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Application of IRX3 protein to prevention and treatment of obesity, metaflammation and related diseases

A protein and disease technology, applied in the field of obesity or metabolic inflammatory diseases, can solve the problem that the function of IRX3 needs to be explored

Pending Publication Date: 2021-12-10
PEKING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the function of IRX3 in other aspects still needs to be discovered

Method used

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  • Application of IRX3 protein to prevention and treatment of obesity, metaflammation and related diseases
  • Application of IRX3 protein to prevention and treatment of obesity, metaflammation and related diseases
  • Application of IRX3 protein to prevention and treatment of obesity, metaflammation and related diseases

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0081] LPS stabilizes IRX3 by inhibiting the ubiquitin-proteasome pathway to increase IRX3 protein levels

[0082] 1. Obtaining of RAW264.7 / pAAV-CAG-IRX3(WT)-Flag transiently transfected cells

[0083] The nucleic acid of mouse source IRX3 and the fusion nucleic acid of the Flag tag fused with it at the 3' end were artificially synthesized, and the obtained fusion nucleic acid was cloned into the pAAV-CAG (Addgene) expression plasmid vector to obtain the recombinant expression plasmid pAAV-CAG-IRX3 (WT )-Flag for overexpression of IRX3 protein with Flag tag (abbreviated as IRX3, IRX3(WT) or IRX3-Flag).

[0084] Cultivate RAW264.7 cells, when the cells grow to >80%, suspend the adherent cells with PBS containing 5mM EDTA to obtain a cell suspension, centrifuge the cell suspension at 800rpm for 5 minutes, discard the supernatant, add medium to fully reconstitute suspension, after cell counting, according to 5×10 5 / ml density into 12-well plates. After 12 hours, when the cell...

Embodiment 2

[0099] LPS can stabilize IRX3 by inhibiting the ubiquitin-proteasome pathway at the K407 site to increase IRX3 protein levels

[0100] The nucleic acid of artificially synthesized green fluorescent protein (GFP) and the fusion nucleic acid of the Flag tag fused with it at the 3' end are cloned into the pAAV-CAG (Addgene) expression plasmid vector to obtain the recombinant expression plasmid pAAV-CAG- GFP-Flag is used to express GFP protein with Flag tag (abbreviated as GPF or GFP-Flag), which is used as the control of pAAV-CAG-IRX3(WT)-Flag.

[0101] The nucleic acid of artificially synthesized ubiquitin protein (nucleic acid Genbank accession number NM_021009.7) and the fusion nucleic acid of the Myc tag fused to it at the 5' end were cloned into the pAAV-CAG (Addgene) expression plasmid vector to obtain the recombinant The expression plasmid pAAV-CAG-Myc-Ub is used to express Myc-tagged ubiquitin protein (abbreviated as Myc-Ub).

[0102] Use one-step PCR to site-directed mu...

Embodiment 3

[0110] LPS is able to phosphorylate IRX3 protein by JNK1 / 2 kinase

[0111] 1. Phos-tag experiment

[0112] The acquisition of RAW264.7 / pAAV-CAG-IRX3(WT)-Flag transiently transfected cells and the LPS treatment of RAW264.7 / pAAV-CAG-IRX3(WT)-Flag transiently transfected cells and subsequent total protein extraction are the same as in the example 1 In Section 1 and Section 2, the total protein extracts of each treatment were obtained.

[0113] The phosphorylation label (Phos-tag) experimental operation is the same as the immunoblotting of the protein in the second subsection of Example 1, the difference is that when preparing a 7% polyacrylamide gel, the 7% polyacrylamide separating gel solution Add a final concentration of 40 μM phos-tag and a final concentration of 100 μM nCl 2 , and then carry out protein electrophoresis to obtain an electrophoresis gel, and then wash the electrophoresis gel 3 times with a transfer solution containing 5mM EDTA for 20 minutes each time. The ...

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Abstract

The invention relates to application of IRX3 protein to prevention and treatment of obesity, metaflammation and related diseases, in particular to application of at least one of IRX3 protein in macrophages, RNA expressing the IRX3 protein and a gene coding the IRX3 protein as a target to preparation of a medicine for preventing and / or treating at least one of obesity, obesity-related diseases, metaflammation and metaflammation-related diseases. The medicine targets at least one of the IRX3 protein in the macrophage, the RNA expressing the IRX3 protein and the gene coding the IRX3 protein, so that the content of the IRX3 protein in the macrophages is reduced or the function of the IRX3 protein in the macrophages is reduced to prevent and / or treat at least one of the obesity, the obesity-related diseases, the metaflammation and the metaflammation-related diseases.

Description

technical field [0001] The invention relates to the field of obesity or metabolic inflammatory diseases, in particular to a protein present in macrophages that affects obesity or metabolic inflammation. Background technique [0002] Inflammation is an important defense response of the body, which can regulate and eliminate various endogenous or exogenous damage factors acting on the body, and also has the function of repairing damage. But excessive or persistent inflammation can also damage tissues and organs. Metabolic disorders or abnormal metabolites caused by various factors are called metabolic inflammation, which participates in and leads to the occurrence of various metabolic diseases such as atherosclerosis, diabetes, and non-alcoholic fatty liver. [0003] The occurrence and development of obesity can cause a variety of metabolic diseases including diabetes, non-alcoholic fatty liver disease, hypertension and atherosclerosis, which seriously threaten human health. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K45/00A61K48/00C12Q1/02A61P3/04A61P5/00A61P29/00A61P3/10A61P5/50A61P1/16A61P9/10A61P3/06A61P9/12
CPCA61K45/00A61K48/005A61P3/04A61P5/00A61P29/00A61P3/10A61P5/50A61P1/16A61P9/10A61P3/06A61P9/12G01N33/5055C12N2503/02G01N2500/10
Inventor 邱义福姚静斐吴冬梅
Owner PEKING UNIV
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