Efficacy of Yangyin Yiqi Huoxue Granule () in Treatment of Ischemic Stroke Patients with Qi-Yin Deficiency and Blood Stasis Syndrome: A Randomized, Double-Blind, Multicenter, Phase-2 Clinical Trial / 中国结合医学杂志

OBJECTIVE@#To evaluate the efficacy and safety of Yangyin Yiqi Huoxue Granule (, YYHG) in the treatment of ischemic stroke (IS) patients with qi-yin deficiency and blood stasis syndrome (QYDBSS), and to explore its effective dosage.@*METHODS@#The total of 288 patients were randomly assigned to the YYHG high-dose, YYHG low-dose, positive control (administered Xiaoshuantong Granule, XSTG, ), or placebo control (administered inert granule) groups (72 cases per group) by software-drived competitive block randomization. The trial was conducted for a 28-day period, with a 180-day follow-up period. The primary outcome was the comprehensive curative evaluation, and secondary outcomes were the National Institute of Health Stroke Scale (NIHSS) score, Barthel activities of daily living (ADL) index score, the quality of life index (QLI) score, and the Chinese medicine syndrome (CMS) score. All analyses were done on an intention-to-treat basis. The clinical safety was also assessed.@*RESULTS@#The total of 288 participants were recruited between June 1, 2008 and September 30, 2009, and 287 patients received intervention; the treatment groups were well balanced at baseline. The comprehensive cure rates of YYHG high-dose, low-dose, positive and placebo control groups were 63.38%, 31.94%, 36.11% and 6.14%, respectively; there was a statistical difference between the two groups (P<0.01), while the high-dose YYHG treatment group was significantly higher than the other 3 groups (P<0.01). The improvement of NIHSS, ADL, QLI and CMS scores of the YYHG high-dose and low-dose groups was significantly better than that of the positive control group and the placebo control group (P<0.05). In terms of improving the classification of the NIHSS scale and the assessment of the ADL scale, the YYHG high-dose group was significantly better than the other three groups (P<0.05), and the YYHG low-dose group was better than the placebo control group (P<0.01). At the same time, except for the QLI score, the high-dose group was better than the low-dose group (P<0.05). In terms of safety, adverse reactions after YYHG treatment were generally mild (3.78%), and no serious adverse reactions have been reported.@*CONCLUSION@#YYHG is safe and effective in the treatment of IS patients with QYDBSS.

Protective Effect of Naoxintong Capsule () Combined with Guhong Injection () on Rat Brain Microvascular Endothelial Cells during Cerebral Ischemia-Reperfusion Injury / 中国结合医学杂志

OBJECTIVE@#To investigate the synergistic effect of Naoxintong Capsule (NXTC, ) and Guhong Injection (GHI, ) on cerebral ischemia-reperfusion (I/R) injury.@*METHODS@#Forty-eight Sprague-Dawley rats were divided into 6 groups: control group, oxygen and glucose deprivation (OGD) group, nimodipine group (9.375 mg/kg), NXTC group (0.5 g/kg), GHI group (5 mL/kg) and NXTC+GHI group (0.5 g/kg NXTC+5 mL/kg GHI), after the onset of reperfusion and once per day for the following 7 days. Blood was collected 1 h after final administration, and the sera were collected. Cultured primary rat brain microvascular endothelial cells (rBMECs) were subjected to OGD to establish a cell injury model. Untreated rBMECs were used as blank control. The cell counting kit-8 assay was used to assess cell viability using the sera. Malondialdehyde (MDA) and superoxide dismutase (SOD) levels were assessed using an enzyme-linked immunosorbent assay. Apoptosis was evaluated after Hoechst33342 staining using fluorescence microscopy and flow cytometry. JC-1 staining was performed to assess changes in mitochondrial membrane potential.@*RESULTS@#Statistical analysis indicated that more than 95% of the cells were rBMECs. Compared with the OGD group, the cellular morphology of the all drug delivery groups improved. In particular, the combined drug group had the most significant effect. Compared with the OGD group, all drug intervention groups induced a decrease in the apoptotic rate of rBMECs, increased the SOD levels, and decreased the MDA levels (all P<0.01). Compared with the mono-therapy groups, the NXTC+GHI group exhibited a significant improvement in the number of apoptotic rBMECs (P<0.01). All drug intervention groups showed different degrees of increase in membrane potential, and the NXTC+GHI group was higher than the NXTC or GHI group (P<0.01).@*CONCLUSION@#The combinationa application of NXTC and GHI on cerebral I/R injury clearly resulted in protective benefits.

Lipid-lowering effect and mechanism of Danhong Injection on hyperlipidemia rats based on lipid metabolism disorder / 中国中药杂志

The animal model of hyperlipidemia in rats was established to investigate the lipid-lowering effect and mechanism of Danhong Injection on hyperlipidemic rats. SD rats were selected as the research object. The rats in normal group were fed with basic diet, and the rats in other groups were fed with high-fat diet to establish hyperlipidemia model. The successfully modeled rats were randomly divided into model group, Danhong Injection low, medium, high dose(1.0, 2.0, 4.0 mL·kg~(-1)) groups, and simvastatin(2.0 mg·kg~(-1)) group. Danhong Injection groups received intraperitoneal administration, and simvastatin group received intragastrical administration, once a day for 4 weeks. At the first, second, third, and fourth weekends after administration, blood was collected from the orbital vein to detect the levels of total cholesterol(TC), triglyceride(TG), low-density lipoprotein cholesterol(LDL-C), and high-density lipoprotein cholesterol(HDL-C), and then the atherosclerosis index(AI) was calculated. After 4 weeks of administration, the animals were sacrificed, and their heart, liver, spleen, lung, kidney and adipose tissue were extracted and weighed respectively to calculate the organ index of each group. The expressions of acyl-coaoxidase 1(Acox1), adenosine 5'-monophosphate(AMP)-activated protein kinase alpha(AMPK-α), bile salt export pump(BSEP), peroxisome proliferator-activated receptor gamma(PPAR-γ), catalase(CAT) and superoxide dismutase(SOD) mRNA in liver tissues were detected by fluorescence quantitative PCR; the content of cholesteryl ester transfer protein(CETP) and lecithin cholesterol acyltransferase(LCAT) in serum was detected by ELISA. The results showed that as compared with the normal group, the levels of serum TC, TG and LDL-C in the model group were significantly increased, and the level of HDL-C was significantly decreased, indicating that the hyperlipidemia rat model was successfully constructed. As compared with the model group, Danhong Injection could decrease the contents of TC, TG, LDL-C and increase the content of HDL-C in hyperlipidemia rats; reduce the body weight of hyperlipidemia rats, and reduce the liver weight, liver index, fat weight and fat index; it had no significant effect on the main organ indexes such as heart, spleen, lung and kidney; but it could increase the expressions of Acox1, AMPK-α, BSEP, PPAR-γ, CAT and SOD mRNA in liver tissues of rats; it could also reduce the level of CETP and increase the level of LCAT in serum; and the regulatory effect of Danhong Injection groups all showed a dose-dependent effect. It can be concluded that Danhong Injection can regulate the blood lipid contents, reduce the blood lipid levels and alleviate the accumulation of body fat in rats with hyperlipidemia. The mechanism may be related to inhibiting lipid metabolism disorder and oxidative stress induced by high-fat diet feeding, and improving the imbalance of lipid transport system.

Effects of Danhong Injection (丹红注射液) and its main components on anticoagulation and fibrinolysis in cultured vein endothelial cells / 中国结合医学杂志

<p><b>OBJECTIVE</b>To observe the effects of Danhong Injection (丹红注射液) and its main components, including daiclzein and hydroxysafflor yellow A (HSYA), on the anticoagulation, fibrinolysis, anti-apoptosis in hypoxia model of vein endothelial cells (VECs).</p><p><b>METHODS</b>VECs were prepared and were put in a hypoxia environment, which consisted of mixed gas of 95% N and 5% CO mixed gas, when reached confluent culture. Five groups used different treatments, including normal control group, hypoxia group, daiclzein group, HSYA group and Danhong Injection group. The VECs were identified by fluorescence double labeling methods. The morphology was observed by a phase contrast microscopy. The effects of Danhong Injection, daiclzein and HSYA on 6 keto prostaglandin F1α (6-keto-PGF1α) level was measured by the method of radioimmunoassay (RIA). Superoxide dismutase (SOD) activity was tested by water soluble tetrazolium salt. The content of malondialdehyde (MDA) was measured by thiobarbituric acid. The activities of tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor (PAI) were measured by the method of chromogenic substrate. The contents of endothelin (ET) and nitric oxide (NO) were detected by non-equilibrium RIA and enzymelinked immunosorbent assay. Cells apoptosis rate was determined by flow cytometry.</p><p><b>RESULTS</b>Compared with the normal control group, the floating cells number, PAI activity, ET and MDA contents, and cells apoptosis rate in the culture solution of hypoxia group were all significantly increased, whereas the 6-keto-PGF1α and NO contents, and t-PA and SOD activities were decreased significantly (P<0.01). Compared with the hypoxia group, Danhong Injection markedly increased the 6-keto-PGF1α content and SOD activity, regulated PAI and t-PA activities, ET and NO contents, and decreased MDA content and cells apoptosis rate (P<0.05 or P<0.01).</p><p><b>CONCLUSIONS</b>Danhong Injection and its main components played an important role in protecting primary VECs from hypoxic damage by regulating the secretion and vasomotor function of VECs. The function of Danhong Injection was most remarkable.</p>

Treating ischemic stroke patients of deficiency of qi and yin syndrome and static blood obstructing collaterals syndrome by Yangyin Yiqi Huoxue Recipe: a clinical study of therapeutic effect / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To observe the clinical effect of Yangyin Yiqi Huoxue Recipe (YYHR, the basic recipe of Yangyin Tongnao Granule) in treatment of ischemic stroke patients of deficiency of qi and yin syndrome (DQYS) and static blood obstructing collaterals syndrome (SBOCS).</p><p><b>METHODS</b>Totally 312 patients were assigned to the control group (86 cases) and the treatment group (226 cases) using strati- fied randomized allocation method. Patients in the treatment group were treated with modified YYHR, while those in the control group took Xueshuan Xinmaining. The treatment course was 4 weeks for all. Constituent ratios of the acute stage and the recovery stage of DQYS and SBOCS and their complicated syndromes were observed in the two groups. Changes of the clinical curative effect, clinical symptoms integral, whole blood viscosity ratio, plasma viscosity ratio, hematocrit, erythrocyte sedimentation rate (ESR), total cho- lesterol (TC), triglyceride (TG), high density lipoprotein cholesterol (HDL-C), and low density lipoprotein cholesterol (LDL-C) were detected in the two groups before and after treatment.</p><p><b>RESULTS</b>There was statistical difference in constituent ratios of the acute stage and the recovery stage of DQYS SBOCS and its complicated syndromes between the two groups (P < 0.01). DQYS and SBOCS was basic syndrome types of the two groups. The cured and markedly effective rate was 71.24%(161/226) in the treatment group and 43.02% (37/86) in the control group. The total effective rate was 91.15% (206/226) in the treatment group, higher than that of the control group (76.74%, 66/86) with statistical difference (P < 0.01). There was statistical difference in the clinical symptoms integral, whole blood viscosity ratio, plasma viscosity ratio, hematocrit, ESR, TC, TG,HDL-C, and LDL-C (P < 0.05, P < 0.01).</p><p><b>CONCLUSIONS</b>Symptoms of ischemic stroke patients could be improved by modified YYHR. Indices such as the whole blood viscosity, plasma viscosity ratio, hematocrit, ESR, abnormal metabolism of blood lipids were also significantly improved. Pathological changes of blood stasis induced by qi-yin deficiency exist in ischemic stroke patients, and DQYS and SBOCS were basic syndrome types.</p>

Effect of Shenxiong injection on inflammation injury of ischemia-reperfusion injury rats / 中国中药杂志

To investigate the effect of Shenxiong injection on the inflammation injury of ischemia-reperfusion injury senile rats. Totally 84 Sprague-Dawley (SD) rats were randomly divided into six groups: the sham operation group, the model group, the Nimodipine group and the Shenxiong injection(low, middle, and high dosage) groups. The rat cerebral ischemia-reperfusion model was established through intraperitoneal injection for 3 d and middle cerebral artery occlusion (MCAO). Ater the reperfusion for 24 h, efforts were made to give neurological score, collect brains for TTC staining, detect tumor necrosis factor-α(TNF-α) and interleukin-1β(IL-1β) content in serum by enzyme-linked immunosorbent assay (ELISA) method and measure IL-1β, ICAM-1 and MMP-9 mRNA expressions in hippocampal area by Real-time PCR (RT-PCR). According to the results, Shenxiong injection could decrease the cerebral infarction volume, greatly improved the neurological function and reduce IL-1β, TNF-α, ICAM-1 and MMP-9 mRNA expressions and IL-1β and TNF-α contents. In conclusion, Shenxiong injection shows the significant protective effect on ischemia-reperfusion injury in rats. Its mechanism may be related to the inhibition of inflammatory factor expression.

Protective effect of Shenxiong injection on cerebral ischemia-reperfusion injury of rats / 中国中药杂志

<p><b>OBJECTIVE</b>To study the protective effect of Shenxiong injection on the cerebral ischemia-reperfusion injury of senile rats.</p><p><b>METHOD</b>Totally 108 Sprague-Dawley (SD) rats were randomly divided into the sham operation group, the model group, the Ni-modipine group and Shenxiong injection groups (low, middle, and high doses). The rat brain ischemia-reperfusion model was established by the middle cerebral artery occlusion (MCAO) method in rats, in order to observe the effect of Shenxiong injection on neurological score and brain infarct volume of rats with cerebral ischemia-reperfusion injury, and determine the contents of NOS, NO, SOD, MDA and LDH in brain tissues. The contents of TNF-alpha and IL-1beta levels in brain tissues were measured by enzyme-linked immunosorbent assay (ELISA) method.</p><p><b>RESULT</b>Shenxiong injection could significantly decrease neurological score, injury degree of brain tissues and brain infarct volume of rats with cerebral ischemia-reperfusion injury, increase the vigor of SOD, decrease the levels of MDA, NO, NOS and LDH, and inhibit IL-1beta and TNF-alpha expressions.</p><p><b>CONCLUSION</b>Shenxiong injection has the obvious protective effect on the brain ischemia-reperfusion injury in rats. Its mechanism may be related to the improvement of neurological function, the reduction of free radical injury, and the inhibition of inflammation factor expression.</p>

Study on pharmacokinetics-pharmacodynamics correlation of Danshensu in rats with focal cerebral ischemia / 中国中药杂志

To study the pharmacokinetic process of Danshensu in cerebal ischemia injury model rats and the correlation with its anti-cerebral ischemia effect. In this study, the middle cerebral artery occlusion (MCAO) model was established, in which all of the rats were intravenously injected of Danshensu at a single dose of 40 mg x kg(-1). The HPLC-DAD method was applied to determine the plasma concentration of Danshensu at different time points and draw the drug-time curve. Meanwhile, the superoxide dismutase (SOD) and the lactate dehydrogenase (LDH) activity were determined to draw the time-effect curve. The DAS 3.2. 6 software was used to process the data, analyze their correlation, compare the pharmacokinetic difference between model and normal rats after the administration of the same doses of Danshensu and the changes in pharmacodynamic indicators of model rats after the administration, and evaluate the effect of Danshensu in treating the cerebral ischemia disease. According to the results, the pharmacokinetic processes of Danshensu in the cerebral ischemia-reperfusion and normal rats were consistent to the two-compartment model. The main pharmacokinetic parameters were: t1/2alpha were (0.267 +/- 0.026), (0.148 +/- 0.020) h;t1/2beta were (1.226 +/- 0.032), (1.182 +/- 0.082) h; AUC0-infinity were (42.168 +/- 4.007), (26.881 +/- 1.625) mg x L(-1) x h. After the cerebral ischemia-reperfusion, the activity of SOD decreased and the activity of LDH increased. Danshensu could inhibit the decrease in the SOD activity and the increase in the LDH activity within a certain period of time. This indicated that Danshensu could stay longer in cerebral ischemia-reperfusion rats than in normal rats and eliminated more slowly, which reflected the rationality of Danshensu in the clinical treatment of cerebral ischemia diseases. Danshensu's effect against the cerebral ischemic injury may be related with its level in vivo. Its plasma concentration is positively related to the SOD activity and negatively related to the LDH activity.

Protective effect of combined administration of active ingredients of Danhong on cerebral micro-vascular endothelial cell injured by hypoxia / 中国中药杂志

To study the protective effect of combined administration of active ingredients of Danhong on cultured primary mice's brain microvascular endothelial cells (rBMECs) injured by hypoxia. Primary mice's brain micro-vascular endothelial cells were cultured to establish the 4 h hypoxia model. Meanwhile, active ingredients (protocatechuic aldehyde, salvianolic acid B, hydroxysafflor yellow A and tanshinol) of Danhong were administered in rBMECs. The non-toxic dosage was determined by MTT. The leakage of lactate dehydrogenase(LDH), cell superoxide dismutase (SOD) activity and MDA level were detected by the colorimetric method. The expressions of ICAM-1, MMP-9, P53 mRNA were detected by RT-PCR method. Changes in rBMECs cell cycle and early apoptosis were detected by flow cytometry. Danhong's active ingredients and prescriptions 1, 2, 3, 7, 8, 9 could be combined to significantly restrain LDH in hypoxic cells supernatant. Prescriptions 1, 2, 3, 7, 8, 9 could significantly enhance SOD activity in anoxic cells; Prescriptions 1, 2, 3, 8, 9 could significantly decrease the MDA level; Prescriptions 1, 2, 6, 7, 9 could significantly inhibit the early rB-MECs apoptosis induced by hypoxia. After hypoxia, the up-regulated P53 mRNA expression could cause retardation in G, phase and promote cell apoptosis. This proved that the regulatory function of P53 gene lay in monitoring of calibration points in G, phase. Prescriptions 1, 2, 5, 6, 7, 8, 9 could significantly down-regulate the P53 mRNA expression; Prescriptions 1, 4, 7, 8, 9 could significantly down-regulate the ICAM-1 mRNA expression; Prescriptions 1, 3, 6, 9 could significantly down-regulate the MMP-9 mRNA expression. The combined administration of Danhong's active ingredients showed a significant protective effect on primary cultured rBMECs injury induced by hypoxia Its mechanism may be related to the enhancement of cellular antioxidant capacity and the inhibition of inflammatory response and cell apoptosis. This study could provide ideas for researching prescription compatibility, and guide the clinical medication.

Effect and mechanism of Guhong injection against cerebral ischemia reperfusion injury / 中国中药杂志

To explore the effect and mechanism of Guhong injection against cerebral ischemia reperfusion injury, SD rats were randomly divided into the sham-operated group, the model group, the nimodipine group, and high, medium and low-dose Guhong injection groups, with 10 rats in each group. The middle cerebral artery embolization (MCAO) model was established to observe neurological deficit symptoms, infarct volume, SOD activity, MDA content, GSH-Px and CAT activity in rats, as well as the contents of t-PA, PAI, TXB2 and 6-keto-PGF1α in serum. The results showed that Guhong injection could obviously promote the recovery of neurological deficit symptoms, narrow the brain infarct volume in rats after surgery, significantlyincrease the activities of SOD, GSH-Px and CAT and decrease the content of MDA. Meanwhile, it also could obviously increase the contents of t-PA and 6-keto-PGF1α and decrease the contents of PAI and TXB2 in serum, indicating that Guhong injection have better antioxidant and antithrombus effects, as well as a significant protective effect against cerebral ischemia reperfusion injury in rats.

Protective mechanism of Danhong injection on brain microvascular endothelial cells injured by hypoxic / 中国中药杂志

To study the protective mechanism of Danhong injection on brain microvascular endothelial cells (rBMECs) injured by hypoxic. In the experiment, primary suckling mouse's rBMECs cells were collected and identified with factor VIII to establish the 4 h injury model. Meanwhile, rBMECs were given Danhong injection (25, 50, 100 mL . L-1), and the superoxide dismutase (SOD) activity and the malonyldialdehyde (MDA) level were detected by the biochemical method. Cell MMP-9, ICAM-1 and P53 mRNA expression levels were detected by RT-PCR method. Changes in cells' microscopic structure were observed by transmission electron microscope. According to the results, primary rBMECs were notably injured by hypoxia. Compared with model group, Danhong injection (50, 100 mL . L-1) could remarkably resist the injury induced by hypoxic, increase intracellular SOD activity, decrease MDA level and significantly down-regulate ICAM-1, MMP-9 and P53 mRNA expressions. Danhong injection (100 mL . L-1) could protect the cells' normal morphology and microscopic structure, maintain the close intercellular junction, and inhibit the hypoxia-induced cell apoptosis. The results showed that Danhong injection plays a significant role in protecting rBMECs injured by hypoxia. Its mechanism may be related to the enhancement of cells' antioxidant capacity, the inhibition of inflammatory response and the cell apoptosis.

The effect and mechanism of Yiqi Tongluo Jiedu capsule against cerebral ischemia reperfusion injury / 药学学报

To observe the effect and mechanism of Yiqi Tongluo Jiedu capsule aganist cerebral ischemia reperfusion injury, the SD rats were randomly divided into following groups: sham-operated group, model group, the group of low, medium and high dose of Yiqi Tongluo Jiedu capsule, and nimodipine group. Using focal middle cerebral artery embolization (MCAO) model, following items were observed: symptoms of neurological deficit score; infarct volume; activity of SOD, content of MDA and NO, activity of NOS of ischemic brain tissue; Bcl-2 and Bax protein expression; content of IL-1beta, IL-6 and TNFalpha in serum; IL-1beta mRNA expression of ischemic brain tissue. Results showed that Yiqi Tongluo Jiedu capsule could significantly reduce the symptoms of neurological deficits, promote the recovery symptoms of neurological deficits; narrow infarct volume of brain tissue obviously, reduce the percentage of infarct volume; raise activity of SOD, reduce content of MDA and NO, reduce activity of NOS; increase Bcl-2 protein, reduce Bax expression; reduce content of IL-1beta, IL-6 and TNFa in serum; reduce IL-1beta mRNA expression of ischemic brain tissue. Yiqi Tongluo Jiedu capsule has significant protective effects against ischemic brain injury, it has significant anti-apoptotic, antioxidant and anti-inflammatory effects.

Study on transport of Yangyin Tongnao granules in main effective fractions / 中国中药杂志

<p><b>OBJECTIVE</b>To investigate the absorption characteristics and transportation mechanism Yangyin Tongnao granules in main effective fractions in Caco-2 cell model.</p><p><b>METHOD</b>The safety concentrations of Yangyin Tongnao granules in main effective fractions in Caco-2 cells. A Caco-2 cell model was established to study the transport situations after the compatibility of Yangyin Tongnao granules in main effective fractions, and the content was determined by high performance liquid chromatography (HPLC).</p><p><b>RESULT</b>P(app) of puerarin, ligustrazine and astragaloside were less than 1.0 x 10(-6) cm x s(-1), and their P(app) were hard to be close to atenolol. The oral absorption in descending order is shown as the following: puerarin, ligustrazine, astragaloside. After the compatibility between saponins and flavonoids, P(app) of astragaloside was improved obviously, which promoted the transport from apical (AP) to basolateral (BL); the compatibility of puerarin, ligustrazine and astragaloside showed a significant effect in the efflux of astragaloside and no change in the absorption transport of ligustrazine and puerarin at the same time. There is a great difference in bidirectional transport of representative component of each effective fraction, and P(app)(B --> A) was significantly greater than Papp(A --> B), which suggested that the efflux transport from BL side to AP side had an advantage in the three representative components of the three effective fractions in Caco-2 cell monolayer model.</p><p><b>CONCLUSION</b>Astragaloside, ligustrazine and puerarin may be malabsorptive compounds, and the three compounds may be discharged by the transport protein in small intestine membrane.</p>