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Isolation of Cortical Microglia with Preserved Immunophenotype and Functionality From Murine Neonates
Published on: January 30, 2014
Extracellular microRNAs modulate human microglial function through TLR8
Hannah Weidling1,2,3, Edyta Motta1,2, Leonard D Kuhrt2,3,4
1Institute of Cell Biology and Neurobiology, Charité - Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Berlin, Germany.
Objective:
MicroRNAs (miRNAs) are abundantly expressed in the brain and are specifically dysregulated in central nervous system (CNS) diseases. They act as post-transcriptional gene regulators but can also serve as ligands for Toll-like receptors (TLRs). This study aims to investigate CNS disease-associated miRNAs as signaling molecules for human microglia.
Methods:
Using a machine learning algorithm and the disease-linked database PhenoMiR, we identified Alzheimer's disease (AD)- and glioma-associated miRNAs as ligands for TLR7 and TLR8. Expression of human TLR7 and TLR8 in iPSC-derived human microglia-like cells (iMGLs) was validated by RT-qPCR. Using ELISA, scratch assay, and FACS, we investigated the miRNAs' potential to modulate iMGL function, including cytokine release, motility, and phagocytosis, respectively. The selective human TLR8 antagonist CU-CPT9a was used to determine the role of this receptor in miRNA-induced modulation of human microglial activity. Co-cultures of iMGLs and iPSC-derived human cortical neurons (iNeurons) were analyzed by Neurotrack imaging to assess the effects of miRNAs on human neurites.
Results:
We identified AD- and glioma-associated miR-9-5p, miR-132-5p, miR-340-3p, miR-30e-3p, miR-501-3p, and let-7b as ligands for human TLR7 and TLR8. Exposure of iMGLs to select miRNAs, including miR-9-5p, miR-132-5p, and miR-340-3p, led to interleukin-6 (IL-6) and tumor necrosis factor (TNF) mRNA expression and protein release in a sequence-dependent fashion. Also, these miRNAs acting as signaling molecules, modulated iMGL motility and phagocytosis activity. The miRNA-induced effects on iMGLs were abolished by CU-CPT9a. Extracellular delivery of miR-132-5p and miR-9-5p to co-cultures of iNeurons and iMGLs resulted in reduced neurite length.
Discussion:
Our data establish that distinct CNS disease-associated miRNAs serve as signaling molecules for human microglia via TLR8, thereby controlling the diverse microglial functions and modulating the neuroinflammatory response.
Insights
Central nervous system (CNS) disease-associated microRNAs (miRNAs) signal through Toll-like receptor 8 (TLR8) to regulate human microglia function. These miRNAs modulate neuroinflammation and neurite outgrowth, offering potential therapeutic targets for CNS diseases.
Area of Science:
- Neuroscience
- Immunology
- Molecular Biology
Background:
- MicroRNAs (miRNAs) are key gene regulators in the brain, with dysregulation implicated in central nervous system (CNS) diseases.
- MicroRNAs can act as ligands for Toll-like receptors (TLRs), influencing immune responses.
Purpose of the Study:
- To investigate CNS disease-associated miRNAs as signaling molecules for human microglia.
- To determine the role of Toll-like receptors (TLRs) in mediating miRNA effects on microglia.
Main Methods:
- Machine learning identified Alzheimer's disease (AD) and glioma-associated miRNAs as TLR7/TLR8 ligands.
- Human microglia-like cells (iMGLs) were used to assess miRNA-induced changes in cytokine release, motility, and phagocytosis.
- A selective TLR8 antagonist (CU-CPT9a) was employed to confirm receptor involvement.
Main Results:
- Specific miRNAs (miR-9-5p, miR-132-5p, etc.) were identified as ligands for TLR7 and TLR8.
- Exposure to certain miRNAs modulated iMGL cytokine production (IL-6, TNF), motility, and phagocytosis.
- TLR8 antagonism abolished miRNA-induced microglial responses, and extracellular miRNAs reduced neurite length in co-cultures.
Conclusions:
- CNS disease-associated miRNAs function as signaling molecules for human microglia via TLR8.
- These miRNAs regulate microglial functions and neuroinflammatory responses, impacting neurite outgrowth.
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