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Study on effect of Baisha Green tea extract on blood pressure in treating

spontaneous hypertension rats

Dao-rui Yu

1

, Li-ping Ji

2

, Yi-hong He

3

, Jing Zhou

3

, Chang-yan Wu

3

, Qi-bing Liu

4

1Research Center of Basic Medicine, Hainan Medical University, School of Basic and Life Science, Haikou 571199, China 2Department of Pharmacology, Xian Jiaotong University Health Science Center, Xian 710049, China

3 School of Clinical Medicine, Hainan Medical University, Haikou 571199, China 4Department of Pharmacology, Hainan Medical University, Haikou 571199, China

4Department of Pharmacology, Hainan Medical University, Haikou 571199, China

Journal of Hainan Medical University

http://www.jhmuweb.net/

ARTICLE INFO ABSTRACT

Article history:

Received 5 Aug 2016

Received in revised form 17 Aug 2016 Accepted 12 Aug 2016

Available online 24 Aug 2016

Keywords:

Baisha Greentea extract Primary hypertensive rats Reducing blood pressure Angiotensin Ⅱ

Corresponding author: Qi-Bin Liu, Department of Pharmacology, Hainan Medical University, Haikou 571199, China.

Tel: 0898-31350701 E-mail: yxybing@163.com

Foundation project: It is supported by grants of Hainan provincial project of modernization of traditional Chinese medicine (ZY201426), by grants from the Key Science and Technology Program of Hainan Province (ZDXM2014070).

1. Introduction

The incidence of people "three high" disease gradually increased when modern living standards gradually improved. essential hypertension as one of the diseases in "three high", what mainly for repeated systemic arterial blood pressure continues to rise and spasticity that the development of the course, often involving the other organs such as the heart, brain, kidney and other diseases

and can make it happen, so people's health and lives have been serious harm . Since the latest relevant information on the survey, high blood pressure will be in developing countries, rising 39.3% rate; The prevention and treatment of hypertension is imminent that mortality rate will reach 79% of the total deaths until 2020[1]. Patients are mostly chemotherapy hypertension currently mainly due to the adverse effects of chemical drugs, limiting its widespread clinical use and long-term. a major issue of research that is Developing high efficiency and low side effects of antihypertensive drugs. With the social development and progress, tea has become a necessity of people's lives, but also the social spiritual civilization jewel , thus tea culture occupied an important part in chinese traditional culture. As the tea contains a lot of medicinal value, so there are some prospects for the development of hypertension prevention, treatment. Many of our research buck tea found that it Objective: To explore the effect and Mechanisms of Baisha Greentea extract on blood pressure in treating spontaneous hypertension rats. Methods:The 60 primary hypertensive rats (SHR) were randomly divided into hypertension of control group, amlodipine besylate group and the high, medium and low dose group of green tea extract of five groups, with twelve rats in each group. All treatments were given through oral, and the control group with distilled water. Each measurement of rat blood pressure was detected in the morning every week. After 7 weeks treatment, blood was collected from the rat abdominal aorta for determination of serum nitric oxide (NO) and superoxide dismutase (SOD) activity and level of plasma angiotensin II. At the same time, heart tissue was embedded in 10% neutral buffered formaldehyde fixation, paraffin and sectioned, And then observe the changes of histopathological after HE staining. Results: Compared with the SHR control group, Systolic and diastolic blood pressure of essential hypertension in high dose group and middle dose group and Amlodipine group had significantly reduced. At the same time, the NO and SOD contents of Baisha Greentea

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relates to the main component-based granules in most current patent. For example rhizome buck tea, etc[2]. study Meicha of HanLing Yao and "Jiuwei buck tea" of patent etc[3]. With antihypertensive effect as mainly Ile and a-aminobutyric acid (GABA) tea etc[4]. But the real tea extract under the antihypertensive effects was less reported. Baisha Greentea is Camellia camellia rue mesh plant, the main active ingredient is polyphenols. Studies have shown that tea polyphenols with a variety of pharmacological activities, such as the prevention of cardiovascular disease, lower blood pressure, blood sugar and blood lipids[5]. In recent years, studies have shown that daily consumption of green tea can lower blood pressure and improve endothelial function in humans, a variety of pharmacologically active prevention of cardiovascular disease and other effects[6]. These are just preliminary exploration, and subsequent reports less.Green tea reduce blood pressure levels and NO and SOD angiotensin Ⅱ (AngⅡ) in primary rat model of hypertension in this study that the period of green tea to provide some references for further research with content relationship.

2. Materials and Methods

2.1. Materials

2.1.1. Test sample preparation of aqueous extract of green tea

Go to Hainan green tea (Hainan State Farms white sand Tea Co., Ltd. Production batch: 2015040418) 500 g, respectively, plus 2 000 mL boiling water 3 times 0.5-1.0 h, until relatively clear TCM decoction. Liquid vacuum filtration, concentrated under reduced pressure to a viscous (50 ℃), vacuum dried and pulverized to obtain extract powder. With 0.9% NaCl solution was formulated

as 200 mg•kg-1, 100 mg•kg-1 drug solution and 50 mg•kg-1

when administered. Doses of green tea extract administration group with reference to the value of commercially available green tea in the human dose for conversion, the whole test dose of 2 times the amount of increment[7]. Amlodipine tablets available in Pharmaceutical of hu yuan of Shanghai Co., Ltd. Production batch: 20150092, Configured to liquid when administered concentration

0.5 mg•kg-1.

2.1.2. Animal Essential hypertension (SHR) rats

60,male, Cleaning stage, body weight: (220 ± 10) g, Beijing Vital

River Laboratory Animal Technology Co., Ltd. Animal license number: SCXK(Jing)2006-0009.

2.1.3 Reagents

0.9% Sodium Chloride Injection, Hainan Hua Tuo Tianya Pharmaceutical Co., Ltd. Production batch: 150818-1. Kits of Nitric

Oxide (NO), superoxide dismutase (SOD) and angiotensin Ⅱ,

Institute of Biological Engineering from Nanjing Jiancheng.

2.1.4 instrument

BP-100A automatic noninvasive blood pressure measurement system from chengdu Thai meng Co., Ltd. Microscope of Olympus from Olympus optical microscope of Japanese company. HH-6

digital temperature water bath from Union Institute of Jiangsu

Province Jintan Instruments. 722 UV spectrophotometer from

Beijing Chen Xi Yong Chuang Co., Ltd. Low-temperature low-speed centrifuge of Eppendorf from Germany Eppendorf China Ltd.

2.2 Method

2.2.1 Animal groups and each group method of

administration

All purchases and acclimated to feeding one week after the blood pressure measurement, normal saline and other daily operations, and acclimated to its operation. The 60 primary hypertensive rats (SHR) were randomly divided into hypertension of control group, amlodipine besylate group and the high, medium and low dose group of green tea extract of five groups, with twelve rats in each group. Three dose groups of Green tea extract respectively will

gavage by rat at a concentration of 200 1,100 mg•kg-1•d-1,50 mg•kg-1•d-1.Yet amlodipine besylate group will gavage by rat at 0.5 mg•kg-1•d-1, Hypertension control group gavage a dose of 1

mL/100 g of 0.9% NaCl solution. It is weighing once a week During the administration, and as adjusted for changes in body weight dose by seven weeks. All rats were fed with normal diet and water ad libitum.

2.2.2 Blood pressure measurement

We are measurement of arterial pressure in rats of each group's awake that it isIndirect measurement method by BP-100A automatic noninvasive blood pressure measurement system. Let rats adapt to the environment Before the official measurement week training load of 1 to 2 times a day, and blood pressure was measured after two hours drench. Rats have been placed before the first load in advance

to set a good (37 ± 1) ℃ electric incubator after preheating 15 min. Then the rats were placed invasive measurement of blood pressure measurement system in the cage, until stable blood pressure curve measured systolic and diastolic rat tail artery, repeated measured three times and averaged.

2.2.3 Serum samples were collected and measured

The animals were fasted for 12 h, but can not help but water and weighed after the last administration in Experimental administration of 7 weeks. Abdominal aortic blood samples were taken at 5 mL centrifuge tube and centrifuged to prepare serum with 3% sodium pentobarbital anesthesia. Take the prepared serum, respectively nitrate reductase and xanthine oxidase serum NO and SOD content.

2.2.4 Determination of angiotensin

(Ang

) content

The animals were taken blood to the abdominal aorta of the previously prepared containing 20 g/L sodium ethylenediaminetetraacetate (EDTA) in cryovials, then freeze the blood in the tube moved into 1.5 mL centrifuge tubes and Into the low-temperature low-speed Eppendorf centrifuged 15 min, then

using competitive enzyme-linked immunosorbent assay AngⅡ

content.

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Take heart tissue with saline wash the blood, while placed in 10% neutral formalin fixed that collected of rat abdominal aortic blood when the rat death. it is HE staining pathological and light microscope and tissue diagnosis by pathological changes of the pathology specialist.

2.3. Statistical method

Experimental data applications Excel software for processing what

are used mean ± standard deviation (Mean ± SD) represent, and

using statistical software SPSS 20.0 t test between sets of data, P value calculated, P<0.05 was statistically significant.

3. Result

3.1. Effect of green tea extract on blood pressure

Blood pressure (systolic and diastolic) in each group (SHR) rats substantially at the same level, there is no significant difference before the experiment, The treatment group had certain level of decline in blood pressure after 7 weeks except blood pressure of control group, and there are differences in hypertension control group, with statistical significance(P <0.01) by Table 1 & 2.

3.2. Effect of green tea extract on the weight of the heart and

kidneys of SHR

Each rat heart and kidney mass administration group showed reduced that compared with control group after Seven weeks, And green tea extract high dose group and the amlodipine group was significantly reduced tendency in heart and kidney mass, Having significant (Respectively, P<0.01 and P<0.05). Green tea extract low-dose group and the control group with hypertension was decreasing trend, But there are small differences, not statistically

significant (P>0.05). Administration of green tea and amlodipine certain period of time to be able to on ventricular remodeling in hypertensive rats have a certain role, Table 3.

Table 3.

Effect of green tea extract on the quality of the heart and kidneys of rats (n=12).

Groups Dose(mg/kg) Heart Weight (g) Kidney weights (g)

Control 1.52±0.39 2.33±0.22

Amlodipine 0.5 1.27±0.04* 2.03±0.03**

Green tea extract 50 1.38±0.34 2.24±0.43

Green tea extract 100 1.29±0.05* 2.07±0.16*

Green tea extract 200 1.25±0.14** 2.02±0.07**

Note: Compared with the control group with hypertension *P<0.05, **P<0.01.

3.3. Effect of green tea extract on serum nitric oxide (NO)

and superoxide dismutase (SOD) content

Each treatment group hypertensive rat serum nitric oxide (NO) and superoxide dismutase (SOD) and the content of high blood pressure compared to the control group, were significantly higher, and the high dose group the effect is significant, with statistical significance after 7 weeks (Respectively, P<0.01 and P<0.05). Amlodipine group and low dose group and the control group, while having significantly increased, but the difference between small, not statistically significant(P>0.05), Table 4.

Table 4.

Green tea extract changes in serum concentrations of NO and SOD in rats administered after seven weeks (n=12).

Groups Dose(mg/kg) NO(µmol/mL) SOD(U/mL)

Control 23.82±5.18 58.52±3.39

Amlodipine 0.5 29.04±7.23 61.42±6.44

Green tea extract 50 27.55±6.56 62.56±8.62

Green tea extract 100 33.08±4.73* 65.36±7.15*

Green tea extract 200 37.81±4.97** 81.38±7.82**

Note: Compared with the control group with hypertension*P<0.05, **P<0.01.

Table 1.

Green tea extract on systolic blood pressure (SBP) impact before and after administration (n=12).

Groups mg/kgDose Pre-dose blood pressure(mmHg) After administration of blood pressure (mmHg)

the first week the second week The third week the fourth week the fifth week the sixth Week the seventh week Control 206.47 ± 11.79 202.39 ± 9.21 203.61 ± 8.06 204.77 ± 10.18 202.39 ± 7.14 202.63 ± 9.65 202.63 ± 9.65 201.38 ± 5.87

Amlodipine 0.5 205.40 ± 7.95 197.07 ± 4.61 192.47 ± 7.66 186.71 ± 7.67* 182.32 ± 8.85* 174.64 ± 6.54**171.59 ± 4.23** 168.86 ± 5.95**

Green tea extract 50 206.59 ± 8.12 199.75 ± 9.52 196.38 ± 8.73 188.41 ± 9.82* 185.49 ± 8.79* 179.68 ± 7.16* 176.46 ± 9.23** 173.34 ± 8.47**

Green tea extract 100 204.32 ± 12.06 199.86 ± 8.26 194.35 ± 7.93 187.68 ± 6.02* 183.79 ± 5.11* 176.82 ± 8.24**173.27 ± 6.87** 169.92 ± 8.91**

Green tea extract 200 204.68 ± 11.56 196.22 ± 9.52 193.34 ± 6.47 184.48 ± 6.53* 181.88 ± 6.36* 173.56 ± 7.66**170.78 ± 6.94** 166.18 ± 8.79**

Note: Compared with the control group with hypertension*P<0.05, ** P<0.01

Table 2.

Green tea extract on systolic blood pressure (DBP) impact before and after administration (n=12).

Groups mg/kgDose Pre-dose blood pressure (mmHg)After administration of blood pressure (mmHg)

the first week the second week The third week the fourth week the fifth week the sixth Week the seventh week Control 149.68 ± 14.09 148.29 ± 13.32 150.76 ± 11.96 147.55 ± 10.35 147.67 ± 9.54 146.36 ± 9.75 143.28 ± 8.29 145.24 ± 8.67

Amlodipine 0.5 148.57 ± 10.41 143.26 ± 2.71 134.56 ± 8.26* 133.85 ± 2.67* 133.56 ± 7.46* 131.52 ± 3.93**130.37 ± 6.45**127.64 ± 2.62**

Green tea extract 50 150.83 ± 10.54 141.48 ± 6.93 139.38 ± 9.37 138.63 ± 9.62 136.73 ± 5.42* 135.68 ± 5.39* 132.57 ± 3.67**130.17 ± 6.58**

Green tea extract 100 144.72 ± 11.82 140.72 ± 8.26 135.46 ± 3.76* 136.56 ± 7.43* 135.79 ± 8.03* 133.82 ± 8.24* 131.49 ± 7.98**128.36 ± 6.73**

Green tea extract 200 147.38 ± 10.34 142.18 ± 3.52 134.98 ± 6.83* 131.48 ± 6.53**131.37 ± 4.17** 130.76 ± 5.82**129.78 ± 4.72**126.54 ± 5.82**

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3.4. Effect of green tea extract on plasma angiotensin

(Ang

) content

The treatment group significant difference except angiotensin (AngⅡ) green tea extract low-dose group were content with statistical significance after 7 weeks when compared with hypertension control group (Respectively, P<0.01 and P<0.05). But also the low dose group than the control group, but no significant difference (P>0.05). AngⅡ content with the green tea extract reduced the increase was dose, The experiments showed that after a period of drinking green tea can reduce the amount of angiotensin Ⅱ.

Table 5.

Effect of green tea extract on plasma AngⅡ content of SHR (n=12). Groups Dose (mg/kg) AngⅡ(mg/L)

Control 1.46±0.05

Amlodipine 0.5 1.27±0.03**

Green tea extract 50 1.36±0.06

Green tea extract 100 1.30±0.03*

Green tea extract 200 1.20±0.02**

Note: Compared with the control group with hypertension*P<0.05, **P<0.01.

3.5. Morphological changes of heart

Shows from Figure 1, myocardial HE staining light microscopy, high blood pressure in the control group most visible disorder cardiac fibroblasts, and occasionally see the muscle fibers break, thicker, cardiomyocyte hypertrophy. Green tea high dose group and positive drug amlodipine myocardial fibers arranged in neat, uniform dyeing, stripes clear, uniform distribution of myocardial cells, no significant increase in the cells. Green tea, the low-dose group is insignificant, a small part of the visible region of thicker muscle fibers, but also disorganized, occasionally there is breakage.

Control Amlodipine

Green tea extract (200mg/kg) Green tea extract (100mg/kg) Green tea extract (50mg/kg)

Figure 1.Effect of green tea extract on SHR Myocardial cell morphology.

4. Discussion

The incidence of cardiovascular disease more quickly with the accelerating pace of people's lives[8]. The primary hypertension is a

common clinical cardiovascular disease, a disease that mainly long course, so as to change the patient's heart, great vessels, kidneys normal physiological function. The incidence of cerebrovascular disease is the ultimate goal of treatment why some reduce high blood pressure caused by the heart[9]. In this study, the model of essential hypertension that feature of internationally recognized human essential hypertension of the closest[10]. Examine changes in Hainan green tea extract on blood pressure of essential hypertension in rats., Each treatment group was to make blood pressure drop after gavage to Hainan green tea extract seven weeks from the results, and Green tea and high blood pressure and the effect of two doses of positive drug amlodipine comparable

Left ventricular hypertrophy quality and glomerular and tubular atrophy and compensatory hypertrophy of a chronic adaptive mechanism of blood pressure affects the heart sustained high load under pathological conditions of hypertension[11]. Therefore, cardiac and renal hypertension is one of the most vulnerable target organ involvement. The results show, each treatment group and positive drug amlodipine group ultimately reduce the quality of the heart and kidneys except the control group of green tea extract. Hainan Baisha green tea and amlodipine have a protective role of target organs to some extent in this study.

Blood pressure is the main regulator of NO synthesis of endothelial nitric oxide synthase and a substance release of vasoactive, In a state of hypertension may be relaxation of vascular smooth muscle cells and blood pressure and inhibiting smooth muscle cell proliferation plays an important role in cardiac and vascular remodeling[12]. Because nitric oxide synthase in blood pressure caused by decreased serum NO degradation is weak, so the content is also reduced accordingly[13]. Superoxide dismutase (SOD) is the body's natural antioxidant enzymes and free radicals can maintain homeostasis that the body's cells to avoid damage. SOD significantly reduced the body under pathological conditions of hypertension[14]. The experimental results show that from each treatment group were serum NO and SOD in rats increased, Wherein the green tea extract high dose group compared with the control of hypertension, it was statistically significant (P<0.01 and P<0.05). The positive antihypertensive drug amlodipine group and low dose group, while also able to lower blood pressure, but the effect is not significant (P>0.05).

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reduce the generation of AngⅡ, reduce blood pressure, weakened left ventricle reconstruction play a role. Green tea extract each treatment group and positive drug amlodipine group decreased AngⅡ content decreased blood pressure in SHR SHR, where green tea high dose group and amlodipine quite effective from this study. This Show Hainan green tea with amlodipine mean inhibition of the angiotensin-converting enzyme activity, so their role in lowering blood pressure.

Essential hypertension lead to cardiac hypertrophy, heart failure and thus, arrhythmias, myocardial ischemia or stroke and other diseases, a great threat to their life and health[16]. The results suggest that, along with blood pressure, green tea extract administration group were also significantly decreased heart weight. Morphology cardiac fibroblasts each treatment group were better than the control group with hypertension. Green tea with high-dose group and positive drug amlodipine group as well on both cardiac cell morphology is insignificant. Long-term consumption of green tea can be seen on hypertension myocardial fibrosis and some improvement. Its cardiac structure has a protective effect, reducing the extent of myocardial injury, reversing ventricular remodeling[17].

Green tea as one of the most local characteristics of Hainan in "Hainan specialty". Green tea has become currently a hot research at home and abroad.It was discussed in terms of lowering blood pressure in this study, I believe that with further research, green tea will better benefit of mankind that Effective and safe drugs for heart and cerebrovascular disease foundation.

References

[1] Fengquan Su, Fenlu Guo. Mountain green tea capsules combined with Western elderly hypertensive efficacy. Treatment Modern Med 2015;

24(11): 1191-1193.

[2] Hong Jiang. Clinical Observation on 156 cases from mild to moderate hypertension. Antihypertensive Ther Pecial Health Tea 2014; (3): 370-370. [3] Guoxing He. Buck Huxin chrysanthemum tea. Family Med 2014; (9):

53-53.

[4] Yan She, Guoming Zhang, Xiaoyuan Lin. Effect of Ilex atherosclerotic vascular endothelial function in cirrhotic rats. Chin Med 2014; 23(2):

220-222.

[5] Jun Li, Qin Xin, Yan Yang. Advances in pharmacological effects of green tea polyphenols. China Trop Med 2014; 14(1): 128-130.

[6] Yin Zhuo Rui, Jiayi Wang, Shuai Wang. Green mountain development.

Liaoning Coll Tradit Chin Med 2015; 17(12): 81-84.

[7] Wei Wei, Xi Mei Wu, Jian Li. Pharmacological experiments methodology. Beijing: People's Medical Publishing House; 2010, p. 69.

[8] Chang Pan, Feng Xu, Qiuhuan Yuan. Research and cardiovascular and cerebrovascular diseases progress aldehyde dehydrogenase. Chin J

Cardiol 2016; (2016): 175-178.

[9] Weiwei Chen, Hui Sui, Liyuan Ma. Prevalence of cardiovascular disease prevention and control in China and progress. Heart Dis Prevent 2016; 2: 001.

[10] Kato T, Mizuguchi N, Ito A. Blood pressure, renal biochemical parameters and histopathology in an original rat model of essential hypertension (SHRSP/Kpo strain). Biomed Res 2015; 36(3): 169-177. [11] Gao T, Zhu Z Y, Zhou X. Chrysanthemum morifolium extract improves

hypertension-induced cardiac hypertrophy in rats by reduction of blood pressure and inhibition of myocardial hypoxia inducible factor-1alpha expression. Pharm Biol 2016; 1-6.

[12] Perros F, Ranchoux B, Izikki M. Nebivolol for improving endothelial dysfunction, pulmonary vascular remodeling, and right heart function in pulmonary hypertension. J Am Coll Cardiol 2015; 65(7): 668-680. [13] Ghosh S, Gupta M, Xu W. Phosphorylation inactivation of endothelial

nitric oxide synthesis in pulmonary arterial hypertension. Am J Physiol

Lung Cell Mol Physiol 2016; 310(11): L1199-L1205.

[14] Dornas WC, Silva M, Tavares R. Efficacy of the superoxide dismutase mimetic tempol in animal hypertension models: a meta-analysis. J

Hyperten 2015; 33(1): 14-23.

[15] Xiao L, Kirabo A, Wu J. Renal denervation prevents immune

cell activation and renal inflammation in angiotensin II–Induced

hypertension. Circulation Res 2015; 117(6): 547-557.

[16] Kuwabara Y, Horie T, Baba O. MicroRNA-451 exacerbates lipotoxicity in cardiac myocytes and high-fat diet-induced cardiac hypertrophy in mice through suppression of the LKB1/AMPK pathway. Circulation Res 2015; 116(2): 279-288.

Figure

Table 3.
Table 5.

References

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