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Method and application of MicroRNA (micro ribonucleic acid) 221-3p in preparation of epidermal cells

A high-expression technology of epidermal cells, applied in the field of preparing skin grafts and promoting the proliferation of epidermal cells, can solve the problems that there is no research and application of MicroRNA-221-3p, achieve low cost, overcome slow proliferation, and promote proliferation

Active Publication Date: 2015-04-29
SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] There are more and more studies on MicroRNA and cell function, but there is no literature report on the study and application of MicroRNA-221-3p overexpression to promote epidermal cell proliferation

Method used

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  • Method and application of MicroRNA (micro ribonucleic acid) 221-3p in preparation of epidermal cells
  • Method and application of MicroRNA (micro ribonucleic acid) 221-3p in preparation of epidermal cells
  • Method and application of MicroRNA (micro ribonucleic acid) 221-3p in preparation of epidermal cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1: Obtaining epidermal cells stably and highly expressing MicroRNA221-3p

[0043] 1. Obtaining epidermal cells

[0044] Adult body skin comes from Plastic Surgery of Changhai Hospital. Epidermal cells are obtained by primary culture.

[0045] After taking the foreskin, put it in a sterile centrifuge tube containing sterile PBS or DMEM medium, and operate as soon as possible. Wipe the foreskin repeatedly with an appropriate amount of 75% ethanol cotton ball in a sterile petri dish for about 1 minute. Change the foreskin to another sterile petri dish, and wash thoroughly with Phosphate Buffered Saline (PBS) (both penicillin and streptomycin concentrations are 100IU / ml) containing bi-antibody phosphate buffered saline. Wash twice with liquid. Cut the skin into strips with a width of 1-2 mm, soak in 0.25% Dispase enzyme solution about 5 times the tissue, and digest at 4°C for 16-18 hours. Tear off the epidermis gently from the removed skin, and wash twice with anti-PB...

Embodiment 2

[0068] Example 2: EC high The functional study and biological characterization of stable and highly expressed MicroRNA221-3p

[0069] 1. Cell growth curve

[0070] The cells of each group were seeded in a 24-well plate with 10,000 cells per well, cultured for 7 days, trypsinized every day to form a single cell suspension, and the cell counting experiment was carried out with the help of a cell counting plate and a microscope. 3 duplicate holes, and the experiment was repeated 3 times. It turns out that EC high Group cell proliferation ability is stronger than EC nc ,Such as figure 2 Shown.

[0071] 2. Immunohistochemical detection of epidermal cell characteristics

[0072] 1. EC vaccination high , EC nc Cell crawler

[0073] 2. Divided into 2 groups, one is the experimental group and the other is the antibody control group

[0074] 3. 4%BSA blocks non-specific binding sites

[0075] 4. Add the primary antibody at a concentration of 1:200

[0076] 5. Incubate at 37°C for half an hou...

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Abstract

The invention relates to the field of biotechnology, in particular to a novel application of MicroRNA (micro ribonucleic acid) 221-3p to promotion of epidermal cell proliferation, and provides an application of MicroRNA 221-3p to preparation of a reagent for promoting epidermal cell proliferation. The reagent comprises but is not limited to MicroRNA 221-3p, a recombinant vector containing a MicroRNA 221-3p coding gene, a recombinant virus containing the MicroRNA 221-3p coding gene, and a recombinant virus vector containing the MicroRNA 221-3p coding gene. The experiment proves that MicroRNA 221-3p can inhibit expression of PTEN (phosphatase and tensin homolog deleted on chromosome ten) and remarkably increase epidermal cell proliferation speed, and the characteristics of epidermal cells are not lost; MicroRNA 221-3p relates to epidermal cell proliferation. The invention provides novel thought and mode for quickly proliferating the epidermal cells and quickly obtaining tissue engineering skin.

Description

Technical field [0001] The present invention relates to the field of biotechnology, in particular to a new use of MicroRNA221-3p in promoting the proliferation of epidermal cells, that is, a method for promoting the proliferation of epidermal cells, and the application of the method to the preparation of skin grafts. Background technique [0002] The skin has an important protective effect. Because it is located on the body surface, various skin injuries often occur. Skin wound repair is an important part of burn wound treatment. Following the use of various skin flaps and microskins for transplantation, in recent years, there has been an emergence of tissue engineering technology for epidermal cell culture and in vitro skin tissue construction, and has become a medical bioengineering Research hotspots in the field (Rhett JM, Ghatnekar GS, Palatinus JA, O'Quinn M, Yost MJ, Gourdie RG. Novel therapies for scar reduction and regenerative healing of skin wounds. Trends Biotechnol. 2...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/88C12N15/85C12N15/86C12N5/10A61L27/38A61L27/60
Inventor 刘厚奇仵敏娟徐辰夏照帆王越徐莎熊俊
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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