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Novel method for evaluation of osteoporosis prevention treatment drug effect

A technology for drug action and osteoporosis, applied in the direction of testing drug preparations, measuring devices, instruments, etc., can solve the problems of single action link of in vitro cell models, inability to efficiently evaluate activity, and difficulty in isolating sufficient compounds, etc. The effect of small amount of test drug, overcoming harsh experimental conditions, and strong operability

Inactive Publication Date: 2013-03-20
JIANGSU PROVINCE INST OF TRADITIONAL CHINESE MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the activity evaluation of bone-strengthening traditional Chinese medicine and its components, especially trace components, cannot be carried out efficiently. There are two bottlenecks: one is the evaluation model problem: the existing animal bone-strengthening model takes a long time, requires a large amount of samples, and is labor-intensive. The efficiency of activity screening is low; the cost of existing cell models is high, the requirements for test conditions are high, and the difficulty of primary culture is increased, which cannot be carried out in general laboratories. In addition, the function of in vitro cell models is relatively single, which cannot reflect the comprehensive effect of in vivo tests. ; The second is the problem of the number of compounds: due to the complex composition of traditional Chinese medicine, it is difficult to separate enough compounds to meet the needs of in vivo research, and a large number of trace components can only be screened by cell models
Domestic research in this area is still blank

Method used

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  • Novel method for evaluation of osteoporosis prevention treatment drug effect
  • Novel method for evaluation of osteoporosis prevention treatment drug effect
  • Novel method for evaluation of osteoporosis prevention treatment drug effect

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] Dexamethasone induces osteoporosis in zebrafish

[0057] 1. Materials and Instruments

[0058] Test drug: dexamethasone

[0059] Preparation method: Accurately weigh an appropriate amount of dexamethasone, add a small amount of DMSO to dissolve, and prepare 0.5% DMSO solutions of 1 μM, 2.5 μM, 10 μM and 25 μM with medium.

[0060] Experimental animals: adult zebrafish (Institute of Model Animals, Nanjing University).

[0061] Reagents: dexamethasone (sigma-aldrich, batch number: 077K10501; ≥97%); Alizarin Red S (Zhengzhou Siji Chemical Products Co., Ltd., batch number: Sj20110806); paraformaldehyde (Chengdu Kelong Chemical Reagent Factory, batch number: 20100504); MS-222 (Acros Organics, lot number: A0288328).

[0062] Instruments: ZEISS fluorescence inverted microscope Axio observer.A1 (ZEISS Optical Instrument International Trade Co., Ltd.); biochemical incubator SPX-80 (Ningbo Haishu Saifu Experimental Instrument Factory).

[0063] Related software: image acquisi...

Embodiment 2

[0081] Predney Songlong induces osteoporosis in zebrafish

[0082] 1. Materials and Instruments

[0083] Drug tested: Prednisolone

[0084] Preparation method: Accurately weigh an appropriate amount of prednisolone, add a small amount of DMSO to dissolve, and prepare 0.5% DMSO solutions of 1 μM, 2.5 μM, 10 μM and 25 μM with medium.

[0085] Reagent: Prednisolone (Suzhou Yake Chemical Reagent Co., Ltd., batch number: YK2012020101); Alizarin Red S, paraformaldehyde and MS-222 are the same as in Example 1.

[0086] Experimental animals and instruments are the same as in Example 1.

[0087] Relevant software is the same as embodiment one.

[0088]

[0089] 2. Experimental method

[0090] 2.1 Administration

[0091] Put zebrafish embryos at 3 dpf (3 days after fertilization) into a 24-well plate filled with embryonic medium, add 1 mL of medium to each well, put 5 juveniles in each well, 10 juveniles / group / 2 Wells were divided into model drug group (prednisolone) and vehicl...

Embodiment 3

[0104] Effects of etidronate disodium on zebrafish skeleton

[0105] 1. Materials and Instruments

[0106] Test drug: etidronate disodium

[0107] Preparation method: Accurately weigh etidronate disodium and place it in a 10 mL volumetric flask, add a small amount of DMSO to dissolve and constant volume, and prepare 2.5 μg / mL, 15 μg / mL, and 30 μg / mL of Etidronate with medium Tidronate disodium solution.

[0108] Reagent: disodium etidronate (National Institute for the Control of Pharmaceutical and Biological Products, batch number: 101174-201001); Alizarin Red S, paraformaldehyde and MS-222 are the same as in Example 1.

[0109] Experimental animals and instruments are the same as in Example 1

[0110] Related software: image acquisition software: AxioVisionRel.4.6, image analysis software: Image J.

[0111] 2. Experimental method

[0112] 2.1 Administration

[0113] Put 3 dpf (3 days after fertilization) zebrafish embryos into 24-well plates filled with embryo medium, d...

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Abstract

The present invention discloses a novel method for model organism zebrafish evaluation of osteoporosis prevention treatment drug effects. The method includes selecting zebrafish as a model animal, grouping and designing experiments by an optimized experimental grouping method, administering by using a preferable drug administration method, using a preferable method to dye zebrafish bones with alizarin red, capturing images by intuitive microscopic observation, and analyzing bone dyed areas to reflect the amount of bone mineralization by using a preferable image analysis software. The entire experimental method is high in operability, can objectively reflect the effects of the drug in zebrafish in vivo on bone mass generation, and can quickly and accurately evaluate the effects of compounds and complex ingredients of traditional Chinese medicine in the prevention and treatment of osteoporosis. The method can overcome the shortcomings that ordinary in-vitro cell experiments are rigor in experimental conditions and difficult to reflect body effect comprehensive results; and overcomes shortcomings that mammal in-vivo experiments are time-consuming, large amount of drug are tested, and labor intensity is high. The method is high in accuracy of experimental results, the experimental method is high in repeatability, the required drug to be tested is relatively less, the cost is low, labor intensity is low, application range is wide, the experimental studies can be conducted in batch, and the working efficiency is high.

Description

technical field [0001] The invention relates to a new method for evaluating the effect of drugs for preventing and treating osteoporosis by model organisms, in particular to a new method for evaluating the effects of drugs for preventing and treating osteoporosis by using model organisms zebrafish. Background technique [0002] Activity screening has always been a key link in drug discovery, and it is also an indispensable means in key fields such as the material basis, mechanism of action, quality control, processing mechanism, and compatibility of traditional Chinese medicine. The research on new models and methods for activity screening is an important part of Chinese medicine research. part. [0003] Osteoporosis is a kind of disease characterized by decreased bone mass, abnormal microstructure of bone tissue and increased risk of fracture. Due to the disadvantages of long-term use of western medicine, such as side effects, the efficacy of traditional Chinese medicine in...

Claims

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Application Information

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IPC IPC(8): G01N33/15G01N21/00
Inventor 韦英杰贾晓斌王长梅蔡雪婷詹扬孙娥舒娈宋捷
Owner JIANGSU PROVINCE INST OF TRADITIONAL CHINESE MEDICINE
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