Construction method of neural tube defect model and application of construction method

A neural tube defect and construction method technology, applied in the field of animal model construction, can solve problems such as unclear neural tube defects

Active Publication Date: 2018-11-23
首都儿科研究所
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Problems solved by technology

Many studies have found that elevated homocysteine ​​(homocysteine, HCY) levels caused by abnormal folic acid metabolism is one of the im

Method used

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  • Construction method of neural tube defect model and application of construction method
  • Construction method of neural tube defect model and application of construction method
  • Construction method of neural tube defect model and application of construction method

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Experimental program
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Effect test

Embodiment 1

[0026] The following materials are mainly used in this embodiment:

[0027] White Laihang chicken eggs (China Agricultural University, China); Homocysteine ​​Thiolactone hydrochloride (Sigma, USA); Tyrode Buffer; Phenol Red (Beijing Chemical Plant); Incubator (Showa Furanki, Japan). The main instruments include optical and dissecting microscopes, automatic incubators, micro-injectors, dental drills and egg openers, etc.

[0028] The specific preparation method of the neural tube defect animal model is as follows:

[0029] Collect artificially inseminated White Legion breeder eggs, select eggs with smooth surface, oval shape, and moderate size, wash off the mucus and feces on the surface with water, wash with 0.1% bromogeramine and put them in the incubator for incubation. The incubation condition is the incubation temperature 38°C, 55-65% relative humidity, 90° rotation angle, automatic egg turning at intervals of 30 minutes, and continued incubation. This time is recorded a...

Embodiment 2

[0035] In order to explore the effect of 0.5mM HTL neural groove injection on chicken embryo neural tube defects, the inventors expanded the sample size and injected 0.5mM HTL neural groove with an injection volume of 5 μl. The control group was injected with bench-top liquid containing phenol red. figure 1 shown. according to figure 2 At the key time points of neural tube development shown, embryos were taken for observation at E2 before neural tube closure, E3, E5 and E8 after neural tube closure.

[0036] At the E2 stage of embryonic development, the malformation rate of embryos was 59.1%, with abnormal brain development and abnormal closure of neural grooves, and some embryos were accompanied by abnormal body shape. At E3, brain development was obviously abnormal, and the deformity rate was 50%. Spina bifida, brachycephaly and abnormal body segment development appeared, and the stillbirth rate was 9.09%. At E5, the malformation rate of embryos was 56.6%, mainly brain dy...

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Abstract

The invention discloses a construction method of a neural tube defect model. The construction method comprises the following steps of step 1, collecting artificially inseminated eggs, cleaning the eggs, and putting the eggs in an incubator for incubation; step 2, performing incubation on the eggs until neural plates of chicken embryos start to close, adopting a chicken embryo neural groove injection method to inject homocysteine thiolactone into the eggs; and step 3, performing continuous incubation on the treated eggs, observing phenotypes of the chicken embryos, and screening a successful neural tube defect model. On the other hand, the invention also discloses application of the construction method in studying pathogenesis of neural tube defects caused by hyperhomocysteinemia. By adopting the construction method of the neural tube defect model, the deformity rate of a nervous system reaches 50 percent, the survival rate of the embryos reaches up to 87.5 percent, and the constructionmethod is a best model making method of observing the neural tube defects at present.

Description

technical field [0001] The invention relates to the technical field of animal model construction, in particular to a construction method and application of a neural tube defect model. Background technique [0002] Neural tube defects (NTDs) are serious congenital malformations of the central nervous system, and the incidence rate in northern my country, especially in Shanxi Province, can be as high as 10.2‰. The etiology of neural tube defects is complex. Many studies have shown that neural tube defects are caused by the interaction of genes and environmental factors, such as environment, nutrition, metabolism, organic solvents, etc., are closely related to the occurrence of neural tube defects. Many studies have found that elevated homocysteine ​​(homocysteine, HCY) levels caused by abnormal folic acid metabolism is one of the important risk factors for birth defects, and the mechanism of high homocysteine ​​leading to neural tube defects is still unclear. unclear. [000...

Claims

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Application Information

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IPC IPC(8): A61K31/381A61P25/00
CPCA61K31/381A61P25/00
Inventor 张勤张霆白宝玲李丹万春雷
Owner 首都儿科研究所
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