0.001. Error bars indicate mean ± SEM.

See this image and copyright information in PMC

Comment in

Astrocytic IL-3 could help microglia protect against Alzheimer disease.
Wood H. Wood H. Nat Rev Neurol. 2021 Sep;17(9):525. doi: 10.1038/s41582-021-00546-0. Nat Rev Neurol. 2021. PMID: 34321636 No abstract available.

Cited by

VEGFD/VEGFR3 signaling contributes to the dysfunction of the astrocyte IL-3/microglia IL-3Rα cross-talk and drives neuroinflammation in mouse ischemic stroke.
Wang S, Guo Y, Cao RQ, Zhu YM, Qiao SG, Du HP, Liu Y, Xu Y, Zhou XY, Sun L, Lu QX, Schoen I, Zhang HL. Wang S, et al. Acta Pharmacol Sin. 2025 Feb;46(2):292-307. doi: 10.1038/s41401-024-01405-6. Epub 2024 Oct 30. Acta Pharmacol Sin. 2025. PMID: 39478160
Saponin components in Polygala tenuifolia as potential candidate drugs for treating dementia.
Li S, Hou Z, Ye T, Song X, Hu X, Chen J. Li S, et al. Front Pharmacol. 2024 Jul 10;15:1431894. doi: 10.3389/fphar.2024.1431894. eCollection 2024. Front Pharmacol. 2024. PMID: 39050746 Free PMC article. Review.
Gut Microbiota Mediates Neuroinflammation in Alzheimer's Disease: Unraveling Key Factors and Mechanistic Insights.
Junyi L, Yueyang W, Bin L, Xiaohong D, Wenhui C, Ning Z, Hong Z. Junyi L, et al. Mol Neurobiol. 2025 Mar;62(3):3746-3763. doi: 10.1007/s12035-024-04513-w. Epub 2024 Sep 25. Mol Neurobiol. 2025. PMID: 39317889 Review.
Novel Development and Prospects in Pathogenesis, Diagnosis, and Therapy of Alzheimer's Disease.
Teng Z. Teng Z. J Alzheimers Dis Rep. 2024 Feb 20;8(1):345-354. doi: 10.3233/ADR-230130. eCollection 2024. J Alzheimers Dis Rep. 2024. PMID: 38405339 Free PMC article. Review.
Targeting tau in Alzheimer's disease: from mechanisms to clinical therapy.
Ye J, Wan H, Chen S, Liu GP. Ye J, et al. Neural Regen Res. 2024 Jul 1;19(7):1489-1498. doi: 10.4103/1673-5374.385847. Epub 2023 Sep 22. Neural Regen Res. 2024. PMID: 38051891 Free PMC article.

See all "Cited by" articles

References

1. Linnerbauer M, Wheeler MA & Quintana FJ Astrocyte Crosstalk in CNS Inflammation. Neuron 1–15 (2020) doi:10.1016/j.neuron.2020.08.012. - DOI - PMC - PubMed
1. Vainchtein ID & Molofsky AV Astrocytes and Microglia: In Sickness and in Health. Trends Neurosci. 43, 144–154 (2020). - PMC - PubMed
1. Castellani G & Schwartz M Immunological Features of Non-neuronal Brain Cells: Implications for Alzheimer’s Disease Immunotherapy. Trends in Immunology (2020) doi:10.1016/j.it.2020.07.005. - DOI - PubMed
1. Fakhoury M Microglia and astrocytes in Alzheimer’s disease: implications for therapy. Curr. Neuropharmacol (2017) doi:10.2174/1570159x15666170720095240. - DOI - PMC - PubMed
1. Long JM & Holtzman DM Alzheimer Disease: An Update on Pathobiology and Treatment Strategies. Cell (2019) doi:10.1016/j.cell.2019.09.001. - DOI - PMC - PubMed

Publication types

Actions
Actions

MeSH terms

Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions

Substances

Actions
Actions

Grants and funding

LinkOut - more resources

Full Text Sources
Other Literature Sources
- H1 Connect
- The Lens - Patent Citations Database
Medical
- MedlinePlus Health Information
Molecular Biology Databases
- Mouse Genome Informatics (MGI)
- NIAID Data Ecosystem - Find datasets on Infectious and Immune-mediated Diseases
Research Materials
- NINDS Human Cell and Data Repository

[1] Linnerbauer M, Wheeler MA & Quintana FJ Astrocyte Crosstalk in CNS Inflammation. Neuron 1–15 (2020) doi:10.1016/j.neuron.2020.08.012. - DOI - PMC - PubMed

[2] Linnerbauer M, Wheeler MA & Quintana FJ Astrocyte Crosstalk in CNS Inflammation. Neuron 1–15 (2020) doi:10.1016/j.neuron.2020.08.012. - DOI - PMC - PubMed

[3] Vainchtein ID & Molofsky AV Astrocytes and Microglia: In Sickness and in Health. Trends Neurosci. 43, 144–154 (2020). - PMC - PubMed

[4] Vainchtein ID & Molofsky AV Astrocytes and Microglia: In Sickness and in Health. Trends Neurosci. 43, 144–154 (2020). - PMC - PubMed

[5] Castellani G & Schwartz M Immunological Features of Non-neuronal Brain Cells: Implications for Alzheimer’s Disease Immunotherapy. Trends in Immunology (2020) doi:10.1016/j.it.2020.07.005. - DOI - PubMed

[6] Castellani G & Schwartz M Immunological Features of Non-neuronal Brain Cells: Implications for Alzheimer’s Disease Immunotherapy. Trends in Immunology (2020) doi:10.1016/j.it.2020.07.005. - DOI - PubMed

[7] Fakhoury M Microglia and astrocytes in Alzheimer’s disease: implications for therapy. Curr. Neuropharmacol (2017) doi:10.2174/1570159x15666170720095240. - DOI - PMC - PubMed

[8] Fakhoury M Microglia and astrocytes in Alzheimer’s disease: implications for therapy. Curr. Neuropharmacol (2017) doi:10.2174/1570159x15666170720095240. - DOI - PMC - PubMed

[9] Long JM & Holtzman DM Alzheimer Disease: An Update on Pathobiology and Treatment Strategies. Cell (2019) doi:10.1016/j.cell.2019.09.001. - DOI - PMC - PubMed

[10] Long JM & Holtzman DM Alzheimer Disease: An Update on Pathobiology and Treatment Strategies. Cell (2019) doi:10.1016/j.cell.2019.09.001. - DOI - PMC - PubMed

Save citation to file

Email citation

Add to Collections

Add to My Bibliography

Your saved search

Create a file for external citation management software

Your RSS Feed

Astrocytic interleukin-3 programs microglia and limits Alzheimer's disease

Affiliations

Astrocytic interleukin-3 programs microglia and limits Alzheimer's disease

Authors

Affiliations

Abstract

Conflict of interest statement

Figures

Comment in

Similar articles

Cited by

References

Publication types

MeSH terms

Substances

Grants and funding

LinkOut - more resources

Full Text Sources

Other Literature Sources

Medical

Molecular Biology Databases

Research Materials

Abstract

Conflict of interest statement

Figures

Comment in

Similar articles

Cited by

References

Publication types

MeSH terms

Substances

Related information

Grants and funding

LinkOut - more resources

Full Text Sources

Other Literature Sources

Medical

Molecular Biology Databases

Research Materials