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Human pluripotent stem cell exosome loaded with resveratrol, and preparation method and application thereof

A technology of human pluripotent stem cells and resveratrol, which is applied in the direction of non-embryonic pluripotent stem cells, artificially induced pluripotent cells, animal cells, etc., can solve the problems that the treatment potential of diseases and injuries has not been fully utilized and needs further confirmation

Active Publication Date: 2019-08-16
上海睿泰生物科技股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

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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  • Human pluripotent stem cell exosome loaded with resveratrol, and preparation method and application thereof
  • Human pluripotent stem cell exosome loaded with resveratrol, and preparation method and application thereof
  • Human pluripotent stem cell exosome loaded with resveratrol, and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0081] Culture of human embryonic stem cells (ESCs) and extraction and identification of exosomes

[0082] Place a layer of embryonic stem cell matrigel (ESC-Qualified BD Matrigel, BD Sparks, MD, USA), move ESCs into the petri dish, and add mTeSR1 serum-free medium (StemCell Vancouver,BC,Canada), in a thermostat (37℃, 5%CO 2 , Saturated humidity) culture, and collect the medium that is changed every day. Filter the medium through a 0.22 micron pore filter membrane and centrifuge at 10000g at 4°C for 30 minutes to remove cell debris; use an ultrafiltration tube with a molecular weight of 100KD and centrifuge (3500g, 15min) to trap exosomes in the concentrated supernatant to obtain exosomes Concentrate; transfer the concentrate to a 30% sucrose / dense water density pad (1.210g / cm 3 ), centrifuge at 100000g for 210 minutes at 4℃, collect 5ml sucrose / dense water density pad at the bottom, add PBS to dilute, transfer to ultrafiltration centrifuge tube that can retain 100KD molecular w...

Embodiment 2

[0087] Cultivation of human induced human pluripotent stem cells (iPSCs) and extraction and identification of exosomes

[0088] Place a layer of embryonic stem cell matrigel (ESC-Qualified BD Matrigel, BD Sparks, MD, USA), move iPSC into the petri dish, add mTeSR1 serum-free medium (StemCell Vancouver,BC,Canada), in a thermostat (37℃, 5%CO 2 , Saturated humidity) culture, and collect the medium that is changed every day. Filter the medium through a 0.22 micron pore filter membrane and centrifuge at 10000g at 4°C for 30 minutes to remove cell debris; use an ultrafiltration tube with a molecular weight of 100KD and centrifuge (3500g, 15min) to trap exosomes in the concentrated supernatant to obtain exosomes Concentrate; transfer the concentrate to a 30% sucrose / dense water density pad (1.210g / cm 3 ), centrifuge at 100000g for 210 minutes at 4℃, collect 5ml sucrose / dense water density pad at the bottom, add PBS to dilute, transfer to ultrafiltration centrifuge tube that can retain ...

Embodiment 3

[0093] Human ESC-derived exosomes (ESC-Exos) encapsulate resveratrol (Res) by co-incubation

[0094] The ESC-Exos solution is from Example 1.

[0095] Determination of the quantitative standard curve of Res: accurately weigh 1mg of Res powder and dissolve it in 1mL of methanol, and then dilute the solution with methanol to prepare 50μg / mL, 20μg / mL, 10μg / mL Res methanol standard solution, HPLC injection. The chromatographic conditions are as follows:

[0096] Column: Zorbax Extend C-18, 150*4.6μm, 5-micro

[0097] Mobile phase: methanol: water = 95:5

[0098] Flow rate: 1mL / min

[0099] Column temperature: room temperature

[0100] Detection wavelength: 305nm

[0101] After obtaining the corresponding experimental results, plot the peak area (PA) of the chromatographic peak as a function of the Res concentration (C, μg / mL) (as attached image 3 ) To obtain the following quantitative standard curve of Res under the chromatographic separation conditions:

[0102] PA=38.42C+71.12, R 2 =0.9991...

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Abstract

The invention relates to a human pluripotent stem cell exosome loaded with resveratrol, and a preparation method and an application thereof. The invention provides the following applications of a human pluripotent stem cell exosome and the human pluripotent stem cell exosome loaded with resveratrol: an application in preparation of medicines or cosmetics for preventing stretch marks, repairing stretch marks or treating stretch marks, an application in preparation of skin anti-aging medicines or cosmetics, and an application in preparation of medicines or cosmetics for treating skin scars, skinfreckles, pox, pimples, acne and the like. Compared with the prior art, the exosome derived from embryonic stem cells and induced human pluripotent stem cells is used as a drug carrier of resveratrol, the biological effect of resveratrol can be greatly improved, and with combination of the self function of the human pluripotent stem cell exosome, a better therapeutic effect on diseases can be played.

Description

Technical field [0001] The invention belongs to the technical fields of cell biology, molecular biology and drug research and development, and particularly relates to human pluripotent stem cell exosomes loaded with resveratrol and a preparation method and application thereof. Background technique [0002] Exosomes are a type of extracellular vesicles with a diameter of about 30-150nm secreted by cells. They carry a variety of biologically active substances derived from donor cells, including DNA, RNA, proteins, etc., into effector cells, and effectively regulate cell signal transduction in the body. guide. Studies have shown that exosomes secreted by stem cells deliver biologically active substances derived from stem cells to effector cells to physiologically repair or eliminate damaged, diseased and senescent cells in the body, play anti-inflammatory, regulate immunity, and promote effector cell proliferation, migration and differentiation , Promote angiogenesis and other func...

Claims

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Application Information

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IPC IPC(8): A61K35/545A61K31/05A61P17/18A61P17/10A61P17/02A61P17/00A61K8/99A61K8/34A61Q19/08A61Q19/00C12N5/074
CPCA61K35/545A61K31/05A61K8/99A61K8/347A61Q19/08A61Q19/00C12N5/0696A61K2300/00
Inventor 汪泱吴复跃何振东
Owner 上海睿泰生物科技股份有限公司
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