Method of preparing osteoporosis model of zebra fish and alizarin red dyeing method thereof

A technology of osteoporosis and zebrafish, applied in the field of animal models of osteoporosis, can solve the problems of not being able to simulate osteoporosis well, zebrafish embryo teratogenicity, and high lethality, and achieve obvious characteristics of osteoporosis, Effects of increased total bone mass damage and decreased expression

Active Publication Date: 2017-05-31
MINNAN NORMAL UNIV
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  • Abstract
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Problems solved by technology

[0004] At present, prednisolone and dexamethasone are the main drugs used in the construction of zebrafish osteoporosis models, but the above two drugs are glucocorticoid drugs, and it is very easy to use them due to inaccurate doses during model building. Excessive immunosuppression leads to teratogenicity and high lethality in zebrafish embryos
At present, some studies have found that deferoxammonium or hepcidin can repair the skeletal development defects of zebrafish caused by high iron, b

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  • Method of preparing osteoporosis model of zebra fish and alizarin red dyeing method thereof
  • Method of preparing osteoporosis model of zebra fish and alizarin red dyeing method thereof
  • Method of preparing osteoporosis model of zebra fish and alizarin red dyeing method thereof

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[0058] Example 1

[0059] 1. Establish a staining method for zebrafish juvenile Alizarin Red:

[0060] (1) Prepare the reagents needed for dyeing: a) Alizarin red solution: weigh 0.5g of Alizarin Red powder dissolved in 100ml of 95% ethanol, the final concentration is 0.5%; b) 0.7% KOH solution: weigh 0.7g The KOH solid was dissolved in 100 ml of distilled water and prepared into a 0.7% KOH solution.

[0061] (2) The specific steps of Alizarin Red staining:

[0062] a) Sample collection and fixation: Collect the juveniles on the 6th day after fertilization, put the juveniles in a 1.5ml EP tube, and fix them in 4% PF at 4°C overnight, not more than one week;

[0063] b) Depigmentation: remove 4% PF, wash 3 times with PBST for 5 min each time; add depigmentation solution (0.5%KOH / 3%H 2 O 2 ) Until the melanin and yellow pigment are not visible by naked eyes, replace the solution for 1 to 2 times as needed. The whole process takes 30min to 1h. In order to prevent the tube from expanding a...

Example Embodiment

[0074] Example 2

[0075] 1. Ferric ammonium citrate induces the establishment of a zebrafish juvenile osteoporosis model

[0076] Refer to the following method to establish an osteoporosis model of zebrafish juveniles induced by ferric ammonium citrate: Ferric ammonium citrate (FAC) is used as a model drug, and the juveniles 48h after fertilization are exposed to different concentrations of FAC solution. FAC concentrations are 50, 100, 200, 500 μg·ml -1 , While setting up ordinary culture medium (Holfreter H 2 0) is the control group, cultured in a 6-well plate at 28.5°C for 4 days until the zebrafish skull is formed, during which the dressing is changed every 12h.

[0077] 2. Evaluation of the osteoporosis model of juvenile zebrafish

[0078] (1) Statistics of teratogenicity and lethality: After modeling, the teratogenicity and lethality of zebrafish juveniles are counted, and the results are shown in Table 1. It can be seen that when using ferric ammonium citrate for modeling, the...

Example Embodiment

[0090] Example 3

[0091] Alendronate can well repair FAC-induced osteoporosis in juvenile fish

[0092] Juvenile repair: Collect the zebrafish juveniles 48h after fertilization and randomly divide them into 4 groups, each with at least 30 pieces: the control group is cultured in normal fish water until the 10th day; the model is 500μg·ml -1 FAC treatment to the 10th day; the negative drug control group is FAC treatment for 4 days, then change to H 2 O was cultured for 4 days; the positive drug repair group was FAC treatment for 4 days, and then replaced with 0.1% positive drug alendronate for 4 days. The 4 groups of juvenile fish on the 10th day after treatment were collected and stained with Alizarin Red, Calcein and Alcian Blue to detect the repair of hard bone and cartilage development. The results are as follows Figure 5 Shown. Experimental results show that whether it is from bone density, degree of mineralization or cartilage development, alendronate can perfectly repair th...

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Abstract

The invention provides a method of preparing an osteoporosis model of a zebra fish and an alizarin red dyeing method thereof. According to the modeling method provided by the invention, a juvenile zebra fish or an adult zebra fish is treated by using a high concentration Fe<3+> solution to obtain the osteoporosis model. The model has obvious and prominent characteristics of osteoporosis, and can be used for screening various drugs for treating osteoporosis in a broad spectrum. The dyeing method provided by the invention optimizes the alizarin red dyeing method of the juvenile zebra fish or the adult zebra fish, and the dyeing effect of the zebra fish, especially the adult zebra fish, is improved, so that the adult zebra fish can be used for preparing the osteoporosis model.

Description

technical field [0001] The invention relates to the field of animal models of osteoporosis, in particular to a method for preparing a zebrafish osteoporosis model and an alizarin red staining method thereof. Background technique [0002] Osteoporosis is a group of disorders characterized by decreased bone mass, abnormal bone microstructure and increased risk of fracture. This disease is a common and frequently-occurring disease of the elderly. With the acceleration of population aging process, the incidence rate of osteoporosis is getting higher and higher. Therefore, constructing an effective osteoporosis drug screening model is of great significance for the drug treatment of osteoporosis and the discovery of safe and effective new drugs. [0003] At present, the animals used to prepare osteoporosis models mainly include rats, mice, and zebrafish; due to their small size, fast growth cycle, and low feeding and maintenance costs, compared with rats and mice, zebrafish is th...

Claims

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Application Information

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IPC IPC(8): A61K33/26A61K31/295A61K49/00A01K61/10
CPCA01K61/00A61K33/26A61K49/0008Y02A40/81
Inventor 薛钰潘裕添张文娟许璟瑾李秀敏
Owner MINNAN NORMAL UNIV
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