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Preparation and application of exosome of toxoplasma gondii infected DC

A technology of toxoplasma gondii and exosomes, applied in the field of medicine and biology, to directly regulate cellular immune response, single source of cells, and reduce toxic and side effects

Active Publication Date: 2021-12-24
SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But there are currently no safe and reliable strains of the parasite for use in cancer therapy

Method used

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  • Preparation and application of exosome of toxoplasma gondii infected DC
  • Preparation and application of exosome of toxoplasma gondii infected DC

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Isolation and culture of mouse bone marrow-derived dendritic cells

[0035] (1) Separate the mouse femur and tibia, wash the bone marrow cells with a medium syringe filled with 1640, then wash twice with PBS, and centrifuge at 250g for 8min. The erythrocyte lysate was lysed for 8 minutes, and counted on a hemocytometer.

[0036] (2) Add 1640 complete medium (10% fetal bovine serum, 1% streptomycin) containing GM-CSF 40ng / ml and IL-420ng / ul, and culture in a carbon dioxide cell incubator for three days.

[0037] (3) After 72 hours, fresh 1640 medium (containing 10% fetal calf serum, 1% streptomycin, 40ng / ml GM-CSF and 20ng / ul IL-4) was supplemented, and the culture was continued for 72 hours.

Embodiment 2

[0039] Toxoplasma culture

[0040] Take out the frozen Toxoplasma gondii strains Me49 and RH from liquid nitrogen, revive them into a 70% Vera T25 cell culture flask, and put them into a 37°C incubator containing 5% CO2 for culture. When releasing about 70% of Toxoplasma gondii, use a cell scraper to scrape off the Vera cells infected by Toxoplasma gondii, then transfer the cell suspension to a 15ml centrifuge tube, filter the cell suspension 3 times with a syringe with a 25-gauge needle, After centrifugation, the supernatant was discarded, and the pellet was resuspended with 10ml PBS and counted under a microscope.

Embodiment 3

[0042] Toxoplasma gondii infected mouse DC cells

[0043] (1) Add complete medium 1640 (10% fetal bovine serum, 1% streptomycin solution) to the mouse DC cells to resuspend the Me49 strain, and mix the parasites and host cells at a ratio of 1:5 Inoculated on the DC monolayer cells at the ratio of 5% CO2, 37 ° C incubator static culture for 24 hours.

[0044] (3) After culturing for 12h, 24h, and 36h, collect the cell culture supernatant.

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Abstract

The invention discloses preparation and application of an exosome of a toxoplasma gondii infected dendritic cell (DC). The DC is infected by toxoplasma gondii, and an exosome derived from the DC is obtained as a strategy for tumor treatment. The exosome from the toxoplasma gondii infection DC is used for treating a colorectal cancer mouse through intratumor injection, tumor volume is reduced, survival rate of the mouse is remarkably increased, and a result shows that the exosome generated by the toxoplasma gondii infection DC has a remarkable tumor inhibition effect. The toxoplasma gondii is utilized to infect DC, the exosome which is controllable in quality and can be prepared on a large scale is prepared through an ultracentrifugation method for tumor treatment, and a new method is provided for tumor treatment.

Description

technical field [0001] The invention belongs to the field of medical biotechnology, and in particular relates to a method for preparing exosomes from DCs infected with Toxoplasma gondii and its application in tumor treatment. Background technique [0002] Tumor is a disease caused by uncontrolled proliferation of cells in cancer patients affected by genetic and environmental factors (Caner, A., et al., Parasites and immunotherapy: Immunostimulatory effect of leishmania spp. In cancer treatment (vol 110 , s33, 2019). European Journal of Cancer, 2019.119: p.199-199). Carcinogenesis can be caused by a variety of factors, among which infectious parasites play an important role in tumor development. Toxoplasma gondii, Echinococcus granulosus, and Trypanosoma cruzi can release the immunosuppression caused by tumors and inhibit tumor growth through Th1-type immune responses; Fathead tapeworms can inhibit the inflammatory response of tumors through the production of anti-inflammato...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0784A61K47/46A61K35/15A61P35/00
CPCC12N5/0639A61K47/46A61K35/15A61P35/00C12N2500/78C12N2509/10Y02A50/30
Inventor 蔺智兵陈兆国陆金苗
Owner SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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