Use of acetylchloinesterase inhibitor for preparing medicine for treating parkinson's disease

A technology for acetylcholinesterase and Parkinson's disease, applied in the treatment of Parkinson's disease, in the field of acetylcholinesterase inhibitors, can solve the problems of inability to use and not very obvious effect, etc.

Inactive Publication Date: 2005-07-20
上海国联干细胞技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Since one of the pathogenesis of PD has always been believed to be caused by the concentration imbalance between dopamine and acetylcholine in the nerve center, only anticholinergic drugs are used clinically instead of acetylcholinesterase (AChE) inhibitors to treat PD patients. The effect of this treatment is not very obvious, and it cannot be used for dementia symptoms and elderly patients

Method used

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  • Use of acetylchloinesterase inhibitor for preparing medicine for treating parkinson's disease
  • Use of acetylchloinesterase inhibitor for preparing medicine for treating parkinson's disease
  • Use of acetylchloinesterase inhibitor for preparing medicine for treating parkinson's disease

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0191] Example 1 Detection of Acetylcholinesterase Expression in Apoptotic Cells in Vitro (Histochemical Method)

[0192] Cell culture: SK-N-SH cells, CHO cells: at 37.5°C, 5% CO 2 Under the moist conditions of the above, the cells were cultured in the RMBI-1640 (Gibco) medium of 10% calf serum (Huamei), adding HEPES and glutamine, 100IU / ml ampicillin, 100μg / ml streptomycin; the cells were treated with Subculture at a split ratio of 1:10 every week, culture for 8-12 days, pass for about 4-20 generations, and inoculate in 48-well culture plates (Costar) 2 days in advance until they are full and ready for use. A DAT-CHO cell line permanently expressing the rat dopamine transporter in a CHO cell line (designated D8 cell line). D8 was the same as CHO cell culture, except that Geneticin (G418) 400 μg / ml (Gibco / BRL) was added; SH-SY5Y cell culture used DMEM / F12 (Gibco) instead of RMBI-1640, and other conditions remained unchanged.

[0193] Induction of dopaminergic neurocytotoxici...

Embodiment 2

[0207] Example 2 Detecting the expression of acetylcholinesterase in dopaminergic neurotoxicity-induced cells from the mRNA level

[0208] The induction method is the same as above, collect the cells induced by dopamine and 6-hydroxydopamine, take the cells induced by serum culture as positive control and the uninduced normal cells as negative control; take 7×10 6 After the cells were lysed with 1ml Trizol, the total RNA was extracted with chloroform, isopropanol, and 75% ethanol; 2 μg of total RNA was taken to a 20 μl reaction system, and random primers were used to perform reverse transcription to generate cDNA; design upstream primers and Downstream primers: 5'-CGGGTCTACGCCTACGTCTTTGAACACCGTGCTTC-3' and 5'-CACAGGTCTGAGCAGCGATCCTGCTTGCTG-3'.

[0209] The product length is: 482bp. Use PE480 type PCR instrument, PCR reaction conditions: denaturation: 94°C, 60sec; annealing: 60°C, 70sec; extension 72°C, 90sec; 30 cycles; β-actin as internal reference.

[0210] result:

[021...

Embodiment 3

[0212] Example 3 MTT method to detect cell viability

[0213] SH-SY5Y cells were diluted to an appropriate concentration and inoculated in a 96-well plate in a volume of 200 μl. Induced treatment after 24 hours of culture. After dopamine induction, change the medium, add 20 μl (5 mg / ml) of MTT, 37 ° C for 4 hours, remove the medium, add 150 μl of dimethyl sulfoxide, shake and dissolve for 15 minutes, measure the O.D. value at 590 nm with a microplate reader, The group was repeated at least 3 times, and the average value was taken.

[0214] result:

[0215] Dopamine (200 μM) induces a large number of cell death, while the addition of huperzine A (1 μM) can partially prevent cell death, and there is a significant difference (p0.05), therefore, the acetylcholinesterase inhibitor in vitro could partially inhibit the cell death caused by dopaminergic neurotoxicity ( Figure 7 ).

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Abstract

The present invention discloses an application of acetylcholinesterase inhibitor in the preparation of medicine for curing Parkinson's disease. The described inhibitor includes the compounds which can inhibit activity of acetylcholinesterase and have the action of curing Parkinson's disease. Said inhibitor can effectively improve motion defect resulted from Parkinson's disease and can protect dopaminergic neuron from injury.

Description

technical field [0001] The invention relates to the field of biomedicine, in particular to the application of an acetylcholinesterase inhibitor in the treatment of Parkinson's disease. Background technique [0002] Parkinson's disease (PD) is an senile degenerative neurological disease in which dopaminergic neurons in the substantia nigra of the midbrain are mainly damaged, and it develops progressively. In the early clinical stage, extrapyramidal motor impairment is the main feature: bradykinesia, resting tremor, muscle rigidity, and postural reflex disorder are the clinical features. As the disease progresses, cognitive impairment, including dementia, depression, and anxiety, may appear. For patients with Parkinson's disease accompanied by dementia, it can account for 30-60%, and even a survey shows that up to 93% of patients have dementia; dopamine replacement therapy is effective; the age of clinical onset is generally around 60 years old, as early as 40 years old Sympt...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/44A61P25/16
Inventor 郭礼和栗超跃暨荀鹤惠国桢
Owner 上海国联干细胞技术有限公司
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