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Method for inducing umbilical cord mesenchymal stem cells to be differentiated into dopaminergic neurons

A technology of mesenchymal stem cells and induction of differentiation, applied in the field of neurobiology, can solve the problems of undisclosed MSC directional induction and differentiation, and achieve the effect of improving efficiency and selectivity

Inactive Publication Date: 2015-07-22
中国人民解放军总医院第三医学中心
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  • Summary
  • Abstract
  • Description
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  • Application Information

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Problems solved by technology

However, this patent document does not disclose how to effectively induce MSCs to differentiate into dopaminergic neurons, and there are very few studies on how to effectively induce MSCs to differentiate into dopaminergic neurons in other prior art

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1: Acquisition and culture of umbilical cord mesenchymal stem cells

[0043] Neonatal umbilical cords authorized with maternal consent were used as a source of mesenchymal stem cells.

[0044] 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, 2...

Embodiment 2

[0046] Example 2: Directed differentiation of umbilical cord mesenchymal stem cells in vitro

[0047] 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 .

[0048] 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.

[0049] The cells obtained from the pre-induction culture were divided into 5×10 4 / ml cell density inoculation for single-layer adherent induction culture, after 2 days of adherent culture, replace the induction medium for induction culture, change the medium after 3 days, stop the induction on the 3rd day, add no inducer to ...

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Abstract

The invention discloses a method for inducing umbilical cord mesenchymal stem cells to be differentiated into dopaminergic neurons. According to the method, a two-step combination method of pre-induction and induction is adopted, optimized pre-induction culture solution and induction culture solution are prepared, and a glycoside compound improved in structure is added as an inducer, so that the umbilical cord mesenchymal stem cells are successfully induced to be differentiated into the dopaminergic neurons and the differentiation ratio is relatively high.

Description

technical field [0001] The invention relates to the field of neurobiology, in particular to a method for inducing differentiation of umbilical cord mesenchymal stem cells into dopaminergic neurons, and the invention also relates to cultured neurons. Background technique [0002] Since Erices et al. reported in 2000 that umbilical cord blood is rich in hematopoietic stem cells and mesenchymal stem cells, umbilical cord blood mesenchymal stem cells have attracted the attention of researchers due to their multi-directional differentiation potential, immunoregulatory effect and no ethical issues. . Subsequently, some scholars reported that umbilical cord blood mesenchymal stem cells could induce the expression of neuronal markers and glial cell markers in vitro. At present, people have been able to successfully isolate and culture umbilical cord blood mesenchymal stem cells in vitro, and have a further understanding of their biological characteristics, surface markers and immun...

Claims

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Application Information

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IPC IPC(8): C12N5/0793C12N5/0775
Inventor 安沂华董健伸
Owner 中国人民解放军总医院第三医学中心
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