RNA Analysis of Laser‐Captured Neurons, Microglia and Astrocyte After Intracerebroventricular Injection of Neural Stem Cell‐Derived Exosomes

Abstract Background Neural stem cells (NSC) and adult neurogenesis are known to modulate synaptic plasticity and cognitive function. Decreased number of NSC have been reported in Alzheimer’s Disease (AD), whereas their increase is associated with improved learning and memory. However, an increased neurogenesis does not correlate with preserved cognition, thus suggesting that other mechanisms involving increased numbers of NSC promote the preservation of memory. NSC are also known for their capacity to secrete exosomes, microvesicles containing cell‐specific cargos of biomolecules, that are taken up by other cells, thus modulating their function. We and others have shown that NSC‐derived exosomes (NSCexo) injected in brains of AD transgenic mice models ameliorated the cognitive decline and decreased the binding of toxic oligomers to synapses. The mechanisms by which NSCexo provide protection to synapse is still under investigation. Our long‐term goal is to find the specific genes modulated by NSCexo that may be responsible for the cognitive resilience and the overall improvement of CNS function. With this goal in mind, here we determine the cell‐specific (neuron vs. glia) uptake of NSCexo and subsequent cell‐specific changes in gene expression. Method C57Bl/6 mice received fluorescent‐labeled NSCexo via intracerebroventricular injection; after collection of brain samples, immunofluorescence on brain slices were performed to identify neurons, astrocytes and microglia. The colocalization of NSCexo with neuronal, astrocytic and microglial markers was evaluated. The single cell types were subsequently laser‐captured, and their RNA was isolated. Single‐cell RNA analysis was performed to investigate differences in RNA expression comparing not‐injected and injected animals. Result We found that NSCexo are taken up preferentially by neuronal cells, but also astrocyte and microglia, thus leading to the modulation of relevant gene expression. Interestingly, we also found a pronounced uptake of NSCexo in the sub‐granular area of the hippocampus dentate gyrus, likely by NSCs. Conclusion Our results suggest targeting of neurons (and to a lesser extent, astrocyte and microglia) by NSCexo. They also suggest important targeting of NSC themselves, in what appear to be an autocrine signaling mechanism deserving further in‐depth investigation. Collectively, these mechanisms may play a key role in the reported involvement of decreased NSC and neurogenesis in AD.