Exosome compound with tumor inhibition effect
A technology of tumor suppression and exosomes, which is applied in the field of biomedicine, can solve the problems of low progression and survival rate of recurrent and metastatic osteosarcoma, inability to effectively optimize the treatment plan and drugs, etc.
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Embodiment 1
[0034] Example 1 Separation and extraction of exosomes and loading of drug rifampicin
[0035] (1) Isolation and extraction of exosomes from bone marrow mesenchymal stem cells (BMSCs): Select human BMSCs as the research object, use serum without exosomes for cell culture, and select ultracentrifugation to extract exosomes. The methods are as follows: Human Source BMSCs were incubated at 37 °C, 5% CO 2 In a cell incubator, collect 50 mL of cell supernatant at 300 g, centrifuge at 4 °C for 15 min, remove impurities and take the supernatant; 3000 g, centrifuge at 4 °C for 15 min, remove cell debris and take the supernatant; 20000 g, centrifuge at 4 °C for 60 min, take the supernatant Filter through a 0.22 μm sieve; centrifuge at 120,000 g for 120 min at 4°C, carefully remove the supernatant and place it upside down on filter paper to blot dry. , stored at 80°C for later use;
[0036] (2) Preparation of rifampicin-encapsulated exosomes: Add 10 mg / mL of rifampicin to the exosomes...
Embodiment 2
[0038] Example 2 Inhibitory effect of exosome complex on osteosarcoma 143B cell line in vitro
[0039] (1) Human osteosarcoma cell line 143B was inoculated in high glucose complete medium DMEM containing 10% FBS and 1% penicillin, and then placed in DMEM containing 5% CO. 2 cultured in a constant temperature incubator at 37°C. Set up blank control group (Control): add the same amount of PBS solution as other groups in the culture medium; no drug-loaded exosome group (EXO): the culture medium contains 10 6 exosomes / mL; exosome complex group (EXO-RIF): culture medium containing 25 μg / mL rifampicin and 10 6 exosomes / mL.
[0040] (2) Detect the activity of cells by CCK-8 experiment: resuspend the cells by centrifugation to make a cell suspension, then count the cell suspension, dilute the cells to 50 cells / μL, and inoculate 100 μL in a 96-well plate, 10 replicates were set for each group of cells. After culturing in medium containing different drugs for 24 h, 10 μL of CCK-8 rea...
Embodiment 3
[0046] Example 3 Tumor growth inhibition effect of exosome complex on tumor-bearing nude mice
[0047] In this example, 18 male BALB / C nude mice aged 4-6 weeks were selected, and each nude mouse was injected with 20 μL containing 10 7 A suspension of 143B osteosarcoma cells was randomly divided into 3 groups 7 days after tumor injection, with 6 cells in each group. Blank control group (Control): daily intravenous injection of the same amount of normal saline solution as other groups; no drug-loaded exosome group (EXO): daily intravenous injection containing 10 8 exosomes / mL saline solution; exosome complex group (EXO-RIF): According to the weight of nude mice, according to the dose of 10 mg / kg rifampicin, daily intravenous injection containing 10 mg / mL rifampicin and 10 mg / mL rifampicin 8 each / mL exosomes in saline solution. After 21 days of continuous injection, the samples were taken and weighed for observation.
[0048] like Figure 16 and Figure 17 As shown, the tumo...
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