A synergistic bactericidal effect of low-frequency and low-intensity ultrasound combined with levofloxacin-loaded PLGA nanoparticles on M. smegmatis in macrophages

Abstract Purpose: Tuberculosis (TB) is a highly infectious disease caused by Mycobacterium tuberculosis (Mtb), which often parasites in macrophages. This study is performed to investigate the bactericidal effect and underlying mechanisms of low-frequency and low-intensity ultrasound (LFLIU) combined with levofloxacin-loaded PLGA nanoparticles (LEV-NPs) on M. smegmatis (a surrogate of Mtb) in macrophages.Methods and results: The LEV-NPs were prepared using a double emulsification method. The average diameter, zeta potential, polydispersity index, morphology, and drug release efficiency in vitro of the LEV-NPs were investigated. M. smegmatis in macrophages was treated using the LEV-NPs combined with 42 kHz ultrasound irradiation at an intensity of 0.13 W/cm2 for 10 min. The results showed that ultrasound significantly promoted the phagocytosis of nanoparticles by macrophages (p < 0.05). In addition, further ultrasound combined with the LEV-NPs promoted the production of reactive oxygen species (ROS) in macrophage, and the apoptosis rate of the macrophages was significantly higher than that of the control (p < 0.05). The transmission electronic microscope showed that the cell wall of M. smegmatis was ruptured, the cell structure was incomplete, and the bacteria received severe damage in the ultrasound combined with the LEV-NPs group. Activity assays showed that ultrasound combined with the LEV-NPs exhibited a 10-fold higher antibacterial activity against M. smegmatis residing inside macrophages compared with the free drug.Conclusion: These data demonstrated that ultrasound combined with LEV-NPs has great potential as a therapeutic agent for TB.