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Application of miR-21 to preparing medicines for promoting healing of skin injury

A mir-21, 1.mir-21 technology, applied in drug combination, skin disease, pharmaceutical formulation, etc.

Inactive Publication Date: 2011-08-31
ARMY MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The role and experimental proof of miR-21 in wound healing have not been reported yet

Method used

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  • Application of miR-21 to preparing medicines for promoting healing of skin injury
  • Application of miR-21 to preparing medicines for promoting healing of skin injury
  • Application of miR-21 to preparing medicines for promoting healing of skin injury

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Example 1 Expression of miR-21 in the process of skin wound healing

[0022] 1.1 Establishment of mouse skin wound healing model

[0023] After Balb / c mice were anesthetized with 1% phenobarbital (30 mg / kg body weight), their backs were clipped and disinfected. The skin on both sides of the spine was resected in full thickness with a diameter of 8 mm. Injured mice were housed in single cages.

[0024] 1.2 Changes in miRNA expression profile during skin wound healing

[0025] On the 7th day after injury, the whole wound and the tissues within 1 mm of the wound margin were cut (n=6). Normal skin samples were used as controls. After the skin sample was crushed in liquid nitrogen, 1ml Trizol reagent was added to 100mg tissue to grind the tissue finely. After further homogenizing into a uniform suspension in a tissue homogenizer, standard methods were used to extract skin total RNA. About 50 μg of RNA from normal skin and wound skin were sent to Beijing Boao Biotechnol...

Embodiment 2

[0028] Example 2 Expression of miR-21 and its target genes in cells treated with TGF-β

[0029] C3H10T1 / 2 cells were incubated at 37°C and 5% CO with IMDM medium containing 10% calf serum (purchased by Shanghai Yingjun Company). 2 cultivated under conditions. When the cells reach 50%-60% confluency, replace with new medium containing 2.5ng / ml TGF-β. The medium was changed every day for 3 consecutive days. Cells were photographed daily. C3H10T1 / 2 cells treated with TGF-β for 3 days were extracted with Trizol reagent (Sigma-Aldrich, St Louis, MO) according to the instructions, and used for Northern blot detection of miR-21 expression and RT-PCR detection of miR-21 target genes expression changes. The results showed that the cells changed from fibroblast-like to multidendritic-like cells under the induction of TGF-β, and TGF-β significantly induced the expression of miR-21 in C3H10T1 / 2 cells (see image 3 ). At the same time, as the direct and indirect regulatory genes of m...

Embodiment 3

[0030] Example 3 Effect of antisense miR-21 on wound surface healing

[0031] 3.1 Antisense miR-21 inhibits TGF-β-induced miR-21 expression

[0032] Locked nucleic acid (LNA) oligonucleotide (Sigma-Aldrich, St Louis, MO) was chemically synthesized, and its center contained 8 consecutive bases containing locked nucleic acid (underline), antisense miR-21 locked nucleic acid LNA-antimiR-21 The sequence is: 5'-TCAACAT CAGTCTGA TAAGCTA-3', random control locked nucleic acid oligonucleotide LNA-scrambled sequence is: 5'-CATTAAT GTCGGACA ACTCAAT-3'.

[0033] First, we transfected C3H10T1 / 2 cells with LNA-antimiR-21 and LNA-scrambled, and extracted RNA for Northern blot detection of miR-21 after 3 days. It was confirmed that LNA-antimiR-21 could significantly inhibit the expression of miR-21, while The randomized control miRNA had no such effect (see Figure 4 ).

[0034] When the C3H10T1 / 2 cells reached 80% confluence, trypsinization prepared 5×10 4 / ml of single cell suspens...

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Abstract

The invention relates to an application of miR-21 to preparing medicines for promoting healing of skin injury, which belongs to the field of biological medicines, and the miR-21 can effectively promote the healing of skin injury. The miR-21 provided by the invention can be constitutively expressed in a normal skin, and has obviously increased expression in an injured skin. An antisense miR-21 with the function of antagonizing miR-21 can significantly inhibit the healing of skin injury, which indicates that the miR-21 can effectively promote the healing of skin injury.

Description

technical field [0001] The present invention relates to the application of non-coding RNA gene miR-21, in particular to the application of miR-21 in the preparation of medicines for promoting skin wound healing, especially the application of miR-21 in the preparation of medicines for promoting skin wound healing. technical background [0002] Fibroblasts are the main functional cells in the process of wound healing and have heterogeneity. During vertebrate wound healing, if dermal fibroblasts can be activated, scarless healing may occur. During epithelial regeneration, no hair grows, and the same is true during sweat gland regeneration. In fetal wound healing, fibroblasts of dermis-related tissues can receive permission signals to develop into different types of hair follicles at corresponding locations, and can also differentiate and regenerate other skin appendages. TGF-β is involved in the whole process of skin healing after trauma. TGF-β can chemoattract and act on fi...

Claims

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Application Information

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IPC IPC(8): A61K48/00A61K31/7105C12N15/113A61P17/02
Inventor 邹仲敏粟永萍王涛冯一梅孙慧勤王军平郝磊
Owner ARMY MEDICAL UNIV
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