Application of berberine in preparation of medicine for treating hypertension

Abstract

The invention discloses an application of berberine in preparation of a medicine for treating hypertension. Experiments prove that berberine can improve changes of intestinal pathology caused by angiotensin II, blood pressure is reduced by adjusting intestinal flora, especially increasing abundance of Ackermann mucilaginophile and remodeling cardiovascular system, and blood pressure comprises systolic pressure and diastolic pressure. The invention has important application value in treatment of hypertension.

Description

Technical Field

[0001] The invention belongs to the field of biomedicine, and specifically relates to the application of berberine in preparing a medicine for treating hypertension. Background Art

[0002] Hypertension is a common cardiovascular disease that is often accompanied by increased vascular tension, increased cardiac burden, and other serious health problems. Patients with hypertension often need to take medications for a long time to maintain normal blood pressure levels, but these drugs may have adverse side effects and are not always effective. In recent years, intestinal flora (gut microbiota) plays an important role in human health, including regulation of the immune system, food digestion, and maintaining the integrity of the intestinal mucosa. More and more studies have shown that an imbalance in the intestinal flora is closely related to the occurrence and development of cardiovascular diseases such as hypertension. Therefore, regulating the intestinal flora has become a new research area for the treatment of hypertension.

[0003] In the intestinal flora, Akkermansia muciniphila (Akk bacteria) is considered to be an important probiotic that can reduce chronic inflammation, improve intestinal mucosal health, and have a positive impact on regulating intestinal flora and influencing host health. Intestinal flora imbalance is closely related to diet, and changing lifestyle and dietary structure plays an important role in intervening in hypertension. Therefore, readjusting the structure of intestinal flora by adjusting dietary structure, supplementing prebiotics, probiotics, and performing fecal microbiota transplantation may become a new way to treat hypertension.

[0004] Berberine is a naturally occurring alkaloid found in some herbs, such as those in the Berberidaceae family. Berberine has been used in traditional medicine to treat intestinal disorders. Summary of the invention

[0005] The purpose of the present invention is to treat hypertension or improve blood pressure levels.

[0006] The present invention firstly protects the application of berberine, which can be A1) or A2) or A3) or A4): A1) Preparation of medicaments for the treatment of hypertension; A2) Preparation of health products for improving blood pressure levels; A3) Preparation of drugs for lowering blood pressure; A4) Regulate intestinal flora.

[0007] In the above application, the regulation of intestinal flora can be manifested as increasing the abundance of Akkermansia muciniphila.

[0008] The present invention also protects the use of a composition containing berberine, which may be A1) or A2) or A3) or A4): A1) Preparation of medicaments for the treatment of hypertension; A2) Preparation of health products for improving blood pressure levels; A3) Preparation of drugs for lowering blood pressure; A4) Regulate intestinal flora.

[0009] In the above application, the regulation of intestinal flora can be manifested as increasing the abundance of Akkermansia muciniphila.

[0010] In the above application, the composition containing berberine can be a health product composition, a pharmaceutical composition or a probiotic composition.

[0011] In the above application, the composition containing berberine further comprises a second active ingredient, which may be a drug different from berberine.

[0012] In the above application, the composition containing berberine may further comprise one or more of telmisartan, captopril, valsartan and amlodipine.

[0013] In the above application, the hypertension may be renal hypertension, spontaneous hypertension, metabolic hypertension caused by combined dietary factors, or isolated systolic hypertension.

[0014] In the above applications, the drug may be a medicine, and its dosage form may be tablets, capsules, granules, pills, ointments, solutions, suspensions, emulsions, creams, sprays, drops, patches or tube feeding preparations.

[0015] In the above application, the health care product may be a health food, and its type may be a dairy product or a fermented cereal food.

[0016] In any of the above applications, the medicine or the health product may contain an effective amount of berberine.

[0017] In any of the above applications, the subject of the drug or the health product can be a mammal. Preferably, the mammal can be a human or a mouse.

[0018] In any of the above applications, lowering blood pressure may be lowering systolic blood pressure and / or diastolic blood pressure.

[0019] In any of the above applications, the blood pressure reduction may be to reduce the blood pressure level of angiotensin II-induced hypertension.

[0020] In any of the above uses, the hypertension may also be induced by angiotensin II.

[0021] The structural formula of any of the above berberine is shown in Formula I; .

[0023] The inventors of the present application have proved through experiments that berberine can lower blood pressure by remodeling the cardiovascular system and improving the intestinal microenvironment, and blood pressure includes systolic pressure and diastolic pressure. The present invention has important application value in treating hypertension. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 These are the measurement results of systolic blood pressure (SBP) and diastolic blood pressure (DBP) of AngII group mice and control group mice in step 4 of Example 1.

[0025] Figure 2 The staining and statistical results of aortic tissue and colon tissue of AngII group mice and control group mice in step 5 of Example 1.

[0026] Figure 3 These are the measurement results of systolic blood pressure (SBP) and diastolic blood pressure (DBP) of the AngII group and the AngII+Bbr group in step 2 of Example 2.

[0027] Figure 4 These are the PAS staining results of colon tissues of the AngII group and the AngII+Bbr group in step 3 of Example 2, and the statistical results of the colon muscle layer thickness and goblet cell ratio of mice.

[0028] Figure 5 These are the Masson staining results of aortic tissues in the AngII group and the AngII+Bbr group in step 3 of Example 2, and the statistical results of the aortic thickness and aortic fibrosis ratio of mice.

[0029] Figure 6 These are the statistical results of the number of genes, Chao1 index, Observed_species index and Shannon index of the metagenomic data of the two groups of mice in Example 3.

[0030] Figure 7 The PCoA results of the metagenomic data of the two groups of mice in Example 3 were based on the Bray-Curtis distance.

[0031] Figure 8 These are the LEFSe analysis results of the metagenomic data of the two groups of mice in Example 3 and the statistical results of Akk bacteria. DETAILED DESCRIPTION

[0032] The present invention is further described in detail below in conjunction with specific embodiments, and the examples provided are only for illustrating the present invention, rather than for limiting the scope of the present invention. The examples provided below can be used as a guide for further improvements by those of ordinary skill in the art, and do not constitute a limitation of the present invention in any way.

[0033] The experimental methods in the following examples, unless otherwise specified, are all conventional methods, and are performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials, reagents, etc. used in the following examples, unless otherwise specified, can all be obtained from commercial channels.

[0034] The healthy subjects and hypertensive patients in the following examples all gave their informed consent to the experiment. Example 1. Preparation and detection of angiotensin II (AngII) hypertensive mouse model 1. Take an implantable osmotic capsule (product of Alzet, model 1004, delivery rate 0.11 µl / hr) and prepare the AngII capsule pump and NaCl capsule pump according to the instructions.

[0035] 2. Take 10 SPF-grade C57BL / 6J mice aged 7-8 weeks and weighing 20±5g (hereinafter referred to as mice, purchased from Beijing Weitong Lihua Experimental Animal Technology Co., Ltd.), anesthetize them with 0.7% pentobarbital solution (solvent is normal saline), then make a small incision about 5mm long on the back of the mouse's neck, insert the AngII capsule pump, and finally suture the back of the mouse. The AngII capsule pump injects AngII through an osmotic micropump for 4 weeks, and the AngII injection dose is 800ng / (kg·min). After the mouse wakes up, the AngII hypertensive mouse model is prepared.

[0036] The AngII hypertension group (referred to as the AngII group) consisted of 10 AngII hypertensive mouse models.

[0037] 3. Take 10 SPF C57BL / 6J mice aged 7-8 weeks and weighing 20±5g, anesthetize them with 0.7% pentobarbital solution (solvent is normal saline), then make a small incision about 5mm long on the back of the mouse's neck, insert the NaCl capsule pump, and finally suture the back of the mouse. The NaCl capsule pump injects 0.9% NaCl through an osmotic micropump for 4 weeks. After the mouse wakes up, the control mouse is prepared.

[0038] The control group consisted of 10 control mice.

[0039] 4. Non-invasive blood pressure measurement in mice (tail-cuff pressure method) After completing steps 2 and 3, mice in the AngII group and control group were taken, and the systolic blood pressure (SBP) and diastolic blood pressure (DBP) were measured using the BP-300A fully automatic non-invasive BP measurement system (Chengdu Taimeng Software Co., Ltd.). Each mouse was measured three times with an interval of more than 30 seconds. The average of the three measurements was taken as the measurement result, that is, the measurement result of week 0. The measurement was carried out every week for 4 consecutive weeks.

[0040] Due to the influence of environmental factors, the blood pressure of mice generally fluctuates within 10 mmHg. Therefore, it is necessary to measure under the same objective conditions as much as possible to reduce measurement errors and ensure the stability of measurement results.

[0041] The measurement results are shown in Figure 1 (A is the statistical result of systolic blood pressure, B is the statistical result of diastolic blood pressure, # represents P Value <0.01). The results showed that the blood pressure of mice in the AngII group gradually increased, and the systolic blood pressure reached 154.6±2.2 mmHg and the diastolic blood pressure reached 132.8±2.5 mmHg at the fourth week, which were significantly higher than the systolic blood pressure (108.4±3.4 mmHg) and diastolic blood pressure (94.85±3.9 mmHg) of the control group at the same time point, respectively. This proves that AngII can induce an increase in blood pressure in mice and prepare a mouse hypertension model.

[0042] 5. After completing steps 2 and 3, the mice were killed and the aortic and colonic tissues of the mice were observed. The aortic tissue was subjected to Masson staining (for specific steps, please refer to the following literature: Liu Yang, Chen Guihai. Effects and mechanisms of Hanjing Decoction on phenotypic transformation of vascular smooth muscle cells, inflammation, and oxidative stress in rats with cold-coagulation type hypertension [J]. Journal of Jinan University (Natural Science and Medicine Edition), 2023, 44 (04): 347-357.). The colon tissue was subjected to PAS staining (for specific steps, please refer to the following literature: Guo Weiling. Effect and mechanism of inosine-producing intestinal bacteria on the relief of colitis [D]. Jiangnan University, 2023.).

[0043] Results Figure 2 (A is the result of Masson staining of aorta and PAS staining of colon, B is the statistical result, the data in the figure are P value, P The results showed that the aorta and colon tissues of the mice in the AngII group showed signs of hypertrophy, while the aorta and colon tissues of the mice in the control group showed no significant changes; compared with the control group mice, the aorta of the mice in the AngII group was significantly thickened, and the proportion of aortic fibrosis area was significantly increased; compared with the control group mice, the thickness of the colon muscle layer of the mice in the AngII group was significantly increased, and the proportion of goblet cells was significantly reduced.

[0044] These results indicate that AngII-induced aortic and colon tissue remodeling in mice is affected by elevated blood pressure and altered intestinal permeability.

[0045] Example 2: Application of berberine (Bbr) in reducing Ang II-induced hypertension Berberine is a product of Shanghai Yuanye Biotechnology Co., Ltd., with a catalog number of B21379. The structural formula of berberine is shown in Formula I; .

[0047] 1. Randomly select 12 AngII hypertensive mouse models prepared in step 2 of Example 1 and divide them into 2 groups (6 mice in each group), namely AngII group and AngII+Bbr group. Systolic blood pressure (SBP) and diastolic blood pressure (DBP) were measured using BP-300A fully automatic non-invasive BP measurement system (Chengdu Taimeng Software Co., Ltd.). Each mouse was measured three times and the interval time was required to be more than 30 seconds. The average of the three measurements was taken as the measurement result, i.e., the blood pressure result of week 0 (0W).

[0048] 2. After completing step 1, each group is processed as follows: AngII group: Each mouse was gavaged with 0.5 ml of deionized water once a day for 28 consecutive days.

[0049] AngII+Bbr group: Each mouse was gavaged with 0.5 ml berberine solution (solvent: deionized water) once a day for 28 consecutive days. The dose of berberine used for each mouse was 100 mg / kg / d.

[0050] During step 2, the systolic blood pressure (SBP) and diastolic blood pressure (DBP) were measured weekly using the BP-300A fully automatic non-invasive BP measurement system (Chengdu Taimeng Software Co., Ltd.). Each mouse was measured three times with an interval of more than 30 seconds. The average of the three measurements was taken as the measurement result, that is, the blood pressure results of the first week (1W), the second week (2W), the third week (3W), and the fourth week (4W) were obtained.

[0051] Test results are shown in Figure 3 (#express P Value < 0.01, the difference is extremely significant; * indicates P The results showed that the blood pressure (both systolic and diastolic) of mice in the AngII+Bbr group were significantly lower than those in the AngII group. This proves that berberine can reduce AngII-induced hypertension and exert its antihypertensive effect.

[0052] 3. After completing step 2, the two groups of mice were killed and the aortic tissue and colon tissue of the mice were taken. The aortic tissue was subjected to Masson staining. The colon tissue was subjected to PAS staining. PAS staining was used to observe the proximal colon single-layer columnar epithelium, colon muscle layer thickness and goblet cell number. The specific staining method is shown in step 5 in Example 1.

[0053] The results of PAS staining and the statistical results of the thickness of the mouse colonic muscle layer and the proportion of goblet cells are shown in Figure 4 (The data in the figure are P value, P The value less than 0.05 indicated significant difference). The results of Masson staining and the statistical results of mouse aortic thickness and aortic fibrosis ratio are shown in Figure 5 (The data in the figure are P value, P Values ​​less than 0.05 indicate significant differences).

[0054] The results showed that compared with the AngII group, the thickness of the colonic muscular layer of mice in the AngII+Bbr group was significantly reduced (i.e., berberine significantly inhibited the thickening of the colonic muscular layer), and the proportion of goblet cells was significantly increased. Compared with the AngII group, the aortic thickness of mice in the AngII+Bbr group was significantly reduced (i.e., berberine significantly reduced aortic thickness), and the proportion of aortic fibrosis area was significantly reduced (i.e., berberine significantly reduced aortic fibrosis).

[0055] It can be seen that berberine can improve the changes in intestinal pathology caused by AngII and has a positive effect on vascular remodeling. Berberine lowers blood pressure by remodeling the cardiovascular system.

[0056] Example 3: Berberine increases the abundance of Akkermansia muciniphila (Akk bacteria) Berberine regulates the intestinal microbial community by inducing apoptosis of harmful microorganisms and promoting the growth of beneficial bacteria (such as Bifidobacterium juvenile and Lactobacillus acidophilus). This therapeutic effect of berberine is similar to that of probiotics. In order to study whether berberine can lower blood pressure by changing the intestinal flora and increasing the abundance of Akkermansia muciniphila (Akk bacteria), the inventors collected feces from the two groups of mice after completing step 2 of Example 2 and extracted genomic DNA, and then performed metagenomic sequencing to obtain metagenomic data.

[0057] The metagenetic data of the two groups of mice were analyzed and the results were as follows: Compared with the AngII group, mice in the AngII+Bbr group expressed fewer genes, and the Chao1 index, Observed_species index, and Shannon index in the AngII+Bbr group were significantly reduced (see Figure 6 The data in the figure are P value, P <0.05 indicates significant difference). It can be seen that berberine affects the α diversity of the intestinal flora in the AngII hypertensive mouse model, affecting its diversity and richness.

[0058] The PCoA results based on Bray-Curtis distance are shown in Figure 7 The results showed that the intestinal microbiota of the two groups of mice were clearly separated between the AngII group and the AngII+Bbr group.

[0059] The results of LEFSe analysis are shown in Figure 8 (The figure shows the dominant bacterial species between the two groups. LDA>2 indicates a significant difference. In addition, the data in the bar graph are P value, P <0.05 indicated significant differences). The results showed that the level of Akkermansia muciniphila (Akk bacteria) in the AngII+Bbr group was significantly increased compared with that in the AngII group.

[0060] These results suggest that berberine lowers blood pressure by increasing the abundance of Akkermansia muciniphila in the intestinal flora.

[0061] The present invention has been described in detail above. For those skilled in the art, without departing from the purpose and scope of the present invention, and without the need to carry out unnecessary experimental conditions, the present invention can be implemented in a wide range under equivalent parameters, concentrations and conditions. Although the present invention provides specific embodiments, it should be understood that the present invention can be further improved. In a word, according to the principles of the present invention, the application is intended to include any changes, uses or improvements to the present invention, including departure from the disclosed scope in the application, and changes made with conventional techniques known in the art.

Claims

1. Application of berberine, which is A1) or A2) or A3) or A4): A1) Preparation of medicaments for the treatment of hypertension; A2) Preparation of health products for improving blood pressure levels; A3) Preparation of drugs for lowering blood pressure; A4) Regulate intestinal flora; The structural formula of berberine is shown in Formula I; 。 2. The use according to claim 1, characterized in that: The modulation of the intestinal flora is manifested as an increase in the abundance of Akkermansia muciniphila.

3. Use of the composition containing berberine as claimed in claim 1, which is A1) or A2) or A3) or A4): A1) Preparation of medicaments for the treatment of hypertension; A2) Preparation of health products for improving blood pressure levels; A3) Preparation of drugs for lowering blood pressure; A4) Regulate intestinal flora.

4. The use according to claim 3, characterized in that: The modulation of the intestinal flora is manifested as an increase in the abundance of Akkermansia muciniphila.

5. The use according to claim 3, characterized in that: The composition containing berberine as claimed in claim 1 is a health product composition, a pharmaceutical composition or a probiotic composition.

6. The use according to claim 5, characterized in that: The composition containing berberine as claimed in claim 1 further comprises a second active ingredient; the second active ingredient is a drug different from berberine as claimed in claim 1; The composition containing berberine as claimed in claim 1 further comprises one or more of telmisartan, captopril, valsartan and amlodipine.

7. The use according to any one of claims 1 to 6, characterized in that: The hypertension is renal hypertension, spontaneous hypertension, metabolic hypertension caused by combined dietary factors or isolated systolic hypertension.

8. The use according to any one of claims 1 to 7, characterized in that: The drug is a medicine, and its dosage form is tablets, capsules, granules, pills, ointments, solutions, suspensions, emulsions, creams, sprays, drops, patches or tube feeding preparations; and / or, The health care products are health care foods, and the types thereof are dairy products or fermented cereal foods.

9. The use according to any one of claims 1 to 8, characterized in that: The medicine or the health product contains an effective amount of berberine as claimed in claim 1.

10. The use according to any one of claims 1 to 9, characterized in that: The subject of the medicine or the health product is a mammal; Preferably, the mammal is a human or a mouse.

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