CN111166757A

CN111166757A - Application of loganin in promoting angiogenesis and treating myocardial ischemia

Info

Publication number: CN111166757A
Application number: CN201811336329.XA
Authority: CN
Inventors: 不公告发明人
Original assignee: Beijing Boya Kejian Technology Co Ltd
Current assignee: Beijing Quanfang Biotechnology Co.,Ltd.
Priority date: 2018-11-12
Filing date: 2018-11-12
Publication date: 2020-05-19

Abstract

The application provides an application of loganin in promoting angiogenesis and treating myocardial ischemia. The loganin is found to have a good protective effect on promoting the recovery of cardiac function after myocardial ischemia for the first time, and can be used for preventing or treating diseases related to myocardial ischemia, such as coronary heart disease, myocardial injury (such as myocardial ischemia-reperfusion injury), angina, arrhythmia, myocardial infarction (such as acute myocardial infarction) or sudden death.

Description

Application of loganin in promoting angiogenesis and treating myocardial ischemia

Technical Field

The present application relates to the field of Chinese herbal medicine. In particular, the application relates to the use of loganin for promoting angiogenesis in the treatment of myocardial ischemia-related diseases.

Background

Myocardial ischemia (myocardial ischemia) refers to a pathological condition in which the blood perfusion of the heart is reduced, the oxygen supply of the heart is reduced, the energy metabolism of the heart muscle is abnormal, and the heart cannot work normally. At present, the most common causes of myocardial ischemia are coronary atherosclerosis, coronary artery luminal stenosis or vessel occlusion, which leads to myocardial ischemia, hypoxia, myocardial infarction and concomitant acute loss of active myocardium, and finally, heart failure, arrhythmia and sudden death. At present, the clinical effective schemes for treating myocardial ischemia caused by coronary atherosclerosis are specifically classified into three types: surgical bypass, percutaneous coronary intervention, and drug thrombolysis. These treatments still do not allow adequate blood recirculation and cardiac function to be achieved. Experts on the American society for cardiology in 2000 have proposed "therapeutic angiogenesis (also known as drug bypass surgery" to provide a new idea for the study of ischemic heart disease. Therapeutic angiogenesis means that the therapeutic means is applied to promote the angiogenesis of ischemic myocardium, establish collateral circulation, provide sufficient oxygen supply for the ischemic myocardium and recover blood flow, realize revascularization of the ischemic myocardium and finally achieve the therapeutic purpose.

The pathological factors of the ischemia and the anoxia of the heart can stimulate the release of endogenous angiogenesis promoting growth factors such as VEGF, FGF-2, Ang-1 and the like and promote the proliferation signal path of endothelial cells. The VEGF signaling pathway can regulate various angiogenesis-related proteins, and PKC regulates the phosphorylation of downstream MEK1/2 and finally regulates the phosphorylation of Erk1/2 under the mediation of VEGF, so that the proliferation of endothelial cells is promoted. However, under pathological conditions, the ability of the body to regulate the angiogenesis promoting factors is limited, and the collateral circulation is not enough to be established to improve blood supply, so that the angiogenesis is stimulated by an exogenous intervention means, the blood flow recovery after MI is promoted, the ischemic cardiac muscle is protected, and finally the effect of protecting the cardiac function is achieved.

Loganin is a compound extracted from Cornus officinalis (Cornus officinalis sieb. et. zucc.) and has the following structure.

Disclosure of Invention

In the application, through a large number of researches, the inventors find that loganin has a promoting effect on angiogenesis after myocardial ischemia, and can be used for treating diseases related to myocardial ischemia, such as coronary heart disease, myocardial injury (such as myocardial ischemia-reperfusion injury), angina, arrhythmia, myocardial infarction (such as acute myocardial infarction) or sudden death.

In the present invention, unless otherwise specified, scientific and technical terms used herein have the meanings that are commonly understood by those skilled in the art. Also, animal disease models, cell culture, molecular genetics, nucleic acid chemistry, immunological laboratory procedures, as used herein, are all conventional procedures that are widely used in the relevant arts. Meanwhile, in order to better understand the present invention, the definitions and explanations of related terms are provided below.

As used herein, the term "myocardial ischemia-related disorders" refers primarily to disorders of cardiac dysfunction associated with reduced cardiac blood perfusion. For example, "Coronary Heart Disease (CHD)", collectively known as atherosclerotic coronary heart disease, also known as Ischemic Heart Disease (IHD), refers to heart disease caused by coronary artery atherosclerosis leading to luminal narrowing or occlusion, resulting in myocardial hypoxia or necrosis.

For example, "Ischemic injury of myocardium" (Ischemic injury) "means that after the blood supply of myocardium is interrupted, myocardial cell acidosis, organelle ultrastructure change and dysfunction occur, resulting in irreversible injury and even death. After the ischemic region is reconstructed, the myocardial cells are more seriously damaged, which can cause the disease deterioration phenomena such as blood pressure drop, cardiac insufficiency, arrhythmia and even sudden death, and the like, namely myocardial ischemia reperfusion injury.

For example, "angina" refers to a group of clinical syndromes with paroxysmal chest pain as the main manifestation, caused by insufficient blood supply to coronary arteries, acute transient ischemia and hypoxia of cardiac muscle.

For example, "myocardial infarction" refers to the acute necrosis of a portion of the myocardium due to prolonged and severe insufficient blood supply to the myocardium as a result of a disruption of coronary arteries flowing to the myocardial region.

As used herein, the term "subject" refers to an animal, particularly a mammal, preferably a human.

As used herein, the term "effective amount" refers to an amount sufficient to obtain, or at least partially obtain, a desired effect. For example, a prophylactically effective amount is an amount sufficient to prevent, or delay the onset of disease; a therapeutically effective amount is an amount sufficient to cure or at least partially arrest the disease and its complications in a patient already suffering from the disease. It is well within the ability of those skilled in the art to determine such effective amounts. For example, an amount effective for therapeutic use will depend on the severity of the disease to be treated, the general state of the patient's own immune system, the general condition of the patient, e.g., age, weight and sex, the mode of administration of the drug, and other treatments administered concurrently, and the like.

Advantageous effects of the invention

According to the application, by establishing a rat myocardial ischemia model and using indexes such as proliferation amount of vascular endothelial cells, number of blood vessels around myocardial infarction, expression of angiogenesis-related protein VEGF and the like, loganin is found to have a good promoting effect on angiogenesis of myocardial ischemia, and finally, recovery of cardiac function is promoted, so that a new way is developed for clinical treatment of diseases related to myocardial ischemia.

Drawings

FIG. 1 shows a schematic diagram of ligation sites in a coronary artery ligation method for establishing a myocardial ischemia model in rats.

FIG. 2 shows a rat electrocardiogram, wherein FIG. 2(a) is a normal rat electrocardiogram; FIG. 2(b) is an electrocardiogram of a rat model of myocardial ischemia.

FIG. 3 shows loganin administration 7d on myocardial ischemia model rat lectin⁺/ki67⁺Effects of vascular endothelial cell proliferation.

FIG. 4 shows the effect of loganin administration for 7 days on the number of peri-infarct vessels in rats in the myocardial ischemia model.

FIG. 5 shows the effect of loganin administration for 14d on the number of peri-infarct vessels in rats in the myocardial ischemia model.

FIG. 6 shows the effect of loganin administration for 7 days on the FGF-2 content in myocardial tissue of rats in myocardial ischemia model.

FIG. 7 shows the effect of loganin administration for 7 days on Ang-1 content in myocardial tissue of rats in myocardial ischemia model.

FIG. 8 shows the effect of VEGFA content in myocardial tissue of rats in the myocardial ischemia model, administered loganin 7 d.

FIG. 9 shows the results of immunoblot analysis of VEGF administration 7d, VEGF downstream regulation of endothelial proliferation associated protein in myocardial tissue of rats in myocardial ischemia model. Wherein, the lanes from top to bottom are P-Src, P-Pkc, Pkc, P-Erk1/2 and Erk1/2 respectively. From left to right are: a sham operation group, a model group and a small, medium and large dose of loganin.

FIG. 10 shows the effect of loganin administration for 14d on left ventricular function in rats with myocardial ischemia model.

FIG. 11 shows the effect of loganin administration for 28d on left ventricular function in rats with myocardial ischemia model.

Detailed Description

Embodiments of the present invention will be described in detail below with reference to examples, but those skilled in the art will appreciate that the following examples are only illustrative of the present invention and should not be construed as limiting the scope of the present invention. The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products commercially available.

1. Materials and methods

1.1 medicaments

The content of the monomeric compound loganin is more than 98.5 percent by high performance liquid chromatography detection. Before use, the medicine is dissolved into medicine liquid with required concentration by distilled water for experiment.

1.2 Experimental animals

SPF grade male Sprague-Dawley rats, 7-8 weeks old, 260-280g in mass. Purchased from sbefu laboratory animal science and technology limited, certificate number: SCKK (Jing) 2011-0004. Feeding in a conventional way, wherein the environmental temperature is 24 +/-1 ℃, the humidity is 55 +/-5%, and the water is not forbidden after 12h of preoperative fasting.

1.3 Main Experimental instruments

Small animal ventilators (harvard instruments, usa); operating microscopes (zeiss, germany); polygraph physiological signal recorder (BIOPAC, USA); scanners (hewlett packard, usa); an ultrasonic cell disruption pulverizer (Ningbo, New Ganoderma science and technology institute, China); bench-top micro-refrigerated centrifuge (Beckman Coulter corporation, usa); a full-wavelength microplate reader (Thermo Fisher, usa); powerpac Basic electrophoresis apparatus (Bio-Rad, USA); chemiluminescent gel imaging systems (Alpha corporation, usa); confocal laser microscopy (Leica, germany); high resolution small animal ultrasound imaging system (VisualSonics Inc, canada); a cold microtome (Thermo Fisher Scientific, USA).

1.4 Primary reagents

RIPA lysate (bio-technical institute, china, bi yun tian); BCA method protein quantification kit (plenilla Gene technology, Inc., China), rabbit-derived VFGFA antibody (Abcam, USA); rabbit-derived Ang-1 antibodies (bosch de bioengineering, ltd, usa); mouse-derived FGF-2 antibodies (Santa Cruz Biotechnology, usa); rabbit derived P-PKC antibody (Cell Signaling Technology, usa); rabbit derived PKC antibodies (Cell Signaling Technology, usa); rabbit derived Src antibodies (Cell Signaling Technology, usa); rabbit derived P-src antibodies (CellSignaling Technology, usa); rabbit-derived Erk1/2 antibody (Cell Signaling Technology, U.S.A.); rabbit derived P-erk1/2 antibody (Cell Signaling Technology, USA); rabbit Ki67 antibody (Millipore, usa); FITC-labeled lectin (Vector, USA); rabbit derived GAPDH antibody (Cell signaling technology, usa); horseradish peroxidase-labeled goat anti-rabbit secondary antibody (china, sequoia, gold bridge biotechnology limited); ECL Western Blotting Kit (THERMO-PIERCE, USA).

1.5 Experimental methods

1.5.1 preparation of myocardial ischemia model

The experiment was conducted to prepare a Myocardial Ischemia Model by coronary ligation (Nagaoka K1, Matoba T1, Mao Y1, et al. A New therapeutic modification for Acute Myocardial Ischemia: Nanoparticle medical delivery of Pitavastatin indexes heart detection from Ischemia-playback tissue Via Activation of PI3K/Akt Pathway and Anti-Ischemia in a Rat Model [ J ]. PLoS one.2015, 10 (7): e 0132451.). 3.5ml/kg of 10% chloral hydrate is used for intraperitoneal injection anesthesia. The rat was fixed on the rat plate in supine position, and the operating lamp was focused on the neck trachea cannula. Then adjusting to lateral position, connecting with a respirator, breathing frequency 50 times/min, breathing ratio 2: 1, tidal volume 2.5ml/100 g. Skin preparation is performed between the axilla and the end of the rib, and iodine is added for disinfection. The most obvious jumping position from the skin surface is that a transverse incision is about 2 cm-3 cm, the pectoralis major and the serratus anterior are fully exposed, the pectoralis major and the serratus anterior are separated bluntly, a small opening is cut from the fourth intercostal space, then the incision is separated and enlarged bluntly by using hemostatic forceps carefully until the incision just can be used for placing a chest opener, the chest opener is opened, the heart is fully exposed, but the intercostal incision is not enlarged intentionally, so that the situation that the wound is too large and the chest closing difficulty is increased is prevented. The pericardium is carefully torn off by using ophthalmological forceps, a needle is held by a left hand of a needle holder, the needle is inserted about 4mm downwards from the junction of the root of the left auricle and the pulmonary artery cone, the depth of the inserted needle is about 1.5mm, the needle is taken out from the junction of the left auricle and the pulmonary artery cone, the needle is parallel to the right edge of the left auricle and spans 3 mm-5 mm (as shown in figure 1), and a loose joint is ligated. The ligation position is too high, the infarct area is too large, and the mortality rate is high; too low, the infarct size is too small or less pronounced. After ligation, the ventricular wall below the ligation part is observed to become pale by naked eyes, and the success of molding can be preliminarily judged. The 8-shaped suture method is used for suturing intercostals, 1 indwelling tube is reserved, then the thoracic cavity is closed and sutured layer by layer, a 10ml injector is used for expanding the lung before the thoracic cavity is completely closed, and after the thoracic cavity is closed, the air and the effusion in the thoracic cavity are completely discharged through the indwelling tube. The ventilator was turned off intermittently until the rat regained spontaneous breathing. The rats were transferred to a postoperative care kit. In sham group rats, only threading was performed, and ligation was not performed, as in the other steps.

1.5.2 electrocardiographic testing

After modeling, rat II-lead electrocardiogram (Ma X1, Zhang K, Li H, ethanol. extracts from Aspergillus cell radius and myocardial function in a rat model of myocardial ischemia [ J ]. JEthnopharmacological.2013, 149 (3): 720-8.) was collected by a multi-lead physiological signal recorder, specifically: preparing skins of the right upper limb and the inner sides of the two lower limbs of a rat, wetting the skins with physiological saline, inserting a negative electrode probe into the inner side skin of the right upper limb, inserting a positive electrode probe into the inner side skin of the left lower limb, inserting a ground electrode probe into the inner side skin of the right lower limb, and paying attention not to insert muscles and insert blood vessels. A complete normal rat electrocardiogram is shown in FIG. 2(a), and includes, from left to right, P wave, QRS complex, and T wave; the rat electrocardiogram after successful modeling shows an elevated S-T band, as shown in FIG. 2 (b). Rats with insignificant elevation of the S-T band of the electrocardiogram were eliminated and the amount lost by elimination and intraoperative death of each group was replenished according to the deficit supplementation method.

1.5.3 animal grouping and administration

Rats were randomly divided into 5 groups, i.e., sham group (sham group), model group (model group, myocardial ischemia model group), loganin group (loganin group, myocardial ischemia model group + loganin) (divided into 45mg/kg low dose group, 90mg/kg medium dose group and 180mg/kg high dose group). Dissolving loganin in distilled water, and performing intragastric administration for 1 time every day for 7 days at doses of 45mg/kg, 90mg/kg and 180mg/kg 3h after molding. The sham and model groups were given equal volumes (1.5ml) of distilled water.

1.5.4 immunofluorescent staining for detecting vascular endothelial cell proliferation and blood vessel density

After perfusion and material drawing, the rat is frozen and sliced. 15um hearts were sectioned, washed 3 times with PBS solution for 5 minutes each, then placed in a wet box and ruptured for 30 minutes in 0.5% PBST solution. 10% donkey serum blocking solution (prepared in 0.5% PBST solution) was blocked for 2 hours at room temperature. Primary anti-dilution (rabbit Ki67 antibody diluted 1: 200, lectin antibody diluted 1: 800) was added and allowed to act at 4 ℃ for 16 hours, with care taken away from light. The PBS solution was washed 3 times for 5 minutes each, and carefully protected from light. Secondary antibody dilutions (Alexa Fluor 594 donkey anti-rabbit IgG diluted 1: 400) were added and allowed to react for 2 hours at room temperature. Adding a sealing agent to seal the piece, taking care of no air bubbles, and drying at room temperature. The images were observed and photographed with an upright fluorescence microscope and analyzed with Image Pro Plus 6.0 software. lecins are microvascular markers that selectively bind to the cell membrane of endothelial cells, and Ki67 is used to label novacells. As shown in FIG. 3, the immunofluorescence double-labeling results showed that the lectin in peri-infarct region in the model group was compared with that in the sham operation group⁺/Ki67⁺Marked neovascular endothelial cells are increased significantly (P < 0.01). Compared with the model group, the loganin is administered at 180mg/kg⁺/Ki67⁺Marked neoendothelial cells were further significantly increased (P < 0.01). The result shows that loganin has obvious promotion effect on the proliferation of endothelial cells in the peripheral area of infarction.

After 7 days from AMI modeling, compared with the sham operation group, the periinfarct lectin in the model group⁺The blood vessel density was significantly reduced (P < 0.001), indicating that AMI surgery significantly disrupted the microvascular network in the peri-infarct area. Compared with the model group, the loganin administration group can increase lectin⁺Blood vessel density, but no significant difference (as shown in figure 4). FIG. 5 shows the results of 14 days after AMI modeling, in comparison with the sham group, the periinfarct lectin in the model group⁺The blood vessel density is obviously reduced (P is less than 0.01); form a model setAdministration of collemin 180mg/kg⁺The blood vessel density is obviously increased (P < 0.05). The results show that loganin can significantly increase the vascular density in the peri-infarct area after 14 days of AMI administration, thereby establishing collateral circulation and restoring the blood supply of ischemic myocardium.

1.5.5Western Blot technique for detecting the influence of loganin on the expression of angiogenesis-related factors after myocardial ischemia in rats

7 days after molding, rats are anesthetized with 4ml/kg of 10% chloral hydrate abdominal cavity, the thoracic cavity is cut open, the heart is quickly cut off, and the rat is placed in 4 ℃ physiological saline while the heart still beats, cut off an infarction focus and a peripheral area, washed clean, sucked dry by filter paper, wrapped in tinfoil paper, placed in liquid nitrogen for freezing and storing for a moment, and stored at-80 ℃. The myocardial tissue frozen at-80 ℃ was weighed in a 5ml EP tube, and 9. mu.l/1 mg fresh tissue of precooled lysate and 100. mu.l RIPA of PMSF 1. mu.l were added thereto, and sufficiently pulverized by ultrasound on ice. Standing at 4 deg.C for 30min, centrifuging at 4 deg.C 12000r/min for 30min, collecting supernatant, determining protein concentration, diluting total protein with 5X sample buffer solution 1: 4, and denaturing at 95 deg.C for 10 min. The gel was separated by 12% SDS-PAGE and concentrated by 5% SDS-PAGE. Electrophoresis: concentrating gel voltage at 60V for 40min, separating gel voltage at 90V for 90min, and separating protein. After electrophoresis, the membrane is transferred by using a 0.45 mu mNC membrane; the strips were blocked in 5% skim milk powder for 2 h; rabbit-derived FGF-2 antibody (1: 1000), rabbit-derived Ang-1 antibody (1: 1000), rabbit-derived VFGFA antibody (1: 1000), rabbit-derived P-PKC antibody (1: 1000), rabbit-derived P-Src antibody (1: 1000), rabbit-derived P-Erk1/2 antibody (1: 1000), rabbit-derived Erk1/2 antibody (1: 1000), and rabbit-derived GAPDH antibody were incubated overnight at 4 ℃ in a refrigerator, respectively. TBST buffer washes 3 times for 10min each. Then incubating with corresponding horseradish peroxidase (HRP) -conjugated secondary antibody at room temperature for 2h, washing with TBST buffer solution for 3 times, each time for 10 min; developing with ECL for 1min, filtering off developing solution, and taking picture with gel imager. The protein bands were subjected to gray scale analysis using quality One software.

7 days after loganin administration, FGF-2 expression was examined and the results are shown in FIG. 6. The expression level of FGF-2 protein in the model group is found to be increased compared with that in the sham operation group, but no significant difference exists; the administration of 45mg/kg, 90mg/kg and 180mg/kg of loganin can further increase the expression of FGF-2, and show dose dependence, and compared with a model group, the expression level of FGF-2 protein of a 180mg/kg loganin administration group is obviously increased (P < 0.05).

7 days after loganin administration, expression of Ang-1 was measured, and the results are shown in FIG. 7. The expression level of Ang-1 in the model group is obviously increased (P is less than 0.01) compared with that in the sham operation group; compared with the model group, the expression level of Ang-1 in the loganin administration groups of 90mg/kg and 180mg/kg is obviously increased, and the difference has statistical significance (P is less than 0.05, and P is less than 0.01).

VEGFA expression was measured 7 days after loganin administration, and the results are shown in fig. 8. The protein expression level of VEGFA in the model group is obviously increased (P is less than 0.05) compared with that in the sham operation group; compared with the model group, the VEGFA protein expression level of the loganin administration group of 180mg/kg is obviously increased (P < 0.05).

7 days after loganin administration, the phosphorylation expression of Src, PKC, Erk1/2 was measured, and the results are shown in FIG. 9. Compared with a sham operation group, the protein expression level of the model group P-Src, P-PKC and P-Erk1/2 is increased, but no significant difference exists; compared with the model group, the protein expression levels of the 180mg/kg loganin administration group P-Src, P-PKC and P-Erk1/2 are all obviously increased (P is less than 0.05, P is less than 0.01, and P is less than 0.01), and the influence of loganin on the expression of the P-Src, P-PKC and P-Erk1/2 proteins shows dose dependence.

1.5.6 echocardiographic monitoring

The M-mode two-dimensional echocardiogram of the rat is monitored 14 days and 28 days after the model is made respectively. The rat is inhaled according to the ratio of isoflurane to oxygen of 1: 1, and the chest hair is scraped. The supine position is fixed on a workbench, a high-resolution small animal ultrasonic image system (Vevo 2100) is used for monitoring a rat heart M-type two-dimensional ultrasonic cardiogram, the probe frequency is 15MHz, the left ventricle end diastolic inner diameter (LVIDd) and the left ventricle end systolic inner diameter (LVIDs) are measured, and the left ventricle Ejection Fraction (EF) and the left ventricle short axis shortening rate (FS) are calculated according to a formula. All measurements were taken as the average of 3 cardiac cycles.

The echocardiogram result shows that after 14 days of modeling, the left ventricular Ejection Fraction (EF) and the short axis shortening rate (FS) of the model group are both obviously reduced (P is less than 0.001 and P is less than 0.001) compared with the false operation group, and the cardiac function is obviously damaged. Compared with the model group, the left ventricular Ejection Fraction (EF) and the short axis shortening rate (FS) of the 180mg/kg loganin treatment group are both obviously increased (P is less than 0.05, and P is less than 0.05), and the cardiac function is obviously improved compared with the model group (as shown in figure 10).

After 28 days of modeling, the left ventricular Ejection Fraction (EF) and the short axis shortening rate (FS) of the model group are obviously reduced (P is less than 0.001 and less than 0.001) compared with the false operation group, and the left ventricle is expanded and is accompanied with heart failure. Compared with the model group, the left ventricular Ejection Fraction (EF) and the short axis shortening rate (FS) of the loganin treatment group of 180mg/kg are obviously increased (P is less than 0.05, and P is less than 0.05). The results show that loganin can effectively improve the long-term ejection function of AMI and has a protective effect on the cardiac function (as shown in figure 11).

1.5.7 data analysis

The experimental data are statistically analyzed by SPSS17.0 statistical software, and the results are averaged to be +/-standard error

And (4) showing. The mean-average comparison of samples between groups was performed using one-way analysis of variance (ANOVA). P < 0.05 indicates statistical significance.

And (4) conclusion: according to the application, a rat myocardial ischemia model is adopted, and loganin is firstly used for researching and treating myocardial ischemia diseases. After myocardial ischemia, loganin establishes collateral circulation by increasing endothelial cell proliferation and blood vessel density in the periinfarction area, restores blood flow perfusion in the ischemic area, has obvious effect of promoting angiogenesis on AMI, and finally improves the cardiac function. The mechanism that loganin promotes angiogenesis after AMI is probably realized by regulating the expression of angiogenesis promoting factors FGF-2, Ang-1, VEGFA and related proteins of proliferation signal pathway P-Src, P-PKC and P-Erk 1/2.

Claims

1. Use of loganin in the manufacture of a medicament for the treatment or prevention of a disease associated with myocardial ischemia.

2. The use according to claim 1, wherein loganin has an angiogenesis promoting effect.

3. The use according to claim 1, wherein the diseases associated with myocardial ischemia are selected from the group consisting of coronary heart disease, myocardial injury, angina pectoris, arrhythmia, myocardial infarction and sudden death.

4. The use of claim 3, wherein the myocardial injury is myocardial ischemia reperfusion injury, and/or the myocardial infarction is an acute myocardial infarction.

5. Use of loganin in the manufacture of a medicament for promoting angiogenesis.

6. Use of loganin in preparing promoter of blood vessel growth factor expression is provided.

7. The use according to claim 6, wherein said pro-angiogenic growth factor is selected from the group consisting of FGF-2, Ang-1, VEGFA.

8. Use of loganin in preparing proliferation signal pathway related protein P-Src, P-PKC or P-Erk1/2 expression enhancer is provided.