A kind of nasal epithelial stem cell culture method and nasal epithelial stem cell proliferation medium

A stem cell culture and stem cell proliferation technology, applied in the field of nasal epithelial stem cell culture method and nasal epithelial stem cell proliferation medium, can solve the problem of reduced differentiation ability, difficulty in differentiation of cells into cilia or goblet epithelial cells, inability to maintain stem cell function for a long time and characteristics and other issues, to achieve the effect of maintaining differentiation ability, breaking through technical bottlenecks, and maintaining proliferation ability

Active Publication Date: 2022-02-15
THE FIRST AFFILIATED HOSPITAL OF SUN YAT SEN UNIV
View PDF8 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The existing culture systems and methods have the following limitations: 1) The commercial medium can only maintain the proliferation and culture of undifferentiated nasal epithelial stem cells for 2-3 generations, and generally after the 2nd generation, the cells are difficult to differentiate into ciliated or goblet epithelial cells
2) Although the culture system developed by the applicant in the early stage can maintain the proliferation and culture of 3-5 generations of undifferentiated epithelial stem cells, it is difficult to guarantee the differentiation ability of the cells after more than 3 generations of proliferation
Although certain stem cell proliferation characteristics can be maintained, the reduction in differentiation ability makes it difficult to maintain the functional characteristics (mucociliary clearance function) of epithelial stem cells
3) In the expected clinical application, in some cases (such as a large area of ​​mucosal injury), a large number of stem cells are required. Therefore, the existing medium cannot provide a sufficient amount under the condition of a limited number of passages (i.e. expansion). stem cells
4) Regardless of whether it is a commercial culture medium or a culture system developed by an individual laboratory, the proliferation of nasal epithelial stem cells is relatively slow, and it will be a challenge to obtain a sufficient number of epithelial stem cells with stem cell characteristics in a short period of time (such as within 4 weeks). problem
[0008] To sum up, the existing nasal epithelial stem cell culture systems and methods are still unable to maintain the function and characteristics of stem cells for a long time, that is, in terms of proliferation and differentiation, it is difficult to meet the needs of clinical stem cell transplantation, which is also a common clinical application of adult stem cells. technical bottleneck

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • A kind of nasal epithelial stem cell culture method and nasal epithelial stem cell proliferation medium
  • A kind of nasal epithelial stem cell culture method and nasal epithelial stem cell proliferation medium
  • A kind of nasal epithelial stem cell culture method and nasal epithelial stem cell proliferation medium

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0093] Nasal epithelial stem cells were derived from sinusitis tissue samples from a patient diagnosed with chronic sinusitis.

[0094] 1) Treat trophoblast cells

[0095] ①The trophoblast cells were 3T3 cells, and the culture conditions were DMEM medium, 10% fetal bovine serum and 1x penicillin-streptomycin complex solution.

[0096] ②The 3T3 cells were grown to 100% density and treated with mitomycin to inhibit the growth of 3T3 cells. The concentration of mitomycin was 0.5-20ug / mL.

[0097] ③The trophoblast cells treated with mitomycin were digested according to a certain density (0.1-1x10 4 cells / cm 2 ) were pre-plated in cell culture dishes.

[0098] ④The 3T3 cells treated with mitomycin can adhere to the wall smoothly within 12 hours to 24 hours.

[0099] 2) Proliferation and cultivation of nasal epithelial stem cells (respectively used as the medium of the present invention and the BEGM medium)

[0100] ① After the trophoblast cells adhere to the wall, the epitheli...

Embodiment 2

[0125] Nasal epithelial stem cells were derived from nasal polyp tissue samples, and the patient was diagnosed with nasal polyps.

[0126] 1) Treat trophoblast cells

[0127] ①The trophoblast cells were 3T3 cells, and the culture conditions were DMEM medium, 10% fetal bovine serum and 1x penicillin-streptomycin complex solution.

[0128] ②The 3T3 cells were grown to 100% density and treated with mitomycin to inhibit the growth of 3T3 cells. The concentration of mitomycin was 0.5-20ug / mL.

[0129] ③The trophoblast cells treated with mitomycin were digested according to a certain density (0.1-1x10 4 cells / cm 2 ) were pre-plated in cell culture dishes.

[0130] ④The 3T3 cells treated with mitomycin can adhere to the wall smoothly within 12 hours to 24 hours.

[0131] 2) Proliferation and cultivation of nasal epithelial stem cells (respectively used as the medium of the present invention and the BEGM medium)

[0132] ① After the trophoblast cells adhere to the wall, the epith...

Embodiment 3

[0156] Nasal epithelial stem cells were derived from inferior turbinate tissue samples, and the patient was diagnosed with inferior turbinate hypertrophy.

[0157] 1) Treat trophoblast cells

[0158] ①The trophoblast cells were 3T3 cells, and the culture conditions were DMEM medium, 10% fetal bovine serum and 1x penicillin-streptomycin complex solution.

[0159] ②The 3T3 cells were grown to 100% density and treated with mitomycin to inhibit the growth of 3T3 cells. The concentration of mitomycin was 0.5-20ug / mL.

[0160] ③The trophoblast cells treated with mitomycin were digested according to a certain density (0.1-1x10 4 cells / cm 2 ) were pre-plated in cell culture dishes.

[0161] ④The 3T3 cells treated with mitomycin can adhere to the wall smoothly within 12 hours to 24 hours.

[0162] 2) Proliferation and cultivation of nasal epithelial stem cells (respectively used as the medium of the present invention and the BEGM medium)

[0163] ① After the trophoblast cells adhe...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention discloses a method for cultivating nasal epithelial stem cells, which comprises the following steps: 1) paving trophoblast cells in a culture dish; 2) isolating primary nasal epithelial stem cells from nasal mucosa tissue; 3) separating primary nasal epithelial stem cells into The generation cells are placed on the trophoblast cells, the culture medium is replaced with nasal epithelial stem cell proliferation medium, and the rest of the tissue cells are removed; 4) when the nasal epithelial stem cells proliferate to 80% density, they are digested with trypsin and passed on to continue to expand; 5) Transfer nasal epithelial stem cells to gas-liquid phase culture system and PneumaCult-ALI differentiation medium for differentiation culture. The invention also discloses a nasal epithelial stem cell proliferation medium, comprising the following components: DMEM / F12, fetal bovine serum, penicillin, streptomycin, insulin, epidermal growth factor, hydrocortisone, 3,3',5 ‑triiodo‑L‑thyronine sodium, ROCK inhibitors. The invention provides a key implementation basis for the transformation application of nasal epithelial stem cell clinical transplantation.

Description

technical field [0001] The invention belongs to the technical field of adult stem cells, and in particular relates to a nasal epithelial stem cell culture method and a nasal epithelial stem cell proliferation medium. Background technique [0002] Adult stem cells refer to undifferentiated cells that exist in tissues. These cells can self-renew, and under certain conditions, differentiate according to a certain program to form new functional cells, so that the tissue or organ maintains a dynamic balance of growth and decline. . Embryonic stem cells are totipotent, can be established and passed on, can be immortalized and have strong proliferation ability, but their disadvantages are obvious, such as immune rejection, non-localized differentiation, and the possibility of causing cancer. In contrast, adult stem cells are derived from the individual, have no histocompatibility problems, avoid transplant rejection, and have tissue-specific proliferation and directional different...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/071
CPCC12N5/0625C12N2501/11C12N2501/33C12N2501/30
Inventor 李春炜洪玥何根谢曦
Owner THE FIRST AFFILIATED HOSPITAL OF SUN YAT SEN UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products