Peptidoglycan-loaded PLGA-PVA nanoparticles sustain NOD2 and IFN-γ transcription in NK-92 cells beyond nanoparticle withdrawal.

Effective immunomodulation of natural killer (NK) cells holds promise for enhancing innate immune responses in cancer and infectious diseases; however, free immunostimulants often induce only transient activation. In this study, we developed peptidoglycan (PG)-loaded poly(lactic-co-glycolic acid)-polyvinyl alcohol nanoparticles (PGN) as a biocompatible platform for sustained immune signalling. PGN fabricated via nanoprecipitation exhibited a spherical morphology with an average diameter of 150 nm, a zeta potential of - 15 mV, and approximately 18% loading efficiency. 50% encapsulation efficiency was achieved, with release studies confirming controlled PG delivery over 48 h. Functional assays in NK-92 cells demonstrated that while both free PG and PGN upregulated key effector molecules, including perforin, granzyme B, and interferon-γ (IFN-γ), PGN treatment uniquely maintained significantly elevated IFN-γ expression at transcriptional level even after an PAMP withdrawal and a subsequent rest period, indicating prolonged immune stimulation compared to the transient effects of free PG. Mechanistically, PGN sustained NOD2 receptor gene expression, suggesting prolonged modulation of pattern recognition receptor signalling and delayed signal attenuation relative to bare PG. Notably, this sustained post-stimulation transcriptional response exhibited features conceptually consistent with innate immune memory-like or trained immunity-associated phenomena previously described in NK cells and monocytes. These findings highlight the potential of PGN-mediated delivery to extend the temporal window of NK cell activation and support its application as a nanocarrier strategy for next-generation NK cell-based immunotherapies.