Induction medium for inducing human adipose tissue-derived stromal cells as nerve cells and method
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A technology for inducing mesenchymal stem cells, which is applied in the field of inducing differentiation medium, can solve the problems of high cell death rate, long induction time, and low induction efficiency, and achieve high induction success rate, short induction time, and high induction efficiency. Effect
Inactive Publication Date: 2013-07-24
QINGDAO RESTORE BIOTECHNOLOGY CO LTD
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[0009] Although the traditional chemical inducer method can efficiently and quickly differentiate adipose-derived mesenchymal stem cells into nerve cells, the cell death rate is relatively high (chemical inducers have certain toxicity, BHA, DMSO, etc. can cause cell mutation, and are used f
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Embodiment 1
[0024] Example 1 An induction medium for inducing human adipose-derived mesenchymal stem cells into neurons in this example is composed of liquid A, liquid B, and liquid C, and is prepared by the following method:
[0025] Solution A: Dissolve 10ug bFGF (basic fibroblast growth factor, peprotech, 96-100-18B-50) in 500ml of human mesenchymal stem cell serum-free medium (LONZA, product number 00190632), filter and sterilize to obtain A solution, in which bFGF concentration is 20ng / ml;
[0026]Solution B: Add 10ml of Xiangdan injection (active ingredient Danshensu, Jiangsu Changshu Leiyunshang Pharmaceutical Co., Ltd.) into 490ml of human mesenchymal stem cell serum-free medium to obtain solution B, wherein the final volume concentration of Xiangdan injection is 2%;
[0028] Example 2 The method for inducing differentiation of human adipose-derived mesenchymal stem cells into neurons by using the induction medium of the present invention, the steps are as follows:
[0029] 1. Preparation of human adipose-derived mesenchymal stem cells:
[0030] 1. To receive adipose tissue, wipe the outer wall of the container containing adipose tissue with 75% alcohol;
[0031] 2. Dispense adipose tissue, each T175 culture bottle is divided into 50ml adipose tissue. With a 10ml pipette, remove the tip, first absorb the lower layer of red liquid in the fat collection bottle and discard it, and mix the remaining upper layer of fat before subpackaging.
[0032] 3. Wash fat tissue and remove blood cells. Add 100ml of sodium chloride injection into the T175 culture bottle, tighten the cap, shake vigorously for 3 minutes to fully wash the adipose tissue, then stand still for 3 to 5 minutes to separate the different phases, and suck off the lower aqueous phase;...
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Abstract
The invention belongs to the technical field of induction and differentiation of stem cells and in particular relates to a medium for inducing differentiation and a method. An induction medium for inducing human adipose tissue-derived stromal cells as nerve cells consists of a solution A, a solution B and a solution C and is prepared by adopting a method comprising the following steps of: preparing the solution A by dissolving 8-12 micrograms of bFGF (Basic Fibroblast Growth Factor) in 500ml of a serum-free medium of the human adipose tissue-derived stromal cells, filtering and degerming to acquire the solution A; preparing the solution B by adding 10-20ml of injection of salviae miltiorrhizae composite into 490-480ml of the serum-free medium of the human adipose tissue-derived stromal cells to acquire the solution B; and preparing the solution C by dissolving 4-8 micrograms of bFGF, 4-8 micrograms of EGF (Epidermal Growth Factor), 40-60 micrograms of BDNF (Brain Derived Neurotrophic Factor) and 80-120 micrograms of GM1 (gangliosides) in 500ml of the serum-free medium of the human adipose tissue-derived stromal cells to acquire the solution C. By adopting the medium for inducing human adipose tissue-derived stromal cells as nerve cells disclosed by the invention, the human adipose tissue-derived stromal cells are differentiated into the nerve cells by adopting the injection of salviae miltiorrhizae composite and combining the growth factors EGF, bFGF, BDNF and GM1; and the induction medium disclosed by the invention is non-toxic in selected induction components, high in induction efficiency and short in induction time; and the exclusion and ethics problems do not exist in the inducted cells after transplanting, so that the induction medium is high in safety.
Description
technical field [0001] The invention belongs to the technical field of induction and differentiation of stem cells, and in particular relates to a medium and a method for inducing differentiation. Background technique [0002] Nervous system damage and neurodegenerative diseases, such as Parkinson's disease, Alzheimer's disease, Huntington's disease, etc., have always been a problem that plagues human health, and cell transplantation is the hope for curing such diseases. At present, embryonic stem cells and neural stem cells are mainly used for transplantation to treat central nervous system injuries. However, due to difficulties in donor sources, ethical objections, immune rejection, and difficulties in the survival of transplanted cells, the development space is limited. Adipose-derived mesenchymal stem cells have a wide range of sources, are easy to obtain, are convenient for autologous transplantation, and have strong proliferation ability. They can always maintain their...
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