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CN-121204157-B - Multisystem atrophy animal model with abnormal blood pressure regulation and construction method thereof

CN121204157BCN 121204157 BCN121204157 BCN 121204157BCN-121204157-B

Abstract

The invention relates to a multisystem atrophy animal model with abnormal blood pressure regulation and a construction method thereof, belonging to the technical field of biology. The invention provides a construction method of a multisystem atrophy animal model with abnormal blood pressure regulation, which comprises the steps of injecting recombinant adeno-associated virus of alpha-synuclein specifically expressed by oligodendrocyte into a brain region of an experimental animal target spot to obtain the multisystem atrophy animal model, wherein the brain region of the multisystem atrophy modeling target spot is a medulla oblongata ventral lateral region. The animal model constructed by the construction method can replicate the expression of orthostatic hypotension in multiple system atrophy, and has typical pathological characteristics and behavioral characteristics of multiple system atrophy. Therefore, the construction method and the animal model obtained by the construction method have wide application prospects in research of multisystem atrophy and screening of related therapeutic drugs.

Inventors

  • QIN CHUAN
  • ZHANG LING
  • ZHU XUCHAO
  • DOU CHANGSONG

Assignees

  • 中国医学科学院医学实验动物研究所

Dates

Publication Date
20260508
Application Date
20251126

Claims (9)

  1. 1. A construction method of a multisystem atrophy animal model capable of simulating significant reduction of pressure difference of orthostatic hypotension is characterized by comprising the steps of injecting recombinant adeno-associated virus expressing alpha-synuclein into a multisystem atrophy modeling target brain region of an experimental animal to obtain the multisystem atrophy animal model, wherein the multisystem atrophy modeling target brain region is a medullary head ventral outside region, and the experimental animal is a mouse.
  2. 2. The construction method according to claim 1, wherein the recombinant adeno-associated virus drives a gene encoding an alpha-synuclein using a PLP promoter as a promoter element, and the nucleotide sequence of the PLP promoter is shown in SEQ ID NO. 1.
  3. 3. The construction method according to claim 1 or 2, wherein the recombinant adeno-associated virus expresses a gene encoding an α -synuclein with an adeno-associated virus as a vector, and the nucleotide sequence of the gene encoding the α -synuclein is shown in SEQ ID No. 2.
  4. 4. The method of claim 3, wherein the recombinant adeno-associated virus further expresses a gene encoding a tag peptide and a reporter gene using the adeno-associated virus as a vector.
  5. 5. The method of any one of claims 1 to 4, wherein the recombinant adeno-associated virus is injected into the animal at a dose of 10 9 ~10 10 vg/animal.
  6. 6. The method according to any one of claims 1 to 5, wherein the injection rate of the recombinant adeno-associated virus in the animal is 0.1 to 0.4 μl/min.
  7. 7. The method of any one of claims 1-6, wherein the recombinant adeno-associated virus is delivered to the target brain region by brain stereotactic injection.
  8. 8. A method for screening a drug capable of preventing and/or treating multiple system atrophy is characterized in that the drug to be screened is used for taking the multiple system atrophy animal model constructed by the construction method according to any one of claims 1-7 before, during and/or after molding, and the effectiveness of the drug to be screened in preventing and/or treating multiple system atrophy is judged according to the change of the multiple system atrophy animal model in pathological characteristics and behavior characteristics after the drug taking is finished.
  9. 9. The application of the multisystem atrophy animal model constructed by the construction method of any one of claims 1-7 in screening medicines capable of preventing and/or treating multisystem atrophy.

Description

Multisystem atrophy animal model with abnormal blood pressure regulation and construction method thereof Technical Field The invention relates to a multisystem atrophy animal model with abnormal blood pressure regulation and a construction method thereof, belonging to the technical field of biology. Background Multisystem atrophy (Multiple system atrophy, MSA) is a neurodegenerative disease with rapid onset and progression in adulthood and fatal properties, mainly involving autonomic nerve function, motor coordination and extrapyramidal function, and its central pathological features are cytoplasmic inclusion bodies (glial cytoplasmic inclusion, GCI) formed by abnormal aggregation of α -synuclein (α -synuclein) in oligodendrocytes. Numerous studies have shown that pathological deposition of α -syn is closely related to the pathogenesis of MSA. The symptoms of MSA are complex and various, and can be classified into extrapyramidal systems, pyramidal systems, cerebellar ataxia and autonomic nerve dysfunction according to clinical manifestations. Among them, autonomic dysfunction, especially orthostatic hypotension, is present in about 78% of MSA patients, has a significant impact on the prognosis of MSA patients, severely affects the quality of daily life of MSA patients, and 32.4% of orthostatic low blood pressure severe patients are at risk of disabling death. However, currently, there is no effective therapeutic drug or method for treating the symptom of orthostatic hypotension in MSA. Thus, at present, there is still a need for extensive basic research through animal models capable of rescuing the manifestations of orthostatic hypotension in multiple system atrophy to obtain potential therapeutic drugs or treatments. Currently, there are three major classes of animal models used to study MSA, including toxin induction models, exogenous protein models, and gene editing models. The first two models are not specific in terms of the disease, and can partially mimic the pathological features of parkinsonism and MSA dyskinesia or α -syn. The professor Erwan, france, tried to target the substantia nigra-striata brain region and set up an alpha-syn pathology model using a brain stereotactic local injection method. Local injection (injection of adeno-associated virus vector or exogenous protein) of the team concentrates brain region targets of the MSA model into a substantia nigra-striata loop, and can only reflect abnormal phosphorylation and aggregation of oligodendrocyte alpha-syn of MSA in pathology, behavioural shows dyskinesia with mild bradykinesia, and cannot reproduce the important clinical manifestation of human orthostatic hypotension. Therefore, it is highly desirable to find an animal model and a construction method thereof capable of effectively rescuing the manifestation of orthostatic hypotension in multiple system atrophy and having typical pathological and behavioral characteristics of multiple system atrophy, for clinical study of multiple system atrophy and its concurrent diseases. In addition, the adeno-associated viral vector promoter used in the existing methods for constructing animal models of MSA is myelin basic protein (myelin basic protein, MBP), which accounts for about 30% of myelin total protein, and the target cells capable of infection are limited, and this problem is also in need of solution (references "Teil M, Dovero S, Bourdenx M, et al. Brain injections of glial cytoplasmic inclusions induce a multiple system atrophy-like pathology. Brain. 2022;145(3):1001-1017."、"Kaji S, Maki T, Ishimoto T, Yamakado H, Takahashi R. Insights into the pathogenesis of multiple system atrophy: focus on glial cytoplasmic inclusions. Transl Neurodegener. 2020;9:7." and MBP "Bassil F, Guerin PA, Dutheil N, et al. Viral-mediated oligodendroglial alpha-synuclein expression models multiple system atrophy. Mov Disord. 2017;32(8):1230-1239."）. Disclosure of Invention In order to solve the problems, the invention provides a construction method of a multisystem atrophy animal model with abnormal blood pressure regulation, which comprises the steps of injecting recombinant adeno-associated virus expressing alpha-synuclein into a multisystem atrophy modeling target brain region of an experimental animal to obtain the multisystem atrophy animal model, wherein the multisystem atrophy modeling target brain region is a medullary head ventral outside region (rostral ventrolateral medulla, RVLM). In one embodiment of the invention, the recombinant adeno-associated virus drives a gene encoding alpha-synuclein by using a PLP promoter as a promoter element, wherein the nucleotide sequence of the PLP promoter is shown as SEQ ID NO. 1. In one embodiment of the invention, the recombinant adeno-associated virus expresses a gene encoding alpha-synuclein by taking adeno-associated virus as a vector, and the nucleotide sequence of the gene encoding alpha-synuclein is shown as SEQ ID NO. 2. In one embodiment of the present in