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Bionic micro-fluidic chip for simulating pathological blood-brain barrier based on fibrous protein gel and construction method of bionic micro-fluidic chip

A fibrin gel and microfluidic chip technology, applied in the field of bionic microfluidic chip and its construction, can solve problems such as the inability to effectively build a pathological blood-brain barrier model, reduce the amount of cells and reagents, and reduce the test cost Effect

Active Publication Date: 2022-03-15
RUIJIN HOSPITAL AFFILIATED TO SHANGHAI JIAO TONG UNIV SCHOOL OF MEDICINE
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] Aiming at the problem that the existing microfluidic blood-brain barrier chip cannot effectively construct a pathological blood-brain barrier model, the present invention provides a bionic microfluidic chip based on fibrin gel to simulate pathological blood-brain barrier and its construction method

Method used

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  • Bionic micro-fluidic chip for simulating pathological blood-brain barrier based on fibrous protein gel and construction method of bionic micro-fluidic chip
  • Bionic micro-fluidic chip for simulating pathological blood-brain barrier based on fibrous protein gel and construction method of bionic micro-fluidic chip
  • Bionic micro-fluidic chip for simulating pathological blood-brain barrier based on fibrous protein gel and construction method of bionic micro-fluidic chip

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Effect test

Embodiment 1

[0049] This embodiment provides a bionic microfluidic chip based on fibrin gel to simulate pathological blood-brain barrier, its structure is as follows figure 1 shown. The microfluidic chip mainly includes three parallel channels and corresponding inlets and outlets, which are the endothelial cell channel, the astrocyte-fibrin gel mixed solution channel, and the microglial cell channel; the endothelial cell channel contains Endothelial cell inlet, endothelial cell side channel and endothelial cell outlet; astrocyte and fibrin gel mixed solution channel includes mixed solution inlet, main channel and outlet; microglia channel includes small Glial cell inlet, microglia side channel and microglia outflow port. The diameter of the endothelial cell inlet and endothelial cell outlet is 5mm, and the width of the endothelial cell channel is 1mm; The width is 1mm; the microglial cell inlet and the microglial cell outflow port are circular, with a diameter of 5mm, and the width of t...

Embodiment 2

[0060] Characterization of endothelial tight junction proteins on fibrin gel-based simulated pathological blood-brain barrier model

[0061] Using the self-designed and produced microfluidic chip above, a pathological blood-brain barrier model based on fibrin gel was established after chip injection, and after 5 days of co-culture, the endothelial barrier structure was formed. The tight junction protein ZO-1 of the cells was immunofluorescently stained, and the method was as follows: prepare the chip to be immunofluorescent, aspirate and discard the culture medium at the injection port, add PBS buffer to the injection port on the side of the chip and wash it 2-3 times, 5 min each time; fix with 4% paraformaldehyde at room temperature for 15 min, and then wash with PBS buffer three times, 5 min each; Prepare the primary antibody solution with the primary antibody diluent containing 1% BSA+0.3% Triton X-100, the primary antibody (rabbit anti-human ZO-1) dilution ratio is 1:100, ...

Embodiment 3

[0063] Simulation of the permeability of pathological blood-brain barrier model based on fibrin gel

[0064] The permeability test was carried out using fluorescein isothiocyanate (FITC)-labeled dextran molecules with a molecular weight of 4 kDa and rhodamine-labeled dextran molecules with a molecular weight of 70 kDa. After the endothelial barrier was formed in the three cell co-culture systems based on fibrin gel, dextran molecules with different molecular weights were added into the endothelial cell channel from the endothelial cell inlet at a concentration of 400 μg / mL, and recorded continuously with a fluorescence microscope. Diffusion of fluorescent molecules in the chip for 30 minutes, and fluorescent images were taken every 5 minutes to characterize the permeability of the pathological blood-brain barrier model to dextran of different molecular weights. The results are as follows Figure 4 shown. Figure 4 The image shown shows that the pathological blood-brain barrie...

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Abstract

The invention relates to a bionic micro-fluidic chip for simulating a pathological blood brain barrier based on fibrous protein gel and a construction method thereof, the chip is provided with three parallel channels and corresponding sample inlets and outlets, and trapezoidal micro-valve structure arrays arranged at equal intervals are arranged between every two adjacent channels. According to the method, a pathological model of blood-brain barrier integrity destruction can be established, tight junction proteins with abnormal expression forms can be established, the barrier function shows the pathological characteristic of leakage, real-time dynamic imaging of cell biological behaviors can be realized, multiple independent and repeated pathological blood-brain barrier models can be constructed at the same time, the test cost is reduced, and the test efficiency is improved. The method has an application prospect in the aspects of disease mechanism exploration and drug screening.

Description

technical field [0001] The invention belongs to the field of microfluidic chips, in particular to a bionic microfluidic chip based on fibrin gel simulating pathological blood-brain barrier and a construction method thereof. Background technique [0002] Existing BBB research models include in vivo animal models, in vitro cell models, and computer models. Although animal models can highly simulate the complexity of human physiology and reflect the real state of the microenvironment, there is no animal model that can fully reproduce all the characteristics of human diseases, and the research results have encountered obstacles in the process of clinical translation. In vitro models include cell culture models, brain slices, fiber-based dynamic in vitro blood-brain barrier models, and microfluidic organ-on-a-chip models. Transwell chamber culture in a petri dish is the most widely used in vitro cell model. One or more cells are cultured in semi-permeable microwells. Although th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12M3/04C12N5/079C12N5/071B01L3/00
CPCC12M23/16C12M23/20C12M25/14C12M29/10C12N5/069C12N5/0622B01L3/502707B01L3/502715C12N2533/90C12N2513/00C12N2502/086C12N2533/56C12N2535/00
Inventor 罗艳刘妍君曾羽连沈宇辉
Owner RUIJIN HOSPITAL AFFILIATED TO SHANGHAI JIAO TONG UNIV SCHOOL OF MEDICINE
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