Application of long-chain non-coding RNA-NEAT1, adeno-associated virus overexpressing RNA-NEAT1 and application

An RNA-NEAT1, long-chain non-coding technology, applied in sgRNA and adeno-associated virus and application fields, can solve the problems of insufficient corneal donors, corneal endothelial dystrophy, and high price

Active Publication Date: 2021-09-07
山东第一医科大学附属青岛眼科医院
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Although keratoplasty has been largely successful in the treatment of Fuchs' endothelial dystrophy, it is still invasive, insufficient in number of corneal donors, and expensive, prompting the development of other treatments

Method used

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  • Application of long-chain non-coding RNA-NEAT1, adeno-associated virus overexpressing RNA-NEAT1 and application
  • Application of long-chain non-coding RNA-NEAT1, adeno-associated virus overexpressing RNA-NEAT1 and application
  • Application of long-chain non-coding RNA-NEAT1, adeno-associated virus overexpressing RNA-NEAT1 and application

Examples

Experimental program
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Effect test

Embodiment 1

[0030] Adeno-associated virus AAV9-Neat1-sgRNA construction

[0031] The present invention uses the long-chain non-coding RNA-NEAT1 to design sgRNA, wherein the ID of the long-chain non-coding RNA-NEAT1 is 66961, and the nucleotide sequence of the sgRNA is shown in SEQ ID NO:1. The sgRNA was ligated into the GV639 adenovirus vector, and the ligated product was transformed with TOP10. Colony PCR obtained a positive clone and was sequenced to obtain the adeno-associated virus AAV9-Neat1-sgRNA expressing the sgRNA with the correct sequence. Among them, sgRNA was synthesized by Jikai Gene Chemical Technology Co., Ltd.; GV639 adenovirus vector and TOP10 competence were purchased from Jikai Gene Chemical Technology Co., Ltd.; colony PCR primers were purchased from Shanghai Jierui Bioengineering Co., Ltd. The final packaged adeno-associated virus AAV9-Neat1-sgRNA had a titer of 2.18×10 12 v.g. / mL.

Embodiment 2

[0033] Investigation on the treatment of Fuchs' corneal endothelial dystrophy

[0034]C57BL / 6J female mice aged 8 to 12 weeks (about 20 g in weight) were selected to construct the Fuchs' corneal endothelial dystrophy model, and the therapeutic effect on Fuchs' corneal endothelial dystrophy was investigated. The method of model construction refers to the following literature:

[0035] Liu et al.,Ultraviolet A light induces DNA damage and estrogen-DNAadducts in Fuchs endothelial corneal dystrophy causing females to be more affected.PNAS Latest Articles.,January 7,2020117(1)573-583.

[0036] In this experiment, C57BL / 6J female mice were randomly divided into three groups, Neat1 overexpression group, NAC (N-acetyl-L-cysteine) group and control group.

[0037] Neat1 overexpression group: 4 weeks before modeling, inject the adeno-associated virus AAV9-Neat1-sgRNA of Example 1 into the anterior chamber of the mice. Long non-coding RNA-Neat1 was overexpressed; for modeling, C57BL / 6J...

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Abstract

The invention provides an application of long-chain non-coding RNA-NEAT1, sgRNA overexpressing the RNA-NEAT1, an adeno-associated virus and an application and belongs to the technical field of medicines. The invention provides the application of long-chain non-coding RNA-NEAT1 (Ribonucleic Acid-NEAT1) to preparation of medicines for treating Fuchs' corneal endothelial malnutrition. Overexpression of the long-chain non-coding RNA-NEAT1 can effectively reduce corneal endothelial injury and relieve corneal edema symptoms, and the long-chain non-coding RNA-NEAT1 can be used for treatment of Fuchs' corneal endothelial malnutrition, operation steps and risks of treatment are greatly simplified, corneal transplantation operation is not needed, the source is sufficient, and the treatment cost is low.

Description

technical field [0001] The invention belongs to the technical field of medicine, and specifically relates to the application of long-chain non-coding RNA-NEAT1, sgRNA and adeno-associated virus overexpressing RNA-NEAT1 and the application thereof. Background technique [0002] Fuchs' endothelial corneal dystrophy (FECD), also known as guttate cornea, is a common hereditary corneal endothelial degeneration disease, mainly manifested by edema of corneal stroma and epithelium, causing significant vision loss. Currently, the clinical treatment for Fuchs' endothelial dystrophy is corneal transplantation. Although keratoplasty has been largely successful in treating Fuchs' endothelial corneal dystrophy, it is still invasive, insufficient in number of corneal donors, and expensive, prompting the development of other treatments. Contents of the invention [0003] In order to solve the above problems, the present invention provides the application of long-chain non-coding RNA-NEAT...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/7088A61K9/00A61K47/46C12N15/113C12N15/864A61P27/02
CPCA61K31/7088A61K9/0048A61K47/46C12N15/113C12N15/86A61P27/02C12N2310/10C12N2750/14143C12N2310/20C12N2310/14
Inventor 史伟云王群周庆军窦圣乾姜慧张彬
Owner 山东第一医科大学附属青岛眼科医院
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