Preparation Of Dihydrotanshinone I Loaded Nanoparticles And Its Mechanism Of Inhibiting The Proliferation Of Hela Cells

Tanshinones are phenanthrenequinone compound with the similar structure.A lot of studies have shown that tanshinones have various physiological activities,such as anti-atherosclerosis,anti-tumor,anti-inflammatory,anti-bacterial and anti-oxidation.Among those,the anti-tumor efficacy is an important part of tanshinones.Dihydrotanshinone I?DI?exhibited the most significant anti-tumor effect.However,DI was insoluble in water and showed low bioavailability in vivo.A large number of novel carriers have been used to improve the aqueous solubility.However,most studies have been focused on tanshinone IIA?TNIIA?.Few reports focused on the research of DI.To solve this problem,DI was selected in this article.New carriers NH₂-PEG-PLGA?PPA?and biotin-PEG-PLGA?BPA?were used to prepare DI-loaded nanoparticles?DI-PPA-NPs and DI-BPA-NPs?.The anti-proliferative of DI/DI-PPA-NPs/DI-BPA-NPs on Hela cells was studied in vitro and the mechanisms of it were investigated.This study is expected to provide reference for utilization of DI in food industry and guidance of DI-BPA-NPs to prevent and treat human cervical cancer.1.The DI-loaded nanoparticle was successfully prepared by the emulsification solvent evaporation method.The optimized preparation process was as follows:the emulsifier PVA concentration was 1%,the weight ratio of DI and PLGA was 1:10,the volume ratio between aqueous and organic phase was 10:1,the ultrasonic time was 5 min,and the ultrasonic power was 60%.Obtained results showed that the particle size,particle size distribution?PDI?and zeta potential of DI-loaded nanoparticles were 169.04±2.73 nm,0.151±0.016 and14.63±1.21 mV,respectively.Images of SEM showed a spherical distribution,uniform particle size and the smooth surface.The red blood cell hemolysis experiments showed that the DI-loaded nanoparticles not only can improve the water solubility of DI,but also can alleviate the red blood cell hemolysis caused by the hydrophobicity of DI.2.Biotin-targeted triblock BPA and non-targeted diblock PPA were synthesized by the carbodiimide method.And the FTIR and ¹H-NMR spectra were used to confirm the synthesis of them.The red blood cell hemolysis experiment showed that the hemolysis rate of the synthesized blocks were less than 5%.In addition,the DI-PPA-NPs and DI-BPA-NPs were prepared with the particle sizes of 180.04±3.12 and 178.57±4.01 nm,respectively.SEM and TEM images showed uniform particle size,spherical distribution and the rough surface.DI-BPA-NPs exhibited sustained-release and pH response properties in PBS solution.3.MTT assay showed that DI,DI-PPA-NPs and DI-BPA-NPs had significant anti-proliferative effects on human cervical cancer Hela cells in a concentration-and time-dependent manner.The IC₅₀0 values of them were 8.20±0.85,6.14±0.31,and 4.55±0.63?M in 72 h,respectively.Cell scratch assay showed that DI-BPA-NPs can significantly inhibit the migration of Hela cells.In addition,the inhibition of cell migration in DI-BPA-NPs group is much higher than that in free DI.Absorption experiments showed that C6-BPA-NPs have better absorption efficiency than C6-PPA-NPs and lysosomes were the uptake site.Results of flow cytometry showed that DI-BPA-NPs inhibited the proliferation of Hela cells through G2/M phase arrest and apoptosis.In addition,western blotting results showed that both the mitochondrial pathway and the death receptor pathway were involved in the apoptosis of Hela cells.In addition,DI-BPA-NPs can scavenge intracellular ROS,further activate AKT,p53 and MAPK and other related signal transduction pathways and regulate the expression of G2/M phase related proteins.In summary,the signaling pathway of Hela cell growth inhibition induced by DI-BPA-NPs was obtained.