Method for simulating mechanical stimulation of abnormal blood flow to calcification of valvular cells

A technology of calcification and stimulation pairs, which is applied in the fields of biochemical equipment and methods, scientific instruments, and microbial determination/inspection, etc., can solve the problems of inability to eliminate, complicated operation, and long macro-mechanical stimulation culture calcification cycle. Achieving the effect of shortening the required time

Inactive Publication Date: 2017-12-19
XIEHE HOSPITAL ATTACHED TO TONGJI MEDICAL COLLEGE HUAZHONG SCI & TECH UNIV
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  • Abstract
  • Description
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Problems solved by technology

The macroscopic mechanical model cannot rule out the influence of cytokines secreted by the valve endothelial cells on the calcification of the valve interstitial cells, so as to simply study the effect of mechanical stimulation on the valve interstitial cells; at the same time, the macroscopic mechanical model cannot completely simulate the extracellular matrix or It is the process by which valve endothelial cells transmit mechanical stimuli to valve interstitial cells
[0004] At the same time, to study the influence of macroscopic mechanical stimulation receptors on calcification, the operation is complicated, and complex devices need to be placed in the incubator for continuous cultivation. The calcification cycle of macroscopic mechanical stimulation culture is longer.

Method used

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  • Method for simulating mechanical stimulation of abnormal blood flow to calcification of valvular cells
  • Method for simulating mechanical stimulation of abnormal blood flow to calcification of valvular cells

Examples

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Embodiment 1

[0032] Example 1: The method of simulating the effect of abnormal hemodynamic stimulation on valve cell calcification

[0033] The embodiment of the present invention provides a method for simulating the calcification of valve cells by abnormal hemodynamic stimulation:

[0034] S1, the interstitial cells of the valve are extracted from the aortic valve tissue by collagenase digestion method to make the cell supernatant medium, which specifically includes the following process:

[0035] S101, take the pig heart, cut off the aortic valve under aseptic conditions as much as possible, and soak it in high-sugar solution;

[0036] S102, after repeated washing of the valve with sterile PBS supplemented with Supsin and amphotericin, the valve was digested in type I collagenase containing Supsin and amphotericin. After no visible tissue residues were found, the upper part was removed by centrifugation. Added to the complete medium with 10% serum concentration to culture;

[0037] S10...

Embodiment 2

[0049] Example 2: Western blot verification method in the method of simulating the effect of abnormal hemodynamic stimulation on valve cell calcification

[0050] This example is used to verify the calcification response in valve interstitial cells after the method of simulating the calcification of valve cells by simulating abnormal hemodynamic stimulation

[0051] S401, use the method shown in Example 1 to stimulate the interstitium of the valve with F=0Pa, F=2.185Pa, F=4.375Pa, F=8.75Pa, F=17.5Pa, F=26.25Pa in the calcification medium Cells (F indicates magnetic force, frequency is 1Hz, and time is 20 minutes), and the valvular interstitial cells cultured in simple ordinary medium and calcification medium are set as the control group at the same time, and cultured in a 5% carbon dioxide incubator at 37 degrees Celsius for 3 days ;

[0052] S402, three days later, the valve interstitial cells were cracked with RIPA and protein was extracted from the valve interstitial cells...

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Abstract

The invention discloses a method for simulating mechanical stimulation of abnormal blood flow to calcification of valvular cells and relates to the technical field of biological model construction. The method comprises the following steps: S1) adopting a collagenase digestion method for extracting valvular interstitial cells in an aortic valve tissue and preparing into a cell supernatant culture medium; S2) mixing a magnetic bead with an RGD peptide stoste and preparing into a RGD peptide attached magnetic bead; S3) adding the RGD peptide attached magnetic bead into the cell supernatant culture medium acquired in the step S1), abandoning supernatant, cleaning with PBS and then adding into a first culture medium for culturing, putting the first culture medium into a magnetic distortion instrument, simulating the mechanical stimulation of the abnormal blood flow to the valvular interstitial cells by regulating the magnitude, frequency and acting time of the magnetic force of the magnetic distortion instrument and observing the calcification state of the valvular interstitial cells. According to the invention, the micro-simulation for the action of the mechanical stimulation of the abnormal blood flow to the calcification of valvular cells is realized, and meanwhile, the time for constructing a calcification model by a pure calcification culture medium can be shortened.

Description

technical field [0001] The invention relates to the technical field of biological model construction, in particular to a method for simulating the calcification of valve cells by abnormal hemodynamic stimulation. Background technique [0002] At present, in the study of mechanical stimulation of aortic valve calcification, bioreactors are mainly used, such as tensile force bioreactors, bending force bioreactors, shear force bioreactors, and composite bioreactors, all of which study macroscopic abnormalities The calcification of valve tissue or valve cells by mechanical stimulation can only simulate part of the macroscopic mechanics, such as pure laminar flow shear force and turbulent flow shear force, and there is no way to completely simulate external abnormal mechanical stimulation. [0003] Specifically, in the normal calcification process, the mechanical stimulus can only directly act on the valve endothelial cells, and the mechanical stimulation is transmitted to the va...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/02C12N13/00
CPCC12N13/00G01N33/5005
Inventor 董念国汪宁姜烨凡陈军见李庚刘义华陈锦杰
Owner XIEHE HOSPITAL ATTACHED TO TONGJI MEDICAL COLLEGE HUAZHONG SCI & TECH UNIV
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