PROBING THE IMPACT OF GAMMA-IRRADIATION ON THE METABOLIC STATE OF NEURAL STEM AND PRECURSOR CELLS USING DUAL-WAVELENGTH INTRINSIC SIGNAL TWO-PHOTON EXCITED FLUORESCENCE

Two-photon excitedfluorescence(TPEF)spectroscopy and imaging were used to investigate the effects of gamma-irradiation on neural stem and precursor cells(NSPCs).While the observed signal from reduced nicotinamide adenine dinucleotide(NADH)was localized to the mitochondria,the signal typically associated with oxidizedflavoproteins(Fp)was distributed diffusely throughout the cell.The measured TPEF emission and excitation spectra were similar to the established spectra of NAD(P)H and Fp.Fpfluorescence intensity was markedly increased by addition of the electron transport chain(ETC)modulator menadione to the medium,along with a concomitant decrease in the NAD(P)H signal.Three-dimensional(3D)neurospheres were imaged to obtain the cellular metabolic index(CMI),calculated as the ratio of Fp to NAD(P)Hfluorescence intensity.Radiation effects were found to differ between low-dose(
50 cGy)and high-dose(50 cGy)exposures.Low-dose irradiation caused a marked drop in CMI values accompanied by increased cellular proliferation.At higher doses,both NAD(P)H and Fp signals increased,leading to an overall elevation in CMI values.Thesefindings underscore the complex relationship between radiation dose,metabolic state,and proliferation status in NSPCs and highlight the ability of TPEF spectroscopy and imaging to characterize metabolism in 3D spheroids.

Evaluating different routes of extracellular vesicle administration for cranial therapies

Aim:Human stem cell-derived extracellular vesicles(EV)provide many advantages over cell-based therapies for the treatment of functionally compromised tissue beds and organ sites.Here we aimed to highlight multiple administration routes for the potential treatment of various forms of brain injury.Methods:Human neural stem cell-derived EV were isolated from conditioned media and administered via three distinct routes:intrahippocampal transplantation,retro-orbital vein injection,and intranasal.EV were administered after which brains were evaluated to determine the capability of EV to translocate into normal tissue.Results:Data showed no significant differences in the amount of EV able to translocate across the brain,indicating the functional equivalence of each administration route to effectively deliver EV to the brain parenchyma.Conclusion:Findings show that both systemic administration routes(retro-orbital vein or intranasal delivery)afforded effective penetrance and perfusion of EV throughout the brain in a minimally invasive manner,and point to a translationally tractable option for treating certain neurological disorders including those resulting from cranial irradiation procedures.