CN114588136A

CN114588136A - Application of honokiol in preparing medicine for treating myocardial infarction

Info

Publication number: CN114588136A
Application number: CN202210362805.5A
Authority: CN
Inventors: 郭艳杰; 刘乃溶; 张金洲; 杨金保; 康伟; 卢涛; 郭颖强; 姚艳妮
Original assignee: Xian International Medical Center Co Ltd
Current assignee: Xian International Medical Center Co Ltd
Priority date: 2022-04-07
Filing date: 2022-04-07
Publication date: 2022-06-07

Abstract

The invention discloses an application of honokiol in preparing a medicine for treating myocardial infarction. The inventors confirmed by both in vivo animal and cell models: when honokiol is given after myocardial infarction, the heart function of mice can be improved, the fibrosis of the far-end myocardium in an infarct area is reduced, and the apoptosis of myocardial cells in the infarct marginal zone is reduced; improving the activity of the simulated ischemic myocardial cells and relieving the injury and the apoptosis of the simulated ischemic myocardial cells. The honokiol has the function of treating the ischemic injury of the heart and has good clinical application prospect.

Description

Application of honokiol in preparing medicine for treating myocardial infarction

Technical Field

The invention belongs to the technical field of medicines, and particularly relates to application of honokiol in preparation of a medicine for treating ischemic heart disease.

Background

Honokiol (HK) is extracted from Magnolia grandiflora, is a powerful lignan, and has pungent and aromatic odor. Honokiol has wide therapeutic effects, such as antidepressant, antibacterial, antitumor, antianxiety, antithrombotic, analgesic, antispasmodic and neuroprotective effects. Honokiol has been proved to be effective in neurodegenerative diseases, diabetes, cancer and other diseases caused by oxidative stress and inflammation.

Disclosure of Invention

The invention provides the application of honokiol in treating myocardial infarction based on new research findings.

After a mouse myocardial infarction operation, a mouse with myocardial infarction is treated by taking honokiol for 4 weeks, the cardiac function of the mouse is obviously improved, the systolic end pressure of the left ventricle is obviously reduced, the maximum rate of pressure rise in the left ventricle in the isovolumetric contraction period of the left ventricle is obviously increased, the length of the heart and the tibia is obviously reduced, the fibrosis index of the far end of the infarction is obviously reduced, the apoptosis of myocardial cells of the infarction marginal tissue is obviously inhibited, and the ratio of apoptosis molecules Bax/Bcl-2 is also obviously reduced.

The inventor establishes a myocardial cell ischemia simulation model, gives honokiol, simulates the increase of cell activity of ischemic primary myocardial cells, increases the release amount of lactate dehydrogenase, reduces the apoptosis ratio, reduces the activity of Caspase-3, and increases the ratio of Bcl-2/Bax.

The invention proves that honokiol can be used for preparing the medicine for treating myocardial infarction.

Drawings

Figure 1 is the results of honokiol protecting heart of myocardial infarction mice; wherein: a is the statistics of the heart ejection fraction of the mouse, B is the end-diastolic pressure of the left ventricle, C is the maximum rate of pressure rise in the left ventricle in isovolumetric contraction, D is the ratio of the heart weight of the mouse to the length of the tibia, E is the end-diastolic inner diameter of the left ventricle of the mouse, F is the end-systolic inner diameter of the left ventricle of the mouse, G is a Masson staining typical picture, H is a TUNEL staining typical picture, I is the statistics of interstitial fibrosis of the myocardium, J is the statistics of apoptosis proportion, K is a protein immunoblot typical picture, and L is Bax/Bcl-2; p < 0.05, P < 0.01, P < 0.001.

FIG. 2 is a graph of the results of honokiol reduction in simulated ischemic cardiomyocyte injury; a is cell activity detection, B is lactate dehydrogenase release, C is TUNEL staining typical graph, D is apoptosis ratio statistics, E is Caspase-3 activity, F is protein immunoblotting typical graph, G is Bcl-2/Bax; p < 0.05, P < 0.01, P < 0.001.

Detailed Description

Unless otherwise specified, the terms or methods herein are understood or implemented using known methods as would be recognized by one of ordinary skill in the relevant art.

Myocardial Infarction (MI) is one of the complex phenotypes of coronary artery disease, and is the result of the interaction of multiple genetic and environmental factors. After myocardial infarction, myocardial cells in the corresponding region die due to persistent ischemia due to a sharp decrease or interruption of coronary blood supply.

Myocardial damage caused by myocardial infarction is mainly characterized by large-area death (including necrosis and apoptosis) of myocardial cells in an ischemic area, apoptosis of myocardial cells around the ischemic area, massive activation and collagen secretion of fibroblasts in the ischemic area and around the ischemic area, and finally cardiac rupture or ischemic heart failure. Myocardial damage caused by myocardial infarction is completely caused by insufficient blood supply, and myocardial cell death in corresponding areas and subsequent consequences caused by myocardial cell death are caused.

Myocardial infarction in animal model is simulated human myocardial infarction by ligating the left anterior descending branch of coronary artery of rat/mouse heart to cause permanent ischemia of partial myocardium; in terms of cell models, the ischemia-hypoxia environment of the myocardium during myocardial infarction is usually simulated by replacing the cell culture medium with a sugar-free and serum-free medium or buffer and placing the cells in an anaerobic environment.

The invention is further illustrated by the following examples. The examples are intended to illustrate the invention in detail and not to limit it. The honokiol used in the following examples was purchased from Shanghai Aladdin Biotech Co., Ltd (cat. H111271).

Example 1: honokiol for protecting heart of myocardial infarction mouse

Mice (purchased from the experimental animal center of the university of air force and military medical science) are randomly divided into two groups, and one group is subjected to myocardial infarction surgery to construct an MI mouse model; one group was subjected to sham surgery; 20 in each group;

construction of MI mouse model: mice were anesthetized with 2% isoflurane and after fixation the heart was exposed by thoracotomy from the third and fourth left intercostals; ligating the left anterior descending branch of the coronary artery by 6-0 silk thread, wherein the whitening of the anterior wall indicates the success of the coronary artery ligation;

post-operatively randomized MI mouse models into MI groups and MI + HK groups; the Sham group (Sham group) performed exactly the same surgical procedure, but no ligation;

the MI + HK group is injected with honokiol in the abdominal cavity immediately after the operation according to the dose of 0.24mg/kg body weight, once a day; after the mouse MI is operated for four weeks, the heart function of the mouse is evaluated by the ultrasonic of the small animal, the existing corresponding methods are adopted for each evaluation and detection method, and reagents and biological materials used in each method are all commercially available products.

And (3) displaying a detection result:

as shown in fig. 1, after honokiol treatment, the heart function of mice was improved from (25.15 ± 2.48)% to (35.31 ± 2.67)% (P < 0.05) (fig. 1A);

similarly, the pressure-volume measurements also showed a 20.22% end-systolic pressure drop in MI mice treated with four weeks of honokiol (P < 0.001) (FIG. 1B);

after four weeks of honokiol treatment, the maximal rate of pressure rise in the left ventricle of MI mice in isovolumetric contraction of the left ventricle was increased to 5321.6 + -166.2 mmHg/s (P < 0.001) (FIG. 1C);

HW/TL of MI mice decreased from 10.02 + -0.25 mg/mm to 8.32 + -0.52 mg/mm (P < 0.05) (FIG. 1D);

the left ventricular end-systolic and end-diastolic internal diameters of the MI mouse hearts were also significantly reduced by honokiol treatment (fig. 1F-F);

after honokiol treatment, the infarct distal fibrosis index of the infarcted mice decreased from (20.65 + -0.98)% to (10.53 + -0.87)% (P < 0.001) (FIG. 1G, I);

after the treatment of honokiol, the apoptosis of the myocardial cells is inhibited, and the apoptosis ratio is reduced from (7.52 +/-0.15)% to (3.54 +/-0.09)% (P is less than 0.001) (figure 1H, J);

honokiol also reduced the ratio of apoptosis marker molecule Bax/Bcl-2 from 2.31 + -0.17 in MI group to 1.67 + -0.12 in treatment group (FIG. 1K-L);

the results show that when honokiol is administered after myocardial infarction, the heart function of a mouse can be improved, the myocardial fibrosis at the far end of an infarct area can be reduced, and the myocardial apoptosis at the infarct marginal area can be reduced, so that the honokiol has the effect of protecting the heart of the myocardial infarction mouse.

Example 2: reduction of simulated ischemic cardiomyocyte injury by honokiol

Obtaining primary myocardial cells: newborn SD rat suckling mice (purchased from laboratory animal center of military university of air force) are cut into small pieces after heart removal; digesting the fragmented tissue in a Phosphate Buffered Saline (PBS) solution containing 1% collagenase-I; at 5 x 10 per ml⁵(ii) density inoculation of individual cells in DME/F-12 medium containing 10% fetal bovine serum, 0.1mM bromodeoxyuridine (BrdU), penicillin (100U/ml) and streptomycin (100U/ml) at 5% CO2, and primary cardiomyocytes were obtained starting after 48 hours of culture at 37 ℃;

the primary cardiomyocytes were divided into a control group (Nor + DMSO group), a control + and magnolol group (Nor + HK group), a simulated ischemia model group (Hypo + DMSO group), a simulated ischemia model + and a magnolol group (Hypo + HK group);

constructing a simulated ischemia model: replacing a normal culture medium with Hanks balanced salt solution, and placing primary cardiomyocytes in a 8000WJ hypoxia (5% CO2 and 95% N2) incubator in Napco for 8 hours to establish a cardiomyocyte simulated ischemia model;

and (3) constructing a honokiol treatment group: and adding honokiol with the final concentration of 10 mu M into a culture medium for constructing the simulated ischemia model, and performing other operations in the same way as the simulated ischemia model construction method.

Performing CCK-8 detection, TUNEL staining, LDH content and protein immunoblotting detection on the cells of each group; the existing corresponding methods are adopted in all evaluation and detection methods, and reagents and biological materials used in the methods are all commercially available products.

And (3) displaying a detection result:

after simulated ischemia, the activity of the myocardial cells is reduced by 37.46 percent (P is less than 0.01), and the activity of the simulated ischemia myocardial cells is increased (P is less than 0.01) by giving honokiol (see figure 2A);

simulating that after ischemic treatment, the release of myocardial cell lactate dehydrogenase increased from (12.02 + -2.34)% to (42.35 + -2.89)%, but after honokiol administration, the release of lactate dehydrogenase decreased to (28.45 + -3.12)% (see FIG. 2B);

a typical plot of TUNEL staining is shown in fig. 2C, which simulates an increase in cardiomyocytes after ischemia (P < 0.001), and following honokiol treatment, the apoptotic ratio of cardiomyocytes decreased from (37.88 ± 2.94)% to (20.51 ± 3.87)% (P < 0.05) (fig. 2D);

after simulated ischemic treatment, the activity of Caspase-3 in the myocardial cells was increased by 2.57 times, and after treatment with honokiol, the activity of Caspase-3 in the simulated ischemic myocardial cells was decreased by 26.96% (FIG. 2E);

similarly, western immunoblots of the pro-apoptotic molecule Bax and the suppressor apoptotic molecule Bcl-2 also showed that the Bcl-2/Bax ratio decreased 65.98% after simulated ischemia, while honokiol treatment increased the ratio by 1.05-fold (FIGS. 2F-G);

these results suggest that honokiol improves the cell viability of the simulated ischemic cardiomyocytes, reduces the damage and apoptosis of the simulated ischemic cardiomyocytes, and indicates that honokiol has the effect of reducing the damage of the simulated ischemic cardiomyocytes.

Claims

1. Application of honokiol in preparing medicine for treating myocardial infarction is provided.

2. The use of claim 1, wherein the medicament is in an injectable or orally administrable formulation.