Redox Response Based Nano Co-delivery System For The Treatment Of Pulmonary Fibrosis

Idiopathic pulmonary fibrosis(IPF)is a "tumor-like" disease with a short survival time and high mortality rate,which has no clear cause and no effective treatment.The mechanism of IPF is not yet clear,currently,exploring new drugs and pathways is an important issues.Various traditional Chinese medicines and active monomers have shown positive effects in the treatment of IPF,and the development of traditional Chinese medicine preparations for IPF treatment is of great significance.However,common injection drugs have a short circulation time in the body and are difficult to achieve lung targeting,which limits the treatment effect.Therefore,the development of a novel fibrosis microenvironment-responsive nanocarrier system based on IPF pathophysiological characteristics for drug co-delivery and targeted delivery has great potential in improving lung fibrosis treatment.This article designs a combined administration strategy of arnebin-1 and ginsenoside Rg3 targeting IL-17,Fox O3 a,p27 and other protein targets in IPF treatment.Based on the microenvironment characteristics of lung fibrosis,an oxidant-responsive liposome with double modification of hyaluronic acid and PEG is designed for drug co-delivery,achieving long circulation,lung targeting,and responsive drug release to improve IPF treatment.Part Ⅰ:OverviewThis part provides an overview of the oxidative stress mechanism and related proteins in IPF.It then reviews new types of liposomes with special properties,such as sensitive release liposomes and long-circulating liposomes.Finally,it discusses the research progress on the use of resveratrol and ginsenoside Rg3 for IPF treatment,providing a theoretical basis for future research.Part Ⅱ : Synthesis and characterization of PEG modified cholesterol and HA modified cholesterolThis part describes the synthesis of PEG-modified cholesterol linked by disulfide bonds through two esterification reactions,and the characterization of the resulting product.Cholesterol chloroformate was used as the starting material to prepare aminized cholesterol,which was then subjected to condensation and acylation reactions to obtain hyaluronic acid-modified cholesterol.The product was also characterized.PartⅢ:Preparation and in vitro characterization of coloaded liposomesThis part establishes an in vitro content determination analysis method for SKN and Rg3,prepares co-loaded liposomes,and characterizes their appearance,morphology,particle size,and potential.The stability,oxidation-reduction sensitivity,in vitro release behavior,and hemolysis phenomenon were investigated.The results showed that the encapsulation rates of SKN and Rg3 were 85.94% and 79.92%,respectively,and the drug loading capacities were 10.19% and 8.02%,respectively.The co-loaded liposomes had a spherical shape with a lipid bilayer on the surface,a particle size of 402.38±3.2 nm,a dispersity index of 0.191,and a zeta potential of-31.63±2.25 m V.The stability of the co-loaded liposome solution was good,and the particle size and turbidity decreased while the in vitro drug release increased under an oxidation-reduction environment.The co-loaded liposomes did not exhibit hemolysis,indicating good biocompatibility.Part Ⅳ:In vitro cell study of coloaded liposomesThis part evaluates the biosafety of blank lipid carriers using MTT assay;the inhibitory effects of free drugs and drug-loaded liposomes on 3T3 cells;and the inhibitory effects of co-loaded liposomes on 3T3 cells under oxidative conditions,elucidating the synergistic effect of SKN and Rg3.The cellular uptake,targeting,and macrophage escape effects of liposomes were examined using coumarin-6(C6)as a fluorescent probe.The results showed that blank lipid carriers had good biocompatibility.The inhibitory effect of SKN and Rg3 on cells was concentration-dependent and exhibited a synergistic effect.Liposomes showed stronger cell inhibitory effects compared to free drugs and exhibited stronger effects under oxidative conditions.Liposomes could enhance the uptake of C6 in 3T3 cells,and HA-modified liposomes showed the strongest uptake effect in vitro targeting.PEG-modified liposomes had good macrophage escape effect.Part Ⅴ:In vivo pharmacokinetics,targeting and pharmacodynamics of coloaded liposomesThis part the in vivo content determination analysis method of SKN and Rg3 was established.The pharmacokinetic parameters were calculated by drawing the drug-time curves of the two drugs.The results showed that PEG-modified co-loaded liposomes could prolong the retention time of drugs in the body and had a sustained-release effect.In addition,examines the expression of CD44 in the lungs of mice using immunohistochemistry.The distribution and targeting of co-loaded liposomes in animals were studied through in vivo imaging of mice.The results showed that CD44 was overexpressed in the pulmonary fibrotic lesion,and HA-modified liposomes had a significant lung-targeting effect.A pulmonary fibrosis animal model was established by tracheal instillation of a solution of bleomycin.The animals were randomly divided into control,model,and treatment groups to investigate the in vivo pharmacology of co-loaded liposomes.After treatment,changes in body weight were assessed,and the content of hydroxyproline in lung tissue was measured.The levels of col-Ⅰ,TGF-β1,and α-SMA in lung tissue were determined by immunohistochemistry,and the collagen fiber level was assessed by Masson staining.The results showed that the weight of the treatment group increased compared to the model group.The content of hydroxyproline in the co-loaded liposome group was significantly decreased,which effectively reduced the expression of col-Ⅰ,TGF-β1,α-SMA,and the level of pulmonary collagen fibers.The mechanism of action was explored through ELISA,which showed that the co-loaded liposomes downregulated IL-17 through indigo and upregulated Fox O3 a while downregulating p27 protein levels through ginsenoside Rg3,resulting in a therapeutic effect on pulmonary fibrosis.Histological studies using HE staining showed that co-loaded liposomes had good biosafety and effectively reduced the pathological morphological changes in pulmonary fibrosis animals.