Application of native copper leachate in preparation of medicine for promoting differentiation of bone marrow mesenchymal stem cells

A bone marrow mesenchyme and natural copper technology, applied in the field of biomedicine, can solve the problems of low bioavailability and low solubility

Pending Publication Date: 2019-04-12
LANZHOU UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the course of long-term clinical application, it was found that natural copper used in traditional Chinese medicine orthopedics has a significant effect on promoting fracture healing. However, the natural copper processed by classical methods has low solubility in water and low bioavailability, which has brought great problems to clinical medicine.

Method used

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  • Application of native copper leachate in preparation of medicine for promoting differentiation of bone marrow mesenchymal stem cells
  • Application of native copper leachate in preparation of medicine for promoting differentiation of bone marrow mesenchymal stem cells

Examples

Experimental program
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Effect test

Embodiment 1

[0015] Example 1 Preparation of Natural Copper Thiobacillus ferrooxidans BY3 Leaching Solution

[0016] Weigh 0.5 g of natural copper, grind it through a 200-mesh sieve, and obtain a powder. In a clean 250mL Erlenmeyer flask, add 100mL of sterilized OK liquid medium and adjust the pH to 1.60 with 5M sulfuric acid. After the pH is stable, inoculate a volume fraction of 10% Thiobacillus ferrooxidans BY3, and extract at 25°C and 130r / min 15 days. During the experiment, 5M sulfuric acid was used to adjust the pH value, and distilled water was used to supplement the evaporated water. Shake the flask for 15 days to stop leaching, let it stand overnight, collect the supernatant, adjust the pH of the solution to neutral, pass through a 0.22m filter membrane to sterilize, and set aside.

Embodiment 2

[0017] Example 2 Culture of Bone Marrow Mesenchymal Stem Cells

[0018] Get about 120-150g SD rats (purchased from the Animal Experiment Center of Gansu University of Traditional Chinese Medicine), kill with 75% alcohol disinfection for 10min after decapitation, peel off the femur tissue, and then use DMEM / F12 culture solution (containing sodium heparin) 500U / mL) syringe to draw out the bone marrow in the bone marrow cavity, and repeated blowing to break up the cells as much as possible. The cell suspension was sieved with 150 mesh, and the cell concentration was adjusted to 1.0×10 7 cells / mL, inoculate in 90mm cell culture dish, and culture the cells with DMEM / F12 medium containing 10% fetal bovine serum. Culture conditions are 37°C, 5% CO 2 Cultivate under the condition of saturated humidity, replace the culture medium every 3 days, rinse twice with sterile PBS (phosphate buffer saline) before changing the culture medium, and subculture when the cells reach 70-80% confluenc...

Embodiment 3A

[0019] Example 3 ALP activity assay

[0020] As a marker of osteoblast maturation and mineralization, ALP is a glycoprotein ubiquitously present on the cell membrane, capable of hydrolyzing phospholipids, and providing the material basis for the mineralization of bone cells to form bones. In this example, ALP activity was used as an index to evaluate the effect of natural copper bioleaching solution on osteogenic differentiation of bone marrow mesenchymal stem cells.

[0021] 3.1 Grouping

[0022] Set up natural copper 0K+A.f+N-pyrite (1:100) group, 0K+A.f+A-pyrite (1:10000) group, positive control group and negative control group.

[0023] 0K+A.f+N-pyrite (1:100) group uses natural natural copper as the original drug, adopts the natural copper leaching solution prepared by the leaching method in Example 1, and cultures with α-MEM with a volume ratio of 1:100 Base is diluted.

[0024] The 0K+A.f+A-pyrite (1:10000) group uses calcined natural copper as the original drug, and...

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Abstract

The invention relates to application of a Thiobacillus ferrooxidans BY3 leachate of native copper in preparation of a medicine for promoting differentiation of bone marrow mesenchymal stem cells. According to the invention, the effect of the Thiobacillus ferrooxidans BY3 leachate of native copper in a process of promoting osteogenic differentiation of bone marrow mesenchymal stem cells is studiedby taking bone marrow mesenchymal stem cells of rats as a target object. Results show that the Thiobacillus ferrooxidans BY3 leachate of native copper can enhance the ALP activity of bone marrow mesenchymal stem cells in vitro, promote deposition of calcium salts in bone marrow mesenchymal stem cells, and promote osteogenic differentiation of bone marrow mesenchymal stem cells.

Description

technical field [0001] The invention relates to the application of natural copper Thiobacillus ferrooxidans BY3 leaching solution in the preparation of medicines for promoting osteogenesis and differentiation of bone marrow mesenchymal stem cells, which belongs to the field of biomedicine. Background technique [0002] Osteoporosis is a systemic and systemic skeletal disease, which is caused by the destruction of the dynamic balance of bone metabolism and bone formation less than bone resorption. The bone metabolism process requires the maintenance of many functional cells to maintain bone homeostasis, among which osteoclasts and osteoblasts are the main functional cells in this process. Osteoblasts secrete calcium salts and bone matrix to form new bones during bone formation, while osteoclasts secrete acidic substances during bone formation to play a role in bone absorption. [0003] Rat bone marrow mesenchymal stem cells (rBMSCs) are stem cells that exist in the bone marr...

Claims

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Application Information

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IPC IPC(8): C12N5/077
CPCC12N5/0654C12N2506/1353C12N2500/72C12N2500/20
Inventor 李红玉黄欣周建
Owner LANZHOU UNIVERSITY
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