Manipulation of miR-124 expression on neuronal APP-SWE cells results in different microglial polarization through paracrine signaling
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1
Research Institute for Medicines (iMed.ULisboa), Portugal
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2
Faculdade de Farmácia, Universidade de Lisboa, Portugal
Neuronal expression of miR-124 is associated to brain regeneration and glial signaling. We found miR-124 overexpression in SH-SY5Y cells transfected with the Swedish mutation APP695 (APP-Swe), as a model of Alzheimer’s disease (AD) [1]. Our recent data suggest that miR-124 upregulation in APP-Swe cells, rather than being pathological, sustains neuronal morphology and inhibits inflammatory-miRNA dissemination via exosomes (unpublished). However, whether/how such miR-124 overexpression by neurons impacts on microglia immunoregulatory properties remains unclear. Here, we investigated the inflammatory dynamic signaling between human AD neurons up-/down-regulated for miR-124 and the activated microglia, as well as the acquired phenotypic diversity. miR-124 modulation in APP-Swe cells was done by transfection with anti/pre-miR-124. Activation of human CHME-3 microglia into the M1-phenotype was performed by IFN-γ stimulation. APP-Swe+CHME-3 co-cultures were maintained for 2, 12 and 24 h. Microglial mRNA/miRNA expression of inflammatory markers was accessed by RT-qPCR. Metalloproteinase activity was determined by gelatin zymography and nitric oxide (NO) by the Griess reaction.
IFN-γ induced a classical M1 phenotype on CHME-3 microglia, with consistent overexpression of classical inflammatory-associated markers like RAGE, HMGB1, TNF-α and MHC-II, as well as up-regulation of miR-21, miR146a and miR-155. When IFN-γ-stimulated microglia were co-cultured with miR-124 downregulated APP-Swe cells, we observed an early overexpression of M1-markers, followed by reduction of the M2-markers Arginase-1 and IL-10, as well as of miR-124 and miR-125b. Contrariwise, the co-culture of APP-Swe cells overexpressing miR-124 with IFN-γ-stimulated microglia led to an initial repression of HMGB1 and MHC-II transcription before their recovery at 24h. At this time point, upregulation of neuronal miR-124 led to iNOS, TNF-α, miR-21 and miR-146a repression in microglia. Noteworthy, APP-Swe cells overexpressing miR-124 also inhibited the release of NO and metalloproteinases 2 and 9 into cell media by IFN-γ-stimulated microglia.
More than the beneficial effects observed on APP-Swe cells, overexpression of neuronal miR-124 results in a strong paracrine signaling and regulation of microglia activation, by counteracting pro-inflammatory cascades and inducing a regenerative/repairing polarization. It remains to be understood how effectively these microglia benefit bystander cells and counteract neurodegeneration, the major bottleneck in the development of efficient therapies for AD.
Acknowledgements
EU-JPND project (grant Nº643417 to DB); Fundação para a Ciência e a Tecnologia (JPco-fuND/0003/2015 to DB; UID/DTP/04138/2013 to iMed.ULisboa and SFRH/BD/128738/2017 to GG).
References
[1] A. Fernandes, et al., Biochimie, 155 (2018).
Keywords:
microglia activation,
Neuroinflammation,
Secretome-mediated Signaling,
Alzheimer’s disease (AD),
Inflammatory microRNAs
Conference:
XVI Meeting of the Portuguese Society for Neuroscience (SPN2019), Lisboa, Portugal, 30 May - 1 Jun, 2019.
Presentation Type:
Poster presentation
Topic:
Glia / Neuroinflammation
Citation:
Garcia
G,
Fernandes
A and
Brites
D
(2019). Manipulation of miR-124 expression on neuronal APP-SWE cells results in different microglial polarization through paracrine signaling.
Front. Cell. Neurosci.
Conference Abstract:
XVI Meeting of the Portuguese Society for Neuroscience (SPN2019).
doi: 10.3389/conf.fncel.2019.01.00021
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Received:
16 Apr 2019;
Published Online:
27 Sep 2019.
*
Correspondence:
Prof. Dora Brites, Research Institute for Medicines (iMed.ULisboa), Lisboa, Portugal, dbrites@ff.ulisboa.pt