(M1230-12-77) Cellular Uptake and Real-Time ROS Sensing of Doxorubicin-Loaded Self-Assembled Albumin Nanoparticles Using Liver Bilayer Microfluidic Chip System

Purpose: To evaluate cellular uptake and real-time ROS sensing of doxorubicin-loaded self-assembled albumin nanoparticles using liver bilayer microfluidic chip system.

Methods: A conjugate of a human serum albumin (HSA) and oleic acid (C18) was synthesized via 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC)/N-hydroxy succinimide (NHS) reaction to form self-assembled nanoparticles. Human serum albumin-oleic acid nanoparticles (AONs) were prepared via de-solvation method. Doxorubicin hydrochloride (DOX) was efficiently loaded into NPs by incubation process via electrostatic interaction. The physicochemical properties of AONs and DOX-AONs such as particle size, zeta potential and surface morphologies were evaluated dynamic light scattering (DLS) and field emission scanning electron microscope (FE-SEM). Human liver bilayer biomimetic microfluidic system was established to evaluate the effect of nano formulation and shear stress on cellular uptake and real time ROS sensing. Cellular uptake and cell viability were evaluated for free drug and DOX-AONs in static or dynamic environments (0.2 /0.5 /1 dyne/cm²). The degree of ROS generation in cells according to the concentration of free DOX and DOX-AONs were simultaneously measured under dynamic conditions.

Results: The average particle size, and zeta potential of the AONs and DOX-AONs were as follows: average particle size, 190.80 ± 1.020 nm and 325.20 ± 1.04nm; PI, 0.289 ± 0.029 and 0.331 ± 0.027; zeta potential, -45.930 ± 0.785mV and -26.41 ± 0.553mV. Loading content and encapsulation efficiency of DOX was found to be 6.90 ± 0.76% and 64.14 ± 2.41%. The surface morphology of the spherical AON was visualized through FE-SEM. The cellular uptake of DOX-AONs under dynamic condition was significantly increased as compared to static condition, resulting in higher cell-killing efficiency. The biomimetic shear stress (0.2 /0.5 /1 dyne/cm²) was also an important factor on the behavior of cellular uptake. As the concentration of DOX increased, cell viability decreased and ROS generation increased accordingly. Furthermore, the incorporation of pharmaceutical excipients in AONs could modify the cellular uptake and cell-killing behaviors.

Conclusion: Biomimetic microfluidic chip system with real-time ROS sensing using human liver bilayer cells were useful to evaluate the cellular uptake and cell viability of free drug and drug-loaded nanoparticles. Furthermore, formulation compositions and biomimetic shear stress should be carefully considered.

Acknowledgements: Supported by a grant from the Korea Evaluation Institute of Industrial Technology (KEIT) funded by the Ministry of Trade, Industry and Energy (20008413), Republic of Korea.