Characterization And Evaluation Of HGF-loaded PLGA Nanoparticles In CCl4-induced Acute Liver Injury Mice Model

The liver is the detoxification organ of human body,which is easily damaged by various factors.Chemotherapy is often used in clinical practice to create opportunities for further treatment of patients,which will inevitably cause liver damage.Therefore,in the course of chemotherapy,it is often accompanied by hepatoprotective drugs.Hepatocyte growth factor(HGF)is a multifunctional growth factor that plays an important role in liver regeneration.However,HGF has a very short half-life and limited sources,which greatly limits its clinical application.Moreover,as a growth factor,rapid entry of HGF into the blood in large quantities may cause systemic toxic effects.Therefore,not only the efficient use of HGF,but also the safe use of HGF is required.Poly(lactic-co-glycolic acid)(PLGA)has good biocompatibility and degradability,and has been approved by FDA for use in pharmaceutical preparations.Studies have shown that growth factor-loaded PLGA nanoparticles can make the growth factor not easy to be degraded,increase the stability in the body and improve the bioavailability.Objective:In this study,PLGA nanoparticles with different concentrations of HGF were prepared using the PLGA nanoparticle slow-release system constructed by our laboratory.The qualities of the prepared nanoparticles were evaluated,and the prepared nanoparticles were applied to treat the acute liver injury model induced by carbon tetrachloride in mice to verify the effect of nanoparticles.Methods:CCK-8 kit was used to determine the effect of HGF on HepG2 cell proliferation,and the concentration with the best effect on HepG2 proliferation was selected for subsequent experiments.HGF-loaded PLGA nanoparticles were prepared with an initial dose of O?g,2?g and 4?g HGF.Particle size,polydisperse index and zeta potential of each kind of nanoparticles were measured by nanoparticle size analyzer.The surface and internal morphology of nanoparticles were observed by TEM.The drug loading and in vitro release curve of nanoparticles were determined by BCA kit and mHGF ELISA kit.The electronic balance was used to weigh the nanoparticles and calculate the recovery rate.Control group,model group,HGF 60ng group,HGF 120ng group,NPs group,HGF-NPs-2 g group and HGF-NPs-4 g group were set.HGF solution and nanoparticles were injected into the tail vein 3 days before modeling,and samples were taken 24 hours after modeling.HGF concentrations in the blood and liver tissues of mice in each group were detected by ELISA kit.The liver coefficient and lesion rate were calculated by observing the liver gross and tissue changes of HE staining sections.The concentrations of AST,ALT,ALP,T-BIL,BUN and Scr in blood were determined by automatic biochemical analyzer.The contents of SOD,GPX and MDA in liver tissues were determined by ELISA kit.Results:1.In a certain concentration range,HGF can promote the proliferation of HepG2.When HGF concentration was 40ng/ml and 80ng/ml,HepG2 proliferation was significantly promoted.2.The three kinds of nanoparticles,including NPs,HGF-NPs-2?g and HGF-NPs-4?g had good and stable physical properties,and the nanoparticles were evenly distributed in size,with single-peak curve distribution and stable recovery rate.Nanoparticles could achieve the effect of sudden release and sustained release of drugs,and the release time can last for 7 days.The drug loads of NPs,HGF-NPs-2?g and HGF-NPs-4?g were(4.21±0.15)%,(0.001268±0.000058)%and(0.002422±0.000030)%,respectively.3.HGF-NPs-2?g and HGF-NPs-4?g had protective effects on CCl4-induced acute liver injury mice model.Compared with the same dose of HGF solutions,HGF nanoparticles could maintain a higher level of HGF in the blood and liver tissues of mice for a longer period of time.HGF nanoparticles could significantly reduce the contents of liver enzymes in blood,including AST,ALT,ALP and T-BIL,and reduce the concentration of BUN and Scr in blood.HGF nanoparticles could increase the content of SOD and GPX,reduce the generation and release of MDA in liver tissue,reduce the necrosis and inflammatory infiltration of liver tissue,and reduce the degree of edema in liver tissue.HGF nanoparticles could protect the liver function and kidney function of mice under the injury of CCl4.Conclusion:HGF-lodaed PLGA nanoparticles containing different drug loads of HGF were successfully prepared,with good and stable physical properties,uniform distribution of sizes,single peak curve distribution,and stable recovery rates.The prepared nanopartilces can achieve the effect of sudden release and sustained release of drugs.Compared with HGF solution,HGF-loaded PLGA nanoparticles can maintain higher levels of HGF in blood and liver tissue in mice in a longer period of time.In this way,HGF-loaded PLGA nanoparticles can more significantly reduce concentrations of AST,ALT?ALP and T-BIL in blood,reduce concentrations of BUN and Scr,improve the content of SOD,GPX in liver tissue,and reduce the synthesis and release of MDA on CCl4-induced acute liver injury mice model.In this way,HGF-loaded PLGA nanoparticles can reduce necrosis and inflammatory infiltration of liver tissue,reduce the degree of edema in liver tissue,and maintain liver and kidney functions of mice.