Application of down-regulated circular gene expression reagent in preparing medicine for preventing and/or treating pulmonary fibrosis and down-regulated circular gene expression medicine

A lung fibrosis and lung fibroblast technology, applied in gene therapy, drug combination, genetic engineering, etc., can solve the problems of limited application, difficult donor source, high cost, etc., and achieve the effect of inhibiting cell transdifferentiation

Active Publication Date: 2020-05-05
BINZHOU MEDICAL COLLEGE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There is no definite curative drug for this disease, and lung transplantation is the only important means of treatment for end-stage pulmonary fibrosis, but the high cost and difficult source of donors limit its application

Method used

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  • Application of down-regulated circular gene expression reagent in preparing medicine for preventing and/or treating pulmonary fibrosis and down-regulated circular gene expression medicine
  • Application of down-regulated circular gene expression reagent in preparing medicine for preventing and/or treating pulmonary fibrosis and down-regulated circular gene expression medicine
  • Application of down-regulated circular gene expression reagent in preparing medicine for preventing and/or treating pulmonary fibrosis and down-regulated circular gene expression medicine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Expression of circRNA-0007535

[0040] 1. Method

[0041] 1 Cell culture and grouping: MRC-5 cells in MEM medium containing 10% fetal bovine serum, A549 cells in F12 medium containing 10% fetal bovine serum, 37°C, 5% CO 2 cultured in a humidified incubator. Cells in the logarithmic growth phase were selected to make a single cell suspension and seeded in a six-well culture plate. A blank control group and a TGF-β1 stimulation group were set up. When the cells grew to 70-80% confluent, the blank control group was cultured in serum-free medium to synchronize cell growth. In the TGF-β1 stimulation group, the final concentration of 5nM TGF-β1 was stimulated for 72 hours to complete the transformation of the cells into fibroblasts.

[0042] 2qRT-PCR validation analysis of expression changes of circRNA-0007535

[0043] (1) Use the RNA extraction reagent Trizol to extract the total cellular RNA, and use the NANO drop 2000 spectrophotometer to measure the absorbance values ...

Embodiment 2

[0049] Effects of siRNA interference fragments specific for circRNA-0007535 on the transdifferentiation of fibroblasts

[0050] 1. Method

[0051] 1. Cell culture and grouping: Select MRC-5 cells in logarithmic growth phase, make single cell suspension with 0.25% trypsin, and inoculate in 6-well plate. When the cells grew to 70-80% confluence, the transfection group and the transfection control group respectively used riboFECTTM CP transfection reagent to transfect cells with siRNA interference fragments, overexpression plasmids and control cells at a final concentration of 50 nM, and the blank control group was given no Cultured in serum medium, the model group was stimulated with 5nM TGF-β1, and the cells were collected after 72 hours of culture.

[0052] 2Westernblot detection of expression changes of pulmonary fibrosis-related proteins

[0053] Strictly follow the instructions of the Western and IP cell lysate kits to extract the protein of each group of cells, follow th...

Embodiment 3

[0057] Effects of siRNA interference fragments specific to circRNA-0007535 and overexpression plasmids on the proliferation and migration of fibroblasts

[0058] 1. Method

[0059] RTCA real-time label-free cell proliferation and migration analysis: select MRC-5 cells in the logarithmic growth phase, make single-cell suspension with 0.25% trypsin, and inoculate in six-well plates. When the cells grew to 70-80% confluence, the transfection group and the transfection control group respectively used the riboFECTTM CP transfection reagent to transfect the siRNA interference fragment, the overexpression plasmid and the control transfection cells, the final concentration was 50nM, and the blank control group was given Cultured in serum-free medium, the model group was stimulated with 5nM TGF-β1, cells were collected after 72 hours of culture, the collected cells were inoculated on E-Plate and CIM Plate test boards and placed in the detector, and parameters were set to detect within ...

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Abstract

The invention relates to an application of a down-regulated circular RNA-0007535 expression reagent in preparing medicine for preventing and / or treating pulmonary fibrosis and down-regulated circularRNA-0007535 expression medicine and belongs to the technical field of medicine preparation. The invention provides the application of the down-regulated circular RNA-0007535 expression reagent in preparing the medicine for preventing and / or treating the pulmonary fibrosis. The nucleotide sequence of circular RNA-0007535 is shown by SEQ ID NO.1. The obtained medicine prepared from the the down-regulated circular RNA-0007535 expression reagent can prevent and / or treat the pulmonary fibrosis. As potential molecules and drug targets for treating the pulmonary fibrosis, the circular RNA-0007535 provide a totally new perspective and field for exploring the regulatory mechanism of gene expression in the occurrence and development of the pulmonary fibrosis and seeking intervention means / medicine and have good application prospects.

Description

technical field [0001] The invention relates to the technical field of medicine preparation, in particular to the application of a reagent for down-regulating the expression of circular RNA-0007535 in the preparation of medicines for preventing and / or treating pulmonary fibrosis and the medicine for down-regulating the expression of circular RNA-0007535. Background technique [0002] Idiopathic pulmonary fibrosis (IPF) refers to lung interstitial disease that occurs in the absence of a clear etiology, and progressively aggravates, eventually leading to respiratory failure and death. The pathogenesis of IPF is still unclear, and it may be related to the damage of alveolar epithelial cells caused by the interaction of genetic factors and environmental exposure, and the fibroplasia caused by abnormal lung repair. It manifests as cough, irreversible dyspnea, decreased lung function, and respiratory failure. Imaging examinations show bilateral reticular opacities of bronchiectasi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/7088A61K31/713A61P11/00
CPCA61K31/713C12N2310/113C12N2320/30C12N15/113A61P43/00C12N2310/14A61P11/00A61K31/7088
Inventor 宋晓冬李明娥张瑾锦徐磐李洪斌李蓉蓉
Owner BINZHOU MEDICAL COLLEGE
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