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Lymphocyte antigen 96 protein-activated blocker and application thereof

A technology of lymphocytes and blocking agents, applied in the field of lymphocyte antigen 96 protein, can solve the problems of blocking MD2 activation, tripterine has not been reported, and achieve the effect of blocking inflammatory reactions

Inactive Publication Date: 2017-05-31
张登海 +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, tripterine has not been reported to block the activation of MD2

Method used

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  • Lymphocyte antigen 96 protein-activated blocker and application thereof
  • Lymphocyte antigen 96 protein-activated blocker and application thereof
  • Lymphocyte antigen 96 protein-activated blocker and application thereof

Examples

Experimental program
Comparison scheme
Effect test

experiment example 1

[0036]Experimental example 1 Computer simulation for molecular docking (Docking)

[0037] Such as figure 2 As shown, download the three-dimensional structure of MD2 molecule (file number: 2e56pdb) from the protein website (RCSB), such as figure 2 Shown in A is the three-dimensional structure of the LPS molecule downloaded from the protein database (PDB) (file number: 4G8A). Such as figure 1 and figure 2 As shown in B, the three-dimensional structure file of tripterifera was downloaded from the National Institutes of Health compound website (PubChem Compound), and then molecular docking simulation was performed with molecular docking software (AutoDock V4.1).

[0038] Analysis of molecular docking simulation results:

[0039] Such as figure 2 As shown in D, LPS binds in a region surrounded by MD2 protein, such as figure 2 As shown in F, the above region into which tripterine enters is hydrophobic. Such as figure 2 C and figure 2 As shown in E, tripterine can als...

experiment example 2

[0040] Experimental Example 2 The experiment of measuring the combination of tripterine and cells by flow cytometry

[0041] (1) Cell culture

[0042] MEM culture medium: purchased from Hyclone Company, which is a mature commodity, containing 2.00 mmol / L L-glutamine and balanced salt solution.

[0043] Cell culture: hepatocytes were cultured in MEM medium (purchased from Hyclone), and grown in 5% CO 2 In a 37-degree incubator, add 10% fetal bovine serum and 1% antibiotics (100IU / mL penicillin and 100ug / mL streptomycin) to the culture medium, and when the confluence of the cells in the culture flask reaches about 80%, digest Spread into 6-well plates, the number of cells per well is about 1×10 6 indivual.

[0044] (2) siRNA Oligo transfection

[0045] Dilute the oligonucleotide (Oligo) sequence of siRNA-siRNA (purchased from Shanghai Gemma Biological Company) mixed with interfering MD2 protein and the siRNA transfection control sequence (purchased from Shanghai Gemma Biolog...

experiment example 3

[0058] Experimental Example 3 The effect of tripterine on the interaction between LPS and MD2

[0059] (1) tripterine blocks cell binding to LPS (MD2 activator)

[0060] Trypsin was added to the cultured hepatocytes to digest the adherent cells from the culture plate, and MEM medium was added to terminate the digestion. Collect the cell suspension, transfer it into a 1.5ml EP tube, centrifuge at 1000g for 5min, and discard the supernatant. After washing 3 times with PBS (phosphate buffered saline), the cells were counted and the results were recorded on the tube. Take 2×10 5 The cells were fixed with 4% paraformaldehyde at 4°C for 1 h. After the fixation, centrifuge at 1000 g for 5 min, discard the supernatant, and wash the cells three times with 1% BSA (bovine serum albumin) to obtain treated cells.

[0061] Collect the above cell pellet, first wash it with PBS three times, then fix it with 4% paraformaldehyde at 4 degrees for 1 hour, after fixing the result, wash it with...

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Abstract

The invention discloses a lymphocyte antigen 96 protein-activated blocker and an application thereof. The blocker comprises tripterine and can be used for preparing drugs or pharmaceutical compositions for treating obesity, diabetes and an acute lung injury and resisting inflammation and atherosclerosis. The lymphocyte antigen 96 protein-activated blocker can be combined with a hydrophobic region combined with MD2 protein, activation of the MD2 protein is inhibited, combination of an MD2 activator and LPS and MD2 can be inhibited and inflammatory reaction is blocked.

Description

technical field [0001] The invention relates to a lymphocyte antigen 96 protein (MD2), in particular to a blocking agent for activation of the lymphocyte antigen 96 protein and its application. Background technique [0002] Lymphocyte antigen 96 (Lymphocyte antigen 96), also known as MD2 protein, is a protein on the human cell membrane, encoded by the LY96 gene. MD2 protein can bind to some compounds, such as bacterial lipopolysaccharide (lipopo-lysaccharide, LPS), which is an important component of the outer membrane of the cell wall of Gram-negative bacteria, and plays an important role in Gram-negative bacterial infection and disease evolution. It is believed to be the main cause of systemic inflammatory syndrome. Bacterial component lipopolysaccharide (LPS) binds to Toll-like receptor 4 protein (Toll-like receptor 4, TLR4) and MD2 protein complex, TLR4 protein activation can further activate nuclear transcription factor (NF-κB), thereby synthesizing a series of inflamm...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/56A61P29/00
CPCA61K31/56
Inventor 张登海张雪彭彬
Owner 张登海
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