阅读说明：本技术 厚朴酚在制备抑制铁死亡药物中的应用 (Application of magnolol in preparation of medicine for inhibiting iron death ) 是由 谭克 樊玉梅 李丹雨 于 2019-10-21 设计创作，主要内容包括：本发明公开厚朴酚在制备抑制Erastin和RSL3诱导铁死亡信号通路表达的药物中的应用,深入研究了不同浓度的厚朴酚对Erastin和RSL3诱导的铁死亡的作用及分子机制,进一步挖掘厚朴酚的药理功能,拓宽了对厚朴酚的认识,并为铁死亡相关疾病的预防和治疗提供理论基础和新途径。(The invention discloses application of magnolol in preparation of a medicine for inhibiting Erastin and RSL 3-induced iron death signal pathway expression, deeply researches the action and molecular mechanism of magnolol with different concentrations on Erastin and RSL 3-induced iron death, further excavates the pharmacological functions of magnolol, widens the knowledge of magnolol, and provides a theoretical basis and a new way for prevention and treatment of iron death-related diseases.)

1. Application of magnolol in preparing medicine for inhibiting iron death is provided.

2. The use of magnolol according to claim 1 in the preparation of a medicament for inhibiting iron death, wherein magnolol is used in the preparation of a medicament for inhibiting expression of an Erastin-induced iron death signaling pathway.

3. The use of magnolol according to claim 1 for the manufacture of a medicament for inhibiting iron death, wherein magnolol is used for the manufacture of a medicament for inhibiting expression of the RSL 3-induced iron death signaling pathway.

Technical Field

The invention relates to the technical field of chemical medicines, in particular to application of magnolol in preparation of a medicine for inhibiting Erastin and RSL3 induced iron death.

Background

Iron death (ferroptosis) was initially defined in 2012 as an iron-dependent mode of cell death that differs from apoptosis, necrosis, and autophagy. During the process of iron death, the free iron pool of cells is obviously increased, mitochondria are reduced and the mitochondrial cristae is disappeared, and meanwhile, a series of changes occur in the biochemical characteristics of the cells, such as accumulation of cell membrane lipid peroxide (lipidoxidation), depletion of reduced Glutathione (GSH) and the like. A large number of researches show that iron death is closely related to diseases such as tumors, nervous system diseases, ischemia-reperfusion injury, kidney injury and the like. In the pathological process of neurodegenerative diseases, the main cause of nerve cell injury may be the massive accumulation of intracellular iron, and is also related to intracellular iron metabolism, amino acid metabolism and lipid metabolism. Therefore, the molecular mechanism and the regulation pathway of iron death are deeply researched, natural small molecular compounds for inhibiting or intensifying iron death are explored, and a new idea and a new target are possibly provided for clinical treatment of various refractory diseases.

Erastin is a typical inducer of iron death. Erastin mainly inhibits the function of a cystine/glutamate antiporter System Xc- (cysteine/glutamate transporter) in cells, reduces the uptake of Cys in cells and further hinders the synthesis of GSH, and the reduction of the content of the GSH can cause the low enzymatic activity of glutathione peroxidase 4(GPX4), so that the lipid peroxidation reaction in the cells is aggravated, the lipid ROS is increased, and finally the cells are induced to generate iron death.

Unlike Erastin, the small molecule compound RAS-selective least small molecule 3(RSL3) also induces cell iron death, and RLS3 induced iron death appears to not require signaling through mitochondria VDAC2 or VADC3 and System Xc-, early studies have shown that RSL3 can bind GPX4 and inhibit its enzymatic activity, thereby interfering with its ability to prevent lipid peroxidation; however, recent studies have shown that both RSL3 and Erastin can induce GPX4 protein degradation leading to the accumulation of ROS and ultimately the induction of pig death by cells.

Magnolol (magnolol) is a main active ingredient extracted from traditional Chinese medicine Mangnolia officinalis, and has wide pharmacological effects of resisting inflammation, bacteria, tumor, aging and the like. Magnolol (C)₁₈H₁₈O₂) Has wide effect, and can eliminate related free radicals, enhance enzyme activity, etcHas antioxidant effect. Magnolol has excellent antifungal effect, and also has obvious antibacterial activity against gram-positive bacteria and acid-fast bacteria. In addition, the prominent anti-tumor effect of magnolol is also more and more concerned and regarded. Magnolol shows a remarkable pro-apoptotic effect by increasing the activity of casepase-3, casepase-8 and casepase-9 and down-regulating the anti-apoptotic protein Bcl-2 in various malignant tumor cells such as glioma, bladder cancer, ovarian cancer, prostate cancer, thyroid cancer and the like. In lung cancer cells, magnolol also blocks the PI3K/PTEN/AKT signaling pathway, inducing tumor cell death by autophagy. Therefore, the magnolol anti-tumor effect has the characteristics of multiple targets and multiple ways.

However, the mechanism of action of magnolol in treating diseases associated with iron death by inhibiting iron death is not clear.

Therefore, how to research the mechanism of inhibiting iron death by magnolol and apply the magnolol in the medicament for inhibiting iron death is an urgent problem to be solved by the technical personnel in the field.

Disclosure of Invention

In view of the above, the invention provides an application of magnolol in preparing a medicine for inhibiting the expression of Erastin and RSL 3-induced iron death signal pathways, and deeply studies the action and molecular mechanism of magnolol with different concentrations on Erastin and RSL 3-induced iron death, further exploits the pharmacological functions of magnolol, widens the knowledge of magnolol, and provides a theoretical basis and a new way for preventing and treating diseases related to iron death.

In order to achieve the purpose, the invention adopts the following technical scheme:

application of magnolol in preparation of medicines for inhibiting Erastin-induced iron death signal pathway expression.

Application of magnolol in preparation of medicine for inhibiting expression of signal pathway of RSL3 induced iron death.

Through the technical scheme, compared with the prior art, the invention discloses and provides the application of magnolol in preparing the medicine for inhibiting the expression of the Erastin and RSL 3-induced iron death signal channel, and finds that the magnolol can inhibit the generation and accumulation of lipid peroxide malondialdehyde MDA induced by Erastin and RSL3, can inhibit the expression of the Erastin and RSL 3-induced glutathione GSH-depleted signal channel, and can inhibit the expression of the Erastin and RSL 3-induced GPX4 signal channel, so that the iron death in cells is inhibited, and the survival rate of the cells is improved.

Drawings

In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, it is obvious that the drawings in the following description are only embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to the provided drawings without creative efforts.

FIG. 1 is a schematic representation of the effect of the addition of different concentrations of magnolol on the viability of HEK293 cells;

FIG. 2 is a graph showing the effect of magnolol at different concentrations on Erastin-induced iron death;

FIG. 3 is a graph showing the inhibitory effect of different concentrations of magnolol on RSL 3-induced iron death;

FIG. 4 is a graph showing the effect of magnolol on Erastin-induced MDA content;

FIG. 5 is a graph showing the effect of magnolol on RSL 3-induced MDA content;

FIG. 6 is a graph of the effect of magnolol on Erastin-induced GSH depletion;

figure 7 is a graph of the effect of magnolol on RSL 3-induced GSH depletion;

FIG. 8 is a graph showing the effect of magnolol on Erastin-induced ROS levels in HEK293 cells;

FIG. 9 is a bar graph showing the effect of magnolol on Erastin-induced ROS levels in HEK293 cells;

FIG. 10 is a graph showing the effect of magnolol on the distribution of ROS levels in HEK293 cells induced by RSL 3;

FIG. 11 is a bar graph showing the effect of magnolol on ROS levels in HEK293 cells induced by RSL 3;

FIG. 12 is a graph showing the effect of magnolol on Erastin-induced distribution of ROS (lipidROS) levels in HEK293 cells;

FIG. 13 is a bar graph showing the effect of magnolol on Erastin-induced lipid ROS levels (lipid ROS) in HEK293 cells;

FIG. 14 is a graph showing the effect of magnolol on RSL 3-induced distribution of ROS (lipidROS) levels in HEK293 cells;

FIG. 15 is a bar graph showing the effect of magnolol on RSL 3-induced lipid ROS levels (lipid ROS) in HEK293 cells;

FIG. 16 is a graph showing the effect of magnolol on Erastin-induced LIP levels in HEK293 cells;

FIG. 17 is a bar graph showing the effect of magnolol on Erastin-induced LIP levels in HEK293 cells;

FIG. 18 is a graph of the effect of magnolol on Erastin-induced key regulator levels of iron death;

figure 19 is a graph of the effect of magnolol on the levels of key regulators of iron death induced by RSL 3.

Detailed Description

The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

The test equipment and materials used in the invention are as follows:

fetal bovine serum (ExCelBio), DMEM medium, CCK-8(MCE), DMSO (Sigma), cell culture incubator (Thermo Fisher), ELX-800 microplate reader (BioTek), inverted microscope (BIO-TEC), Magnolol (MCE), RLS3(MCE), Erastin (seleck), 96-well cell culture plate (JET), human embryonic kidney cells (HEK293 cells, purchased from Shanghai cell of the academy of China), detection kit (Nanjing MDA), 10cm cell culture dish (JET), GSH detection kit (Nanjing Mak), flow cytometer FACS Calibur (BD), basic electrophoresis apparatus power supply (Bio-rad), electrophoresis apparatus (JUNYI), transfer bath (Bio-rad), PVDF membrane (Roche Applied Science), GPX4(ab 1256, acam), SL 06 4(K004812P, GAarbio 547, Wash HRP, Wal-01H (PDH + PDH).

The following tests were carried out using the above test equipment to verify the mechanism of action of magnolol in inhibiting iron death: