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Method of screening action target of cardiac glycoside medicine using protein composition technology

A proteomics and drug action technology, applied in the field of medicine, can solve the problems of affecting cell signal transduction, can not fully explain the inhibition or activation of Na-KATPase, etc., and achieve the effect of huge economic and social benefits

Inactive Publication Date: 2006-06-28
SHANDONG UNIV
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, it is believed that this type of drug exerts positive inotropic effect through the inhibitory pathway of Na-KATPase on the myocardial cell membrane, but this cannot fully explain the opposite effects such as inhibition or activation of Na-KATPase at different drug concentrations. Recent studies have found that the drug participates in apoptosis at the level of cardiomyocytes and affects cell signal transduction and intervention to improve cardiac function, and part of the mechanism is not related to the inhibition of Na-KATPase

Method used

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  • Method of screening action target of cardiac glycoside medicine using protein composition technology
  • Method of screening action target of cardiac glycoside medicine using protein composition technology
  • Method of screening action target of cardiac glycoside medicine using protein composition technology

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] (1) To establish a model of cardiac glycosides on cardiomyocytes.

[0032] Newborn 1-2 day old Wistar rats, 10, were thoracotomy, the heart was removed, washed with D-Hanks solution, 0.125% pancreatin and 0.02% disodium diethylenetetraacetate were added, water bath at 37°C for 5 minutes, pipetting for 1 2 minutes, naturally precipitated, discard the supernatant; add 0.125% trypsin, put it in a 37°C water bath for 10 minutes, then pipette for 1-2 minutes, naturally precipitate, suck the supernatant, filter it with a 400-mesh metal filter, and transfer it into a 10ml centrifuge. Tube, add an equal volume of DMEM medium to the centrifuge tube. Repeat the digestion 5 times until the myocardial tissue blocks are basically digested into single cells; then add DMEM medium, which contains 15% fetal bovine serum, and break up the cell mass into a single cell suspension; adjust the number of cells to 1 × 105 / ml, put the The single cell suspension was inoculated in a culture flas...

Embodiment 2

[0045] (1) Establish a model of the effect of digitalis drugs on cardiomyocytes.

[0046] The cardiomyocyte culture was as described in Example 1, except that in the drug action group, digoxin was replaced by convolvulus, and the drug concentration was 1 × 10 -9 mol / L, respectively, for 24 hours, resulting in the myocardial cell model of cardiac glycosides.

[0047] (2) Preparation of protein samples for cardiomyocyte two-dimensional electrophoresis, the same as in Example 1.

[0048] (3) Two-dimensional gel electrophoresis, same as Example 1

[0049] (4) Image acquisition and analysis, same as Example 1

[0050] After the gel was stained with Coomassie brilliant blue, it was scanned with a gel scanner to obtain a digital image, which was analyzed with PDQUEST software. The image analysis process included map clipping, protein spot detection, matching between different gels, quantitative analysis. The relative molecular weight and isoelectric point of the protein spots in t...

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Abstract

A method for selecting action target of cardiac glucocide medicine by utilizing protein multistudy technique setting up model of cadiocytes culture and action on cadiocytes by cardiac glucocide, preparing protein sample, carrying two ¿C way gel electrophoresis and collecting image for analysis to obtain difference expression protein, using mass spectrum to determine difference protein peptide mass finger atlas to obtain DNA series for selecting out difference protein, verifying biological property of difference expression protein and using protein multistudy technique to select target of cardiac glucocide medicine.

Description

(1) Technical field [0001] The invention relates to a method for screening the target protein of cardiac glycoside drugs acting on cardiomyocytes by using proteomics technology, in particular to a method for screening the target protein that cardiac glycoside drugs act on cardiac muscle cells, and belongs to the technical field of medicine. (2) Background technology [0002] Coronary heart disease, hypertension, chronic valvular disease, congenital heart disease and other important cardiovascular system diseases can develop to a certain extent and heart failure can occur, becoming the main cause of death of the above diseases. Heart failure has a poor prognosis, and the overall mortality rate is about 15-20%, and the annual mortality rate of severe patients can reach 50%. Therefore, effective treatment of heart failure is a global public health problem. Cardiac glycosides are currently the first choice for the treatment of heart failure, but the effective range and safety r...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N35/00B01D57/02G01N1/34G01N21/84
Inventor 高海青邱洁贾继辉陈春燕马亚兵
Owner SHANDONG UNIV
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