A method for generating a ho-1/app/psen1 triple transgenic mouse model of Alzheimer's disease

A technology of Alzheimer's disease and PSEN1, applied in the field of biotechnology, can solve the problems of incompatible AD pathological mechanism and lack of therapeutic effect.

Active Publication Date: 2018-11-30
HARBIN MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the APP / PSEN / TAU triple transgenic animal is currently the best AD model and has a lot of application value, it does not conform to the real pathological mechanism of AD
If such a model is used to study the pathogenesis of AD, conclusions that are inconsistent with reality may be drawn. Using it to develop therapeutic targets or evaluate preclinical drug efficacy may not receive the expected curative effect

Method used

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  • A method for generating a ho-1/app/psen1 triple transgenic mouse model of Alzheimer's disease
  • A method for generating a ho-1/app/psen1 triple transgenic mouse model of Alzheimer's disease
  • A method for generating a ho-1/app/psen1 triple transgenic mouse model of Alzheimer's disease

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] Embodiment 1: Construction of Tg (mHO-1) transgenic mouse model

[0053] 1. Obtain the target gene:

[0054] Primers containing EcoRI restriction endonuclease sites were designed to amplify the cDNA sequence of heme oxygenase 1 (mHO-1, Gene ID: 15368) gene in the mouse genome, using EcoRI restriction The amplified product was digested with an endonuclease, and the digested product was inserted into the EcoRI site of the pCAGG vector to obtain a pCAG recombinant vector containing the cDNA of Hmox1, which was named pCAG-mHO-1. mHO-1 cDNA gene is initiated by chickenβ-actin promoter and CMV enhancer, and mHO-1 is connected with rabbitβ-globin poly(A) termination signal. The pCAG-mHO-1 plasmid was digested with SalI and DraI to obtain the linear transgene sequence, and the structure diagram of the constructed Tg(mHO-1) mouse transgene fragment is as follows figure 1 Purified recovery was used for microinjection as indicated.

[0055] 2. Produce fertilized eggs of Tg(mHO-...

Embodiment 2

[0059] Example 2: Generation of Tg(HO-1 / APP / PSEN1) transgenic mice

[0060] Schematic diagram of the method for obtaining Tg(HO-1 / APP / PSEN1) transgenic mice by crossing Figure 5 Shown: the Tg(mHO-1) transgenic mouse constructed in Example 1 was crossed with the APP / PSEN1 double transgenic AD mouse model (Tg(APP / PSEN1), 2TG) to obtain HO-1 / APP / PSEN1 triple transgenic Positive offspring, ie Tg(HO-1 / APP / PSEN1) transgenic mice (3TG). Tg(APP / PSEN1) mice (stock number: 004462) were obtained from the National Jackson Animal Center (Bar Harbor, ME, USA) and introduced by Nanjing University-Nanjing Institute of Biomedicine.

[0061] The heterozygous progeny was extracted from the mouse tail DNA and screened for the heterozygotes of the three transgenes by PCR. The APP / PSEN1 primers (provided by the National Jackson Animal Center of the United States), and the HO-1 primers are shown in Table 1 below:

[0062] Table 1

[0063]

Embodiment 3

[0064] Example 3: Cognitive function characteristics of Tg(HO-1 / APP / PSEN1) transgenic mice

[0065]Morris water maze (Morris R.Developments of a water-maze procedure for studying spatial learning in the rat.J Neurosci Methods.1984May; 11(1):47-60.) is a classic test for detecting the cognitive function of the hippocampus in Alzheimer's disease In the experiment, the learning, memory and spatial cognition abilities of Tg(HO-1 / APP / PSEN1) transgenic mice were evaluated by water maze behavioral experiments. It was verified that the learning and spatial memory abilities of Tg(HO-1 / APP / PSEN1) triple transgenic mice were significantly reduced compared with wild-type mice (WT), and decreased compared with Tg(APP / PSEN1) mice (2TG) more obvious.

[0066] result:

[0067] The detection results of escape latency in the first 5 days of water maze experiment training are as follows: Image 6 As shown, in the water maze experiment, the swimming routes of mice in each group were traced as ...

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Abstract

The invention discloses a method for generating an HO-1 / APP / PSEN1 triple-transgenic Alzheimer disease mouse model. According to the method, a tauP301L mutant gene in an APP / PEEN1 / TAU triple-transgenic model is replaced by the HO-1 through the transgenic technology, the HO-1 / APP / PSEN1 triple-transgenic Alzheimer disease (AD) mouse model is built, and the transgenic mouse conforms to the pathological state of the Alzheimer disease. The HO-1 / APP / PSEN1 transgenic mouse conforms to the characteristic phenotype of the Alzheimer disease in the respects of the cognitive function, brain tissue pathology and marks and can be taken as an Alzheimer disease animal model through the test of the ethology, the morphology and the molecular biology. In addition, the HO-1 / APP / PSEN1 triple-transgenic mouse is the animal module capable of covering the Alzheimer disease phenotype and being conforming to the pathogenetic mechanism of the Alzheimer disease pathology. Compared with an APP / PSEN1 double-transgenic mouse, the Alzheimer disease phenotype of the HO-1 / APP / PSEN1 triple-transgenic mouse is remarkable. The method can be applied to the discussion of pathogenesis of the Alzheimer disease, drug new target discovery and preclinical efficacy evaluation.

Description

technical field [0001] The invention relates to a biological technology, in particular to a method for establishing a biological model, which is also a method for establishing a biological model of a disease. Background technique [0002] Alzheimer's disease (AD) model is an essential tool for studying AD pathogenesis and treatment strategies. Currently, the best way to establish AD model is transgenic technology. [0003] The existing transgenic models of Alzheimer's disease mainly transfer the mutated genes in familial AD (FAD), the most commonly used are: β-amyloid precursor protein (APP) and presenilin (PSEN) mutated genes. [0004] Because the murine Aβ polypeptide is difficult to aggregate at a high level in vivo to form obvious plaques even in an overexpressed state (Jankowsky, J.L., Younkin, L.H., Gonzales, V., Fadale, D.J., Slunt, H.H., Lester, H.A., Younkin, S.G., and Borchelt, D.R. (2007) Rodent Ab modulates the solubility and distribution of amyloid deposits in ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/10C12N15/53C12N15/85A01K67/027
CPCA01K67/0275A01K2217/056A01K2217/30A01K2227/105A01K2267/0312C12N9/0083C12Y114/99003
Inventor 惠洋高旭彭亚会王大勇李冀宏李婧司紫珍周游张竹君王金鑫
Owner HARBIN MEDICAL UNIVERSITY
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