Application of berberine hydrochloride and mulberry leaf extract in preparation of composition for preventing, delaying or treating diabetic cardiomyopathy related symptoms

Abstract

The invention provides an application of berberine hydrochloride and mulberry leaf extract in preparation of a composition for preventing, delaying or treating diabetic cardiomyopathy related symptoms. According to the application, berberine hydrochloride and a mulberry leaf extract are combined for use, and the composition for treating or improving the symptoms related to the diabetic cardiomyopathy is prepared. The combination of berberine hydrochloride and mulberry leaf extract can significantly relieve the symptoms of diabetic cardiomyopathy, including but not limited to promotion of structural growth and functional protein synthesis of myocardial cells of patients with diabetic cardiomyopathy, and optimization of stress response and electrophysiological homeostasis of myocardial cells. The traditional Chinese medicine composition can regulate and control energy metabolism disorder, myocardial apoptosis, cardiac microcirculation disorder and the like of patients, and has great potential in the fields of diabetes mellitus cardiac function protection and whole body health management as a composition component.

Description

TECHNICAL FIELD

[0001] The application relates to the technical field of diabetes diseases, in particular to application of berberine hydrochloride and mulberry leaf extract in preparation of a composition for preventing, delaying or treating symptoms related to diabetic cardiomyopathy. BACKGROUND

[0002] Diabetic cardiomyopathy (DCM) is one of the most serious complications of diabetes, and is a key factor leading to heart failure and death in diabetic patients. Its typical pathological features are systolic dysfunction and myocardial fibrosis, and its pathogenesis is unrelated to traditional cardiovascular risk factors such as hypertension, coronary atherosclerosis and heart valve disease. The pathophysiological mechanism of DCM is complex, involving mitochondrial dysfunction, oxidative stress, inflammatory response, myocardial cell apoptosis, myocardial fibrosis, autophagy disorder and endoplasmic reticulum stress.

[0003] Myocardial fibrosis is a key pathological environment for the development of DCM. Its essence is that when the myocardial tissue is subjected to continuous damage such as high glucose toxicity, oxidative stress, inflammation and hemodynamic load, myocardial fibroblasts are excessively activated, leading to excessive deposition of extracellular matrix, replacing normal functional myocardial cells. This further leads to increased stiffness and decreased compliance of the heart, severely damaging the systolic and diastolic functions of the heart, and there is currently no specific reversal method.

[0004] Existing treatment strategies for DCM or myocardial fibrosis include controlling blood glucose and blood pressure, or using drugs that inhibit the adrenergic-angiotensin system. Traditional hypoglycemic drugs (such as metformin) can effectively regulate blood glucose, but have limited intervention ability for myocardial cell apoptosis and calcium homeostasis imbalance induced in a high-sugar high-fat environment. Standard drugs for heart failure (such as angiotensin-converting enzyme inhibitors, ACEI) mainly alleviate hemodynamic abnormalities. These therapies have a single target and cannot simultaneously address the two core problems in the pathological process of DCM: the fibrosis process induced by continuous stress damage, and the lack of effective means to promote myocardial self-repairing ability to replace damaged areas.

[0005] Therefore, there is an urgent need in the art to develop a new treatment regimen that can simultaneously solve the two core pathological problems of DCM, i.e., protecting the myocardium from stress damage while promoting myocardial cell repair, thereby efficiently reversing myocardial fibrosis. SUMMARY

[0006] The main purpose of the present application is to provide application of berberine hydrochloride and mulberry leaf extract in preparation of a composition for preventing, delaying or treating symptoms related to diabetic cardiomyopathy, so as to solve the problem of poor treatment effect on diabetic cardiomyopathy in the prior art.

[0007] In order to achieve the above object, according to a first aspect of the present application, there is provided a use of berberine hydrochloride and mulberry leaf extract in the preparation of a composition for treating or improving symptoms associated with diabetic cardiomyopathy, the use comprising: combining the berberine hydrochloride and the mulberry leaf extract to prepare the composition for treating or improving symptoms associated with diabetic cardiomyopathy.

[0008] Further, the use further comprises: the mulberry leaf extract containing 1-deoxynojirimycin.

[0009] Further, the mulberry leaf extract contains 1-10% of 1-deoxynojirimycin by mass fraction.

[0010] Further, the symptoms associated with diabetic cardiomyopathy comprise: cardiac systolic dysfunction and / or myocardial fibrosis.

[0011] Further, the symptoms associated with diabetic cardiomyopathy comprise: elevated levels of inflammatory factors in serum, elevated respiratory exchange ratio, VCO2 / VO2, reduced cardiac ejection fraction, elevated expression levels of myocardial injury markers, myocardial cell oxidative stress, and myocardial cell apoptosis.

[0012] Further, the elevated and reduced criteria are whether the relevant parameters of diabetic cardiomyopathy patients treated with the combination of berberine hydrochloride and mulberry leaf extract are significantly different from the relevant parameters of diabetic cardiomyopathy patients without treatment.

[0013] Further, the inflammatory factors comprise: any one or more of IL-1β, IL-6, and TNF-α.

[0014] Further, the myocardial injury markers comprise: any one or more of troponin and B-type natriuretic peptide.

[0015] Further, when the berberine hydrochloride and the mulberry leaf extract are used in combination, the berberine hydrochloride is used in an amount of 1-100 mg / kg / day and the mulberry leaf extract is used in an amount of 5-500 mg / kg / day, based on the body weight of the mouse.

[0016] Further, when the berberine hydrochloride and the mulberry leaf extract are used in combination for a subject, the berberine hydrochloride is used in an amount of 100-3000 mg / day and the mulberry leaf extract is used in an amount of 100-1500 mg / day.

[0017] Further, the mass ratio of the use amount of the berberine hydrochloride to the mulberry leaf extract is 1:1-10.

[0018] In order to achieve the above object, according to a second aspect of the present application, there is provided a method for preventing, delaying or treating diabetic cardiomyopathy related symptoms, the method comprising: administering to a subject a pharmaceutically effective amount of a composition comprising berberine hydrochloride and mulberry leaf extract.

[0019] Further, the application further comprises that the mulberry leaf extract contains 1-deoxynojirimycin.

[0020] Further, the mulberry leaf extract contains 1-10% of 1-deoxynojirimycin by mass fraction.

[0021] Further, the diabetic cardiomyopathy related symptoms include cardiac systolic dysfunction and / or myocardial fibrosis.

[0022] Further, the diabetic cardiomyopathy related symptoms include elevated levels of inflammatory factors in serum, elevated respiratory exchange ratio, VCO2 / VO2, reduced cardiac ejection fraction, elevated expression levels of myocardial injury markers, myocardial cell oxidative stress, and myocardial cell apoptosis.

[0023] Further, the elevated and reduced criteria are whether the related parameters of the diabetic cardiomyopathy patients treated with the combination of berberine hydrochloride and mulberry leaf extract are significantly different from the related parameters of the diabetic cardiomyopathy patients without treatment.

[0024] Further, the inflammatory factors include any one or more of IL-1β, IL-6 and TNF-α.

[0025] Further, the myocardial injury markers include any one or more of troponin and B-type natriuretic peptide.

[0026] Further, when the berberine hydrochloride and the mulberry leaf extract are used in combination, the amount of the berberine hydrochloride used is 100-3000 mg / day, and the amount of the mulberry leaf extract used is 100-1500 mg / day.

[0027] Further, in the composition, the mass ratio of the berberine hydrochloride to the mulberry leaf extract is 1:1-10.

[0028] In order to achieve the above object, according to a third aspect of the present application, there is provided a pharmaceutical composition for preventing, delaying or treating diabetic cardiomyopathy related symptoms, the pharmaceutical composition comprising berberine hydrochloride and mulberry leaf extract.

[0029] Further, the mulberry leaf extract contains 1-deoxynojirimycin.

[0030] Further, the mulberry leaf extract contains 1-10% of 1-deoxynojirimycin by mass fraction.

[0031] Further, the symptoms related to diabetic cardiomyopathy include: cardiac systolic dysfunction and / or myocardial fibrosis.

[0032] Further, the symptoms related to diabetic cardiomyopathy include: elevated content of inflammatory factors in serum, elevated respiratory exchange ratio, VCO2 / VO2, reduced cardiac ejection fraction, elevated expression level of myocardial injury markers, myocardial cell oxidative stress, myocardial cell apoptosis.

[0033] Further, the judgment criteria for the increase and decrease are: whether the relevant parameters of the diabetic cardiomyopathy patients treated by the combination of berberine hydrochloride and mulberry leaf extract are significantly different from the relevant parameters of the diabetic cardiomyopathy patients without treatment.

[0034] Further, the inflammatory factors include: any one or more of IL-1β, IL-6 and TNF-α.

[0035] Further, the myocardial injury markers include: any one or more of troponin and B-type natriuretic peptide.

[0036] Further, when the berberine hydrochloride and the mulberry leaf extract are used on the subject, the amount of the berberine hydrochloride used is 100-3000 mg / day, and the amount of the mulberry leaf extract used is 100-1500 mg / day.

[0037] Further, in the pharmaceutical composition, the mass ratio of the berberine hydrochloride to the mulberry leaf extract is 1:1-10.

[0038] By using the technical solution of the present application, the combination of the berberine hydrochloride and the mulberry leaf extract can significantly relieve the symptoms of diabetic cardiomyopathy, not only protecting the myocardium from stress injury, but also promoting the repair of myocardial cells, so as to achieve the effect of efficiently reversing myocardial fibrosis. In addition, the combination of the two can promote the structural growth of myocardial cells and the functional protein synthesis of diabetic cardiomyopathy patients, optimize the stress response and electrical physiological homeostasis of myocardial cells, and regulate the energy metabolism disorder, myocardial cell apoptosis and cardiac microcirculation disorder of patients on multiple levels. As a composition ingredient, it has great potential in the field of diabetic cardiac function protection and overall health management. BRIEF DESCRIPTION OF DRAWINGS

[0039] The drawings accompanying the specification of this application are used to provide further understanding of the present application, the illustrative embodiments of the present application and the description thereof serve to explain the present application, and do not constitute an improper limitation on the present application. In the drawings:

[0040] Figure 1 FIG. 1 shows the ROS level fluorescence microscope observation results of the HGPA group and the BSA group in Example 1 of the present application after staining by the reactive oxygen species (ROS) detection kit. Figure 1A) and quantitative statistics of ROS levels ( Figure 1 B);

[0041] Figure 2 The statistical graph of apoptosis ratios of the HGPA group and the BSA group detected by the Annexin V-FITC apoptosis detection kit in Example 1 of the present application is shown;

[0042] Figure 3 The quantitative statistical graph of blood glucose of the type 2 diabetes mouse model and normal mice in Example 1 of the present application is shown;

[0043] Figure 4 The results of fluorescence microscopy observation of ROS levels in HGPA cells treated differently in Example 1 of the present application after staining with a reactive oxygen species (ROS) detection kit are shown ( Figure 4 A) and quantitative statistics of ROS levels ( Figure 4 B);

[0044] Figure 5 Shown is a statistical graph of different inflammatory factor levels detected in the type 2 diabetes mouse model with different treatments in Example 1 of the present application;

[0045] Figure 6 Statistical graphs showing the detection of cardiac troponin (cTNT) and B-type natriuretic peptide (BNP) levels in type 2 diabetic mouse models treated with different methods in Example 1 of the present application are shown;

[0046] Figure 7 The images obtained by performing cardiac ultrasound on the type 2 diabetes mouse model with different treatments in Example 1 of the present application are shown ( Figure 7 A) and cardiac ejection fraction statistics ( Figure 7 B);

[0047] Figure 8 The diagram shows the results of staining observation of tissue sections of the heart of the type 2 diabetes mouse model treated with different methods in Example 1 of the present application;

[0048] Figure 9 The figure shows the statistical graph of the myocardial fibrosis degree observed by staining tissue sections of the hearts of the type 2 diabetes mouse models with different treatments in Example 1 of the present application. DETAILED DESCRIPTION

[0049] It should be noted that, in the absence of conflict, the embodiments and features of the embodiments in this application can be combined with each other. The present invention will be described in detail below with reference to the accompanying drawings and in combination with the embodiments.

[0050] As mentioned in the background, due to the complexity of the pathology of diabetic cardiomyopathy, which involves multiple indicators throughout the body, it is difficult to achieve ideal therapeutic effects by using single mechanism drug therapy in the prior art, and it is urgent to develop new therapeutic strategies that can specifically improve the energy metabolism disorder of patients, myocardial cell apoptosis and cardiac microcirculation disorder and other aspects to regulate multiple aspects of symptoms. At present, there is no report on the combination of berberine hydrochloride and mulberry leaf extract to treat or improve symptoms related to diabetic cardiomyopathy. Based on this, the present application provides a drug application capable of improving various indicators of patients with diabetic cardiomyopathy in multiple aspects.

[0051] In a first typical embodiment of the present application, the use of berberine hydrochloride and mulberry leaf extract in the preparation of a composition for treating or improving symptoms related to diabetic cardiomyopathy is provided. The application includes: the use of berberine hydrochloride and mulberry leaf extract in combination to prepare a composition for treating or improving symptoms related to diabetic cardiomyopathy.

[0052] Berberine hydrochloride, also known as berberine (Berberine, BBR), is a naturally occurring alkaloid compound widely distributed in various plants such as Berberis, Coptis, and Phellodendron. Berberine can specifically up-regulate at least one of the following signal pathways: cardiac myosin binding, amino acid transport, cardiac myocyte development, myosin complex level or tissue homeostasis. The core mechanism of berberine is to promote the structural growth of myocardial cells and the synthesis of functional proteins (myosin and actin), and to actively guide the heart to adapt to functional reconstruction rather than pathological fibrosis.

[0053] In a preferred embodiment, the mulberry leaf extract contains 1-deoxynojirimycin. 1-Deoxynojirimycin (1-Deoxynojirimycin, DNJ) is a naturally occurring amino sugar alcohol compound mainly found in mulberry leaves and is a characteristic active ingredient of mulberry leaves. Mulberry leaf extract can specifically up-regulate at least one of the following signal pathways: cation transmembrane transport, response to hypoxia, response to mechanical stimulation, or myocardial cell calcium signal regulation. The core mechanism of mulberry leaf extract is to optimize the stress response system and electrical physiological homeostasis of myocardial cells, stabilize cell electrical physiology, optimize calcium signals, and enhance the resistance of cells to hypoxia and mechanical stress, thereby providing sufficient protection to myocardial cells, significantly reducing cell damage and death in a diabetic environment, and inhibiting the generation of pro-fibrosis signals from the source.

[0054] The composition obtained by using berberine and mulberry leaf extract has a significant synergistic gain effect, especially for myocardial fibrosis, which is much greater than the effect of using berberine alone or mulberry leaf extract alone, and is not a simple additive effect of the two, but has a significant synergistic effect.

[0055] The heart function optimization function of mulberry leaf extract and the heart structure enhancement effect of berberine are combined to break the vicious cycle of "myocardial injury → functional injury → compensatory remodeling → accelerated fibrosis → further injury" of diabetic cardiomyopathy patients. A benign cycle of stable function and optimized structure is established. The combination of berberine and mulberry leaf extract can not only inhibit myocardial fibrosis, but also reverse the formed fibrosis, restore heart function, and even restore the degree of myocardial fibrosis to the level similar to that of the non-diseased population. This is difficult to achieve with single mechanism drugs, and it is also a technical effect that cannot be achieved by the compositions in the prior art. Myocardial fibrosis can cause other diseases such as heart failure and arrhythmia, and the improvement of myocardial fibrosis also has a certain positive effect on other diseases caused by it.

[0056] Any mulberry leaf extract containing 1-deoxynojirimycin is suitable for the present application. In a preferred embodiment, the mulberry leaf extract contains 1-10% 1-deoxynojirimycin, specifically 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, or 10%.

[0057] Since the pathology of diabetic cardiomyopathy is complex, any symptom related to the diagnosis of diabetic cardiomyopathy is suitable for the present application. In a preferred embodiment, the symptoms related to diabetic cardiomyopathy include: cardiac systolic dysfunction and / or myocardial fibrosis. The above two symptoms are the main symptoms of this disease.

[0058] Any symptom related to the diagnosis of diabetic cardiomyopathy is suitable for the present application. In a preferred embodiment, the symptoms related to diabetic cardiomyopathy include: elevated levels of inflammatory factors in serum, increased respiratory exchange rate (VCO2 / VO2), decreased cardiac ejection fraction, elevated expression levels of myocardial injury markers, myocardial cell oxidative stress, and myocardial cell apoptosis.

[0059] In order to further objectively judge the indicators, in a preferred embodiment, the judgment criteria for the increase and decrease are: whether the relevant parameters of diabetic cardiomyopathy patients treated with a combination of hydrochloric acid berberine and 1-deoxynojirimycin are significantly different from the relevant parameters of diabetic cardiomyopathy patients without treatment.

[0060] Any inflammatory factor that can indicate the degree of inflammation is suitable for the present application. In a preferred embodiment, the inflammatory factors include any one or more of IL-1β, IL-6, and TNF-α.

[0061] Any marker that can indicate the degree of myocardial injury is suitable for the present application. In a preferred embodiment, the myocardial injury markers include any one or more of troponin (cTNT) and B-type natriuretic peptide (BNP).

[0062] In order to efficiently exert the effects of berberine hydrochloride and mulberry leaf extract, the actual application amount thereof needs to be controlled within a proper range. In a preferred embodiment, when the berberine hydrochloride and mulberry leaf extract are used in combination, the amount used is 1-100 mg / kg / day of berberine hydrochloride and 5-500 mg / kg / day of mulberry leaf extract, based on the body weight of the mouse, in the case of mouse experiments, by gavage or drinking water. In clinical studies, the amount used is 100-3000 mg / day of berberine hydrochloride and 100-1500 mg / day of mulberry leaf extract, in the case of oral administration in 2-3 doses per day. In a preferred embodiment, the mass ratio of the amount of berberine hydrochloride to that of mulberry leaf extract is 1:1-10.

[0063] In a second exemplary embodiment of the present application, a method for preventing, delaying or treating a diabetic cardiomyopathy-related symptom is provided, the method comprising administering to a subject a pharmaceutically effective amount of a composition comprising berberine hydrochloride and mulberry leaf extract.

[0064] The "pharmaceutically effective amount" of the present application refers to an amount sufficient to produce the intended prophylactic, delaying or therapeutic effect, which can achieve (i) a detectable modulation of a target protein / receptor / pathway in a mammal (especially a human), (ii) an improvement of at least 10% in one or more clinical indicators of the relevant disease or symptom compared to the baseline level, or (iii) a statistically significant difference (p<0.05) compared to a blank control or placebo. The effective amount can be a single administration dose or a cumulative dose after multiple administrations. The specific dose can be determined by routine experiments by those skilled in the art according to the administration route, disease type and severity, patient age, body weight, liver and kidney function, and concomitant medication. Therefore, any administration amount that can achieve the above-mentioned prophylactic, delaying or therapeutic results without producing unacceptable toxicity falls within the scope of the "pharmaceutically effective amount" of the present application.

[0065] In a preferred embodiment, the application further comprises that the mulberry leaf extract contains 1-deoxynojirimycin.

[0066] In a preferred embodiment, the mulberry leaf extract contains 1-10% of 1-deoxynojirimycin by mass fraction.

[0067] In a preferred embodiment, the diabetic cardiomyopathy-related symptom comprises cardiac systolic dysfunction and / or myocardial fibrosis.

[0068] In a preferred embodiment, the symptoms related to diabetic cardiomyopathy include: elevated levels of inflammatory factors in serum, elevated respiratory exchange ratio, VCO2 / VO2, reduced cardiac ejection fraction, elevated expression levels of markers of myocardial injury, oxidative stress in cardiomyocytes, and apoptosis of cardiomyocytes.

[0069] In a preferred embodiment, the criteria for determining the elevation and reduction are whether the relevant parameters of the diabetic cardiomyopathy patients treated with the combination of berberine hydrochloride and mulberry leaf extract are significantly different from those of the diabetic cardiomyopathy patients without treatment.

[0070] In a preferred embodiment, the inflammatory factors include any one or more of IL-1β, IL-6, and TNF-α.

[0071] In a preferred embodiment, the markers of myocardial injury include any one or more of troponin and B-type natriuretic peptide.

[0072] In a preferred embodiment, when the berberine hydrochloride and the mulberry leaf extract are used in combination for the subject, the amount of berberine hydrochloride used is 100-3000 mg / day, and the amount of mulberry leaf extract used is 100-1500 mg / day; optionally, the above composition is administered orally; and optionally, the above composition is administered 2-3 times per day.

[0073] In a preferred embodiment, in the composition, the mass ratio of berberine hydrochloride to mulberry leaf extract is 1:1-10.

[0074] The application will be further described in detail below in conjunction with specific embodiments, which should not be construed as limiting the scope of the application.

[0075] Human myocardial cells AC16 are from ATCC (CRL-3568); reactive oxygen species (ROS) detection kit is from Biyun Tian Biotechnology Co., Ltd. (S0033S); Annexin V-FITC cell apoptosis detection kit is from Biyun Tian Biotechnology Co., Ltd. (C1062L); male C57BL / 6J mice are from Sibeifeng Biotechnology Co., Ltd. (B204-02).

[0076] The mulberry leaf extract used in the embodiments of the application is a commercial product Mulberry leaf extract. According to GB / T 40642, the 1-deoxynojirimycin in the mulberry leaf extract is detected, i.e., the content of 1-deoxynojirimycin is detected by high-performance liquid chromatography, and the mass content of 1-deoxynojirimycin in the mulberry leaf extract used below is measured to be 5%. The preparation method of the mulberry leaf extract is as follows:

[0077] A. To the raw material mulberry leaf powder, add water with a weight of 10-20 times the weight of the mulberry leaf powder, extract 1-3 times at 60-100°C, combine the extract and pass through a 200-mesh sieve to obtain a filtrate; wherein the mulberry leaf powder is obtained by crushing mulberry leaves (Mulberry leaf) and passing through a 20-40-mesh sieve;

[0078] B. Pass the filtrate through a 732 cation resin chromatographic column at a flow rate of 0.5-2 times the resin volume / h, remove impurities with deionized water or 0.05-0.5 mol / L ammonia water, and the eluent is 0.5-2 mol / L ammonia water at a flow rate of 1.5 times the column volume / h;

[0079] C. Collect the eluent, concentrate, spray dry, and crush through an 80-100-mesh sieve to obtain a mulberry leaf extract.

[0080] Example 1 Berberine hydrochloride and mulberry leaf extract significantly inhibit the progression of diabetic myocardial fibrosis and promote the recovery of cardiac function

[0081] This example studies the combined use of berberine hydrochloride (BBR) and mulberry leaf extract on diabetic myocardial fibrosis and its effect on promoting the recovery of cardiac function.

[0082] This example uses human myocardial cells AC16 as a model to construct a high glucose and high fat (HGPA) treatment group and a control bovine serum albumin (BSA) group. AC16 cells in the logarithmic growth phase were evenly inoculated in a 96-well plate and incubated at 37°C, 5% CO2 for overnight. To simulate the mechanism of diabetes on myocardium, a high glucose and high fat (HGPA) treatment group (glucose 25.5 mmol / L + palmitic acid 200 μmol / L) and a control bovine serum albumin (Ctrl) group (glucose 5.5 mmol / L + BSA 200 μmol / L) were set up and induced at 37°C, 5% CO2 for 48 hours.

[0083] The above two groups of cells (after 48 hours of treatment) were stained by using an active oxygen (ROS) detection kit, and the fluorescence intensity of the cells was detected under a fluorescence microscope (excitation wavelength 488 nm, emission wavelength 525 nm), as shown in Figure 1 A- Figure 1 B, it can be seen that the ROS level of myocardial cells AC16 significantly increased after 48 hours of HGPA treatment, and the fluorescence intensity increased by 8 times. This indicates that the high glucose and high fat environment significantly induced oxidative stress in the cells.

[0084] In addition, the Annexin V-FITC apoptosis detection kit was used to detect the apoptosis rate (Q2+Q3) of the above two groups of cells (after 48 hours of treatment). As shown in Figure 2As shown, almost no apoptotic cells were detected in the AC16 cells of the control group, while the proportion of early and late apoptotic cells in the HGPA group was more than 80% after 48 hours of treatment.

[0085] As can be seen from the above, the simulated diabetic environment of the HGPA group can significantly induce oxidative stress and apoptosis of cardiomyocytes.

[0086] Through cell experiments, we treated the HGPA-treated cells with 10 μM of berberine alone (HGPA+Berberine), 10 μM of mulberry leaf extract alone (HGPA+Mulberry leaves), and 5 μM of berberine and 5 μM of mulberry leaf extract in combination (HGPA+Berberine+Mulberry leaves), respectively, and detected the reactive oxygen species (ROS) levels of the above three treatments, the control group, and the HGPA group after 24 hours.

[0087] As shown in Figure 4 A- Figure 4 As shown in B, there was no significant difference in the effect of 10 μM of berberine alone or 10 μM of mulberry leaf extract alone on the ROS level of the cells, while the combined use of 5 μM of berberine and 5 μM of mulberry leaf extract could significantly reduce the ROS level of the cells to 30% of the level of the two treatments alone, and there was no significant difference from the control bovine serum albumin (BSA) group. The specific data are shown in Table 1. As can be seen, the combined use of berberine and mulberry leaf extract can exert a synergistic effect and effectively reduce the oxidative stress of cardiomyocytes.

[0088] Table 1:

[0089] Group Active oxygen level (fold) Control (BSA) 1.00±0.11 HGPA 9.83±1.46 HGPA + Berberine 5.50±0.79 HGPA + Mulberry leaves 6.65±0.90 HGPA + Berberine + Mulberry leaves 1.86±0.64

[0090] In addition, in this embodiment, a type 2 diabetic mouse model was successfully constructed by high-fat diet combined with low-dose intraperitoneal injection of streptozotocin (STZ).

[0091] The specific operation of construction is as follows: after 3 months of high-fat diet feeding of 3-month-old male C57BL / 6J mice, the initial value of fasting blood glucose was 6 mmol / L, and then after 5 days of intraperitoneal injection of 60 mg / kg STZ, the fasting blood glucose was significantly increased to 15 mmol / L, indicating that the type 2 diabetic mouse model was successfully constructed (as shown in Figure 3 ).

[0092] In the animal experiment, the diabetic mice were treated with 99% pure berberine (10 mg / kg / day) alone (DM+Berberine), mulberry leaf extract (100 mg / kg / day) alone (DM+Mulberryleaves), and 5 mg / kg / day of 99% pure berberine and 50 mg / kg / day of mulberry leaf extract (DM+Berberine+Mulberry leaves) respectively, and intervened for 3 months. The inflammatory factor content of the mice treated with the above three different treatments and male C57BL / 6J control mice and diabetic mice was detected.

[0093] The results are shown in Figure 5 Compared with single drug, the combination of drugs significantly reduced the levels of IL-1β, IL-6, and TNF-α in the serum of diabetic mice, and the combination of drugs could reduce the verification level of diabetic mice to about 0-1 times of normal mice (as shown in Table 2), further improving the inflammatory state.

[0094] Table 2:

[0095]

[0096]

[0097] In addition, the levels of cTNT and BNP in the serum of the above three different treated mice and male C57BL / 6J control mice and diabetic mice were further detected, and the results are shown in Figure 6 The combination of berberine and mulberry leaf extract significantly reduced the levels of cTNT and BNP in the serum of diabetic mice, and could reduce the myocardial injury marker level of diabetic mice to the normal level (as shown in Table 3), indicating that the combination treatment has obvious advantages in improving myocardial injury.

[0098] Table 3:

[0099] Group cTNT level (ng / L) BNP level (ng / L) Normal group 5.08±0.78 50.59±11.65 Diabetic mice (DM) 15.94±2.14 115.81±14.43 DM + Berberine 8.89±1.49 79.39±7.36 DM + Mulberry leaves 8.92±1.43 80.92±4.59 DM + Berberine + Mulberry leaves 5.67±1.13 57.50±6.33

[0100] The cardiac color Doppler ultrasound analysis of the above three different treated mice and male C57BL / 6J control mice and diabetic mice further confirmed that the combination of berberine hydrochloride and mulberry leaf extract could significantly improve the ejection fraction (EF) of diabetic mice, and could restore the ejection fraction of diabetic mice to the normal level (as shown in Table 4), and enhance the systolic function of the heart (as shown in Figure 7 A- Figure 7 B).

[0101] Table 4:

[0102] Group Ejection fraction (%) Normal group 75.22±2.65 Diabetic mice (DM) 45.08±2.99 DM + Berberine 52.63±4.67 DM + Mulberry leaves 54.98±3.77 DM + Berberine + Mulberry leaves 69.70±3.41

[0103] It is worth noting that by slicing the heart of the normal group (Ctrl), the diabetes group (DM), the diabetes + berberine group (DM + Berberine), the diabetes + mulberry leaf extract group (DM + Mulberry leaves), the diabetes + berberine + mulberry leaf extract group (DM + Berberine + Mulberry leaves) mice and performing picrosirius staining, the cardiac fibrosis observation was performed, and the results are shown in Table 5. Figure 8 - Figure 9 As shown in Table 5, compared with the normal group, the cardiac fibrosis of the diabetic mice was significantly increased, and the use of berberine alone or the use of mulberry leaf extract alone could effectively reduce the myocardial fibrosis, and the combined use of berberine and mulberry leaf extract could greatly reduce the myocardial fibrosis of the diabetic mice, and the degree approached that of the normal mice.

[0104] Table 5:

[0105] Group Myocardial fibrosis level (%) Normal group 0.96±0.42 Diabetic mice (DM) 6.15±1.20 DM + Berberine 3.70±0.44 DM + Mulberry leaves 4.04±0.46 DM + Berberine + Mulberry leaves 1.31±0.38

[0106] As can be seen from the above results, the combined use of berberine hydrochloride and mulberry leaf extract can significantly inhibit the progression of diabetic myocardial fibrosis and promote the recovery of cardiac function, and the effect is much better than that of the use of the two alone, the combined use of berberine hydrochloride and mulberry leaf extract can restore the problem indicators of diabetic cardiomyopathy patients to the level of the non-diseased population, and has a certain potential for the treatment of diabetic cardiomyopathy.

[0107] As can be seen from the above description, the above embodiments of the present application achieve the following technical effects: the combined use of berberine and mulberry leaf extract can significantly relieve the symptoms of diabetic cardiomyopathy.

[0108] The combined use of berberine and mulberry leaf extract can effectively reduce the ROS level in the diabetic model, significantly reduce myocardial cell apoptosis, and restore / improve mitochondrial function, relieve oxidative stress. The combined use of berberine and mulberry leaf extract has a significant synergistic effect, can effectively reduce the myocardial fibrosis of the diabetic model, maintain the structural integrity of the myocardial tissue, and significantly improve the cardiac contraction ability. It has a synergistic regulation on the energy metabolism disorder, oxidative stress, myocardial cell apoptosis, myocardial fibrosis and cardiac microcirculation disorder of diabetic cardiomyopathy patients, and can be seen that berberine and mulberry leaf extract have great potential in the field of diabetic heart function protection and overall health management.

[0109] The above merely provides the preferred embodiments of the present application, and is not used to limit the present application. For those skilled in the art, the present application can have various modifications and changes. Any modifications, equivalent replacements, improvements, etc. made within the principles and technical scope of the present application shall fall into the scope of the present application.

Claims

1. A use of berberine hydrochloride and mulberry leaf extract in the preparation of a composition for preventing, delaying or treating symptoms related to diabetic cardiomyopathy, characterized in that: The applications include: The berberine hydrochloride and the mulberry leaf extract are used in combination to prepare a composition for treating or improving symptoms related to diabetic cardiomyopathy.

2. The use according to claim 1, characterized in that The application also includes: the mulberry leaf extract contains 1-deoxynojirimycin.

3. The use according to claim 2, characterized in that Calculated by mass fraction, the mulberry leaf extract contains 1-10% of 1-deoxynojirimycin.

4. The use according to any one of claims 1 to 3, characterized in that The diabetic cardiomyopathy-related symptoms include: cardiac systolic dysfunction and / or myocardial fibrosis.

5. The use according to any one of claims 1 to 3, characterized in that The symptoms related to diabetic cardiomyopathy include: increased levels of inflammatory factors in serum, increased respiratory exchange rate, VCO2 / VO2, decreased cardiac ejection fraction, increased expression levels of myocardial injury markers, myocardial cell oxidative stress, and myocardial cell apoptosis.

6. The use according to claim 5, characterized in that The judgment criteria for the increase and decrease are: whether the relevant parameters of the diabetic cardiomyopathy patients treated with the berberine hydrochloride and the mulberry leaf extract are significantly different from the relevant parameters of the diabetic cardiomyopathy patients who have not been treated.

7. The use according to claim 5, characterized in that The inflammatory factors include any one or more of IL-1β, IL-6 and TNF-α.

8. The use according to claim 5, characterized in that The myocardial injury markers include any one or more of troponin and B-type natriuretic peptide.

9. The use according to any one of claims 1 to 3, characterized in that When the berberine hydrochloride and the mulberry leaf extract are used in combination, the dosage is based on the body weight of the mouse, and the dosage of the berberine hydrochloride is 1-100 mg / kg / day, and the dosage of the mulberry leaf extract is 5-500 mg / kg / day; Optionally, when the berberine hydrochloride and the mulberry leaf extract are used in combination on a subject, the dosage of the berberine hydrochloride is 100-3000 mg / day, and the dosage of the mulberry leaf extract is 100-1500 mg / day.

10. The use according to any one of claims 1 to 3, characterized in that The mass ratio of the berberine hydrochloride to the mulberry leaf extract is 1:1-10.

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