Application of miR-199a-3p in preparation of medicine for resisting hepatitis B virus

A mir-199a-3p, 1.mir-199a-3p technology, applied in antiviral agents, drug combinations, pharmaceutical formulations, etc., can solve problems such as incomplete elucidation of the mechanism of action

Inactive Publication Date: 2011-01-26
TIANJIN MEDICAL UNIV
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  • Abstract
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Problems solved by technology

At present, the mechanism of HBV persistent infection and the mechanism of interaction between host and HBV have not been fully elucidated

Method used

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  • Application of miR-199a-3p in preparation of medicine for resisting hepatitis B virus
  • Application of miR-199a-3p in preparation of medicine for resisting hepatitis B virus
  • Application of miR-199a-3p in preparation of medicine for resisting hepatitis B virus

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Embodiment 1

[0023] 1. Cell Culture

[0024] HepG2 2.2.15 and HEK293 cell lines were cultured in DMEM and MEM-α (GIBCO, USA) basal medium respectively, both mediums were supplemented with 10% fetal bovine serum, 20mM HEPES, 2mM glutamine, 100 units / ml penicillin and 100 μg / ml streptomycin, all cells were placed in 5% CO 2 In a 37°C incubator, subculture every 2-3 days.

[0025] 2. Cell Transfection

[0026] The 2'methoxy (2'-OMe) modified antisense oligonucleotide (ASO) that is completely complementary to human mature miRNA was synthesized by IDT, and the cells were prepared in 10 4Each well was inoculated in a 96-well plate, and the next day, the cells were transfected with 50nM miRNA 2'-OMe-ASO or 5ng / μL miRNA expression vector with Lipofectamine 2000, and each group of experiments was done in 3 replicate wells, according to the laboratory Conventional methods and refer to instructions. After 48 hours, the culture medium was changed, and after 24 hours, the cell supernatant was coll...

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Abstract

The invention discloses application of miR-199a-3p in the preparation of a medicine for resisting hepatitis B virus (HBV). MiR-199a-3p is a nucleotide sequence described in the sequence table SEQ ID NO:1. Experimental results show that miR-199a-3p can effectively reduce the expression of HBsAg, but can not influence the proliferation of HepG2 2.2.15 cells. Real time PCR (Polymerase Chain Reaction) to HBV DNA quantitatively displays that miR-199a-3p inhibits the replication of the virus. Bioinformatic analysis shows that the HBsAg coding area has a binding site of miR-199a-3p; fluorescence report carrier experiments verify the influence of miR-199a-3p on HBV transcription products; and meanwhile, real time PCR and western blot are used for further verifying that miR-199a-3p has inhibition action on the expression of HBsAg. The results imply that miR-199a-3p plays an important role in regulating and controlling the replication of HBC and can be prepared into a medicine for resisting HBV.

Description

technical field [0001] The invention belongs to the technical field of biomedical materials. It relates to a use of miRNA. Background technique [0002] miRNA is an endogenous small RNA of about 22 nucleotides, expressed in almost all known eukaryotic cells, regulates gene expression through the miRNA-mediated RNA silencing pathway, and regulates cell proliferation, differentiation and apoptosis. At the same time, miRNA also plays an important role in regulating virus replication and viral gene expression. miR-122 can increase the replication of hepatitis C virus (Jopling CL, et al. Modulation of hepatitis C virus RNA abundance by aliver-specific MicroRNA. Science 2005; 309, 1577-1581.), while miR-32 inhibits primate Translation of foamy virus type 1 (PFV-1) (Lecellier CH,. et al. A cellular microRNA mediates antiviral defense in human cells. Science 2005; 308, 557-560.). miRNA can also regulate the interaction between HIV and host T cells, affecting latent or persistent ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K31/7105C12N15/113A61P1/16A61P31/20
Inventor 汤华章广玲李怡璇王芳刘涛万海英李欣刘民
Owner TIANJIN MEDICAL UNIV
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