Application of SAFit2 and culture medium in promoting differentiation of human induced pluripotent stem cells to dopaminergic neurons

A pluripotent stem cell and dopaminergic technology, which is applied in the application field of promoting the differentiation of human-derived induced pluripotent stem cells into dopaminergic neurons, and can solve problems affecting the differentiation of dopaminergic neurons

Active Publication Date: 2022-05-24
ACADEMY OF MILITARY MEDICAL SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

SAFit2 affects the differentiation of dopamine neurons, so far there is no relevant report at home and abroad

Method used

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  • Application of SAFit2 and culture medium in promoting differentiation of human induced pluripotent stem cells to dopaminergic neurons
  • Application of SAFit2 and culture medium in promoting differentiation of human induced pluripotent stem cells to dopaminergic neurons
  • Application of SAFit2 and culture medium in promoting differentiation of human induced pluripotent stem cells to dopaminergic neurons

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] Culture and passaging of hiPSCs

[0056] hiPSC is a commercially purchased cell line, purchased from the Cell Bank of the Type Culture Collection Committee of the Chinese Academy of Sciences (https: / / www.cellbank.org.cn / ), catalog number: DYR0100, which induces human foreskin cells into induced pluripotency Stem cells, hiPSCs established by induction of reprogramming transcription factors OCT4, SOX2, KLF4, MYC. Cells were derived from the ATCC website ( ACS-1011 TM ), is an internationally accepted cell line for hiPSC research.

[0057] According to the instructions when the cells were purchased, the cells were cultured in a feeder-free mode, and the medium was mTeSR1 (Stem Cells, product number: 05850), and the medium was changed in full every day, 2 mL / well (6-well plate). Every 4 days, the cells were digested with Accutace (StemCells, Cat. No.: 07920) for 5 minutes, the cell suspension was transferred to a 15 ml centrifuge tube, centrifuged at 1000 rpm for 10 min...

Embodiment 2

[0059] Differentiation of hiPSCs into dopaminergic neurons

[0060] DAP differentiation medium containing SAFit2 (DAP differentiation medium + SAFit2): When preparing 100 mL of medium, use DMEM / F-12 medium (Gibco, catalog number: 11320033) and neural basal medium (Gibco, catalog number: 21103049) as Basic medium (the two mediums were mixed in a volume ratio of 1:1, in which the amount of DMEM / F-12 medium and neural basal medium was 47.8885 mL, respectively), and the following components: 1 mL of N2 supplement (N2 The mass ratio of supplement and basal medium is 1:100, that is, according to the instructions, the volume ratio is 1%), 2 mL of vitamin A-free B27 supplement (the volume of vitamin A-free B27 supplement and basal medium) The ratio is 1:50, that is, the volume ratio is 2% according to the instructions), 1 mL of 20M ascorbic acid (Sigma, product number: 1043003), 100 μL of 5mM doxorphine hydrochloride (DM, purchased from Tocris, product number: 3039) , ), 100 μL of 10...

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Abstract

The invention belongs to the technical field of cell culture, and particularly relates to application of SAFit2 and a culture medium in promoting differentiation of human induced pluripotent stem cells to dopaminergic neurons. The invention provides an application of SAFit2 in promoting efficient differentiation of human induced pluripotent stem cells to dopaminergic neurons. The SAFit2 provided by the invention can promote efficient differentiation of human induced pluripotent stem cells into dopaminergic neurons.

Description

technical field [0001] The invention belongs to the technical field of cell culture, in particular to the application of SAFit2 and a culture medium in promoting the differentiation of human induced pluripotent stem cells into dopaminergic neurons. Background technique [0002] At this stage, the research methods for human neuropsychiatric diseases are limited, mainly because it is difficult to obtain nerve cell samples from normal people and patients, and only a few studies are derived from nerve cells in autopsy of patients. However, due to the differences in the evolutionary level of species, animal models cannot be established for non-conservative gene defects between animals and humans, and animal models cannot ideally reflect the nature of human disease symptoms and pathological changes, resulting in most animal experiments. Effective drugs fail in preclinical trials due to poor safety and efficacy of the drug. Therefore, finding a suitable humanized cell model has al...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/074C12N5/0793
CPCC12N5/0619C12N2506/45C12N2501/13C12N2501/01C12N2500/30C12N2501/999C12N2501/15C12N2500/46C12N2501/00Y02A50/30
Inventor 李红李锦吴宁李斐卢关伊方婷
Owner ACADEMY OF MILITARY MEDICAL SCI
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