Application of Lin28a gene overexpression AAV (adeno-associated virus) in promotion of wound healing

A technology for wound repair and bone damage repair, applied in the field of biomedicine, can solve problems such as positioning and specific function differences, and achieve the effects of inhibiting scar formation, promoting repair, and promoting dermal regeneration

Inactive Publication Date: 2016-03-30
重庆高圣生物医药有限责任公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although Lin28a and Lin28b have a high similarity in nucleic acid sequence, their location and specific functions in cells are different

Method used

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  • Application of Lin28a gene overexpression AAV (adeno-associated virus) in promotion of wound healing
  • Application of Lin28a gene overexpression AAV (adeno-associated virus) in promotion of wound healing
  • Application of Lin28a gene overexpression AAV (adeno-associated virus) in promotion of wound healing

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Example 1 Preparation of recombinant adeno-associated virus.

[0019] Construction of recombinant adeno-associated virus vectors.

[0020] The Lin28a gene was synthesized from the whole gene by Shanghai Bioengineering Co., Ltd., and constructed into the pUC57 vector to obtain the pUC57-Lin28a plasmid (see SEQ ID NO.1 for the sequencing result).

[0021] Restriction endonucleases EcoRI and BamHI (purchased from Takara Company) were used to double-digest pUC57-Lin28a and pAAV-MCS adeno-associated virus vector (purchased from Stratagene Company), and the restriction enzyme digestion system was: pAAV-MCS / pUC57-Lin28a plasmid: 10ul, EcoRI: 3ul, BamHI: 3ul, buffer: 4ul, H 2 O: 20ul. Bath in water at 37°C for 30 minutes, detect the digested products by agarose gel electrophoresis, and recover the Lin28a gene fragment and pAAV-MCS vector backbone (for the operation steps, refer to the instructions of the Takara Agarose Nucleic Acid Purification and Recovery Kit).

[0022] In...

Embodiment 2

[0050] Example 2 The effect of adeno-associated virus overexpressing Lin28a gene on skin wound healing.

[0051] Experimental animal trauma model making.

[0052] Healthy rats of Wistar species (purchased from the Experimental Animal Center of Nanfang Hospital) were randomly divided into 3 groups, 10 rats in each group, and were weighed and marked. The WISTAR rats were fed for 1 week, depilated and weighed the day before the experiment. After intraperitoneal anesthesia with 2% sodium pentobarbital solution (40 mg / kg body weight), three round skin full-thickness wounds with a diameter of 1 cm were made on both sides of the back, and the distance between the centers of the wounds was 1.5 cm. Cover the wound with sterile gauze. The dressing was removed every other day to expose the wound, and intramuscular penicillin was injected for one week after the operation to prevent anti-inflammatory and the local conditions were closely observed. After surgery, 0.2×10 was injected into...

Embodiment 3

[0058] Example 3 Effects of adeno-associated virus overexpression of Lin28a gene on burn repair.

[0059] laboratory animal Model making of severe burns.

[0060] The C57BL / 6 mice (Shanghai Experimental Animal Center, Chinese Academy of Sciences) were randomly divided into 3 groups with 10 mice in each group. Mice were fed adaptively for 1 week, shaved and weighed the day before the experiment. After intraperitoneal anesthesia with 2% sodium pentobarbital solution (40 mg / kg body weight), the limbs and tail base were fixed on the clean table with ventilation tape. Soak the water-soaked self-made cardboard to control the burn area (take about 1mm thick cardboard, cut it into 7cm×9cm size, and cut 2cm×1.5cm=3cm in the center of the paper with a blade 2 The area of ​​the rectangular window) was placed on the back of the mouse to expose the shaved area. 95% alcohol was dripped into the paper window with a dropper, soaked in the skin, pressed against the paper with large forceps...

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Abstract

The invention belongs to the field of biomedicine and particularly relates to an application of a recombinant AAV (adeno-associated virus) carrying a Lin28a gene in preparation of a wound healing promoting drug. The Lin28a gene is inserted to an AAV vector, and the recombinant AAV carrying the Lin28a gene is acquired. A preparation method of the recombinant AAV is simple, skin wound healing can be effectively promoted, regeneration of part of deeply burnt dermis can be remarkably promoted, bone injury and soft tissue injury healing can be promoted, and further, scar formation can be inhibited.

Description

technical field [0001] The invention belongs to the field of biomedicine, and in particular relates to the application of a Lin28a gene overexpression adeno-associated virus in promoting wound repair. Background technique [0002] At present, the healing of various wounds, burns, and surgical incisions is still a major clinical problem, especially the healing of local radiotherapy wounds, wound healing of diabetic patients, and healing of difficult-to-heal wounds such as large area burns or decubitus ulcers. Great pain, but also a huge burden on medical work. Wound repair is a complex and gradual process, but previous studies basically stayed at the stage of waiting for wound self-healing, and lacked active measures to promote wound repair. [0003] Lin28 is a conserved RNA-binding protein in higher eukaryotes. It is one of the four pluripotency factors that can reprogram human somatic cells into pluripotent stem cells. play an important role in terms of performance. In t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K38/17A61P17/02A61P19/08
CPCA61K38/1709A61K48/0008
Inventor 周勇申友锋
Owner 重庆高圣生物医药有限责任公司
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