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Application of evodiamine in preparation of medicine for enhancing activation of NLRP3 inflammasomes

A technology of inflammasome and evodiamine, which is applied in the field of biomedicine and medicine to achieve the effect of increasing IL-1β secretion, reducing death and promoting pyroptosis

Inactive Publication Date: 2018-11-30
JINAN UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0012] At present, there is no report on the function of evodiamine in enhancing the activation of NLRP3 inflammasome

Method used

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  • Application of evodiamine in preparation of medicine for enhancing activation of NLRP3 inflammasomes
  • Application of evodiamine in preparation of medicine for enhancing activation of NLRP3 inflammasomes
  • Application of evodiamine in preparation of medicine for enhancing activation of NLRP3 inflammasomes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] The test of evodiamine activating NLRP3 inflammasome includes the following steps:

[0049] (1) Preparation of L929 cell culture supernatant:

[0050] L929 cells were purchased from the Cell Bank of Kunming Institute of Zoology, Chinese Academy of Sciences. The cell culture method refers to the operating procedure of Monack laboratory (Stanford University), and 1.5 × 10 7 cells cultured in 75cm 2 In the culture flask, the culture medium is DMEM complete medium (30mL / bottle), and after culturing for 14 days, the supernatant is collected and frozen for future use. (2) Culture method of mouse bone marrow-derived macrophages (BMDM):

[0051] C57BL / 6 mice, 6-8 weeks old, female, were purchased from the Experimental Animal Center of Southern Medical University. After being sacrificed by cervical dislocation, the hind leg bones were removed, and the bone marrow cells were flushed out with cold PBS using a syringe. The obtained cells were cultured in a culture dish (densit...

Embodiment 2

[0057] The test of evodiamine activating NLRP3 inflammasome includes the following steps:

[0058] (1) Detection of ASC plaque formation:

[0059] BMDM cells were seeded in glass-bottom culture dishes (at a density of 5 × 10 5 After overnight, induce 4 hours with 500ng / mL LPS, add evodiamine 5μmol / L to act for 1 hour, then add 2mmol / LATP to stimulate for 30 minutes, finally suck out the medium, add 1mL 4% (w / v ) paraformaldehyde, fixed at room temperature for 15 minutes, then permeabilized each well with 2 mL of cold methanol at -20°C for 10 minutes, then blocked with blocking solution for 1 hour at room temperature, and added primary antibody (ASC dilution ratio of 1:300, 100 μL / well), incubate overnight at 4°C, add non-cross-reactive CF568-goat anti-rabbit IgG (purchased from Biotium, USA) after washing, incubate at room temperature for 1 hour, stain the nucleus with Hoechst 33342 for 10 minutes and protect from light, observe with a Zeiss inverted fluorescence microscope ...

Embodiment 3

[0063] The test of evodiamine activating NLRP3 inflammasome includes the following steps:

[0064] (1) Detection of ASC plaque formation:

[0065] The method was the same as "Example 3", except that after the cells were plated overnight, they were induced with 500ng / mL LPS for 4 hours, added 5 μmol / L evodiamine for 1 hour, and then added 5 μmol / L nigericin to stimulate for 1 hour.

[0066] (2) Results:

[0067] The formation of ASC plaque (ASC speck) is another important indicator of NLRP3 inflammasome activation. image 3 Evodiamine on the formation of ASC speck (ASC speck) in mouse bone marrow-derived macrophages (BMDM) induced by LPS+nigericin by immunofluorescence microscopy. It can be seen from this figure that evodiamine can significantly promote the formation of ASC plaques in BMDM cells induced by LPS+nigericin, which further illustrates that it can promote the activation of NLRP3 inflammasome. *** indicates P<0.001.

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Abstract

The invention discloses application of evodiamine and rutaecarpine in preparation of a medicine for enhancing the activation of NLRP3 inflammasomes. The evodiamine and the rutaecarpine can generate active Caspase-1 by enhancing activiation of macrophage inflammasomes so as to release manure inflammatory mediators such as IL-1beta and promote pyroptosis of sensitized macrophages. The activity of the evodiamine and the rutaecarpine can be used for enhancing the inflammatory response intensity of a human body and treating related diseases of insufficient activation of inflammasome including pulmonary tuberculosis and oral tuberculosis caused by intracellular infection of mycobacterium tuberculosis, diseases caused by listeria monocytogenes, related diseases caused by other intracellular infection bacteria, related diseases caused by pathogenic fungal infection, furunculosis, dental-root periapical diseases, chronic gastritis, chronic enteritis, dysentery and diarrhea and the like.

Description

technical field [0001] The invention belongs to the field of biomedicine and medicine, and specifically relates to evodiamine and / or evodiamine as an enhancer of NLRP3 inflammasome activation and its application in the preparation of drugs for treating diseases related to insufficient activation of NLRP3 inflammasome. Background technique [0002] Evodiamine and its derivatives, rutaecarpin, etc. are active ingredients extracted from the fruits of Rutaceae traditional Chinese medicine plants Evodia rutaecarpa, Shihu and Evodia lenticus. [0003] The molecular weight of evodiamine is 303.36, molecular formula C 19 h 17 N 3 O; the molecular weight of evodiamine is 287.32, and the molecular formula is C 18 h 13 N 3 O, the chemical structures of the two are shown in the following formula. [0004] [0005] Evodiamine is a yellow powder, insoluble in water, but easily soluble in organic solvents such as chloroform and acetone. Evodiamine has the effects of invigorating ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/519A61P29/00A61P31/04A61P37/04
CPCA61K31/519A61P29/00A61P31/04A61P37/04Y02A50/30
Inventor 欧阳东云何贤辉查庆兵李陈广徐丽慧曾琼祯
Owner JINAN UNIVERSITY
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