Novel glycosaminoglycan analogue and synthetic method thereof, and application of novel glycosaminoglycan analogue in invitro embryonic stem cell proliferation and directional neural differentiation

A technology for glycosaminoglycan and analogs, which is applied to novel glycosaminoglycan analogs and their synthesis and their application in in vitro embryonic stem cell proliferation and directed neural differentiation, and can solve the problems of limited yield and not widely obtained. application, cumbersome synthesis and purification steps, etc., to achieve the effects of high biological activity, easy process control, and simple process

Active Publication Date: 2015-09-23
SUZHOU UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this technology requires tedious synthesis and purification steps during operation, and the yield is limited, so it has not been widely used.

Method used

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  • Novel glycosaminoglycan analogue and synthetic method thereof, and application of novel glycosaminoglycan analogue in invitro embryonic stem cell proliferation and directional neural differentiation
  • Novel glycosaminoglycan analogue and synthetic method thereof, and application of novel glycosaminoglycan analogue in invitro embryonic stem cell proliferation and directional neural differentiation
  • Novel glycosaminoglycan analogue and synthetic method thereof, and application of novel glycosaminoglycan analogue in invitro embryonic stem cell proliferation and directional neural differentiation

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preparation example Construction

[0033] A kind of synthetic method of novel glycosaminoglycan analogue, comprises the following steps:

[0034] (1) Select the molecule SS containing sulfonic acid groups and the molecule MAG containing sugar groups as monomers, and by adjusting the relative ratio of the two, use the RAFT method to prepare homopolymers and copolymers with different functional groups (molecular weight in 6000-20000 range).

[0035] (2) The obtained polymers were added to the embryonic stem cell culture medium to culture the embryonic stem cells, and after a certain period of time, the proliferation of cells in different polymer environments was studied.

[0036] (3) The obtained polymers were added to the embryonic stem cell culture medium respectively, and the embryonic stem cells were cultured, and after a certain period of time, the directed neural differentiation of the cells in different polymer environments was studied.

[0037] The principle of the present invention is: adopt the method ...

Embodiment 1

[0042] Preparation of monomer 2-methacrylamide glucopyranose (MAG): Glucosamine hydrochloride and methacryloyl chloride were reacted in a methanol solution containing potassium carbonate under ice bath conditions for 3 to 4 hours, then suction filtered and rotary evaporated to obtain a precipitate , and the precipitate was purified and dried to obtain the monomer MAG.

[0043] SS and MAG were selected as monomers, RAFT polymerization was carried out in H2O / DMF (1 / 1) mixed solvent, and the initial feeding ratio of monomers was adjusted to obtain homopolymer pSS, pMAG and copolymer pSG, respectively.

[0044] The obtained polymer was characterized by NMR and IR.

[0045] The result is as Figure 1-3 Shown, the 1H-NMR spectrum and FT-IR spectrum of polymer pSG.

[0046] The polymer molecule is used to promote the proliferation of embryonic stem cells in vitro, comprising the following steps:

[0047] The embryonic stem cells were seeded on a culture plate covered with ...

Embodiment 2

[0050] The embryonic stem cells were seeded on a culture plate pre-spread with gelatin, and cultured in a stem cell culture medium environment containing 1 μg / mL of the three polymers or heparin respectively for 10 days and 20 days.

[0051] Discard the solution in the culture, treat the embryonic stem cells with the cell culture solution containing 3-(4,5-dimethylthiazole-2)-2,5-diphenyltetrazolium bromide (MTT) for 4 hours, and suck Remove the MTT cell culture medium and add dimethyl sulfoxide (DMSO) to dissolve the formazan crystals generated in living cells.

[0052] The DMSO solution in which formazan crystals were dissolved was transferred to a new culture plate, and the absorbance value of each well was read at 490 nm on a microplate reader.

[0053] The result is as Figure 4 Shown are the proliferative behaviors of embryonic stem cells cultured in the medium containing the obtained polymer for 10 days and 20 days respectively.

[0054] The polymer molecule is ...

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Abstract

The invention provides a novel glycosaminoglycan analogue and a synthetic method thereof, and an application method of the novel glycosaminoglycan analogue in invitro embryonic stem cell proliferation and directional neural differentiation. The synthetic method is characterized by comprising the following steps: adding a sodium p-styrenesulfonate monomer or / and a 2-methacrylamide glucopyranose monomer into a mixed solvent according to a certain proportion, wherein the mixed solvent is a mixture of water and N,N-dimethylformamide; and carrying out polymerization by applying a method of reversible addition-fragmentation chain transfer(RAFT) so as to respectively obtain a plurality of polymers of the sodium p-styrenesulfonate monomer or / and the 2-methacrylamide glucopyranose monomer in different proportions. According to the invention, a macromolecule substance capable of promoting an embryonic stem cell to proliferation in invitro and inducing the embryonic stem cell to realizing highly-efficient directional neural differentiation is obtained with a RAFT synthetic technology by utilizing a sulfonate-group-contained monomer and a sugar-contained monomer and controlling relative proportion of the sulfonate-group-contained monomer and the sugar-contained monomer to adjust the structure of a polymer.

Description

technical field [0001] The invention belongs to the field of polymer synthesis and its application in biomedicine, and specifically relates to a method for synthesizing novel glycosaminoglycan analogues. The RAFT synthesis technology is used to utilize monomers containing sulfonic acid groups and sugar-containing monomers, and by controlling their By adjusting the relative ratio of the polymer structure, a polymer substance that can promote the proliferation of embryonic stem cells in vitro and induce their high-efficiency and directional neural differentiation is obtained. Background technique [0002] Glycosaminoglycan (GAG) is a class of multifunctional active molecules produced by organisms, which play a key role in physiological processes such as cell adhesion, proliferation, differentiation and anticoagulation. In recent years, it has been discovered that glycosaminoglycan substances can induce the directional neural differentiation of embryonic stem cells under in vit...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C08F212/14C08F220/58C08F112/14C08F120/58C12N5/0735C12N5/079
Inventor 陈红袁琳吕仲林陈高健王萌萌丁开国
Owner SUZHOU UNIV
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