Reversion pulmonary fibrosis nano preparation and preparation method thereof
A pulmonary fibrosis and nano-preparation technology, which is applied in the direction of pharmaceutical formulations, medical preparations of non-active ingredients, non-effective ingredients of polymer compounds, etc., can solve the problems of low delivery efficiency and poor targeting, and achieve high-efficiency lung delivery, Achieve specific targeting and prolong circulation time
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Embodiment 1
[0053] Embodiment 1 Synthesis and preparation of nano-preparation components, such as figure 1 Shown:
[0054] 1. Preparation of double-drug / single-drug PLGA-PEG-MAL nanoparticles
[0055] Polymer nanoparticles loaded with anti-pulmonary fibrosis drugs or fluorescent dyes (as a model instead of drugs for formulation tracking) can be prepared by thin film dispersion method, direct titration method and reverse solvent method. In the present invention, the direct titration method is preferably used to prepare PLGA-PEG-MAL nanoparticles loaded with anti-fibrosis drug double / single drug (trametinib and / or astaxanthin). The specific preparation method is as follows:
[0056] Weigh 100 mg PLGA-PEG-MAL and 10 mg double-drug / single-drug and dissolve in 2 mL DMSO solution respectively, and dissolve by ultrasonication. Under stirring conditions, the DMSO solution was added dropwise to 100 mL of pure water, and after stirring was continued for 2 h at room temperature, the unencapsulate...
Embodiment 2
[0080] Example 2 Uptake analysis of nano-preparation at the level of A549 cells and quantitative analysis by flow cytometry
[0081] Nano-preparations of PP / coumarin 6, PPE / coumarin 6, PPR / coumarin 6 and PPER / coumarin 6 were prepared as described in Example 1. A549 cells were divided into 5×10 5 / mL inoculated in 24-well plate, at 37 ℃, 5% CO 2 After 24 hours of adherent growth in the cell culture incubator, the culture medium was sucked off, TGF-β (5 ng / mL) diluted in serum-free medium was added to stimulate 24 hours, and free coumarin 6, PP / coumarin 6 and PP / coumarin diluted in serum-free medium were added respectively. Coumarin 6, PPR / coumarin 6, PPC / coumarin 6 and PPCR / coumarin 6, the concentration of coumarin 6 in each well is 1 μg / mL, 500 μL, and three auxiliary wells are set for each group. Continue culturing for 4 h, wash three times with PBS,
[0082] (1) The uptake of nanoparticles and free coumarin 6 by cells was photographed by laser confocal LSM-700;
[0083...
Embodiment 4
[0085] Example 4 Analysis of lung-specific delivery of nano-preparations coated with fluorescent dyes (DiR) in mice with pulmonary fibrosis
[0086] PP / DiR, PPE / DiR, PPR / DiR and PPER / DiR nano-formulations were prepared as described in Example 1.
[0087] First, 6-8 week-old male C57BL / 6 mice were used for the modeling experiment of the pulmonary fibrosis model, and the lungs of the mice were directly modeled by tracheal intubation. Bleomycin hydrochloride was used as a mouse lung fibrosis inducer during modeling, and the concentration was 2 USP / Kg. After three weeks, the mouse lung fibrosis was formed, and then the nanoparticle tracer experiment was continued. For judging the modeling time of the mouse pulmonary fibrosis model, a large number of literatures have reported that the pulmonary fibrosis of the mouse is in the rapid development stage after three weeks, and the proliferation of circulating fibroblasts in the body is the most obvious. The three-week-old mice were ran...
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