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Application of hsa-mir-382-5p in preparation of kits for diagnosing retinal degenerative diseases

A technology of kits and reagents, applied in the field of biomedicine, can solve problems such as poor curative effect, increased patient pain, and large differences

Active Publication Date: 2021-12-14
JIANGSU PROVINCE HOSPITAL THE FIRST AFFILIATED HOSPITAL WITH NANJING MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, anti-VEGF drug therapy also has a series of defects, often requiring multiple intraocular injections, which increases the suffering of patients and increases the economic burden, and the therapeutic effect of anti-VEGF drugs varies greatly among different patients. After repeated anti-VEGF treatments, the patient's vision is still not improved or the improvement is not obvious

Method used

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  • Application of hsa-mir-382-5p in preparation of kits for diagnosing retinal degenerative diseases
  • Application of hsa-mir-382-5p in preparation of kits for diagnosing retinal degenerative diseases
  • Application of hsa-mir-382-5p in preparation of kits for diagnosing retinal degenerative diseases

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Experimental program
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Effect test

Embodiment 1

[0040] In the early stage, we used circRNA chips to detect the expression of circRNA in induced pluripotent stem cells-retinal pigment epithelium (iPSC-RPE) at different differentiation time points, and found that the expression of hsa_circ_0001543 increased with The degree of differentiation of RPE cells increased, which highly suggested the potential important role of hsa_circ_0001543 in the process of RPE cell differentiation. In order to further clarify the pathological significance of hsa_circ_0001543 in retinal degenerative diseases, we first detected the expression of hsa_circ_0001543 in the retinal tissues of normal controls and patients with retinal degenerative diseases.

[0041] experimental method:

[0042] 1. Collection of blood samples from normal controls and patients with retinal degenerative diseases:

[0043] Blood samples from 9 patients with clinically diagnosed retinal degenerative diseases and 6 normal controls were collected for RNA extraction. 5-8 mL ...

Embodiment 2

[0060] Example 2 hsa_circ_0001543 can act as a miRNA sponge to adsorb hsa-miR-382-5p and regulate its expression

[0061] We found a potential interaction site between hsa_circ_0001543 and miRNA hsa-miR-382-5p through online sequence alignment software ( figure 2 ), suggesting that hsa_circ_0001543 may act as a miRNA sponge to adsorb hsa-miR-382-5p. In order to confirm this point of view, we used dual luciferase reporter gene experiments to detect the interaction between the two, and referred to Example 1 to detect the expression regulation effect of hsa_circ_0001543 on hsa-miR-382-5p by Q-PCR.

[0062] experimental method:

[0063] 1. Construction of reporter gene vector:

[0064] Using human genomic DNA as a template, the hsa_circ_0001543 sequence of the wild type (WT) was amplified by PCR, the PCR product was purified and recovered, and the T-A cloning kit of Promega Company was used to clone and construct the vector with the hsa_circ_0001543 sequence of WT, and the reac...

Embodiment 3

[0089] Example 3 hsa-miR-382-5p intracellular localization analysis

[0090] The expression of hsa-miR-382-5p in ARPE19 cells was localized by the quantitative detection experiment of nucleocytoplasmic separation RNA, U6 was set as the reference for the specific localization of the nucleus, GAPDH was the reference for the specific localization of the cytoplasm. is PARIS TM Kit (Invitrogen Company), RNA quantitative detection method refers to Example 1. Operate in strict accordance with the kit instructions.

[0091] Experimental results:

[0092] Nucleoplasmic separation and RNA extraction found that hsa-miR-382-5p was mainly expressed in the cytoplasm of ARPE19 cells ( Figure 5 ).

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Abstract

The invention discloses the application of hsa-miR-382-5p in preparing a kit for diagnosing retinal degenerative diseases. A diagnostic kit for retinal degenerative diseases, comprising a reagent for detecting the expression level of hsa-miR-382-5p. Application of a substance inhibiting the expression of hsa-miR-382-5p in the preparation of a drug for treating retinal degenerative diseases. The study found that the increased expression of hsa-miR-382-5p can be used as an auxiliary diagnosis basis for retinal degenerative diseases and help realize the early diagnosis of retinal degenerative diseases. The damaging effect of hsa-miR-382-5p on the retina suggests that it can be used to prepare and screen drugs for treating retinal degenerative diseases. The invention provides a new detection marker and therapeutic target for the retinal degenerative disease, and provides brand new molecular information and biological basis for the diagnosis and treatment of the disease.

Description

technical field [0001] The invention belongs to the field of biomedicine and relates to the application of microRNA hsa-miR-382-5p in preparing a kit for diagnosing retinal degenerative diseases. Background technique [0002] Blinding eye disease is the third largest disease affecting human health reported by the World Health Organization. There are about 250 million visually disabled people in the world, and China accounts for nearly 20%. There is no effective treatment for retinal degenerative diseases. It is an important blinding eye disease, a scientific problem and a major public health problem that needs to be solved urgently. The pathogenesis of retinal degenerative diseases is diverse. According to the etiology, they can be divided into hereditary retinal degenerative diseases caused by single gene defects and complex retinal degenerative diseases caused by multifactorial damage. [0003] Hereditary retinal degenerative diseases are closely related to genetic factor...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6883A61K45/00A61P27/02
CPCC12Q1/6883A61K45/00A61P27/02C12Q2600/136C12Q2600/158C12Q2600/178
Inventor 陈雪刘庆淮蒋超杨戴帝孙汝许赵晨计江东
Owner JIANGSU PROVINCE HOSPITAL THE FIRST AFFILIATED HOSPITAL WITH NANJING MEDICAL UNIV
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