Construction and applications of anti-influenza-virus or anti-inflammatory drug screening model based on in vitro culture of human lung particles

An anti-influenza virus and anti-inflammatory drug technology, applied in the field of medicine, can solve problems such as no screening of anti-influenza virus drugs or anti-inflammatory drugs, and achieve the effects of simple and easy operation of drug dosage, accurate drug evaluation, and less drug dosage.

Inactive Publication Date: 2018-10-23
WUHAN INST OF VIROLOGY CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no evaluation and report on the method of screening anti-influenza virus drugs or anti-inflammatory drugs using the in vitro culture method of human lung granule tissue

Method used

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  • Construction and applications of anti-influenza-virus or anti-inflammatory drug screening model based on in vitro culture of human lung particles
  • Construction and applications of anti-influenza-virus or anti-inflammatory drug screening model based on in vitro culture of human lung particles
  • Construction and applications of anti-influenza-virus or anti-inflammatory drug screening model based on in vitro culture of human lung particles

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Establishment of in vitro culture model of porcine lung particles for screening anti-influenza virus drugs

[0040] 1. Experimental materials

[0041] 1.1 Porcine lung tissue and virus

[0042] The pig lung tissue was obtained from the vegetable market of Wuhan University, and it was freshly obtained by the sales staff. Influenza strain A / PuertoRico / 8 / 34 (H1N1) was stored frozen at -80°C.

[0043] 1.2 Reagents

[0044] DMEM / F12 medium was purchased from GIBCO; 2'-(4-methylumbelliferyl)-α-D-acetylneuraminic acid (MUNANA) was purchased from Sigma.

[0045] 1.3 Instruments and equipment

[0046] Ophthalmic scissors, ophthalmic forceps, Perkin Elmer Wallac Envision multi-label reader, 37°C CO 2 Humidified incubator.

[0047] 2. Experimental methods and results

[0048] 2.1 Preparation, subpackage and culture of porcine lung granule tissue

[0049] Take a sterile 4mL EP tube and add 3mL DMEM / F12 medium. Wash the obtained pig lung tissue with Hanks buffer 5 times, cu...

Embodiment 2

[0055] Establishment of human lung granule in vitro culture model method for screening anti-influenza virus drugs and their anti-inflammatory drugs

[0056] 1. Experimental materials

[0057] 1.1 Human lung tissue and viruses

[0058] The human lung tissue was obtained from Zhongnan Hospital of Wuhan University, and it was fragmentary lung tissue that was partially surgically removed from lung cancer tissue. Influenza strain A / PuertoRico / 8 / 34 (H1N1) was stored frozen at -80°C.

[0059] 1.2 Reagents

[0060] DMEM / F12 medium was purchased from GIBCO; 2'-(4-methylumbelliferyl)-α-D-acetylneuraminic acid (MUNANA) was purchased from Sigma. AlphaLISA inflammatory factor detection kits for human IL-6, IL-8, IP-10, MCP-1 and CCL-5 were purchased from Perkin Elmer.

[0061] 1.3 Instruments and equipment

[0062] Ophthalmic scissors, ophthalmic forceps, Perkin Elmer Wallac Envision multi-label reader, 37°C CO 2 Humidified incubator.

[0063] 2. Experimental methods and results

...

Embodiment 3

[0073] Application of human lung granule in vitro culture model in the screening and evaluation of anti-influenza virus and anti-inflammatory drugs

[0074] 1. The materials, reagents and equipment used in this experiment are as above, and the anti-influenza virus drugs and anti-inflammatory drugs used are: Ribavirin was purchased from Sigma-Aldrich Company; EGCG was purchased from Sichuan Weikeqi Biotechnology Co., Ltd.

[0075] 2. To verify the effect of drugs on this model: After human lung granule tissue was cultured at 37°C for 3 days, use 200 microliters per well, 10 6 TCID 50 / mL of PR8(H1N1) for 2 hours. After 2 hours, the virus solution was discarded, washed twice with PBS, and the drug solution after gradient dilution was added. In order to compare the levels of influenza virus and inflammatory factors in the drug-dosed group and the control group, determine the antiviral and anti-inflammatory effects of the drug. One column was set as negative control wells, one ...

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Abstract

The invention belongs to the technical field of medicine, and specifically discloses construction and applications of an anti-influenza-virus or anti-inflammatory drug screening model based on in vitro culture of human lung particles. According to the invention, lung tissue is cut into small particles with a size of 0.05-0.2 mm<3>, the small particles are uniformly sub-packaged in the form of a suspension, and the sub-packaged small particles are infected with influenza virus after in vitro culture is performed for a certain time; the experiment results prove that influenza virus can replicateefficiently in pig lung particle tissue and human lung particle tissue, and initiate the up-regulation of a series of inflammatory factors, and the known anti-influenza-virus drugs and the known anti-inflammatory drugs can inhibit influenza virus and inflammatory factors in the constructed lung particle tissue models and have dose effects; the construction method has characteristics of simple andrapid operation, time saving and labor saving, and the constructed model can substantially retain the tissue activity and reduce the waste of tissue, and is close to the physiological conditions of animals; and after the method is optimized, the anti-influenza virus or anti-inflammatory drug screening model can achieve the micro, rapid and uniform effect, and can accurately evaluate drugs.

Description

technical field [0001] The invention belongs to the technical field of medicine, and relates to model construction and application of human lung tissue cultured in vitro in the form of particles and used for screening anti-influenza virus or anti-inflammatory drugs. Background technique [0002] Influenza viruses seriously threaten human health and have caused huge losses worldwide. In the 1918 influenza pandemic, about 500 million people were infected worldwide, resulting in 50-100 million deaths. The swine flu outbreak in 2009 spread to more than 200 countries and killed about 18,000 people. In 2013, the emergence of highly pathogenic avian influenza H7N9 in China drew widespread attention to influenza viruses and how to control them. In 2017, the H7N9 virus broke out again and caused nearly 100 deaths in China, with a mortality rate of 45%, causing great panic among people. [0003] For the pathogenic mechanism of influenza virus, there have been two arguments. One is...

Claims

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Application Information

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IPC IPC(8): C12N5/071C12Q1/02
CPCC12N5/0688G01N33/5008G01N33/5044
Inventor 陈绪林张丽
Owner WUHAN INST OF VIROLOGY CHINESE ACADEMY OF SCI
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