Pre-stimulated stem cells and preparation method and application thereof

A technology of stem cells and pre-stimulation, applied in biochemical equipment and methods, embryonic cells, animal cells, etc., can solve the problems of complex microenvironment and reduction, and achieve the effect of enhancing secretion function, mild treatment conditions, and strengthening secretion function

Active Publication Date: 2021-10-19
JINAN UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the microenvironment around the damaged tissue is quite complex, and none of these treatment methods can truly restore it

Method used

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Examples

Experimental program
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Effect test

preparation example Construction

[0032] A method for preparing pre-stimulated stem cells of the present invention comprises the following steps:

[0033] (1) selecting non-target stem cells as auxiliary cells, inducing pyroptotic auxiliary cells, and obtaining pyroptotic cell extracts and / or pyroptotic cells;

[0034] (2) using pyroptotic cell secretions, extracts and / or pyroptotic cells for the cultivation of target stem cells;

[0035] (3) Obtain pre-stimulated stem cells with stress state.

[0036] The principle of the technical solution of the present invention will be described in detail below.

[0037] The steps that the present invention comprises are as follows:

[0038] a. Auxiliary cell culture and induction of pyroptosis:

[0039]In the present invention, non-target stem cells are selected as auxiliary cells, and pyroptosis is induced by physical, chemical or biological methods. Because cell pyroptosis is a series of changes that occur autonomously after cells are stimulated by the outside worl...

Embodiment 1

[0062] Example 1: Preparation of Pre-stimulated Limbal Stem Cells for Rat Tissue Engineering Lamellar Cornea Construction

[0063] 1. Induction of pyroptosis in rat corneal stromal cells

[0064] Rat corneal stromal cells were selected as auxiliary cells, and 1*10 6 cells / cm 2 Cells were seeded in a 10cm diameter plastic culture dish and cultured in an incubator for 24 hours. The culture medium was high-glucose DMEM+10% fetal bovine serum+1% L-glutamine+1% NEAA+1% penicillin-chain Mycin solution (100X)+0.02% EGF+0.1% transferrin+0.1% insulin, the culture condition is 37°C, 5% carbon dioxide. After 24 hours, the culture solution containing 1 mol / ml lipopolysaccharide (LPS) / nigericin mixture was replaced for 48 hours.

[0065] 2. Collection of pyroptotic cell secretions

[0066] Add the above-mentioned LPS / nigericin mixture and cultivate for 48 hours, then collect the culture medium, wash it twice with PBS, blow gently to blow off the cell surface substances and collect the ...

Embodiment 2

[0072] Example 2: Preparation of pre-stimulated human bone marrow mesenchymal stem cells for the treatment of bone defects caused by arthritis

[0073] 1. Induction of pyroptosis in human peripheral blood mononuclear cells

[0074] Human peripheral blood mononuclear cells were selected as auxiliary cells, and 2*10 6 cells / cm 2 Cells were seeded in 10 cm diameter plastic culture dishes and cultured in an incubator for 24 hours. The culture medium was RPMI1640 + 10% fetal bovine serum, and the culture conditions were 37°C and 5% carbon dioxide. After 24h, fresh culture medium was replaced, and 50mJ / cm2 of ultraviolet light was irradiated for 24h.

[0075] 2. Extract pyroptotic bodies

[0076] After 24 hours, culture was continued for 24 hours, the culture medium was collected, rinsed twice with PBS, blown gently to blow off the cell surface substances and collect PBS. The collected culture medium and PBS were subjected to ultracentrifugation at 80,000 rpm, and the supernatan...

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Abstract

The invention relates to a pre-stimulated stem cell and its preparation method and application. The preparation method comprises the following steps: selecting non-purpose stem cell cells as auxiliary cells, inducing auxiliary cell pyroptosis, and obtaining pyroptotic cells and secretion of pyroptotic cells extracts, and / or pyroptotic cells for culturing stem cells. The preparation method of the present invention can obtain stem cells pre-stimulated by secretions of pyroptotic cells. The stem cells can adapt to the natural inflammatory environment and improve the ability of stem cells to repair damaged areas. It is a new breakthrough in the pretreatment of seed materials in the field of tissue engineering. At the same time, the pre-stimulated stem cells also provide a feasible solution for the treatment of clinical diseases.

Description

technical field [0001] The invention belongs to the field of tissue engineering, in particular to a pre-stimulated stem cell and its preparation method and application. Background technique [0002] Stem cells, as a kind of cells with self-replication ability and differentiation potential, can repair damaged areas through proliferation, differentiation and exocrine functions, and have always been considered as a good choice for tissue engineering seed cells. Lesions contain a large number of necrotic cells, hypoxia and ischemia, accompanied by a large number of inflammatory cell infiltration. After the stem cells are implanted into the lesion, most of the stem cells will die due to the harsh environment, which greatly reduces the therapeutic effect of the stem cells. In order to enhance the adaptation of transplanted stem cells to the inflammatory environment, researchers proposed a method of pre-stimulating stem cells before transplantation. Compared with the direct appli...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/071C12N5/0735C12N5/0775C12N5/079C12N5/0797
CPCC12N5/0606C12N5/0621C12N5/0623C12N5/0663C12N5/0668C12N5/0692C12N2500/80
Inventor 武征陈宝欣樊泽培陈剑赵巨鹏张建华
Owner JINAN UNIVERSITY
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