Dermatan Sulfate/Chitosan nanomaterials loaded with IRW modulate human endothelial sterile inflammatory response.

A. BLACHMAN; A. M. BIROCCO; N. ZLOTOLOW; A. EIGUREN ; S. L. SAAVEDRA; S. A. CAMPERI; R. GLISONI; G. C. CALABRESE

Congreso; Reunión Anual de Sociedades de Biociencias; 2019

Blachman Agustín, Birocco Ariadna, Zlotolow Nicole, Eiguren Ana Clara, Camperi Silvia , Glisoni Romina, Calabrese Graciela.Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Ciencias Biológicas, Junín 956, C1113AAD Ciudad Autónoma de Buenos Aires, Argentina.Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Cátedra de Biotecnología. NANOBIOTEC-CONICET, Junín 956, C1113AAD Ciudad Autónoma de Buenos Aires, Argentina.Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Cátedra de Tecnología Farmacéutica II. NANOBIOTEC-CONICET, Junín 956, C1113AAD Ciudad Autónoma de Buenos Aires, Argentina.The present work describes a novel delivery system for the selective targeting of egg-derived anti-inflammatory tripeptide Ile-Arg-Trp (IRW) to modulate human endothelial cells inflammation, in the context of high levels of oxidized triglyceride-rich lipoproteins (VLDLox). IRW is produced by solid phase peptide synthesis. Dermatan sulfate/Chitosan nanoparticles loaded with IRW (8.00% w/w) (DS/CS-IRW) are prepared by ionotropic gelification method and characterized by dynamic light scattering (DLS) and transmission electron microscopy (TEM). VLDLs were isolated from healthy human volunteers by density-gradient ultracentrifugation and oxidized by 5µM CuSO4 for 2 at 37°C. To analyze the selective binding and uptake, human endothelial cells (EA.hy926) and human macrophages were incubated in the presence of FITC-nanoparticles at the biologically active concentration of DS, 10 µg/mL. Endothelial sterile inflammatory response was evaluated by NFκB subcellular distribution through immunofluorescence analysis and zymography studies. The incorporation of IRW results in a stable nanoparticle dispersion with a single size population of 539 ± 75 nm (n= 6). TEM shows that IRW inclusion resulted in compact spherical-like particles. Co-cultures between endothelial cells and macrophages confirm the selective interaction of fluorescent DS/CS-IRW with EA.hy926. After incubating endothelial cells with 100 µg protein/mL of VLDLox for 24h, NFκB is localized both at the cytoplasmic and nuclear compartment. Nevertheless, the transcriptional factor is restricted to the cytoplasm in the presence of DS/CS-IRW nanoparticles. NFκB subcellular distribution was correlated with endothelial inflammatory response through the evaluation of the effect of DS/CS-IRW IRW nanoparticles on matrix metalloproteinases activity-9. Zymographic analysis reveal no detectable MMP-9 activity after DS/CS-IRW treatment. We report here on the capability of these IRW-loaded complexes to modulate endothelial inflammatory response by as simple and potentially scalable nanotechnological platform.