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Method for evaluating lung injury therapeutic agent and inductive agent by use of zebra fish bladder injury inflammation model

An inflammation model, zebrafish technology, applied in the direction of material inspection products, measuring devices, instruments, etc., can solve problems such as drug evaluation of inflammation models that have not yet been seen, and achieve the effect of simple experimental process operation, less dosage, and reliable predictability

Active Publication Date: 2017-02-22
HANGZHOU HUANTE BIOLOGICAL TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In 2013 and 2014, Cass and Robertson et al. respectively detected pulmonary surfactant in the zebrafish swim bladder [21,23] Although it has been reported in the literature that the zebrafish swim bladder can be applied to the basic research of some human diseases, for example, in 2013, Gratacap et al. explored the relationship between Candida albicans, epithelial cells and immunity by studying the response of zebrafish to the swim bladder infected by Candida albicans. The interaction between the three cells; in 2015, Voelz et al. explored the interaction between mucormyces spores and the body's immune system through the response of zebrafish to mucormyces spores infecting the posterior ventricle and swim bladder of zebrafish; however, so far, There have been no reports on the use of the zebrafish swim bladder as an animal model for the study of lung diseases
Therefore, the current methods for evaluating therapeutic agents and inducers of lung injury are still based on traditional experimental animal models (such as mice, dogs, etc.), and there are no literature reports on drug evaluation based on the zebrafish swim bladder injury inflammation model.

Method used

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  • Method for evaluating lung injury therapeutic agent and inductive agent by use of zebra fish bladder injury inflammation model
  • Method for evaluating lung injury therapeutic agent and inductive agent by use of zebra fish bladder injury inflammation model
  • Method for evaluating lung injury therapeutic agent and inductive agent by use of zebra fish bladder injury inflammation model

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0086] Example 1 Determining the best developmental stage of zebrafish

[0087] 1 Zebrafish selection

[0088] Take 4 to 5 pairs of neutrophil green fluorescence transgenic strain zebrafish (MPO) parents to mate, according to Westerfield [26] method for hatching embryos. When the zebrafish develops to 4.5dpf, the swim bladder has been inflated and formed. The 4.5dpf, 5dpf, and 6dpf zebrafish were observed under a dissecting microscope, and the normally developed zebrafish were picked and transferred to three 6-well plates, with 30 fish in each well. (Note: dpf=day post fertilization among the present invention, Chinese refers to the number of days after fertilization of zebrafish, as 5dpf refers to five days after fertilization of zebrafish.)

[0089] 2 Inducer LPS treatment

[0090] Set up 3 experimental groups (4.5dpf, 5dpf, 6dpf zebrafish in each group), each experimental group includes 1 inducer treatment group, 1 solvent control group and 1 blank control group. In th...

Embodiment 2

[0100] Example 2 Determining the optimal treatment time length and injection dose of the inducer LPS

[0101] 1 Zebrafish selection

[0102] The 5dpf zebrafish were observed under a dissecting microscope, and the normally developed zebrafish were picked and transferred to five 6-well plates, 30 in each well.

[0103] 2 Compound treatment

[0104] Set up 6 experimental groups: 4 inducer LPS treatment groups, 1 solvent injection group, and 1 blank control group. Different doses of LPS (lipopolysaccharide) were injected into the swim bladder cavity of the zebrafish in the inducer group. LPS was injected into the swim bladder (5nL respectively); the solvent injection group was injected with 5nL PBS into the swim bladder; the zebrafish of the blank control group did not receive any treatment. Each experimental group was added with the same volume of aquaculture water according to the specifications of the microplate, and the microplate was placed in a constant temperature incuba...

Embodiment 3

[0130] Example 3 Construction method of zebrafish swim bladder injury inflammation model

[0131] Based on the optimal developmental stage of zebrafish, the optimal treatment time of the compound and the injection dose in Examples 1 and 2, an inflammatory model of zebrafish acute lung injury was established by optimizing the concentration of LPS. The design scheme is as follows:

[0132] 1 Zebrafish selection

[0133] The 5dpf zebrafish were observed under a dissecting microscope, and the normally developed zebrafish were picked and transferred into six-well plates, 30 in each well.

[0134] 2 Inducer LPS treatment

[0135] Set up 3 experimental groups: 1 inducer treatment group, 1 solvent injection group, and 1 blank control group. The zebrafish swim bladder of the inducer group was injected with 50ng LPS (5nL at a concentration of 10mg / mL); the solvent control group was injected with 5nL PBS; the zebrafish of the normal control group were not treated. Each experimental g...

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Abstract

The invention provides a method for establishing a zebra fish bladder injury inflammation model and a method for evaluating efficacies of a lung injury therapeutic agent and inductive agent by use of the zebra fish bladder injury inflammation model. The zebra fish bladder injury inflammation model provide by the invention has the advantages of simplicity and convenience, rapidness, economical efficiency, high efficiency, high flux and the like, can accurately reflect the real in-vivo condition of the lung injury therapeutic agent or inductive agent, and can realize the in-vivo high-flux evaluation on the efficacy of a medicament in treating acute lung injury inflammation, and the method for evaluating the efficacy of the medicament by use of the zebra fish bladder injury inflammation model has the advantages of reliability, rapidness, high efficiency, low cost, high cost performance and the like, and has great significance.

Description

technical field [0001] The invention belongs to the technical field of drug evaluation and screening, and in particular relates to the application of a simple, economical and high-throughput zebrafish swim bladder injury inflammation model, especially a method for evaluating the efficacy of a lung injury therapeutic agent and the toxicity of an inducer using the zebrafish model . Background technique [0002] Acute and chronic lung diseases, especially lung injury, lung inflammation, and lung tumors are one of the diseases that seriously endanger human health. Atmospheric fine particulate matter (PM 2.5 ), smoking, radiation, chemotherapy and other physical and chemical factors are risk factors for acute and chronic lung injury and lung cancer, among which PM 2.5 Chronic obstructive pulmonary injury caused by smoking and other causes is an incompletely reversible, airflow-limiting respiratory disease that produces an abnormal inflammatory response to noxious particles and ...

Claims

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Application Information

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IPC IPC(8): G01N33/50
CPCG01N33/5088
Inventor 刘洪翠柯越海朱晓宇张雪黄燕烽程洪强李春启
Owner HANGZHOU HUANTE BIOLOGICAL TECH CO LTD
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