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Tandem cell model and preparation method thereof

A cell model, cell technology, applied in biochemical equipment and methods, animal cells, tumors/cancer cells, etc., can solve the problems of incomplete research, inability to investigate the influence of vascular endothelial layer, lack of important processes of drugs, etc., to achieve structural integrity Effect

Inactive Publication Date: 2016-10-26
PEKING UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The current use of cell models to study the process of drug absorption is limited to the simple application of a single cell model, focusing on the transport of drugs across the intestinal epithelial cell layer, lacking the important process of drugs entering the blood circulation across the vascular endothelial cell layer, resulting in a similarity with the real human intestinal tract. There are certain differences in the transport process, and it is impossible to investigate the impact of nutrients or pathogens on the vascular endothelial layer after crossing the epithelial layer, resulting in incomplete research

Method used

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  • Tandem cell model and preparation method thereof
  • Tandem cell model and preparation method thereof
  • Tandem cell model and preparation method thereof

Examples

Experimental program
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Embodiment 1

[0052] Embodiment 1, the establishment of the tandem cell model of the present invention

[0053] 1 Establishment of Caco-2 cell model

[0054] 1.1 Recovery and passage of Caco-2 cells

[0055] Remove Caco-2 cells from liquid nitrogen and immediately place them in a 37°C water bath. After rapid thawing, transfer the Caco-2 cells to a centrifuge tube with 5ml DMEM-10% culture solution, mix well, centrifuge at 1000 rpm for 5min, discard the supernatant, add 5ml DMEM-10% culture solution, Obtain Caco-2 cell suspension. Transfer the Caco-2 cell suspension to a cell culture flask, place it in a constant temperature incubator for culture, change the medium the next day, and change the medium every other day thereafter. The confluence of the cells in the culture flask is about 80%, discard the old culture medium, wash three times with 37°C pre-warmed sterile phosphate buffer solution, and add 0.25% trypsin-0.02% EDTA solution for digestion. Observe the morphological changes of th...

Embodiment 2

[0077] Example 2 Using the tandem cell model of the present invention to study the transmembrane transport of paracellular transport markers

[0078] Fluorescein FITC-Dextran 4,000 (FD4) was used as a marker compound, Caco-2 model and EAhy926 model were used as controls, and the tandem cell model was established as described above.

[0079] Fluorescein FD4 is a marker of paracellular transport and is commonly used to verify the integrity of cell junctions to determine the integrity of cell monolayers.

[0080] In this example, the transport characteristics of the model were investigated through the transport amount of the FD4 trans-tandem model, and the Caco-2 model and the EAhy926 model were used as the control group.

[0081] 2 groups and administration

[0082] 2.1 Series model (ie the series model of the present invention) group: 0.1ml 0.1% FD4 standard solution was given to the apical side of the Caco-2 cells, and samples from the basal side of the EAhy926 cells were col...

Embodiment 3

[0092] Embodiment 3 Utilizes the tandem cell model of the present invention to investigate Fe 3 o 4 Transmembrane transport behavior of nanoparticles.

[0093] Oral absorption of nano-drugs is the focus of nano-formulation research. Fe 3 o 4 Nanoparticles have the advantages of stable particle size, good monodispersity, and easy quantification and modification. Choose Fe 3 o 4 Nanoparticles as model drugs to investigate Fe 3 o 4 Characterization of nanoparticle transport across tandem models.

[0094] 1Fe 3 o 4 Nanoparticle standard solution preparation: Fe 3 o 4 The aqueous solution was dispersed in BSA solution, incubated at 37°C for 30 minutes, centrifuged at 13000g at 4°C to discard the supernatant, and resuspended to 400ug / ml with MEM culture medium.

[0095] 2Fe 3 o 4 Nanoparticle Characterization

[0096] Dynamic Light Scattering: Fe 3 o 4 The nanoparticle standard solution is measured by a particle size analyzer, the particle size is 60.91nm, PDI=0.15...

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Abstract

The invention discloses a tandem cell model, comprising an epidermal cell model and an endothelial cell model, wherein the epidermal cell model and the endothelial cell model are separately inoculated on different semi-permeable membranes. The invention also discloses a preparation method for the tandem cell model and application of the tandem cell model to simulation the process of transferring of a nanometer drug in an intestinal tract. The tandem cell model provides the invention also can simulate an intestinal tract-blood vessel barrier multilayer structure and detect the transferring behaviors and influence of a drug in different single layers of intestinal cells, and has good application prospect and development potential.

Description

technical field [0001] The invention relates to a series cell model, which is formed by series connection of Caco-2 cell model and EAhy926 cell model. It specifically involves the characteristics and preparation methods of the tandem model, as well as the application in the study of the permeability difference of different layered structures in the intestinal tract and the transport of nano-medicines in the intestinal tract. Background technique [0002] Nanoformulations are a promising approach in the study of oral formulations of poorly soluble drugs. Due to the special surface properties of nanoparticles, their transport mode and characteristics in cells are different from those of traditional drug molecules, and the transport mechanism is still unclear. The use of cell models to assess drug oral transport and its mechanism in vitro can simplify the research process and save development costs. [0003] As the main site of digestion and absorption for oral administration...

Claims

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Application Information

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IPC IPC(8): C12N5/09C12Q1/02
CPCC12N5/0693C12N2503/00G01N33/502
Inventor 张强卫备代文兵何冰王学清张华袁兰
Owner PEKING UNIV
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