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Method for inducing and culturing umbilical cord mesenchymal stem cells into nerve cells

A technology of stem cells and nerve cells, applied in the field of neurobiology, can solve the problems of low differentiation efficiency of nerve cells, long induction period and low cell differentiation, and achieve the effect of high cell differentiation uniformity and short induction period.

Inactive Publication Date: 2014-08-27
广州雅安生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] There are a few foreign research institutions conducting experimental research on MSC directional induction and differentiation into nerve cells, but there are few domestic studies, and the existing reported induction differentiation methods have a long induction period, and the obtained nerve cells have low differentiation efficiency and low uniformity of cell differentiation.

Method used

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  • Method for inducing and culturing umbilical cord mesenchymal stem cells into nerve cells
  • Method for inducing and culturing umbilical cord mesenchymal stem cells into nerve cells
  • Method for inducing and culturing umbilical cord mesenchymal stem cells into nerve cells

Examples

Experimental program
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Effect test

Embodiment 1

[0028] Example 1. Acquisition and cultivation of umbilical cord mesenchymal stem cells

[0029] Neonatal umbilical cords authorized with maternal consent were used as a source of mesenchymal stem cells.

[0030] The neonatal umbilical cord was washed three times in physiological saline containing 1% double antibody to remove the blood stains on the surface, and then cut into small pieces about 1 cm long. Use ophthalmic scissors to cut the umbilical cord longitudinally along the direction parallel to the blood vessels, and peel off the 2 umbilical arteries and 1 umbilical vein from the umbilical cord. Peel off the amniotic membrane on the surface, wash the Huatong glue part with normal saline containing 1% double antibody for 3 times, and cut it into pieces to about 1mm 3 size. Evenly spread the shredded tissue blocks on the 75cm 2 Place in the culture bottle at room temperature for 5-10 minutes to make the tissue pieces stick tightly. Add 5ml of medium containing DMEM / F12,...

Embodiment 2

[0032] Example 2. Directed differentiation of umbilical cord mesenchymal stem cells in vitro 1

[0033] Freshly passaged P3-6 umbilical cord mesenchymal stem cells were used in 5×10 4 Inoculate in a 24-well plate with poly-lysine-coated coverslips per ml, 1ml / well, replace the pre-induction culture medium after 3 days of adherent culture, change the medium in half after 3 days, and terminate the induction on the 3rd day .

[0034] The pre-induction culture medium is DMEM as the base medium, which also contains 50nmol vitamin C, 20U / ml human leukocyte inhibitory factor, 8mM L-glutamine, 5mM thioglycerol (MTG), 80U / ml penicillin, 80μg / ml streptomycin.

[0035] The cells obtained from the pre-induction culture were divided into 5×10 4 Inoculate at a cell density of / ml for monolayer adherent induction culture, replace the induction medium for induction culture after 2 days of adherence culture, change the medium for half of the amount after 3 days, terminate the induction on ...

Embodiment 3

[0038] Example 3. Directed differentiation of umbilical cord mesenchymal stem cells in vitro 2

[0039] Freshly passaged P3-6 umbilical cord mesenchymal stem cells were used in 5×10 4 Inoculate in a 24-well plate with poly-lysine-coated coverslips, 1ml / well, replace the pre-induction culture medium after 3 days of adherent culture, change the medium in half after 3 days, and terminate the induction on the 4th day .

[0040] The pre-induction culture medium is DMEM as the base medium, which also contains 20nmol vitamin C, 10U / ml human leukocyte inhibitory factor, 1mM L-glutamine, 2mM thioglycerol (MTG), 100U / ml penicillin, 100μg / ml streptomycin.

[0041] The cells obtained from the pre-induction culture were divided into 5×10 4Inoculate at a cell density of / ml for monolayer adherent induction culture, replace the induction medium for induction culture after 2 days of adherence culture, change the medium in half after 3 days, terminate the induction on the 6th day, and obta...

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Abstract

The invention provides a method for inducing and culturing umbilical cord mesenchymal stem cells into nerve cells. According to the invention, a combined method of two steps of pre-induction and induction is adopted, and an optimized pre-induction culture solution and an induction culture solution are prepared. The induction mode is short in induction period and high in cell differentiation homogeneity. Electrophysiological detection finds that the obtained nerve cells have the function of induced electric discharge, which indicates that the induced nerve cells are obtained successfully.

Description

technical field [0001] The invention relates to the field of neurobiology, in particular to a method for inducing and culturing umbilical cord mesenchymal stem cells into nerve cells, and the invention also relates to the cultured nerve cells. Background technique [0002] Umbilical cord mesenchymal stem cells (MSCs) are mesoderm-derived, capable of self-renewal, and have the potential to differentiate into three types of mesoderm lineages: osteoblast, cartilage, and fat. In recent years, studies have found that MSCs can also transdifferentiate beyond the mesoderm, such as nerve cells and epithelial cells of the ectoderm line, and liver cells and pancreatic cells of the endoderm line. Because MSC has a wide range of sources, is easy to isolate and culture, has low immunogenicity, and does not have the ethical issues faced by embryonic stem cells, this makes its application in regenerative medicine have advantages that other stem cells do not have. Neurodegenerative diseases...

Claims

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Application Information

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IPC IPC(8): C12N5/079
Inventor 安沂华董健伸
Owner 广州雅安生物科技有限公司
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