iPSC-Derived Microglia for Modeling Human-Specific DAMP and PAMP Responses in the Context of Alzheimer’s Disease
Abstract
:1. Introduction
2. Results
2.1. CHRNA7 and CHRFAM7A Expression in Functional Microglia Like Cells
2.2. CHRFAM7A Mitigated the α7-nAChR-Dependent Aβ1–42 Uptake
2.3. Immune Surveillance to DAMP: CHRFAM7A Facilitated Microglia Activation
2.4. CHRFAM7A Heightened the Immune Responsiveness to PAMP
2.5. CHRFAM7A Affected the NF-κB Translocation Dynamics, Resulting in Innate Immune Activation
3. Discussion
4. Materials and Methods
4.1. Cell Culture and Microglia Differentiation
4.2. Transfection
4.3. Reverse Transcription Quantitative Polymerase Chain Reaction (RT-qPCR)
4.4. Immunocytochemistry
4.5. Total Cell Lysate Preparation and Immunoblotting
4.6. Phagocytosis Assay
4.7. Cytokine Profiler
4.8. Amyloid Beta Uptake and Cell Counts
4.9. Treatment with LPS and α7 nAChR Agonist/Antagonist
4.10. ELISA
4.11. NF-κB Binding Assay
4.12. NF-κB Translocation and Quantification
4.13. Statistical Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
Abbreviations
α7 nAChR | Alpha7 nicotinic acetylcholine receptor |
AD | Alzheimer’s disease |
AB | Amyloid beta |
DAM | Disease-associated microglia |
DAMPs | Damage-associated molecular patterns |
PAMPs | Pathogen-associated molecular patterns |
MLA | Methyllycaconitine |
LPS | Lipopolysaccharides |
References
- Zhang, Z.G.; Li, Y.; Ng, C.T.; Song, Y.Q. Inflammation in Alzheimer’s Disease and Molecular Genetics: Recent Update. Arch Immunol. Ther. Exp. (Warsz) 2015, 63, 333–344. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Schmitz, T.W.; Soreq, H.; Poirier, J.; Spreng, R.N. Longitudinal Basal Forebrain Degeneration Interacts with TREM2/C3 Biomarkers of Inflammation in Presymptomatic Alzheimer’s Disease. J. Neurosci. 2020, 40, 1931–1942. [Google Scholar] [CrossRef] [PubMed]
- Wang, H.; Yu, M.; Ochani, M.; Amella, C.A.; Tanovic, M.; Susarla, S.; Li, J.H.; Wang, H.; Yang, H.; Ulloa, L.; et al. Nicotinic acetylcholine receptor alpha7 subunit is an essential regulator of inflammation. Nature 2003, 421, 384–388. [Google Scholar] [CrossRef] [PubMed]
- Shytle, R.D.; Mori, T.; Townsend, K.; Vendrame, M.; Sun, N.; Zeng, J.; Ehrhart, J.; Silver, A.A.; Sanberg, P.R.; Tan, J. Cholinergic modulation of microglial activation by alpha 7 nicotinic receptors. J. Neurochem. 2004, 89, 337–343. [Google Scholar] [CrossRef]
- De Simone, R.; Ajmone-Cat, M.A.; Carnevale, D.; Minghetti, L. Activation of alpha7 nicotinic acetylcholine receptor by nicotine selectively up-regulates cyclooxygenase-2 and prostaglandin E2 in rat microglial cultures. J. Neuroinflamm. 2005, 2, 4. [Google Scholar] [CrossRef] [Green Version]
- Kalkman, H.O.; Feuerbach, D. Modulatory effects of alpha7 nAChRs on the immune system and its relevance for CNS disorders. Cell Mol. Life Sci. 2016, 73, 2511–2530. [Google Scholar] [CrossRef] [Green Version]
- Pohanka, M. Alpha7 nicotinic acetylcholine receptor is a target in pharmacology and toxicology. Int. J. Mol. Sci. 2012, 13, 2219–2238. [Google Scholar] [CrossRef] [Green Version]
- Wang, H.Y.; Lee, D.H.; Davis, C.B.; Shank, R.P. Amyloid peptide Abeta(1-42) binds selectively and with picomolar affinity to alpha7 nicotinic acetylcholine receptors. J. Neurochem. 2000, 75, 1155–1161. [Google Scholar] [CrossRef]
- Lewis, A.S.; van Schalkwyk, G.I.; Bloch, M.H. Alpha-7 nicotinic agonists for cognitive deficits in neuropsychiatric disorders: A translational meta-analysis of rodent and human studies. Prog. Neuropsychopharmacol. Biol. Psychiatry 2017, 75, 45–53. [Google Scholar] [CrossRef] [Green Version]
- Zhou, Y.; Song, W.M.; Andhey, P.S.; Swain, A.; Levy, T.; Miller, K.R.; Poliani, P.L.; Cominelli, M.; Grover, S.; Gilfillan, S.; et al. Human and mouse single-nucleus transcriptomics reveal TREM2-dependent and TREM2-independent cellular responses in Alzheimer’s disease. Nat. Med. 2020, 26, 131–142. [Google Scholar] [CrossRef]
- Fortna, A.; Kim, Y.; MacLaren, E.; Marshall, K.; Hahn, G.; Meltesen, L.; Brenton, M.; Hink, R.; Burgers, S.; Hernandez-Boussard, T.; et al. Lineage-specific gene duplication and loss in human and great ape evolution. PLoS Biol. 2004, 2, E207. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Sudmant, P.H.; Kitzman, J.O.; Antonacci, F.; Alkan, C.; Malig, M.; Tsalenko, A.; Sampas, N.; Bruhn, L.; Shendure, J.; Genomes, P.; et al. Diversity of human copy number variation and multicopy genes. Science 2010, 330, 641–646. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Charrier, C.; Joshi, K.; Coutinho-Budd, J.; Kim, J.E.; Lambert, N.; de Marchena, J.; Jin, W.L.; Vanderhaeghen, P.; Ghosh, A.; Sassa, T.; et al. Inhibition of SRGAP2 function by its human-specific paralogs induces neoteny during spine maturation. Cell 2012, 149, 923–935. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Florio, M.; Albert, M.; Taverna, E.; Namba, T.; Brandl, H.; Lewitus, E.; Haffner, C.; Sykes, A.; Wong, F.K.; Peters, J.; et al. Human-specific gene ARHGAP11B promotes basal progenitor amplification and neocortex expansion. Science 2015, 347, 1465–1470. [Google Scholar] [CrossRef]
- Dennis, M.Y.; Eichler, E.E. Human adaptation and evolution by segmental duplication. Curr. Opin. Genet. Dev. 2016, 41, 44–52. [Google Scholar] [CrossRef] [Green Version]
- Bitar, M.; Kuiper, S.; O’Brien, E.A.; Barry, G. Genes with human-specific features are primarily involved with brain, immune and metabolic evolution. BMC Bioinform. 2019, 20 (Suppl 9), 406. [Google Scholar] [CrossRef]
- Sinkus, M.L.; Graw, S.; Freedman, R.; Ross, R.G.; Lester, H.A.; Leonard, S. The human CHRNA7 and CHRFAM7A genes: A review of the genetics, regulation, and function. Neuropharmacology 2015, 96, 274–288. [Google Scholar] [CrossRef] [Green Version]
- Costantini, T.W.; Chan, T.W.; Cohen, O.; Langness, S.; Treadwell, S.; Williams, E.; Eliceiri, B.P.; Baird, A. Uniquely human CHRFAM7A gene increases the hematopoietic stem cell reservoir in mice and amplifies their inflammatory response. Proc. Natl. Acad. Sci. USA 2019, 116, 7932–7940. [Google Scholar] [CrossRef] [Green Version]
- Jiang, Y.; Yuan, H.; Huang, L.; Hou, X.; Zhou, R.; Dang, X. Global proteomic profiling of the uniquely human CHRFAM7A gene in transgenic mouse brain. Gene 2019, 714, 143996. [Google Scholar] [CrossRef]
- Ihnatovych, I.; Nayak, T.K.; Ouf, A.; Sule, N.; Birkaya, B.; Chaves, L.; Auerbach, A.; Szigeti, K. iPSC model of CHRFAM7A effect on alpha7 nicotinic acetylcholine receptor function in the human context. Transl. Psychiatry 2019, 9, 59. [Google Scholar] [CrossRef]
- Szigeti, K.; Ihnatovych, I.; Birkaya, B.; Chen, Z.; Ouf, A.; Indurthi, D.C.; Bard, J.E.; Kann, J.; Adams, A.; Chaves, L.; et al. CHRFAM7A: A human specific fusion gene, accounts for the translational gap for cholinergic strategies in Alzheimer’s disease. EBioMedicine 2020, 59, 102892. [Google Scholar] [CrossRef] [PubMed]
- Kabbani, N.; Nichols, R.A. Beyond the Channel: Metabotropic Signaling by Nicotinic Receptors. Trends Pharmacol. Sci. 2018, 39, 354–366. [Google Scholar] [CrossRef] [PubMed]
- Rosas-Ballina, M.; Tracey, K.J. Cholinergic control of inflammation. J. Intern. Med. 2009, 265, 663–679. [Google Scholar] [CrossRef] [PubMed]
- Venkataraman, L.; Fair, S.R.; McElroy, C.A.; Hester, M.E.; Fu, H. Modeling neurodegenerative diseases with cerebral organoids and other three-dimensional culture systems: Focus on Alzheimer’s disease. Stem. Cell Rev. Rep. 2020. [Google Scholar] [CrossRef]
- Garcia-Leon, J.A.; Caceres-Palomo, L.; Sanchez-Mejias, E.; Mejias-Ortega, M.; Nunez-Diaz, C.; Fernandez-Valenzuela, J.J.; Sanchez-Varo, R.; Davila, J.C.; Vitorica, J.; Gutierrez, A. Human Pluripotent Stem Cell-Derived Neural Cells as a Relevant Platform for Drug Screening in Alzheimer’s Disease. Int. J. Mol. Sci. 2020, 21, 6867. [Google Scholar] [CrossRef]
- Lee, C.T.; Bendriem, R.M.; Wu, W.W.; Shen, R.F. 3D brain Organoids derived from pluripotent stem cells: Promising experimental models for brain development and neurodegenerative disorders. J. Biomed. Sci. 2017, 24, 59. [Google Scholar] [CrossRef]
- Cenini, G.; Hebisch, M.; Iefremova, V.; Flitsch, L.J.; Breitkreuz, Y.; Tanzi, R.E.; Kim, D.Y.; Peitz, M.; Brustle, O. Dissecting Alzheimer’s disease pathogenesis in human 2D and 3D models. Mol. Cell Neurosci. 2020, 103568. [Google Scholar] [CrossRef]
- Muffat, J.; Li, Y.; Yuan, B.; Mitalipova, M.; Omer, A.; Corcoran, S.; Bakiasi, G.; Tsai, L.H.; Aubourg, P.; Ransohoff, R.M.; et al. Efficient derivation of microglia-like cells from human pluripotent stem cells. Nat. Med. 2016, 22, 1358–1367. [Google Scholar] [CrossRef] [Green Version]
- Kierdorf, K.; Prinz, M. Factors regulating microglia activation. Front. Cell Neurosci. 2013, 7, 44. [Google Scholar] [CrossRef] [Green Version]
- Smith, A.M.; Gibbons, H.M.; Oldfield, R.L.; Bergin, P.M.; Mee, E.W.; Faull, R.L.; Dragunow, M. The transcription factor PU.1 is critical for viability and function of human brain microglia. Glia 2013, 61, 929–942. [Google Scholar] [CrossRef]
- Lane, K.; Van Valen, D.; DeFelice, M.M.; Macklin, D.N.; Kudo, T.; Jaimovich, A.; Carr, A.; Meyer, T.; Pe’er, D.; Boutet, S.C.; et al. Measuring Signaling and RNA-Seq in the Same Cell Links Gene Expression to Dynamic Patterns of NF-kappaB Activation. Cell Syst. 2017, 4, 458–469.e5. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Costantini, T.W.; Dang, X.; Coimbra, R.; Eliceiri, B.P.; Baird, A. CHRFAM7A, a human-specific and partially duplicated alpha7-nicotinic acetylcholine receptor gene with the potential to specify a human-specific inflammatory response to injury. J. Leukoc. Biol. 2015, 97, 247–257. [Google Scholar] [CrossRef] [Green Version]
- Maldifassi, M.C.; Martin-Sanchez, C.; Atienza, G.; Cedillo, J.L.; Arnalich, F.; Bordas, A.; Zafra, F.; Gimenez, C.; Extremera, M.; Renart, J.; et al. Interaction of the alpha7-nicotinic subunit with its human-specific duplicated dupalpha7 isoform in mammalian cells: Relevance in human inflammatory responses. J. Biol. Chem. 2018, 293, 13874–13888. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Ries, M.; Sastre, M. Mechanisms of Abeta Clearance and Degradation by Glial Cells. Front. Aging Neurosci. 2016, 8, 160. [Google Scholar] [CrossRef] [Green Version]
- Jefferson, W.K.; Shane, M.B.; Andrew, S.M.; Amanda, M.L.; Arnold, M.S.; Bruce, T.L. Inflammation as a central mechanism in Alzheimer’s disease. Alzheimers Dement. (N. Y.) 2018, 4, 575–590. [Google Scholar]
- Benfante, R.; Antonini, R.A.; De Pizzol, M.; Gotti, C.; Clementi, F.; Locati, M.; Fornasari, D. Expression of the alpha7 nAChR subunit duplicate form (CHRFAM7A) is down-regulated in the monocytic cell line THP-1 on treatment with LPS. J. Neuroimmunol. 2011, 230, 74–84. [Google Scholar] [CrossRef] [PubMed]
- Cirrito, J.R.; May, P.C.; O’Dell, M.A.; Taylor, J.W.; Parsadanian, M.; Cramer, J.W.; Audia, J.E.; Nissen, J.S.; Bales, K.R.; Paul, S.M.; et al. In vivo assessment of brain interstitial fluid with microdialysis reveals plaque-associated changes in amyloid-beta metabolism and half-life. J. Neurosci. 2003, 23, 8844–8853. [Google Scholar] [CrossRef] [Green Version]
- de Lucas-Cerrillo, A.M.; Maldifassi, M.C.; Arnalich, F.; Renart, J.; Atienza, G.; Serantes, R.; Cruces, J.; Sanchez-Pacheco, A.; Andres-Mateos, E.; Montiel, C. Function of partially duplicated human alpha77 nicotinic receptor subunit CHRFAM7A gene: Potential implications for the cholinergic anti-inflammatory response. J. Biol. Chem. 2011, 286, 594–606. [Google Scholar] [CrossRef] [Green Version]
- Bencherif, M.; Lippiello, P.M.; Lucas, R.; Marrero, M.B. Alpha7 nicotinic receptors as novel therapeutic targets for inflammation-based diseases. Cell Mol. Life Sci. 2011, 68, 931–949. [Google Scholar] [CrossRef] [Green Version]
- Walana, W.; Wang, J.J.; Yabasin, I.B.; Ntim, M.; Kampo, S.; Al-Azab, M.; Elkhider, A.; Dogkotenge Kuugbee, E.; Cheng, J.W.; Gordon, J.R.; et al. IL-8 analogue CXCL8 (3-72) K11R/G31P, modulates LPS-induced inflammation via AKT1-NF-kbeta and ERK1/2-AP-1 pathways in THP-1 monocytes. Hum. Immunol. 2018, 79, 809–816. [Google Scholar] [CrossRef]
- Falvo, J.V.; Tsytsykova, A.V.; Goldfeld, A.E. Transcriptional Control of the TNF Gene. Curr. Dir. Autoimmun. 2010, 11, 27–60. [Google Scholar] [PubMed] [Green Version]
- Laffer, B.; Bauer, D.; Wasmuth, S.; Busch, M.; Jalilvand, T.V.; Thanos, S.; Zu Horste, G.M.; Loser, K.; Langmann, T.; Heiligenhaus, A.; et al. Loss of IL-10 Promotes Differentiation of Microglia to a M1 Phenotype. Front. Cell Neurosci. 2019, 13. [Google Scholar] [CrossRef] [PubMed]
- Liu, M.; Saredy, J.; Zhang, R.; Shao, Y.; Sun, Y.; Yang, W.Y.; Wang, J.; Liu, L.; Drummer, C., IV; Johnson, C.; et al. Approaching Inflammation Paradoxes-Proinflammatory Cytokine Blockages Induce Inflammatory Regulators. Front. Immunol. 2020, 11, 554301. [Google Scholar] [CrossRef] [PubMed]
- Lobo-Silva, D.; Carriche, G.M.; Castro, A.G.; Roque, S.; Saraiva, M. Balancing the immune response in the brain: IL-10 and its regulation. J. Neuroinflamm. 2016, 13. [Google Scholar] [CrossRef] [Green Version]
- Zambrano, S.; Bianchi, M.E.; Agresti, A. High-throughput analysis of NF-kappaB dynamics in single cells reveals basal nuclear localization of NF-kappaB and spontaneous activation of oscillations. PLoS ONE 2014, 9, e90104. [Google Scholar] [CrossRef]
- Maity, A.; Wollman, R. Information transmission from NFkB signaling dynamics to gene expression. PLoS Comput. Biol. 2020, 16, e1008011. [Google Scholar] [CrossRef]
- Dorrington, M.G.; Fraser, I.D.C. NF-kappaB Signaling in Macrophages: Dynamics, Crosstalk, and Signal Integration. Front. Immunol. 2019, 10, 705. [Google Scholar] [CrossRef]
- Zambrano, S.; De Toma, I.; Piffer, A.; Bianchi, M.E.; Agresti, A. NF-kappaB oscillations translate into functionally related patterns of gene expression. Elife 2016, 5, e09100. [Google Scholar] [CrossRef]
- Lin, Y.T.; Seo, J.; Gao, F.; Feldman, H.M.; Wen, H.L.; Penney, J.; Cam, H.P.; Gjoneska, E.; Raja, W.K.; Cheng, J.; et al. APOE4 Causes Widespread Molecular and Cellular Alterations Associated with Alzheimer’s Disease Phenotypes in Human iPSC-Derived Brain Cell Types. Neuron 2018, 98, 1141–1154.e7. [Google Scholar] [CrossRef] [Green Version]
- Wang, Y.; Xiao, C.; Indersmitten, T.; Freedman, R.; Leonard, S.; Lester, H.A. The duplicated alpha7 subunits assemble and form functional nicotinic receptors with the full-length alpha7. J. Biol. Chem. 2014, 289, 26451–26463. [Google Scholar] [CrossRef] [Green Version]
- Warren, L.; Manos, P.D.; Ahfeldt, T.; Loh, Y.H.; Li, H.; Lau, F.; Ebina, W.; Mandal, P.K.; Smith, Z.D.; Meissner, A.; et al. Highly efficient reprogramming to pluripotency and directed differentiation of human cells with synthetic modified mRNA. Cell Stem Cell 2010, 7, 618–630. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Hu, X.; Crick, S.L.; Bu, G.; Frieden, C.; Pappu, R.V.; Lee, J.M. Amyloid seeds formed by cellular uptake, concentration, and aggregation of the amyloid-beta peptide. Proc. Natl. Acad. Sci. USA 2009, 106, 20324–20329. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Trask, O.J., Jr. Nuclear Factor Kappa B (NF-kappaB) Translocation Assay Development and Validation for High Content Screening. In Assay Guidance Manual; Markossian, S., Sittampalam, G.S., Grossman, A., Brimacombe, K., Arkin, M., Auld, D., Austin, C.P., Baell, J., Caaveiro, J.M.M., Chung, T.D.Y., et al., Eds.; Bethesda: Rockville, MD, USA, 2004. [Google Scholar]
- Bagaev, A.V.; Garaeva, A.Y.; Lebedeva, E.S.; Pichugin, A.V.; Ataullakhanov, R.I.; Ataullakhanov, F.I. Elevated pre-activation basal level of nuclear NF-kappaB in native macrophages accelerates LPS-induced translocation of cytosolic NF-kappaB into the cell nucleus. Sci. Rep. 2019, 9, 4563. [Google Scholar] [CrossRef] [PubMed] [Green Version]
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Ihnatovych, I.; Birkaya, B.; Notari, E.; Szigeti, K. iPSC-Derived Microglia for Modeling Human-Specific DAMP and PAMP Responses in the Context of Alzheimer’s Disease. Int. J. Mol. Sci. 2020, 21, 9668. https://doi.org/10.3390/ijms21249668
Ihnatovych I, Birkaya B, Notari E, Szigeti K. iPSC-Derived Microglia for Modeling Human-Specific DAMP and PAMP Responses in the Context of Alzheimer’s Disease. International Journal of Molecular Sciences. 2020; 21(24):9668. https://doi.org/10.3390/ijms21249668
Chicago/Turabian StyleIhnatovych, Ivanna, Barbara Birkaya, Emily Notari, and Kinga Szigeti. 2020. "iPSC-Derived Microglia for Modeling Human-Specific DAMP and PAMP Responses in the Context of Alzheimer’s Disease" International Journal of Molecular Sciences 21, no. 24: 9668. https://doi.org/10.3390/ijms21249668
APA StyleIhnatovych, I., Birkaya, B., Notari, E., & Szigeti, K. (2020). iPSC-Derived Microglia for Modeling Human-Specific DAMP and PAMP Responses in the Context of Alzheimer’s Disease. International Journal of Molecular Sciences, 21(24), 9668. https://doi.org/10.3390/ijms21249668