Application of resveratrol-loading human multipotential stem cell exosome in preparing drug for treating refractory skin wound related diseases

A technology of human pluripotent stem cells and resveratrol, applied in non-embryonic pluripotent stem cells, artificially induced pluripotent cells, skin diseases, etc., can solve the problems that the treatment potential of diseases and injuries has not been fully utilized and needs to be further confirmed

Inactive Publication Date: 2018-08-03
汪洪
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, since milk exosomes contain a large number of non-human genes and proteins, their safety through intravenous systemic injection still needs to be further confirmed, and the focus of this study is mainly on the drug carrier function of exosomes. The therapeutic potential of the substances carried by exosomes to diseases and injuries has not been fully utilized

Method used

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  • Application of resveratrol-loading human multipotential stem cell exosome in preparing drug for treating refractory skin wound related diseases
  • Application of resveratrol-loading human multipotential stem cell exosome in preparing drug for treating refractory skin wound related diseases
  • Application of resveratrol-loading human multipotential stem cell exosome in preparing drug for treating refractory skin wound related diseases

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0094] Culture of human embryonic stem cells (ESCs) and extraction and identification of exosomes

[0095] A layer of embryonic stem cell Matrigel (ESC-Qualified BD Matrigel, BD Sparks, MD, USA), ESCs were moved into the dish, and mTeSR1 serum-free medium (StemCell Vancouver, BC, Canada), in an incubator (37°C, 5% CO 2 , saturated humidity) culture, and collect the culture medium changed every day. Filter the medium through a 0.22 micron pore size filter membrane and centrifuge at 10,000g at 4°C for 30 minutes to remove cell debris; use a 100KD molecular weight ultrafiltration tube and centrifuge (3500g, 15min) to intercept exosomes in the concentrated supernatant to obtain exosomes concentrate; transfer the concentrate to a 30% sucrose / heavy water density pad (1.210 g / cm 3 ), centrifuge at 100,000g at 4°C for 210 minutes, collect the 5ml sucrose / heavy water density pad at the bottom, add PBS to dilute, transfer to an ultrafiltration centrifuge tube with a molecular weigh...

Embodiment 2

[0100] Culture of human induced pluripotent stem cells (iPSCs) and extraction and identification of exosomes

[0101] A layer of embryonic stem cell Matrigel (ESC-Qualified BD Matrigel, BD Sparks, MD, USA), iPSCs were moved into the dish, and mTeSR1 serum-free medium (StemCell Vancouver, BC, Canada), in an incubator (37°C, 5% CO 2 , saturated humidity) culture, and collect the culture medium changed every day. Filter the medium through a 0.22 micron pore size filter membrane and centrifuge at 10,000g at 4°C for 30 minutes to remove cell debris; use a 100KD molecular weight ultrafiltration tube and centrifuge (3500g, 15min) to intercept exosomes in the concentrated supernatant to obtain exosomes concentrate; transfer the concentrate to a 30% sucrose / heavy water density pad (1.210 g / cm 3 ), centrifuge at 100,000g at 4°C for 210 minutes, collect the 5ml sucrose / heavy water density pad at the bottom, add PBS to dilute, transfer to an ultrafiltration centrifuge tube with a mol...

Embodiment 3

[0106] Human ESC-derived exosomes (ES-exo) encapsulated resveratrol (Res) by co-incubation method

[0107] ES-exo solution is from Example 1.

[0108] Quantitative standard curve determination of Res: Accurately weigh 1mg of Res powder and dissolve it in 1mL of methanol, then dilute the solution with methanol to prepare 50μg / mL, 20μg / mL, 10μg / mL Res in methanol standard solution, HPLC injection. The chromatographic conditions are as follows:

[0109] Chromatographic column: Zorbax Extend C-18, 150*4.6μm, 5–micro

[0110] Mobile phase: methanol: water = 95:5

[0111] Flow rate: 1mL / min

[0112] Column temperature: room temperature

[0113] Detection wavelength: 305nm

[0114] After obtaining the corresponding experimental results, the peak area (PA) of the chromatographic peak was plotted as a function of the Res concentration (C, μg / mL) (see attached image 3 ), obtain the quantitative standard curve of Res under this chromatographic separation condition as follows:

[0...

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Abstract

The invention relates to application of resveratrol-loading human multipotential stem cell exosome in preparing a drug for treating refractory healing skin wound related diseases. Compared with the prior art, embryonic stem cells and exosome inducing a human multipotential stem cell source are used as a drug carrier of the resveratrol, so that the biological effect of the resveratrol can be greatly improved; and by combining the function of the human multipotential stem cell exosome, the effect for treating the refractory healing skin wound related diseases can be greatly improved.

Description

technical field [0001] The invention belongs to the technical fields of cell biology, molecular biology and drug research and development, and in particular relates to the use of human pluripotent stem cell exosomes loaded with resveratrol in the preparation of drugs for treating refractory skin wound-related diseases. Background technique [0002] Exosomes are a type of extracellular vesicles secreted by cells with a diameter of about 30-150nm, which carry a variety of biologically active substances from donor cells, including DNA, RNA, protein, etc., into effector cells, effectively regulating the body's cell signal transduction guide. Studies have shown that exosomes secreted by stem cells can physiologically repair or remove damaged, diseased and aging cells in the body by delivering biologically active substances derived from stem cells to effector cells, and play anti-inflammatory, immune-regulating, and promote effector cell proliferation, migration and differentiatio...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K35/545A61K31/05A61P3/10A61P17/02A61P31/00C12N5/074
CPCA61K31/05A61K35/545A61P3/10A61P17/02A61P31/00C12N5/0696A61K2300/00
Inventor 汪洪
Owner 汪洪
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