Intranasal Human NSC-derived EVs Therapy in Late Middle Age Can Restrain the Activation of NLRP3 Inflammasome and cGAS-STING Signaling in the Aged Hippocampus

Age-related cognitive impairments are linked to detrimental alterations in the hippocampus, which include increased oxidative stress and chronic neuroinflammation known as inflammaging. Inflammaging comprises the activation of the nucleotide-binding domain leucine-rich repeat (NLR) family pyrin domain-containing 3 (NLRP3) inflammasomes, and the cyclic GMP-AMP synthase, and the stimulator of interferon genes (cGAS-STING) pathway that triggers type 1 interferon (IFN-1) signaling. Recent studies have shown that extracellular vesicles from human induced pluripotent stem cell-derived neural stem cells (hiPSC-NSC-EVs) contain therapeutic miRNAs and proteins capable of alleviating oxidative stress and neuroinflammation. This study examined the effects of male and female C57BL6/J mice receiving two doses of intranasal (IN) hiPSC-NSC-EVs (12 x 10 ⁹ EVs/dose, aged-EVs group) or vehicle (aged-Veh group) in late middle age (i.e., at 18 months) on the extent of oxidative stress and chronic neuroinflammation in the hippocampus at 20.5 months of age. Compared to the Aged-Veh group, the hippocampus in the aged-EVs group displayed diminished astrocyte hypertrophy and microglial clusters. Furthermore, the concentrations of oxidative stress markers were reduced, associated with elevated levels of the nuclear factor erythroid 2-related factor 2 and superoxide dismutase and enhanced expression of genes encoding proteins that maintain mitochondrial respiratory chain integrity. Moreover, the hippocampus in the aged-EVs group displayed reduced concentrations of mediators and end products of NLRP3 inflammasome and the downstream p38/mitogen-activated protein kinase activation, and proteins involved in the activation of cGAS-STING-IFN-1 signaling, and the consequent Janus kinase and signal transducer and activator of transcription signaling pathway that leads to the transcription of interferon-stimulated genes. These antioxidant and antiinflammatory molecular changes in the aged-EVs group also improved ability to form recognition and location memories. The results provide the first evidence that IN administrations of hiPSC-NSC-EVs in late middle age can effectively reduce oxidative stress and major neuroinflammatory signaling cascades in the aged hippocampus, leading to better cognitive function in old age.