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Placenta source filling dry cell and production thereof

A stromal stem cell and stem cell technology, applied in the field of placenta-derived mesenchymal stem cells and their preparation, can solve the problems of lack of large-scale separation, purification and expansion methods of placenta-derived mesenchymal stem cells, and achieve low immunogenicity, easy Amplification, high yield effect

Inactive Publication Date: 2006-03-15
INST OF BASIC MEDICAL SCI ACAD OF MILITARY MEDICAL SCI OF PLA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there are only sporadic literatures at home and abroad about the small amount of placenta-derived adherent cells, which are mainly used for hematopoietic support research, but there is a lack of detailed technical information on the large-scale separation, purification and expansion of placenta-derived mesenchymal stem cells

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0015] Example 1 Perfusion method to obtain placenta-derived mesenchymal stem cells

[0016] Immediately after delivery, the placenta is placed in a sterile tray or container, injected with balanced salt solution containing anticoagulants and antibiotics via the umbilical artery, exsanguinated, and perfused. The anticoagulant can be heparin at 1~100u / mL, and the antibiotic can be penicillin and streptomycin at 100u / mL. Collect the perfusate, centrifuge to obtain the cell components, then resuspend the cell components, use percoll with a density of 1.073g / mL, centrifuge at a centrifugal force of 500g for 30 minutes, harvest the interfacial cell layer, wash with PBS and inoculate culture bottles or culture plates, or directly for other purposes. It is also possible to use D7-FIB or NGFR-labeled immunomagnetic beads to separate mesenchymal stem cells with certain special markers.

Embodiment 2

[0017] Example 2 Obtaining Placenta-derived Mesenchymal Stem Cells by Enzyme Digestion

[0018] Immediately after delivery, the placenta is placed in a sterile tray or container, injected with balanced salt solution containing anticoagulants and antibiotics via the umbilical artery, exsanguinated, and perfused. The anticoagulant can be heparin at 1~100u / mL, and the antibiotic can be penicillin and streptomycin at 100u / mL. After the initial perfusion, the placental leaflets were cut aseptically, and the placental tissue was digested with trypsin or collagenase to obtain a cell suspension, and then the cell suspension was passed through a cell sieve to remove tissue fragments and blood cells, and the obtained cells were resuspended at a density of 1.073 g / mL percoll, centrifuge at 500g for 30 minutes, harvest the interfacial cell layer, wash with PBS and inoculate culture flasks or culture plates, or directly use for other purposes. It is also possible to use D7-FIB or NGFR-lab...

Embodiment 3

[0019] Example 3 Identification of placenta-derived mesenchymal stem cells

[0020] According to literature reports, the method of inducing the differentiation of bone marrow mesenchymal stem cells in vitro induces placenta-derived mesenchymal stem cells to differentiate into osteoblasts, chondrocytes or adipocytes. Flow cytometry was used to detect the phenotypes of placenta-derived mesenchymal stem cells, such as CD29, CD34, CD44, CD45, CD105, CD166, HLA-DR, etc. At the same time, the cell cycle is detected to determine whether it is in G 0 -G 1 The proportion of cells in phase.

[0021] The results showed that the placenta-derived mesenchymal stem cells induced in vitro could differentiate into osteoblasts, chondrocytes, and adipocytes. Flow cytometry showed that the cells were positive for CD29, CD44, CD105, and CD166, and negative for CD34, CD45, and HLA-DR. The cells in G0-G1 phase accounted for more than 95%.

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PUM

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Abstract

A placenta original filled stem cell and its production are disclosed. The process is carried out by bleeding after delivering placenta from uterus, perfusing to obtain single-cell soliquoid from continuous perfusion method or collagenase digestion method and acquiring filled stem cell from gradient and density centrifugation or immune magnetic bead separation. It has better external enrichment potentiality and multidirectional differential potentiality and low immunogenicity.

Description

field of invention [0001] The invention relates to placenta-derived mesenchymal stem cells and preparation thereof, belonging to the field of stem cells and tissue engineering. Background technique [0002] The source of seed cells is the primary issue in tissue engineering and cell therapy. Due to the lack of sources of autologous tissue cells, the difficulty of expansion, and the easy loss of phenotypes during in vitro culture, people have gradually focused on stem cells in recent years. These cells have a certain ability of self-renewal in vitro, and can differentiate into one, multiple, or even all cell types of the human body under specific induction conditions, thus showing a good prospect for clinical application. Stem cells isolated so far include two types, namely embryonic stem cells and adult stem cells. Embryonic stem cells have omnipotent differentiation ability and unlimited proliferation ability, but due to ethics and lack of human understanding and control ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/08C12N5/06A61K35/12A61K48/00A61K35/28C12N5/0735
Inventor 王常勇郭希民周晓东段翠密江红董灵芝李晶
Owner INST OF BASIC MEDICAL SCI ACAD OF MILITARY MEDICAL SCI OF PLA
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