Nghiên cứu độc tính, tác dụng điều trị nhồi máu não giai đoạn cấp trên thực nghiệm và lâm sàng của chế phẩm trúng phong hoàn tt tiếng anh

1 INTRODUCTION Of the disorders of the nervous system, stroke is the third leading causes of death, followed heart disease and cancer death About 20% of people is affected by stroke Approximately 80% of strokes are infarction The percentage of patients with infarction was between 80% and 85% of stroke in and infarction is a leading cause of disability In addition to methods of modern medicine, there are many kinds of drugs from traditional medicine which is very effective in treating acute stroke, for example, Angongniuhuang, Huatuo Zaizao Pills However, these imported drugs are very expensive while the domestic demand of them have been increased rapidly Thus, there exists a need for safe folk drugs which are more effective and easily available at low cost “Trung phong hoan” is a traditional medicine derived from the ancient Vietnamese remedy in the name of “Thần dược cứu mệnh”, in which the dried mandarin peel is added to “Trung phong hoan” was processed to granules so as to treat wind strike which is characterized by numbness in the mouth and eyes, drooling, limb spasms, aphasia, hemiplegia This drug is composed of four ingredients, namely, earthworm, black bean, sweet leaf and dried mandarin peel, which completely meets the above criteria In addition, although it is commonly used in folk, there has not been any complete and systematic study on yet Therefore, this study was conducted aiming to: Determine the acute toxicity and semi-chronic toxicity and evaluate the effectiveness of treatment for “Trung phong hoan” on experimental animals Evaluate the supporting effectiveness of the “Trung phong hoan” composition on patients with acute ischemic stroke PRACTICAL SIGNIFICANCE AND NEW DISTRIBUTION OF THE STUDY In the search for new ways to treat stroke, pre-clinical studies are required Thus, it is necessary to conduct pre-clinical researches on animals In this study, the modeling infarction in mice was successfully carried out by photochemical method which combines between Krypton laser 568nm, a laser power of 6mW and Rose Bengal The laser dose was 20mg/kg body weight Although modeling infarction using photochemical method have been used for a long time and applied in rats, it maybe the first successful rat model in Vietnam Moreover, we changed some steps during modeling, in particular, the way of introduce Rose Bengal into the circulatory system Especially, Krypton laser system was set for this study only to simplify the procedure, shorten the time of manipulation and increase the success rate of stroke modeling Research on therapeutic effects of traditional medicines for acute ischemic stroke is a new direction According to current statistics on study in Vietnam, this is the second research on acute ischemic stroke treatment followed by the research namely, “Angongniuhuang” It is always needed to continue to seek effective and safe herbal medicines, and as such it is attractive to scientists from all over the world Clinical trials and experiments of this research were performed in a rigorously systematic way The remedy was processed to pills composed of available four ingredients used to treat acute ischemic stroke Studying a remedy in order to apply it in ischemic stroke treatment will contribute to illuminate the effects of treatment of traditional medicine, which has practical and scientific significance TABLE OF CONTENTS Introduction CHAPTER 1: Literature Review 37 pages CHAPTER 2: Methodology 18 pages CHAPTER 3: Results 46 pages CHAPTER 4: Discussion 31 pages Conclusion This study includes 49 tables, 16 charts, 14 figure, 01 drawing, 14 appendix pages and 141 references (Vietnamese 70, English 36 and Chinese 35) CHAPTER LITERATURE REVIEW 1.1 Ischemic stroke * Cerebral infarction Cerebral infarction is caused by the sudden loss of blood circulation because of a sudden lack of blood flow to whole or a part of cerebral arteries or carotid arteries or a vein of the brain in rare Clinically Clinically cerebral infarction suddenly or acutely exhibits focal neurologic signs, in which hemiplegia is common 3 Supratentorial ischemic stroke includes cerebral hemisphere injury caused by lesions in anterior brain arteries, middle brain arteries, anterior choroidal arteries, posterior communicating arteries or branches of these arteries and a part of posterior cerebral artery or branches of posterior cerebral artery * Causes and pathogenic mechanisms of cerebral infarction: Cerebral infarction includes coagulation, embolus, and lacunar infarct * Drug treatment of acute ischemic stroke Stroke is a medical emergency and urgent treatment is essential The sooner a patient receives treatment for a stroke, the less percentage of death and disability is likely happen Ideally, patients with stroke should be treated in in the intensive and comprehensive care units According to the current trend in Vietnam, patients with stroke are often treated at Resuscitation & Emergency Departments or stroke center under the following principles: The ABC’s of first aid: Airway, Breathing, and Circulation Managing cerebral edema; maintaining water and electrolyte balance Specific treatment depending on stroke types 4.Symptomatic treatment, treatment complication, anticonvulsants; Controlling the blood pressure; not reducing the blood pressure too fast; keeping blood pressure at a high limitation level: from 160/90 mmHg to < 185/100 mmHg (recommended by the American Stroke Association) Nursing care for stroke (to prevent superinfection of the lungs and the urinary tract), ensuring adequate daily nutrient requirements of patients, early rehabilitation (prevention of ulcer, muscular atrophy, joint stiffness, and so on) Surgical treatment, methods for assessing intervention mechanical tool, remove blood clots from the circulation by recombinant plasminogen Early relapse prevention therapy 1.2 The concept of stroke in traditional medicine * “Wind Strike” The term ”stroke” has not been mentioned in traditional medicine However, based on the clinical manifestations of stroke, it is labeled a wind disease It is characterized by sudden collapse, insensibility, hemiplegia, numbness in the mouth and eyes, limb spasms and aphasia A stroke often occurs suddenly and quickly with complicate conditions * Clinical type according to traditional medicine According to traditional medicine books and the modern medical literature, wind strike is divided into two types: wind on acupuncture spots and wind on each of the five viscera + Wind on acupuncture spots: The symptoms is mild, patients are not in a coma, damp wind comes to meridian passage, collateral passage causing numbness in the mouth, hemiplegia, a tongue with a thin white coating + Wind on each of the five viscera: The symptoms is severe due to damp wind comes to zang organs, fu organs, representing in two levels as belows: ° Blocking syndrome: Stupor, hemiplegia, incomprehensible sounds, wheeze, numbness in the mouth and eyes, warm limb, floating pulse ° Escaping syndrome: coma carus, wheeze, sweating, cold limbs, fecal incontinence, pulled back tongue, moderate pulse * The study drug “Trung phong hoan” is a remedy derivered from the ancient Vietnamese remedy namely ”Thần dược cứu mệnh”, in which the dried mandarin peel is added to “Trung phong hoan” is mentioned in the book in the name of “Những thuốc vị thuốc Việt Nam”, written by Prof Do Tat Loi At present, it is used to treat ”wind strike” characterized by numbness in the mouth and eyes, drooling, limb spasms, aphasia, hemiplegia 20 patients with stroke in Military Institute of Traditional Medicine were also administered by “Trung phong hoan” in the fluid extract form Initial results showed that the remedy had positive effects in motor functional recovery using the NIHSS and MRC score CHAPTER SUBJECTS AND METHODS 2.1 Research materials - Study drug: “Trung phong hoan” of 8.5g/ pill was prepared at the Center for Eastern medicine research and production - Military Institute of Traditional Medicine, which reached the standard basis - Drugs for the foundation regimen: Cerebrolysin 10 ml; Gliatilin 1g Nootropyl 1g) Blood pressure medications are used by the recommendation of patients with cerebral infarction stroke’ attitude and there was no indication of thrombolytic agent (AHA/ASA 2005 Guideline Update) Aspirin 100mg and Cebrex 40mg were used as experimental controls 2.2 Research subjects 2.2.1 Animal testing Healthy Swiss mice (male an female, 25 ± 2g weight) were provided by National Institute Of Hygiene And Epidemiology Healthy Wistar mice (male an female, 150 – 180g weight) were provided by Vietnam Military Medical University 2.2.2.Patient group 120 inpatient regardless of gender with acute ischemic stroke, age 18 to 90, were treated at Military Institute of Traditional Medicine, 103 Military Medical Hospital, Military Medical University in the period from - 2016 to 6-2017 2.3 Research methods 2.3.1 Evaluate acute toxicity and semichronic toxicity Acute toxicity: The acute toxicity was studied base on guidance of the Ministry 27 of Health and OECD The study drug was administered single ordered dose progression via orally route of 10.4g/kg to 52g/kg by 10 mice per lot The health status as well as the number of dead mice was recorded at 72 hours interval The LD 50 was calculated by using the method of Litchfield- Wilcoxon The semichronic toxicity of “Trung phong hoan” in mice was studied base on guidance of the Ministry of Health and OECD, WHO Thirty mice were divided randomly into three groups of 10 The mice in group control had access to distilled water in oral dose of ml/kg/day The mice in group had access to “Trung phong hoan” in oral dose of 7.14g/kg (equivalent to human dose) The mice in group had access to granulated medicine in the name of “Trung phong hoan” in oral dose of 21.42g/kg/day (equivalent to times the human dose) All mice were administered distilled water or study drugs in weeks continuously, one time per day at AM Inclusion criteria for the study population before, after weeks and after weeks of administering medication were: general health status, weight, hematopoietic function, liver function, kidney function, liver histology and kidney histology 2.3.2 Evaluate the treatment effectiveness of “Trung phong hoan” for aute ischemic stroke in experimental animals a) Mice model of ischemic stroke - Establishment of an rat model of focal cerebral infarction using a photochemical method: To induce intravenous coagulation, a colored photosensitive was injected to the circulation Then, a light with a suitable wavelength was used to radiate the vein that is needed to block from the outside This leaded to photosensitive activation at the position of irradiation, which caused free radical formation The free radicals formed caused oxidation and injury vascular endothelial cells located at the position of irradiation and then activated platelet aggregation mechanism, which formed blood clots at this position b) Evaluate the treatment effectiveness of “Trung phong hoan” in mouse model 36 mice were divided randomly into four groups as follows: - Normal group (n = 6): The mice in this group were not treated to form blood clots, which leads to stroke Motor measurement results and motor decline points of this group were used to compare with the results of mice with stroke - Non-treatment stroke-induced group (n = 10): Movement of strokeinduced mice were evaluated at day 1, 3, 7, 14 after stroke inducing and they were examined the brain specimens at day 3, 7, 14 after stroke inducing - Stroke-induced Cebrex group (n = 10): Mice were administered orally Cebrex in dose of 38.4mg/kg weight/day after stroke inducing inducing Movement of stroke-induced mice were evaluated at day 1, 3, 7, 14 after stroke inducing and they were examined the brain specimens at day 3, 7, 14 after stroke inducing - Stroke-induced group treated with TPH (n = 10): Mice were administered orally TPH after stroke inducing inducing Movement of stroke-induced mice were evaluated at day 1, 3, 7, 14 after stroke inducing and they were examined the brain specimens at day 3, 7, 14 after stroke inducing * Evaluation criteria - Evaluate the level of decline in motor: Mice of post-stroke inducing groups were evaluated the level of decline in motor at day 1, 3, and 14 after surgery The group that was not induced stroke were evaluated once only - The total length of movement distance and the average velocity for minutes of groups were measured at day 1, 3, and 14 after stroke inducing 7 * TTC staining: In order to measure the infarct size and since then, the foci of infarction could be calculated, we used TTC staining method * Lesion volume measuring: Brain slices has different colors after TTC staining Red or pink staining indicates normal tissue, white indicates infarction * Study place and study time: Experimental testing and effectiveness evaluation of the remedy in the name of “Trung phong hoan” in mice model was performed at the Department of Pathophysiology – Military Medical University in the period from 25 November, 2015 to 30 December, 2015 2.3.2 Evaluate effects of the drug “Trung phong hoan” on patients with acute ischemic stroke a) Treatment method - Control group (Group 2): was treated at the A12 Department/ Military Institute of Traditional Medicine and Stroke Department A14, 103 Military Hospital using recommended regimen of WHO as follows: - Study group (Group 1): was treated as the same as the group with “Trung phong hoan” combination by oral administration, pills/ times daily Maintenance of patients’ life using the ABC rule: Ensure breathing and circulation are adequate to deliver oxygen to the body (heart rate adjustment, using blood pressure medications to reduce the blood pressure if it is higher than 180/100 mmHg and to increase if it is lower than 100/60 mmHg) Using Manitol to reduce cerebral edema; antiplatelet drug like Aspirin 100mg orally, once per day after the breakfast; neuroprotective and nutritional support drug of Cerebrolysin, 10ml per day (02 ampoules), that is injected into veins slowly in the morning/afternoon Gliatilin 1g in dose of 02 ampoules/day, that is injected into muscles in the morning/afternoon Nootropyl 1g in dose of 02 ampoules/day, that is injected into veins in the morning/afternoon * Adjusting water and electrolyte balance b) Follow up the patients’ conditions - Patients’ conditions were followed up and evaluated All study criteria were recored in the medical file at day and 15 - Side effects at patient clinic were monitored daily and continuously during the study time 8 c) Monitoring evaluation criteria Clinical monitoring and evaluation criteria Clinical criteria according to modern medicine: Participants were neurological examined, monitored and evaluated at day and day 15 Clinical criteria were assessed by Glasgow score, MRC scale, and NIHSS (Appendix 5, 6, 7) b Approaches to Criteria in traditional medicine: Participants were examined and evaluated by quadrangular Syndrome Differentiation is based on the eight diagnostic principles Diseases are differentiated via fu-organs Diseases are classified according to traditional medicine: catching meridian and collateral c Paraclinical monitoring criteria At day 01 and 15: Total blood tests were performed, which included erythrocyte, leukocyte, white blood cell, platelets counting, hemoglobin test; biochemical blood test (Glucose, total protein, AST, ALT, GGT, total bilirubin, Urea, Creatinine); a cranial CT scan Side effects monitoring at patients: - Clinic: fatigue, headache, dizziness, itching, and other manifestations on gastrointestinal, cardiovascular, respiratory, kidney, urinary organ were monitered - Paraclinic testal: Changes in hematopoiesis function, liver function and kidney function were monitered d Standards for evaluating treatment effects Results were assessed by Glasgow score, MRC scale and NIHSS score e Evaluating the results by characteristics of traditional medicine Participants were examined and evaluated for changes in symtoms (before and After stroke treatment of study groups) via quadrangular (inspection, listening and smelling examination, inquiry, and palpation), including tongue textile, tongue coating, and pulse rate 2.4 Data collection and analysis: After collection, data was analyzed using statistical methods medical methods with SPSS statistical software v.20.0 for Window 2.5 Ethical considerations in clinical research: After studying the the toxicity of the remedy and its effects on experimental animals, proofing its safety and positive effects, this study was approved to human volunteer testing by the Military Institute of Traditional Medicine Council for Science and Medical Ethics Committee of 103 Military Medical Hospital 9 CHAPTER RESULT 3.1 Experimental research results 3.1.1 Results from acute toxicity tests - Mice were administered “Trung phong hoan” in highest dose of 52g/kg weight, (50 folds of dose for human) but no lethal mouse was detected as well as no abnormal symtoms within 72 hours after administering and during days of monitoring were found 3.1.2 Semichronic toxicity 3.1.2.1 General health status During the experiment, rats in all lots still ate as usual They were in normal activity and agile They also had nice eyes, silky fur, and dry stool No abnormal characteristics in all lots were found during the study time 3.1.2.2 Changes in mice body weight: The body weight of mice in all groups (control group and study groups) was increased in comparison with before the test There was no statistically significant difference with respect to the degree of change in mice body weight between the control group and study groups (p>0.05) 3.1.2.3 Effects of “Trung phong hoan” on hematopoiesis function There was no statistically significant difference with respect to the quantity of erythrocytes, hemoglobin amount, the avarage volume of erythrocytes, the quantity of leukocytes and the quantity of platelets of both study group and study group in comparison with the control group before and after administering study drugs (p > 0.05) 3.1.2.4 Effects of TPH on liver function There was no statistically significant difference with respect to AST and ALT activity in mice blood of both study group and study group in comparison with the control group and comparison between before, after administering study drugs (p > 0.05) at the time of weeks and weeks 3.1.2.5 Effectiveness of “Trung phong hoan” on kidney function There was no statistically significant change with respect to urea and creatinine concentration in mice blood of both study group and study group in comparison with the control group (p > 0.05) 3.1.2.6 Histological results of weeks after administering Macroscopy: No abnormal change in macroscopic scale was found in heart, lungs, liver, spleen, pancreas, kidneys and digestive organ of all mice (both the control group and two study groups) 10 3.1.2.7 Microscopic morphology of liver and kidney Microscopic morphology of livers: + Control group: Normal liver image characterized by hepatic cells arranges radically in normal The central veins of liver and liver sinusoid were mildly congestive Portal tracts were thin, some areas were infiltrated mononuclear cells which consists of lymphocytes + Study group 1: Normal liver image with intact hepatitic cells, the central veins of liver and liver sinusoid were mildly congestive Portal tracts were thin and fibers were found scattered through them + Study group 2: Normal liver image- characterized by hepatic cells arranges radically The central veins of liver and liver sinusoid were mildly congestive Portal tracts were thin, lymphocytes and fibers were found scattered through them Microscopic morphology of kidneys: + Control group: Normal kidney image with enlargement of the glomerulus, blood vessels were congestive which were consisted of erythrocytes inside, the Bowman capsules were enlarged and contained clear liquid Blood vessels of the renal interstitium were mildly congestive + Study group 1: Normal kidney image with enlargement of the glomerulus, blood vessels were congestive which were consisted of erythrocytes inside, the Bowman capsules were enlarged and contained clear liquid Blood vessels of the renal interstitium were mildly congestive + Study group 2: Normal kidney image with enlargement of the glomerulus, the Bowman capsules were enlarged and contained clear liquid Tubule epithelial cells had normal morphology Blood vessels of the renal interstitium were mildly congestive 3.1.3 Evaluation results of positive effects of “Trung phong hoan” on experimental model 3.1.3.1 Results of performing experimental a animal model in stroke research Results shows that laser irradiation in two minutes did not cause loss of circulation of the middle brain arteries of all mice of study groups In groups wherein mice were radiated by laser in the period from to minutes, it is realized circulation of the middle brain arteries was lost was lost after the first laser irradiation After the second laser irradiation at the position that is below the position of the first laser irradiation, the circulation capacity of the middle brain arteries was lost in both periods of minutes and minutes 11 Fig 1: The middle brain arteries imaging of mice in coagulation process caused foci of infarction and the method of induction of ischemic stroke a The middle brain arteries imaging after cranial exposure b The middle brain arteries imaging while Krypton laser irradiation c Loss of circulation of the middle brain arteries imaging when blood clots appearing d The method of induction of ischemic stroke: after the first laser at the division position of the middle brain arteries, circulation of the middle brain arteries was lost, the second laser irradiation was performed at the position that is below the position of the first laser irradiation in the same period To ensure the exist of lesion, TTC staining was performed with mice brain days after laser irradiation Results show that in groups in which mice were radiated by laser within minutes, no lesions in brain was found in all slices While in both mice groups radiated with laser within minutes and minutes, injuries were found in all mice Fig 2: Mice brain imaging at day after inducing ischemic stroke 12 a Mice brain were irradiated by Krypton laser in minutes b Mice brain were irradiated by Krypton laser in minutes c Mice brain were irradiated by Krypton laser in minutes The volume of blood clots was measured and compared between two laser irradiation groups for minutes and minutes Results show that there was no statistically significant difference with respect to the volume of blood clots between groups Table 1: The percentage of success of the method of inducing stroke by photochemical method Ischemic stroke volume (mm3) The volume of blood clots was also was measured by method described in the study method part Results of the volume of blood clots (mm 3; Mean ± SEM) at day 3, day and 14 were 267.7 ± 42.7; 219.7 ± 4.4; and 205.7 ± 35.1, respectively (Fig 3.6) Thus, the volume of blood clots tended to reduce over time, but there was no statistically significant difference at the time study Fig 1: Ischemic stroke volume was measured at day 3, and 14 after stroke inducing 13 3.1.3.2 Evaluation results of positve effects of “Trung phong hoan” on the model Fig 2: Evaluation scores of decline in motor over time after stroke inducing of each group Results show that the scores of the group treated with TPH equivalent to the Cebrex group They were always statistically significantly lower in comparison with the stroke – non treatment group at day 1, 3, after stroke inducing, and there was no statistically significant difference in comparison with normal group at day 14 TPH has positive effects on reducing the level of movement disorders stroke-induced mice, the positive effects of this drug was equivalent to Cebrex b) Motor ability of mice was improved by Trung phong hoan We also evaluated the positve effects of Trung phong hoan by calculating distance traveled and speed of mice in groups after stroke inducing One day after stroke inducing, and there was no statistically significant reduction with respect to distance traveled in comparison with the normal group in all groups with stroke-inducing There was no statistically significant difference with respect to the distance traveled and speed of the TPH group and the Cebrex group in comparison with the non-treatment group at one day after stroke inducing It is increased in statistical difference in comparison with the non-treatment group at days and days after stroke inducing and still tended to be larger than this of the non-treatment group at 14 days after stroke inducing In conclusion, TPH had clearly positive effect on improving the mobility of stroke-inducing mice in comparison with the non-treatment group, and the effect was equivalent to the Cebrex group We also compared the distance traveled and speed of each group mice over time so as to evaluate effects of the drug of prolonged use Results show 14 that in the group treated with TPH, the distance traveled and speed of mice had statistically significant increase at 3, and 14 days in comparison with one day after stroke inducing Therefore, in case of prolonged use, TPH had positive effects on improving the mobility at least until day 14 ngày after stroke inducing Fig 3: Distance traveled (a) and speed (b) of each group over time after stroke inducing c) “Trung phong hoan” drug had positive effects on reducing the volume stroke mass Fig 4: Ischemic stroke volume over time From the results of assessing the ischemic stroke volume, the Cebrex had positive effects fast and reducing the volume of stroke at day after administering (in comparison with the non-treatment group) However, these effects did not improve significantly over time While the group treated with TPH were also improved as the same as this treated with Cebrex, characterized by the clearly reduced volume of blood clots at day after drug administration (in comparison with the non-treatment group) The volumes of blood clots then increased over time and were highest at day although the volumes of blood clots were still statistically smaller in comparison with the non-treatment group Then 15 the volumes of of blood clots reduced rapidly and reached the smallest value (in comparison with the non-treatment group and The Cebrex group) at day 14 after stroke inducing 3.2 CLINICAL EVALUATION OF RESULTS EFFECTS OF THE DRUG 3.2.1 Clinical characteristics of patient groups - Characteristics of age: The average of patients with ischemic stroke was mainly 50-70 In which the highest percentage of 38/120 (31.67%) was mainly 61-70 there was no statistically significant difference with respect to age (p > 0.05) between the two group patients - Characteristics of genders: The percentage of men was higher than this of women with the percentage of male/female was 1.5/1 (in “Trung phong hoan” group was 1.59/1 and in the control group was 1.38/1), there was no statistically significant difference with respect to the percentage of male/female hai Study groups (p > 0.05) - Characteristics of generation time of the two groups: The generation time of stroke mainly from the midnight to dawn in the TPH treatment group was 46/70 patients (65.7%), in the control group was 29/50 (58%), in both groups was 75/120 (62.50%) and the difference was not statistically significant between two groups (p > 0.05) - From the beginning of stroke till using study drug: Most hospitalized patients with ischemic stroke in this study were acute, which was within - days of 70.84%, and < 24h of 29.14% - Risk factors of the patient groups: The percentage of patients with hypertension was highest, of 80.8% The next was patients with lipid metabolism disorders of 75.8% There was a similarity between the two groups The third risk fator was other diseases, such as cardiovascular and asthma with 12.5% Other risk factors with less frequency often mentioned were weather, alcoholism and smoking There was no statistically significant difference with respect to the percentage of risk factors in the two patient groups - Characteristics of localised nerve damage of patients before stroke treatment: The main feature of localised nerve damage often seen in ischemic stroke in both groups were hemiplegia, which was highest percentage of 100% The percentage of patients with central facial palsy was of 85.8%, language disorders was of 74.2%, sensory disorders was of 77.5%, pathological reflexes chiếm 75.0%, tendon reflexes was of 76.7% These are most common symtoms in the study groups The less 16 common symtom was round muscle paralysis of 43.4% There was no signigicant difference between the study groups - Characteristics of foci of infarction Before stroke treatment in CT scan film: The data in the table 3.26 shows that the volume of foci of infarction was between 20-40mm in both groups, of 61 patients (50.8%) There was no signigicant difference between the study groups There was no signigicant difference between the study groups (p > 0.05) 3.2.2 Treatment results of patients acute ischemic stroke after 15 days treatment 3.2.2.1 Treatment results according to morden medicine Table Changes in blood pressure, pulse rate, and temperature of patients with acute ischemic stroke after 15 days treatment Criteria Systolic Before blood After pressure P (mmHg) Diastolic Before blood After pressure P (mmHg) Before Pulse rate (times / After minute) P Before Temperature After (˚C) P TPH group (n=70) 148.5 ± 12.08 118.3 ± 11.95 Control group (n=50) 149.5 ± 11.88 120.2 ± 12.41 P 0.05 83.4 ± 9.24 71.3 ± 8.86 p> 0.05 36.9 ± 0.38 36.7 ± 0.19 p> 0.05 > 0.05 > 0.05 p >0.05 >0.05 >0.05 >0.05 Commence:: After 15 days treatment, the “Trung phong hoan” group and the control group, there was a statistically significant reduction in systolic and diastolic blood pressure (p< 0.05) However, there was no statistically significant difference between the two groups (p>0.05) There was no statistically significant change with respect to the pulse rate and temperature between the two groups (p > 0.05) * The level of consciousness recovery using Glasgow day 15 after treatment Table The level of consciousness recovery using Glasgow score TPH group (n=70) A Control group (n=50) B 17 Glasgow score 6-8 - 12 13 - 14 15 Total pA-B Before stroke treatment (1) n % 0 62.8 44 21.4 15 15.7 11 70 100 After stroke treatment (2) n % 0 58 70 p2-1 4.28 12.8 82.8 100 0.05) The positive effects on lowering the blood pressure of "Trung phong hoan" drug are considered due to the fact that it contains earthworm which has positive effects on lowering the blood pressure, proved by the study of Tran Thi Hong (MD capsule) Level of recovery from consciousness: "Trung phong hoan" drug had positive effects on improving the consciousness of patients with stroke According to traditional medicine, acute ischemic strokeis mainly caused by phlegm is blocked at middle jiao, fluctuated ascending and descending leads liver to loss of excretion, which leading to gathering qi to produce fire Lurking qi is blocked that leads to so congestion, bursts clogged phlegm, excess viscera and bowels and so on In which, phlegm syndrome is prominent TPH had positive effects on improving motor function: From the viewpoint of traditional medicine, the muscle strength is closely related to the spleen and stomach according to the theory considered that the spleen is responsible for muscle and four limbs The spleen brings nutrients of food to feed muscles If the spleen qi is efficent, muscles will hard and four limbs will be flexible Whereas,if the spleen qi is deficient, muscles will soft and the muscular tonicity reduces that leads to tiredness TPH had clearly positive effects on improving NIHSS scores because the average NIHSS score of participants was 14.57 ± 4.39, which is as the same as the patient group who are with moderately conscious, motor, and sensitive disorders Lesions observed on X-ray radiographs in the table 3.26 were mainly small and medium Morover, participants were treated early, particularly within < 24h to days leaded to the good results as shown in the table 3.23 These results are rather similar to Glasgow Coma score and MRC scale 4.2.2 The positive effects of "Trung phong hoan" drug on ischemic stroke according to traditional medicine Causes, mechanisms of wind strike are very complicated However, wind strike in general and cerebral infarction are all come from the inside and the outside, in which the inside cause is the mainly reason The remedy "Trung phong hoan" contains the following ingredients: earthworm, sweet leaf that spitting up the blood, unblocking the meridians and collaterals, disperse wind-heat, remove phlegm together 23 with the effects of dried mandarin peels which warm the middle-jiao, regulate qi and remove phlegm; black beans which tonify the liver and the kidney Therefore, it can be considered that this remedy has positive effects of dispersing wind-heat, counteracting toxicity, promote blood circulation and removing blood stasis, unblocking the meridians and collaterals, removing phlegm, tonify the liver and the kidney Wind strike is mainly caused by liver and ying kidney deficiency, heart-heat wind moving, phlegm-heat inside, phlegm-heat Thus, there was string, string-like, rapid, and rapid Tongue coating is mainly yellow, tongue textile has congestive points These characteristics are totally suitable with syndromes liver kidney ying deficiency, phlegmheat and poor circulation Therefor, in this study, using the remedy “Trung phong hoan” which had positive effects on promote blood circulation and remove blood stasis, disperse wind-heat remove phlegm, tonify the liver and the kidney to treat post-acute ischemic stroke CONCLUSION By performing the evaluation of acute toxicity, semi-chronic toxicity and positive effects of “Trung phong hoan” drug on experimental animals and clinical trials, the results are as belows: Acute toxicity, semi-chronic toxicity and positive effects of “Trung phong hoan” drug on experimental animals - In highest dose of 52.0g/kg can be administered by mice and no lethal mouse was detected The LD50 of TPH was not determined - In dose of 7.14g/kg/day and in dose of 21.42g/kg/day in weeks continuously did not affect on the general health status, hematopoietic function, function and morphology of liver and kidneys of mice administered this drug - The results of the stroke model in mice showed that TPH has positive effects on reducing the level of movement disorders immediately after administering and this effect was prolonged during the study time TPH also significantly reduced the volume of the blood clots compared to the Cebrex group The supporting effectiveness of the “Trung phong hoan” composition on patients with acute ischemic stroke - "Trung phong hoan" drug had positive effects on improving the consciousness of patients with stroke via improving Glasgow Coma 24 score The percentage of good and pretty treatment was of 82.85% and 14.28%, respectively - TPH group had positive effects on clearly improving muscle strength of patients using the MRC grade Good and pretty treatment was of 74.29% and 18.57%, respectively - TPH group had positive effects on improving NIHSS score At day 15 after treatment: NIHSS score ≤ was of 38.57%, NHISS score – 15 was of 52.86% and NIHSS score 16 – 20 was of 8.57% In which well and pretty treated patients were of 75.72% Before stroke treatment, the average NIHSS score was 12.57 ± 3.39 points; after treatment was 7.24 ± 3.16 points - TPH group had positive effects on reducing the black tongue status and increasing the pink tongue percentage as well as increasing motivation of tongue in the study group It also had positive effects on reducing yellow level and viscousity of tongue coating, increasing the bright of tongue coating; reducing the number of rapid, racing, and string-like pulse type in the study group - TPH did not cause side effectiveness in clinical trials and paraclinic test: + Clinic trials: Some patients had symtoms such as arllegy, diarrhea, nausea However, it was not needed to treat them due to the disappearing of these symtoms after days + Paraclinical test: “Trung phong hoan” did not effect on liver and kidney functions as well as hematopoietic organs of participants RECOMMENDATION By observing in clinical practice, we realized that “Trung phong hoan” drug has postitive effects on stroke Thus, this remedy shoud be applied to patients with acute ischemic stroke Moreover, the ingredients of “Trung phong hoan” drug are available and cheap Therefore, we recommend as follows: Continously evaluating the effectiveness of “Trung phong hoan” drug in ischemic stroke in a larger scale and the study should be performed to post-acute ischemic stroke Study the mechanism of action of “Trung phong hoan” drug, such as mechanism of vasodilation and reducing the blood pressure, the mechanism of the cholesterol‐lowering effect, its effectiveness on oxidants ... The study drug “Trung phong hoan” is a remedy derivered from the ancient Vietnamese remedy namely ”Thần dược cứu mệnh”, in which the dried mandarin peel is added to “Trung phong hoan” is mentioned... group had access to “Trung phong hoan” in oral dose of 7.14g/kg (equivalent to human dose) The mice in group had access to granulated medicine in the name of “Trung phong hoan” in oral dose of... 3, and 14 after stroke inducing 7 * TTC staining: In order to measure the infarct size and since then, the foci of infarction could be calculated, we used TTC staining method * Lesion volume