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Novel coronavirus SARS-CoV-2 infected rodent model and construction method and application thereof

An animal model, coronavirus technology, applied in animal husbandry and other fields, can solve the problems of the new coronavirus model that cannot normally express human ACE2 receptor protein, lung physiological state, etc., to achieve easy operation, high infection efficiency, and low risk of tumor formation Effect

Active Publication Date: 2021-04-20
TONGJI UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Therefore, the technical problem to be solved by the present invention is to overcome the defects that the new coronavirus model constructed in the prior art cannot normally express human ACE2 receptor protein and simulate the physiological state of normal human lungs, thereby providing a new coronavirus SARS- Rodent animal model of CoV-2 infection and its construction method and use

Method used

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  • Novel coronavirus SARS-CoV-2 infected rodent model and construction method and application thereof
  • Novel coronavirus SARS-CoV-2 infected rodent model and construction method and application thereof
  • Novel coronavirus SARS-CoV-2 infected rodent model and construction method and application thereof

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Embodiment 1

[0049] This embodiment provides a method for constructing a rodent model of novel coronavirus SARS-CoV-2 infection using a bleomycin injury model and Tdtomato fluorescent protein-labeled human bronchial basal cells, comprising the following steps:

[0050] (1) Labeling of human bronchial basal layer cells: Mix lentiviral packaging plasmids pHIV-dTomato, psPAX2 and pMD2.G at a mass ratio of 5:3.75:1.25, transfect packaging cells 293T, and co-culture for 72 hours , collect the culture supernatant containing Tdtomato lentivirus, and centrifuge at 2000g for 10 minutes at 4°C to remove cell debris. The supernatant was filtered through a 0.45 μm filter and collected in an ultracentrifuge tube. Concentrate by ultracentrifugation at 25,000 g at 4°C to obtain a Tdtomato lentivirus suspension. After determining the titer of Tdtomato lentivirus, according to every 1×10 6 Individual bronchial basal cells added 10 7 The Tdtomato lentivirus of TU was co-cultured, and the medium was chang...

Embodiment 2

[0064] The present embodiment provides a kind of method adopting bleomycin injury model and human bronchial basal layer cell to construct the rodent model infected by novel coronavirus SARS-CoV-2, comprising the following steps:

[0065] (1) Construction of lung injury model: prepare 6-8 weeks old NOD.CB17-Prkdc scid / NcrCrl mice were divided into no-transplant control group, 4-day post-infection group and 21-day post-infection group, with one mouse in each group. After inhalational anesthesia with isoflurane, the trachea was intubated, and the bleomycin-PBS solution was administered to the lungs of the three groups of mice through the catheter through a syringe, with a volume of 50 μL for each mouse. The dose is 50 μg bleomycin / monkey.

[0066] (2) Cell transplantation: 7 days after administration of bleomycin, the mice in the 4-day post-infection group and the 21-day post-infection group were anesthetized with isoflurane again and then intubated. Get the human bronchial ba...

Embodiment 3

[0070] This embodiment provides a method for constructing a rodent model of novel coronavirus SARS-CoV-2 infection using lipopolysaccharide combined with porcine trypsin-induced lung injury model and human bronchial basal layer cells, comprising the following steps:

[0071] (1) Construction of lung injury model: prepare three 6-8 week old NOD.CB17-Prkdc scid / NcrCrl mice. After anesthetized with isoflurane, each mouse was given a premixed solution of 40 μL porcine trypsin (10 U / mL) and 0.8 μL lipopolysaccharide (10 mg / mL) through the glottis into the trachea (the solvent was sterile PBS). All mice were injured by continuous instillation for 3 days, once a day, and the volume of each instillation premix was 40.8 μL, and the lung injury model was established.

[0072] (2) Cell transplantation: Cell transplantation was carried out on the second day after the lung injury model was constructed. Cell labeling step), instilled into the trachea through the glottis, and the dose of ...

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Abstract

The invention relates to the technical field of animal model construction, and particularly provides a novel coronavirus SARS-CoV-2 infected rodent model and a construction method and application thereof. The construction method comprises the following steps of constructing a lung injury animal model, specifically, selecting an immunodeficient animal as an object to construct the lung injury animal model; conducting cell transplantation, specifically, transplanting human lung epithelial cells into the lung of the lung injury animal model; and conducting SARS-CoV-2 virus infection, specifically, transplanting SARS-CoV-2 viruses into the animal lung treated in the step of cell transplantation for infection, and establishing the novel coronavirus SARS-CoV-2 infected rodent model. By means of the method, the construction process is simple and convenient, operation is easy, the construction cost is low, and a normal human lung tissue structure and a human ACE2 receptor protein expression situation can be simulated to the greatest extent.

Description

technical field [0001] The invention relates to the technical field of animal model construction, in particular to a rodent animal model infected by a novel coronavirus SARS-CoV-2 and its construction method and application. Background technique [0002] Novel coronavirus disease (coronavirus disease 2019, COVID-19) is a new infectious disease, a respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (severe acute respiratory syndrome coronavirus 2, SARS-CoV-2) , broke out on a global scale, and its high infection rate and death rate threaten the whole world. After SARS-CoV-2 infects the human body, the most common symptoms are cough, fever, shortness of breath, and dyspnea. Severe infection may lead to severe acute respiratory syndrome, renal failure, and even death. Studies have found that SARS, MERS, SARS-CoV-2 and other coronavirus infections can cause an imbalance in the body's immune regulatory network, trigger cytokine storm syndrome (cytokine ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01K67/027
Inventor 左为张婷
Owner TONGJI UNIV
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