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Construction method of an adeno-associated virus cross-blood brain barrier model

A construction method and technology of the blood-brain barrier, applied in the field of bioengineering, can solve problems such as the inability to replicate the BBB shear force, and achieve the effect of reducing the amount of samples

Pending Publication Date: 2021-11-02
HUAQIAO UNIVERSITY
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Problems solved by technology

The in vitro blood-brain barrier model is mostly established in a static two-dimensional culture system, but this model cannot replicate key features such as the shear force of the BBB

Method used

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  • Construction method of an adeno-associated virus cross-blood brain barrier model
  • Construction method of an adeno-associated virus cross-blood brain barrier model
  • Construction method of an adeno-associated virus cross-blood brain barrier model

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Experimental program
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Effect test

Embodiment 2

[0078] Research on AAV crossing the blood-brain barrier.

[0079] Analyze the efficiency of various serotypes of AAV across the blood-brain barrier;

[0080] After diluting various serotypes of AAV to an appropriate titer with culture medium, take 200 μL and add it to the lower chamber of the chip, and add 100 μL of DMEM high-glucose medium to the upper chamber;

[0081] The various serotypes of adeno-associated virus are type 2 adeno-associated virus (AAV2), type 9 adeno-associated virus (AAV9) and a recombinant adeno-associated virus AAV-PHP.eB, and the concentration of various serotypes of adeno-associated virus is 1 ×10 10 GC / mL, the total amount of virus added to the lower chamber is 200 μL, and the upper layer is supplemented with 100 μL of DMEM high-glucose medium;

[0082] At 12h, 24h, 36h, and 48h, 15 μL of samples were taken from the upper layer of the chip, and 15 μL of DMEM high-glucose medium was supplemented at the same time. The samples were measured by real-t...

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Abstract

The invention discloses a construction method of an adeno-associated virus cross-blood brain barrier model. The construction method comprises the following steps: inoculating prepared astrocytes to the upper layer of a chip; inervsely inoculating prepared human brain microvascular endothelial cells to the lower layer of the chip; placing the chip in a matched precise shaking table so as to simulate the fluid shear force in the blood vessel of a human body; and performing continuous culture on the double-layer co-culture chip so that brain microvascular endothelial cells and astrocytes grow to form a blood brain barrier structure. According to the technical scheme provided by the invention, the in-vitro blood-brain barrier model is successfully constructed, the characteristics of the in-vivo blood-brain barrier are reflected more accurately and more closely, and the method can be used for related research of AAV crossing the blood-brain barrier. According to the invention, fluid shear force is added, so that a real in-vivo environment is better met, high throughput can be realized, and consumption of cells and reagents is reduced.

Description

technical field [0001] The invention relates to the field of bioengineering, in particular to the establishment of a blood-brain barrier model by dynamic co-cultivation of double-layer cells in vitro and the use of the model to conduct related research on adeno-associated virus crossing the blood-brain barrier. Background technique [0002] Central nervous system disease is one of the most difficult diseases to treat, and its associated disability rate is high, and it is usually difficult to be completely cured. The main reason for the difficulty in treatment is the existence of the blood-brain-barrier (BBB). The blood-brain barrier is a selectively permeable barrier that selectively restricts or blocks the permeation of many hydrophilic molecules or macromolecules. Therefore, while protecting the brain tissue, the blood-brain barrier also limits the delivery of many therapeutic drugs, seriously affecting the therapeutic effect. [0003] Adeno-associated virus (AAV) as a ve...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071C12N5/079
CPCC12N5/069C12N5/0622C12N2513/00
Inventor 刘丹朱明洋刁勇黄若兰王琨
Owner HUAQIAO UNIVERSITY
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