High-magnesium microenvironment bone marrow stem cell microsphere carrier as well as preparation method and application thereof

A bone marrow stem cell and microenvironment technology, applied in prostheses, microcapsules, pharmaceutical formulations, etc., can solve problems that have not been reported in the literature, and achieve the effect of promoting bone defect repair and cell proliferation.

Active Publication Date: 2020-05-26
SHANGHAI NINTH PEOPLES HOSPITAL SHANGHAI JIAO TONG UNIV SCHOOL OF MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In order to further improve the effect of regeneration and repair, magnesium ions with the ability to induce osteogenic differentiation are proposed to be compounded into microspheres to construct high-magnesium microenvironment microspheres and deliver stem cells for bone regeneration at the same time. This aspect of research has not been reported in the literature.

Method used

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  • High-magnesium microenvironment bone marrow stem cell microsphere carrier as well as preparation method and application thereof
  • High-magnesium microenvironment bone marrow stem cell microsphere carrier as well as preparation method and application thereof
  • High-magnesium microenvironment bone marrow stem cell microsphere carrier as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1. Detection of Magnesium Ion Toxicity to Rat Bone Marrow Stem Cells

[0030] Step 1. Isolation and culture of rat bone marrow stem cells

[0031] Sprague Dawley rats aged 3 to 4 weeks were killed by neck dislocation, and the dead bodies of the rats were disinfected by immersing them in 75% alcohol solution for 3 minutes. After the disinfection was completed, the bone marrow mesenchymal stem cells were obtained in the ultra-clean bench. Cut the skin at the groin of the rat, and bluntly dissect the tissue to expose the tibia and femur. Use scissors to cut off the knee joint, take out the tibia and femur, remove the muscle and other soft tissues on the bone surface, and cut off the epiphysis at both ends of the tibia; absorb an appropriate amount of DMEM complete culture solution with a syringe, insert the needle of the syringe into the bone marrow cavity, and use DMEM Pull up and down while washing the culture medium to ensure that the content of the bone marro...

Embodiment 2

[0034] Example 2. Magnesium ions affect the biological activity of cells

[0035] Step 1. Effect of different concentrations of magnesium ions on cell proliferation

[0036] According to the magnesium ion concentration screened in Example 1, configure the DMEM complete culture solution with magnesium ion concentration of 1.6, 2.5, 5, 10, 20mM respectively for subsequent use, take rat bone marrow stem cells, and inoculate them on a 96-well plate at a density of 1000 / well In medium (n=6), after the cells adhered to the wall, 100 μL of high-magnesium culture medium was replaced, and normal DMEM complete culture medium was used as the control group, and this time was recorded as day 0. On the 1st, 4th, and 7th day, add 10 μl MTT solution to each well, incubate for 4 hours, suck off the original liquid in the plate, add 150 μl DMSO to each well, and shake for 10 minutes to fully dissolve the crystals. The absorbance value was detected by a microplate reader to analyze the cell pro...

Embodiment 3

[0047] Example 3. Detection of changes in the expression of magnesium ion channel MagT1 in stem cells induced by different concentrations of magnesium ions

[0048] Step 1. Detection of MagT1 expression changes in stem cells after osteogenic induction

[0049] Rat bone marrow stem cells with a confluence of more than 90% were taken, digested and centrifuged, counted, inoculated in 6-well plates and confocal small dishes, and when the cell confluence reached 80%, the osteogenic induction medium (provided by Saiye Biotech) was replaced. At the same time, normal DMEM culture medium was used as a control (n=3) for osteogenesis induction. After 3 days and 5 days of induction, samples were collected for qPCR detection and immunofluorescence detection. The immunofluorescence staining method was the same as above; for qPCR samples, 1 mL of trizol was added to lyse the cells, and the total RNA of each group of samples was collected. Determine the concentration and purity of the extrac...

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Abstract

The invention provides a high-magnesium microenvironment bone marrow stem cell microsphere carrier as well as a preparation method and an application thereof, magnesium ions are compounded into the microsphere carrier to form a local high-magnesium environment, the size of the microsphere carrier is within 500 [mu]m, and the concentration of the magnesium ions is 2.5 mM to 10 mM. According to theconcept and the preparation method for constructing the stem cell microsphere carrier of the local high-magnesium microenvironment, on one hand, when the magnesium ion concentration is screened out tobe 2.5 mM to 5mM, the osteogenesis induction effect is optimal; secondly, the diameter of the prepared stem cell microsphere carrier is within 500 microns, so that the loading rate and survival rateof stem cells are effectively increased, osteogenic differentiation of the loaded stem cells is promoted by utilizing a local high-magnesium environment, and the defects of high cost, difficulty in slow release and the like caused by using protein factors are reduced; finally, when the prepared high-magnesium microenvironment stem cell microsphere carrier is used for bone defect regeneration and repair, newly born bones in a defect area are obviously increased, and the repair effect is remarkably improved.

Description

technical field [0001] The invention belongs to the field of tissue engineering and regenerative medicine, and more specifically relates to a high-magnesium microenvironment bone marrow stem cell microsphere carrier and its preparation method and application. Background technique [0002] Stem cells have the ability of self-renewal and multi-directional differentiation. In theory, they can be expanded to a sufficient number in vitro and induced to differentiate into specific adult cells, which can then be used for tissue regeneration therapy. Directly loading cells on the scaffold material is a common method, but due to the influence of material pore size and specific surface area, the number of transplanted cells is obviously limited: too small pore size of the porous scaffold material will limit the penetration of cells into the deep part of the scaffold, and the enlarged pore size will increase the specific surface area. Will decrease accordingly, thus affecting the numbe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61L27/38A61L27/20A61L27/04A61L27/50A61L27/52
CPCA61L27/3834A61L27/047A61L27/52A61L27/20A61L27/3895A61L27/50A61L2400/06A61L2300/622A61L2430/02C08L5/04
Inventor 蒋欣泉张文杰林思涵杨光正
Owner SHANGHAI NINTH PEOPLES HOSPITAL SHANGHAI JIAO TONG UNIV SCHOOL OF MEDICINE
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