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Induced differentiation method for human adipose-derived stem cells

An adipose stem cell and induced differentiation technology, applied in the field of human adipose stem cell induction and differentiation, can solve the problems of unsatisfactory motor neuron function and specificity, low directional transdifferentiation efficiency, etc., to improve the transdifferentiation efficiency and specificity. , good curative effect, high differentiation yield effect

Inactive Publication Date: 2016-05-11
厚朴生物科技(苏州)有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The technical problem to be solved by the present invention is that in the prior art, the directional transdifferentiation efficiency of adipose stem cells is low, and the functionality and specificity of the differentiated motor neurons are not ideal, so as to provide a high-efficiency human-derived adipose Method for inducing differentiation of stem cells into functional motor neuron cells

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  • Induced differentiation method for human adipose-derived stem cells
  • Induced differentiation method for human adipose-derived stem cells
  • Induced differentiation method for human adipose-derived stem cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] The human-derived adipose-derived stem cells in this example are isolated from adipose tissue obtained by human surgery or plastic liposuction, and the isolation, culture, and identification methods are as follows:

[0031] The human-derived adipose-derived stem cells were isolated and screened from adipose tissue by means of adherent culture, and the isolated human-derived adipose-derived stem cells were characterized from the spindle morphology and cell surface markers, and the isolated and obtained adipose-derived stem cells were analyzed by RT-PCR. Human-derived adipose-derived stem cells were identified, wherein the cell surface markers were CD immune surface markers; the universal markers for identifying the human-derived adipose-derived stem cells by RT-PCR were Sox2, Oct4, c-Myc, Nanog, and KLF4. The obtained human adipose stem cells are used for future use.

[0032] The method for inducing differentiation derived from human adipose stem cells in this embodiment...

Embodiment 2

[0036] The human-derived adipose-derived stem cells in this example are isolated from adipose tissue obtained by human surgery or plastic liposuction, and the isolation, culture, and identification methods are as follows:

[0037] Digest the adipose tissue in 0.075% type Ⅰ collagenase at a temperature of 37°C, and shake it every 5 minutes during the digestion process. After 40 minutes, use DMEM containing 10% serum and 1% P / S / F12 medium to neutralize collagenase, then centrifuge at 1500rpm / min for 10 minutes, discard the upper layer of fat and supernatant, suspend the cell mass at the bottom of the centrifuge tube with serum-containing medium, and let it pass through the 70um cells filter to remove tissue debris;

[0038] Spread the above-mentioned harvested cell filtrate into a 10 cm petri dish, and place it at 37°C in an atmosphere containing 5% CO 2 After culturing in a humidified incubator for 24 hours, change the medium and rinse with PBS, then add fresh medium to conti...

Embodiment 3

[0046] The human-derived adipose-derived stem cells in this example are isolated from adipose tissue obtained by human surgery or plastic liposuction, and the isolation, culture, and identification methods are as follows:

[0047] Digest the adipose tissue in 0.075% type Ⅰ collagenase at a temperature of 37°C, and shake it every 5 minutes during the digestion process. After 40 minutes, use DMEM containing 10% serum and 1% P / S / F12 medium to neutralize collagenase, then centrifuge at 1500rpm / min for 10 minutes, discard the upper layer of fat and supernatant, suspend the cell mass at the bottom of the centrifuge tube with serum-containing medium, and let it pass through the 70um cells filter to remove tissue debris;

[0048] Spread the above-mentioned harvested cell filtrate into a 10 cm petri dish, and place it at 37°C in an atmosphere containing 5% CO 2 After culturing in a humidified incubator for 24 hours, change the medium and rinse with PBS, then add fresh medium to conti...

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Abstract

The invention relates to an induced differentiation method for human adipose-derived stem cells. The method comprises the following steps: carrying out induced differentiation on the human adipose-derived stem cells by adopting SHH and RA together, so as to obtain motor neurons; carrying out in-vitro culture on the obtained motor neurons, so as to obtain mature functional motor neurons with an electrophysiological function. According to the induced differentiation method provided by the invention, the easy and feasible and high-differentiation-yield preparation of the neurons is achieved, and the transdifferentiation efficiency and specificity of directed differentiation of the adipose-derived stem cells to the motor neurons are improved.

Description

technical field [0001] The invention belongs to the field of biomedicine, and in particular relates to a method for inducing differentiation of human adipose stem cells. Background technique [0002] Cell replacement therapy is an effective strategy to rebuild the tissue structure of the damaged nervous system and restore the function of the nervous system. In cell replacement therapy, the two most critical issues are the source of seed cells and the survival rate of seed cells in vivo. Although stem cells have great application potential as seed cells, many kinds of stem cells involve many problems and cannot be applied clinically. Due to ethical, tumorigenic and immune rejection issues, they cannot be used as seed cells for clinical treatment; although human Fibroblast Induced Pluripotent Stem Cells (iPSCs) induced by human fibroblasts avoid ethical and immune rejection issues, but due to the The tumorigenicity and other so far unpredictable biosafety issues caused by th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0793C12N5/0775A61K35/30A61P25/28A61P25/00
CPCC12N5/0619A61K35/30C12N2501/385C12N2501/41C12N2506/1384
Inventor 孙振华曹晖
Owner 厚朴生物科技(苏州)有限公司
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