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Method for improving proliferation ability and performance of umbilical cord blood OECs (outgrowth endothelial cells)

An endothelial cell proliferation and endothelial cell technology is applied in the field of improving the proliferation ability and performance of umbilical cord blood outwardly growing endothelial cells to achieve the effects of increasing the number of cells, improving the therapeutic effect and reducing the amount of cells

Inactive Publication Date: 2014-10-15
广州市天河诺亚生物工程有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Although umbilical cord blood outward growth endothelial cells have many advantages over peripheral blood outward growth endothelial cells, but because most of its indications are degenerative diseases, such as cardiovascular and cerebrovascular diseases, diabetic foot, etc., long-term multiple infusions are required. volume becomes the bottleneck of its application

Method used

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  • Method for improving proliferation ability and performance of umbilical cord blood OECs (outgrowth endothelial cells)
  • Method for improving proliferation ability and performance of umbilical cord blood OECs (outgrowth endothelial cells)
  • Method for improving proliferation ability and performance of umbilical cord blood OECs (outgrowth endothelial cells)

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Experimental program
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Effect test

Embodiment 1

[0054] Umbilical cord blood and umbilical cord specimens were obtained from placentas delivered by healthy puerperas through aseptic procedures. The tests of hepatitis B, hepatitis C, syphilis, AIDS, cytomegalovirus, TORCH, mycoplasma, chlamydia, G-6PD and thalassemia were all negative. After the specimens were collected, they were transported back to the blood bank to maintain the transport conditions at 4-8°C. Follow the method below.

[0055] 1. Preparation of umbilical cord mesenchymal stem cells

[0056] Mesenchymal stem cells are extracted from the umbilical cord tissue of healthy people, and the obtained mesenchymal stem cells are cultured and expanded in vitro. After the primary cells were collected, they were subcultured at a ratio of 1:3. When the cells covered the bottom of the bottle with a density of about 80%, they were digested and collected with trypsin, and subcultured at the same ratio, up to the third generation. Collect the cells of the 3rd generation. A...

Embodiment 2

[0087] With reference to the method of Example 1, the collected endothelial cells were adjusted to a density of 1 × 10 with endothelial cell culture medium. 5 cells / mL, marked as cell suspension A; the collected mesenchymal stem cells were adjusted to a density of 1×10 5 Cells / mL, marked as cell suspension B, take 2 mL each of cell suspension A and cell suspension B and add to 6 mL endothelial cell culture medium, that is, the mixed group.

[0088] At the same time, the endothelial cell group was defined as the OEC group, the mesenchymal stem cell group was defined as the MSC group, and 2 mL of cell suspension A was added to 8 mL of endothelial cell culture medium in the OEC group. In the MSC group, 2 mL of cell suspension B was added to 8 mL of endothelial cell culture medium.

[0089] The three groups were respectively added to T75 culture flasks pre-plated with collagen for culture.

Embodiment 3

[0092] Referring to the method of Example 2, the cells were harvested after 4 days, and the harvested cells were counted, and the amount of cells in the three groups was compared, and the multiplication factor was calculated using the following formula.

[0093] Multiplication factor = Harvested cell volume / Inoculated cell volume

[0094] The results are shown in Table 1: the cell proliferation rate of the mixed group was higher than that of the endothelial cell group and the mesenchymal stem cell group, and the difference was statistically significant, meeting the requirements for rapid expansion in this application.

[0095] Table 1

[0096]

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Abstract

The invention discloses a method for improving proliferation ability and performance of umbilical cord blood OECs (outgrowth endothelial cells). The method comprises the steps as follows: pure umbilical cord blood-derived OECs are obtained by separation from fresh umbilical cord blood and purification, and mixed with umbilical cord-derived mesenchymal stem cells for culture, and a culture medium is a EGM-2BulletKit complete medium containing the fetal bovine serum with the volume fraction of 10%. By the aid of the method, the proliferation efficiency and the vascularization ability of the OECs are improved. After the method is applied, due to the immune down-regulatory effect of the mesenchymal stem cells and the expansion of the total treatment cell amount, the cell loss proportion caused by immune reactions is reduced, the treatment effect is guaranteed, and the application possibility of the umbilical cord blood-derived OECs is increased. The mesenchymal stem cells also have the ability of assisting in maintaining blood vessels, so that the long-term treatment effect is guaranteed. The method is mainly applied to blood vessel repair treatment, tissue engineering and the like.

Description

technical field [0001] The invention belongs to the field of cell therapy, in particular to a method for improving the proliferative ability and performance of umbilical cord blood outwardly growing endothelial cells. Background technique [0002] Outgrowth endothelial cells (OECs), which belong to the precursor cells of vascular endothelial cells, grow like paving stones when cultured in vitro, forming a monolayer of cells, which is consistent with the morphology of mature endothelial cells, but they have rapid expansion and angiogenesis The ability to make it a new favorite in the field of cell therapy, researchers hope to use it for the treatment of cardiovascular and cerebrovascular diseases, heart disease, limb ischemia and other diseases. Jumi Kim et al. (Jumi Kim, Young-Joo Jeon. Comparative proteomic analysis of endothelial cells progenitor cells derived from cord blood-and peripheral blood for cell therapy. Biomaterials. Volume34, Issue6, February2013, Pages1669–168...

Claims

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Application Information

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IPC IPC(8): C12N5/071C12N5/0775
Inventor 嵐山芮魏伟郭磊
Owner 广州市天河诺亚生物工程有限公司
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