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Method for enriching cord blood hematopoietic stem cells by using mesenchymal stem cells cultured in low-oxygen three-dimensional environment

A technology of hematopoietic stem cells and mesenchymal stem cells, which is applied in the field of enriching umbilical cord blood hematopoietic stem cells from mesenchymal stem cells, can solve the problems of high cost and reduced cell activity, and achieve the effects of low cost, improved proliferation ability, and elimination of uncontrollable variables

Pending Publication Date: 2022-06-07
重庆市铂而斐细胞生物技术有限公司
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  • Application Information

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Problems solved by technology

[0006] Aiming at the deficiencies in the prior art, the present invention provides a method for enriching umbilical cord blood hematopoietic stem cells by using mesenchymal stem cells cultured in a hypoxic three-dimensional environment, which solves the problem of the high cost of the prior art and the problems of cells being enriched during the enrichment process. The problem of reduced activity in

Method used

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  • Method for enriching cord blood hematopoietic stem cells by using mesenchymal stem cells cultured in low-oxygen three-dimensional environment
  • Method for enriching cord blood hematopoietic stem cells by using mesenchymal stem cells cultured in low-oxygen three-dimensional environment
  • Method for enriching cord blood hematopoietic stem cells by using mesenchymal stem cells cultured in low-oxygen three-dimensional environment

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1 Enrichment of cord blood hematopoietic stem cells

[0047] 1. Preparation of 3D Cell Culture Substrate

[0048] (1) Use SolidWorks software to design a 3D model of a cylindrical micro-island structure with a diameter of 5-100 μm and a height of 20-200 μm, and then convert the 3D data of the model into a 3D printing code, and then input it into a 3D printer. Using a 3D lithography system (PhotonicProfessional GT, Nanoscribe GmbH) equipped with a 780 nm femtosecond laser and objective lens (25×, 0.8NA, Zeiss, Plan Apochromat), the designed micro-islands were prepared using negative photoresist IP-S as raw material. The structure is printed layer by layer on top of the substrate. During the printing process, the movement of the laser focus is controlled by a current scanner, and the movement of the substrate is controlled by a high-resolution xyz platform, so that the positions of each micro-island structure are randomly arranged and cover more than 90% of the c...

Embodiment 2

[0056] Example 2 Enrichment of cord blood hematopoietic stem cells

[0057] Similar to Example 1, the difference is that the diameter of the micro-island is 50 μm and the height is 100 μm.

Embodiment 3

[0058] Example 3 Enrichment of cord blood hematopoietic stem cells

[0059] Similar to Example 1, the difference is: the concentration of oxygen is 5%, and the incubation time is 12h.

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Abstract

The invention provides a method for enriching cord blood hematopoietic stem cells by using mesenchymal stem cells cultured in a low-oxygen three-dimensional environment. Comprising the following steps: inoculating mesenchymal stem cells on a cell culture substrate with a micro-topological structure on the surface, and culturing under a low-oxygen condition; umbilical cord blood mononuclear cells are inoculated to mesenchymal stem cells for co-culture, so that hematopoietic stem cells are enriched. According to the method, the CD34 positive hematopoietic stem cells in the cord blood mononuclear cells are enriched by the mesenchymal stem cells in a specific low-oxygen three-dimensional environment, and the hematopoietic stem cells in the cord blood mononuclear cells are directly separated through the interaction between the mesenchymal stem cells and the hematopoietic stem cells. The method is low in cost and simple and convenient in experimental steps, and the dryness and activity of the hematopoietic stem cells can be maintained in vitro.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for enriching cord blood hematopoietic stem cells by utilizing mesenchymal stem cells cultured in a low-oxygen three-dimensional environment. Background technique [0002] Hematopoietic stem cells (HSCs) maintain a unique capacity for self-renewal and pluripotent differentiation throughout life and have the potential to differentiate into all types of blood cells, including myeloid cells (monocytes, macrophages, neutrophils) granulocytes, basophils, eosinophils, erythrocytes, megakaryocytes / platelets and dendritic cells) and lymphoid cells (T cells, B cells, NK cells). HSCs are essential for maintaining blood and immune function. In the past few decades, hematopoietic stem cell transplantation has been used as a standard treatment for various blood disorders such as severe combined immunodeficiency, congenital neutropenia, and malignancies. [0003] A certain number of HSC...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0789C12N5/0775
CPCC12N5/0647C12N2513/00C12N2502/1388C12N2500/02C12N2509/00C12N2533/30
Inventor 李栋付强戴晓宇刘抗刘玲玲邓恩杰白希
Owner 重庆市铂而斐细胞生物技术有限公司
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