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Preparation method of stem cell factor oral product

A stem cell factor, stem cell technology, applied in biochemical equipment and methods, animal cells, vertebrate cells, etc., can solve the problems of inefficient cytokine secretion process and small culture scale, and achieve the goal of enhancing human immunity and strengthening the body. , the effect of enhancing survival and proliferative activity

Active Publication Date: 2021-12-17
安徽科门生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The pharmacological effects of stem cell factors are obviously reliable, but the culture medium or cell lysate collected from mesenchymal stem cells in vitro has problems such as small scale of culture and insufficient efficiency in the process of inducing cytokine secretion.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] A kind of preparation method of stem cell factor oral product proposed by the present invention comprises the following steps:

[0022] S1. Obtain the fetal bovine umbilical cord, wash it with PBS solution, remove the arteries and veins, cut the umbilical cord into tissue pieces with a diameter of about 1-2 mm with scissors, add 0.4% collagenase II to digest for 2 h, centrifuge at 2000 r / min for 10 min, collect the precipitate, and pour Add 2% trypsin to digest for 30 minutes, stop the digestion, centrifuge at 2000r / min for 15min, collect the precipitate; use PBS to blow into a suspension, filter with a 200μm filter, and centrifuge at 1800r / min for 7min to obtain bovine umbilical cord cells;

[0023] S2. Inoculate bovine umbilical cord cells into T25 culture flasks, coat the culture flasks with fibronectin, add 10% FBS, 100U / mL penicillin, and 100U / mL streptomycin to the α-MEM culture medium at 37°C , 5%CO 2 Cultivate in an incubator, replace the fresh medium every 3 d...

Embodiment 2

[0029] Compared with embodiment 1, the difference is only that step S3 is different; specifically as follows:

[0030] S3. When the P3 generation bovine umbilical cord stem cells are cultured until the cell confluence reaches 80-90%, the supernatant is discarded, washed with PBS, and then cultured with a secretion-promoting culture medium at an inoculation density of 1.5×10 5 / mL, collect the supernatant after induction culture for 48h, filter it with a 0.1 μm hollow fiber microfiltration membrane, concentrate the permeate through an ultrafiltration membrane (molecular weight cut-off 5kD), and collect the retentate;

[0031] The secretory-stimulating culture fluid includes basal culture fluid ɑ-MEM, and glutamine, 10 ng / mL norepinephrine, 2 ng / mL transforming growth factor, 60 ng / mL of glucosamine, 10ng / mL of astragalus polysaccharide.

Embodiment 3

[0033] Compared with embodiment 1, the difference is only that step S3 is different; specifically as follows:

[0034] S3. When the P3 generation bovine umbilical cord stem cells are cultured until the cell confluence reaches 80-90%, the supernatant is discarded, washed with PBS, and then cultured with a secretion-promoting culture medium at an inoculation density of 1.5×10 5 / mL, collect the supernatant after induction culture for 48h, filter it with a 0.1 μm hollow fiber microfiltration membrane, concentrate the permeate through an ultrafiltration membrane (molecular weight cut-off 5kD), and collect the retentate;

[0035] The secretion-stimulating culture fluid includes basal culture fluid DMEM / F12, and adding glutamine, 5 ng / mL norepinephrine, 4 ng / mL transforming growth factor, 80 ng / mL glutamine, 5 ng / mL final concentration in the base culture fluid / mL of glucosamine, 5ng / mL of astragalus polysaccharide.

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Abstract

The invention discloses a preparation method of a stem cell factor oral product, and relates to the technical field of biology. The preparation method comprises the following steps of obtaining primary bovine umbilical cord mesenchymal stem cells; carrying out subculture on the primary bovine umbilical cord mesenchymal stem cells; when the P3-P5-generation bovine umbilical cord stem cells are cultured until the cell fusion degree reaches 80-90%, adopting a secretion-promoting culture solution for culture, collecting a supernate after culture is conducted for 48 h, filtering the supernate with a 0.1-micron filter membrane, and performing ultrafiltration and concentration to obtain a concentrated solution, wherein the secretion-promoting culture solution comprises a basic culture solution, and glutamine, noradrenaline, transforming growth factors, glucosamine and astragalus polysaccharide which are added into the basic culture solution; and pre-freezing the concentrated solution, and then carrying out ultralow-temperature freeze drying to obtain freeze-dried powder. According to the invention, the amplification of the umbilical cord mesenchymal stem cells can be effectively improved, the secretion of stem cell factors can be induced, and the obtained product can condition the body from the cellular level, comprehensively improve the body function, improve the sub-health and repair and strengthen the immune system.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a preparation method of an oral product of stem cell factor. Background technique [0002] The umbilical cord is the connecting structure between the mammal and the fetus. It is shaped like a rope, has a smooth and transparent surface, and contains connective tissue, an umbilical vein and a pair of umbilical arteries. In recent years, with the continuous development of research, it has been possible to extract stem cells from umbilical cord, umbilical cord blood and placenta. [0003] Umbilical cord mesenchymal stem cells refer to a kind of multifunctional stem cells existing in umbilical cord tissue. Umbilical cord mesenchymal stem cells are a kind of low immunogenic cells with multidirectional differentiation potential and strong immune regulation function. easy. An important function of umbilical cord mesenchymal stem cells is to secrete a variety of cytokines and growth factors...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K35/28A61K9/19A61P37/04C12N5/0775
CPCA61K35/28A61K9/19A61P37/04C12N5/0665C12N2500/32C12N2501/81C12N2501/90C12N2501/148C12N2501/15C12N2500/34
Inventor 张正亮
Owner 安徽科门生物科技有限公司
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