Recombinant adipose-derived stem cell for overexpression of Hpgds and preparation method and application thereof

An adipose stem cell and overexpression technology, applied in the field of genetic engineering drugs, can solve problems that affect the quality of life of patients, are not easy to control, and aggravate the skin, and achieve good healing effects, reduce the number, and increase the content of new blood vessels and collagen tissue.

Active Publication Date: 2020-08-18
PLASTIC SURGERY HOSPITAL CHINESE ACAD OF MEDICAL SCI
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  • Summary
  • Abstract
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Problems solved by technology

For people with diabetes, once skin infection occurs, the symptoms of skin ulceration will increase, and it is not easy to control. If the treatment is not timely, it will develop in depth, and even eventually lead to amputation, seriously affecting the quality of life of the patient

Method used

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  • Recombinant adipose-derived stem cell for overexpression of Hpgds and preparation method and application thereof
  • Recombinant adipose-derived stem cell for overexpression of Hpgds and preparation method and application thereof
  • Recombinant adipose-derived stem cell for overexpression of Hpgds and preparation method and application thereof

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preparation example Construction

[0039] The invention provides a method for preparing recombinant adipose stem cells, comprising the following steps: constructing a lentiviral vector expressing Hpgds gene; extracting adipose stem cells; infecting adipose stem cells with the lentiviral vector expressing Hpgds gene to obtain recombinant adipose stem cells.

[0040] In the present invention, the construction of the lentiviral vector expressing the Hpgds gene preferably uses the lentiviral vector to package the Hpgds gene to obtain the lentiviral vector expressing the Hpgds gene. In the present invention, the construction method of the lentiviral vector expressing the Hpgds gene refers to the French Polyplus transfection kit.

[0041] In the present invention, the extraction of adipose stem cells is preferably performed by collagenase digestion. In the specific implementation process of the present invention, the following steps are included: S1) collecting the adipose tissue in liposuction; S2) cleaning and shre...

Embodiment 1

[0053] 1. Extraction of adipose-derived stem cells (ADSCs)

[0054] Take the fat tissue after liposuction (men and women are not limited), put it in a culture dish containing 1% double antibody and wash it 3-4 times, try to wash the blood vessels and residual substances in it, and then transfer the fat tissue into a small In a beaker, shred fully (until it becomes a paste), for about 15 minutes. Transfer the shredded adipose tissue into a 50ml centrifuge tube, add 2 times the volume of 1% (v / v) type Ⅰ collagenase to it, seal the tube with parafilm and put it in a 37°C water bath for digestion for 1 hour. Vibrate in a shaker, and the rotation speed is preferably in the range of 80-100 rpm. After the adipose tissue is completely digested (just observe that there is no blocky tissue), an equal amount of 10% FBS DMEM complete medium is added to terminate the digestion. Filter through a 200-mesh sieve, centrifuge at 1000rpm for 10min, add 5ml of complete medium to resuspend, inoc...

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Abstract

The invention provides a recombinant adipose-derived stem cell over-expressing an Hpgds gene and a preparation method and application thereof, and belongs to the technical field of genetic engineeringmedicines. The recombinant adipose-derived stem cell is obtained by infecting an adipose-derived stem cell with a lentiviral vector expressing the Hpgds gene. The preparation method comprises the following steps of: constructing a lentiviral vector for expressing the Hpgds gene; extracting adipose-derived stem cells; and infecting the adipose-derived stem cells with the lentiviral vector expressing the Hpgds gene to obtain recombinant adipose-derived stem cells. The recombinant adipose-derived stem cells proliferate on a three-dimensional biomaterial scaffold, and can safely and durably express the immunomodulatory gene Hpgds so as to accelerate healing of diabetic damaged skin and shorten the healing time.

Description

technical field [0001] The invention relates to the technical field of genetic engineering drugs, in particular to a recombinant fat stem cell expressing Hpgds, a preparation method and its application in diabetic wound repair. Background technique [0002] In the course of the onset of diabetes, 30% of patients will have skin infection, and often repeated attacks, lasting unhealed. For people with diabetes, once skin infection occurs, the symptoms of skin ulceration will increase, and it is not easy to control. If the treatment is not timely, it will develop in depth, and even eventually lead to amputation, seriously affecting the quality of life of the patient. According to statistics, amputations caused by diabetic ulcers account for more than 50% of patients with non-traumatic amputations. Therefore, skin ulcers in diabetic patients have attracted more and more attention from medical workers and researchers. [0003] In diabetes, the number and function of neutrophils a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/10C12N15/861A61K35/28A61P17/02A61P3/10
CPCC12N9/90C12Y503/99002C07K14/4705C12N5/0667C12N15/86A61K35/28A61P17/02A61P3/10C12N2510/00C12N2710/10043
Inventor 肖苒欧阳龙邱道静傅歆杨志岗
Owner PLASTIC SURGERY HOSPITAL CHINESE ACAD OF MEDICAL SCI
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