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Application of miR-155 in preparing medicine for curing gastricism

1. The technology of mir-155 and gastritis, applied in gene therapy, drug combination, pharmaceutical formula, etc., can solve the research reports on the diagnosis and treatment of inflammation related to miRNAs and Hp infection, etc.

Inactive Publication Date: 2010-03-17
ARMY MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] After searching the domestic and foreign literature of the prior art, there has been no research report on the diagnosis and treatment of miRNAs and Hp infection-related inflammation.

Method used

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  • Application of miR-155 in preparing medicine for curing gastricism
  • Application of miR-155 in preparing medicine for curing gastricism
  • Application of miR-155 in preparing medicine for curing gastricism

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Example 1: Establishment of gastric epithelial cell model infected by Hp

[0018] The Hp 26695 standard strain was cultivated to the logarithmic growth phase, the bacteria were collected by centrifugation at 5000×g, and the Hp bacterial suspension was prepared with RPMI 1640 medium without antibiotics, and the bacterial concentration was adjusted by measuring the A600 absorbance value (1A600=1×108cfu / ml). Use RPMI 1640 (penicillin 100U / ml; streptomycin 100U / ml) containing 10% calf serum as the complete medium to culture the human gastric epithelial cell line GES-1. When the growth rate reaches 80%, discard the medium . According to the multiplicity of infection (multiplicity of infection, MOI) (number of bacteria: cell number) is 100:1, add Hp, 37 ℃, 5% CO 2 co-cultured for 24 hours under the same conditions.

Embodiment 2

[0019] Example 2: Changes of miRNAs expression profile in Hp infected gastric epithelial cells detected by miRNAs chip

[0020] Total RNA was extracted by Trizol method, 1×10 7 Add 1ml Trizol to the cells, pipette vigorously repeatedly with the tip of the pipette, oscillate fully on the vortex to completely lyse the cells, and let stand at room temperature for 5 minutes; add chloroform at a ratio of 0.2ml chloroform / 1ml Trizol, vortex fully, and let stand at room temperature for 10 minutes; 4°C, 12000 Centrifuge at ×g for 15 minutes, take the upper layer of colorless liquid into a new 1.5ml centrifuge tube; add isopropanol according to the ratio of 0.5ml isopropanol / 1ml Trizol, mix well, and place at room temperature for 5-10min to form RNA precipitation; 4 Centrifuge at 12000×g for 10min at ℃, discard the supernatant; add at least 1ml 75% ethanol / 1ml Trizol to suspend the RNA pellet; centrifuge at 7500×g for 5min at 4℃, discard the supernatant; after air drying for 5-10min,...

Embodiment 3

[0025] Example 3: Real-time PCR technology to detect the expression of miRNAs

[0026] In this example, TaqMan miRNA Assay Kit (Applied Biosystems) was used for Real-time PCR detection of miRNAs.

[0027] First, the total cellular RNA was extracted according to the Trizol method described in Example 1, and 10 ng of the above-mentioned total RNA was used as a template, and then cDNA was synthesized using the miRNAs-specific primers included in the kit. The reaction system was as follows: dNTPs (100 mM) 0.15 μl; MultiScribe reverse transcriptase 1 μl; 10×RT buffer 1.5 μl; RNase inhibitor (20U / μl) 0.19 μl; RT primer 3 μl; total RNA 5 μl; DEPC H 2 O 4.16 μl. Reaction conditions: 16°C for 30 minutes, 42°C for 30 minutes, 85°C for 5 minutes, and store at 4°C.

[0028] Then use the cDNA as a template, and use the miRNAs-specific primers and reverse primers that come with the kit to perform TaqMan probe method Real-time PCR. The system is as follows: 2×PCR mix 10μl; cDNA 2μl; Probe...

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Abstract

The invention relates to an application of a non-coding small RNA gene miR-155 in preparing a medicine for curing gastricism. The gastricism can be caused by helicobacter pylori infection. The miR-155is highly expressed in helicobacter pylori infected gastric epithelial cells and human gastric mucosa tissues; furthermore, after the gastric epithelial cells GES-1 are transfected by a miR-155 simulator or a miR-155 inhibitor respectively, the miR-155 simulator can obviously reduce the protein secretion level of inflammatory factor lnterleukin 8, and growth-related oncogene Alpha, thus indicating that the miR-155 can effectively inhibit inflammatory response related to the helicobacter pylori infection.

Description

technical field [0001] The present invention relates to the application of non-coding small RNA gene miR-155, in particular to the application of miR-155 in the preparation of medicine for treating gastritis, especially the application of miR-155 in the preparation of medicine for treating gastritis caused by Helicobacter pylori infection. Background technique [0002] MicroRNAs (microRNAs, miRNAs) are a class of endogenous non-coding RNA molecules with a length of about 20-25 nt, which are ubiquitous in eukaryotes. ) 3' untranslated region complementary binding, induce target mRNA degradation or inhibit target mRNA translation, so as to achieve post-transcriptional gene regulation. Many evidences show that miRNAs play an important regulatory role in various biological processes such as cell proliferation, development, and differentiation, and are closely related to tumorigenesis. [0003] In recent years, more and more attention has been paid to the relationship between mi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61P1/04
Inventor 肖斌邹全明刘真毛旭虎黎伯胜唐彬朱恩东李娜郭刚
Owner ARMY MEDICAL UNIV
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