ETHNOPHARMACOLOGICAL RELEVANCE:Vascular dementia (VaD) is the common cognitive disorder derived mainly from lacunar stroke (LS). The oxidative stress induced neurovascular coupling (NVC) dysfunction involves in the pathogenesis of VaD. Currently, there is no specific drug for VaD. Ling-Yang-Gou-Teng -Decoction (LG), a well-known traditional Chinese formula, has been used for preventing VaD in clinic. AIM OF THE STUDY:In this study, we aimed to investigate the underlying mechanism of LG on VaD in rats. MATERIALS AND METHOD:VaD was replicated with autologous micro-thrombi against the background of hypercholesterolemia induced with high fatty diet. PTX (68.90 mg/kg/day), LG with three dosages (2.58, 8.14, 25.80 g/kg/day) was orally administrated to VaD rats, respectively. The NVC sensitivity was defined as the ratio of the microcirculative cerebral blood velocity (CBV) to the electroencephalograph (EEG) before and after penicillin stimulation. Behavioral performance, pathological changes of brain and oxidation related molecules were detected to assess the effects of LG on VaD. RESULTS:LG exhibited beneficial effects on the VaD, which was demonstrated as improved exploratory, learning and memory abilities, relieved vascular or neural pathological changes in cerebral cortex or hippocampus. LG maintained NVC sensitivity, which was confirmed as significantly increased ΔCBV and the elevated ratio of ΔCBV/ΔqEEG. The underlying mechanisms of LG was associated with antioxidant effects, which was confirmed as significantly decreased nicotinamide adenine dinucleotide phosphate oxidase 2 (NOX2) expression, and increased superoxide dismutase 3 (SOD3) expression. LG also reduced iNOS, increased nNOS and eNOS expression to restore NO bioavailability. CONCLUSIONS:The results suggested that LG prevented VaD may associate with inhibiting oxidative stress, protecting NO bioavailability, and then maintaining NVC sensitivity.

Vascular dementia (VaD) is the common cognitive disorder derived mainly from lacunar stroke. The neurovascular coupling (NVC) dysfunction involves in its pathogenesis. VaD lacks suitable animal models for developing preventive therapies. This study aimed to confirm a model for preventing VaD via maintaining NVC sensitivity in rats. The model was replicated with autologous microthrombi against the background of hypercholesterolemia. A phosphodiesterase inhibitor (pentoxyfylline) was preventively administrated to confirm the role of NVC sensitivity. Cognitive function was evaluated as exploratory, learning and memorizing abilities. NVC sensitivity was defined as the ratio of microcirculative cerebral blood flow (∆CBF) to the quantitative electroencephalograph (∆qEEG) before and after penicillin stimulation. The pathogenesis of NVC dysfunction was explored as expressions of neuronal (nNOS), inducible (iNOS) and endothelial nitric oxide synthase (eNOS) in cerebral cortex. The model rats showed cognitive impairment, microvascular edema (2.54 ± 0.30%, P < 0.01), neuronal edema (1.24 ± 0.48%, P < 0.01) and nissl body loss (0.03 ± 0.003%, P < 0.01) in cerebral cortex, and neuronal necrosis in hippocampal CA1 region (neuronal cell number 41.76 ± 10.04 cells, P < 0.01) compared with sham group. The NVC dullness in model rats was confirmed as significantly decreased ratio of ∆CBF/∆qEEG (0.05 ± 0.02%, P < 0.01) compared with sham group (0.20 ± 0.06%). The underlying mechanism of NVC dysfunction was found as imbalanced NOS expressions (decreased nNOS and eNOS, while increased iNOS levels in cerebral cortex). The NVC dullness was significantly relieved in pentoxyfylline administrated rats (0.12 ± 0.06%, P < 0.01). It indicated that this model was suitable to evaluate candidates for preventing VaD via maintaining NVC sensitivity.

Ethnopharmacological relevance: Qingdu granule (QDG), a traditional Chinese herbal prescription, had antitumor effect on breast cancer. However the underlying mechanism of QDG was unclear. The aim of this study: The present study aimed to investigate whether QDG could inhibit angiogenesis of breast cancer via acting on nuclear factor of activated T-cells (NFAT) signaling pathway. This was implicated in human umbilical vein endothelial cells (HUVECs) in vitro and breast cancer xenograft model in vivo. Materials and methods: The VEGF(165) (15.58 ng/mL) induced human umbilical vein endothelial cells (HUVECs) were treated with serum samples containing tamoxifen (TAM), tacrolimus (FK506), or QDG with three dosages. The migration and canalization capacities of HUVECs were evaluated by transwell migration and tube formation assay. In 72 h-cultured HUVECs, The gene expression, protein amount, and nuclear translocation of NFATc3 were measured. The anti-tumor and anti-angiogenic effects of QDG in vivo were investigated in breast cancer xenograft model. The serum VEGF levels, microvessel density, and protein expressions (immunohistochemistry and western blot) of VEGF, VEGFR2 and NFATc3 were detected. Results: The results showed that, QDG significantly inhibited HUVEC migration and tube formation. It down regulated NFATc3 gene expression, decreased NFATc3 protein amount, and reduced the ratio of NFATc3 nuclear translocation in HUVECs. In breast cancer xenograft model, QDG treatment significantly suppressed tumor growth, inhibited VEGF release, and decreased microvessel density. QDG reduced protein expressions of VEGF, VEGFR2 and NFATc3. Conclusion: The results suggested that QDG showed anti-angiogenic effects of breast cancer both in vitro and in vivo. The mechanism might be partially associated with inhibiting NFAT signaling pathway.

Hepatopulmonary syndrome (HPS) has a fatal hypoxemia from pulmonary shunts. Superoxide dismutase 3 (SOD3) deficiency involves in this pathogenesis. The purpose of this study was to investigate the underlying mechanisms of diammonium glycyrrhizinate (DG) on HPS via SOD3. Carbon tetrachloride induced HPS rats were treated with captopril or DG for 56 days. Blood gas, pulmonary artery pressures, and histological changes were measured. Molecule dynamics of inducible (iNOS), endothelial (eNOS), neuronal nitric oxide synthase (nNOS) and SOD3 were assessed by immunohistochemistry, quantitative RT-PCR and western blot. The results showed that DG significantly increased partial pressure of oxygen (P < 0.01), decreased alveolar-arterial oxygen gradient (P < 0.01), and improved hypoxemia. In HPS model rats, anatomical pulmonary shunts were demonstrated as both constricted arterioles and dilated metarterioles, while physiological shunts were demonstrated by lowered pulmonary artery pressure in vivo. DG significantly reversed the vascular pathological changes. Elevated iNOS or eNOS, and decreased SOD3 expression in model rats indicated imbalance of nitric oxide (NO) bioavailability. Partial SOD3 potencies correlated with circulative events and NOSs, indicating that restorable SOD3 regulated arteriole constriction and metarteriole dilatation. DG reduced iNOS or eNOS, increased SOD3 expression, especially significantly increased the partial SOD3 located in pulmonary arteries (P < 0.05), arterioles (P < 0.05) and alveolus (P < 0.05). These results suggested that DG relieved HPS shunts and limited HPS pathogenesis may associate with restoring SOD3 activity.

To develop a new model of vascular dementia for evaluating Chinese medicine prescriptions.

Vascular endothelial growth factor receptor 2 (VEGFR2) mediated calcineurin/nuclear factor of activated T-cells (NFAT) pathway is crucial in the angiogenesis of human breast cancer. Quercetin (Qu), a flavonoid known to possess anti-angiogenesis and antitumor properties, inhibited calcineurin activity in vitro. Herein, we performed a study in vivo to evaluate the effects of Qu on the angiogenesis in breast cancer. Female BALB/c nude mice were injected with MCF-7 cells into the mammary fat and were randomly divided into four groups. The animals were treated with vehicle solution, tamoxifen (TAM, 5.6mg/kg), tacrolimus (FK506, 3mg/kg), or Qu (34mg/kg) for 21 days, respectively. The results showed that, similar to TAM and FK506, Qu decreased tumor growth, limited oncocyte proliferation and promoted tumor necrosis. Anti-angiogenic actions of Qu were demonstrated as decreased serum VEGF (P<0.01), and sparse microvessel density (P<0.05). Qu significantly inhibited tumor calcineurin activities, and the inhibitory rate was 62.73% in Qu treated animals, compared to that was 72.90% in FK506 group (P>0.05). Effects of Qu on calcineurin/NFAT pathway were confirmed as decreased subcellular located levels of VEGF (P<0.05), VEGFR2 (P<0.05) and NFATc3 (P<0.01), downregulated gene expression of VEGF (P<0.05), VEGFR2 (P<0.05) and NFATc3 (P<0.01), reduced protein levels of VEGF (P<0.05), VEGFR2 (P<0.05), and NFATc3 (P<0.01) in tumor tissues. These findings indicate that Qu inhibit angiogenesis of human breast cancer xenograft in nude mice, which was associated with suppressing calcineurin activity and its regulated pathway activation.

To study the effect and mechanism of Qingdu granules on the tumor growth of 7, 12-dimethyl-benz[ a] an-thracene ( DMBA)-induced breast cancer in Sprague-Dawley rats. Methods: DMBA was used to induce breast cancer in rats. The tumor inhibition of Qingdu granules was observed. Pathological features were observed after hematoxylin-eosin staining, the distribution and content of Ki-67 in tumor were tested by IHC and the content of IL-12, IFN-γ, IL-4 and IL-10 in serum was determined by ELISA. Results:The inhibitory rate of Qingdu granules at low, middle and high dose and saikosaponin a was 30. 93%,43. 84% and 44. 17% and 43. 48%, respectively. The expression of Ki-67 was reduced in Qingdu granules groups and saikosaponin a group, the content of IL-12 and IFN-γ in serum was increased and the level of IL-4 and IL-10 was reduced in the above groups. Conclusion:Qingdu granules can inhibit breast cancer obviously, and the mechanism is probably related to the ability of immune system adjustment, which can enhance the antitumor effect.

目的：研究丹皮酚（Pae）对二乙基亚硝胺（DEN ）诱发大鼠肝癌前病变的预防作用，并探讨其作用机制。方法：采用腹腔注射DEN 诱发大鼠肝癌前病变模型。雄性w istar大鼠按照体质量随机分成3组：对照组、模型组、丹皮酚组。造模给药16周后处死大鼠，观察各组大鼠一般状况，进行肝脏病理组织学检查，检测肝功能指标ALT、AST、GST、TBIL ，脂质过氧化指标LPO、SOD、INOS、GSH‐PX。结果：病理组织学显示，丹皮酚可以显著降低肝细胞异型增生灶和异型增生结节数目；降低血清中异常升高的ALT、AST、TBIL、GST、INOS、LPO水平，升高血清中降低的SOD、GSH‐PX水平。结论：丹皮酚对于大鼠肝癌前病变具有一定程度的预防保护作用，其保护机制可能与改善肝功能，提高抗氧化能力，抑制脂质过氧化反应有关。

AIM To investigate the effects of diammonium glycyrrhizinate (Gly) on portal hypertension (PHT) in isolated portal perfused rat liver (IPPRL) with carbon tetrachloride (CCl₄)-induced chronic hepatitis. METHODS PHT model was replicated with CCl₄ in rats for 84 d. Model was identified by measuring the ascetic amounts, hepatic function, portal pressure in vivo, splenic index, and pathological alterations. Inducible nitric oxide synthase (iNOS) in liver was assessed by immunohistochemistry. IPPRLs were performed at d₀, d₂₈, d₅₆, and d₈₄. After phenylephrine-induced constriction, Gly was geometrically used to reduce PHT. Gly action was expressed as median effective concentration (EC₅₀) and area under the curve (AUC). Underlying mechanism was exploited by linear correlation between AUC values of Gly and existed iNOS in portal triads. RESULTS PHT model was confirmed with ascites, splenomegaly, serum biomarkers of hepatic injury, and elevated portal pressure. Pathological findings had shown normal hepatic structure at d₀, degenerations at d₂₈, fibrosis at d₅₆, cirrhosis at d₈₄ in PHT rats. Pseudo lobule ratios decreased and collagen ratios increased progressively along with PHT development. Gly does dose-dependently reduce PHT in IPPRLs with CCl₄-induced chronic hepatitis. Gly potencies were increased gradually along with PHT development, characterized with its EC₅₀ at 2.80 × 10⁻¹⁰, 3.03 × 10⁻¹¹, 3.77 × 10⁻¹¹ and 4.65×10⁻¹¹ mol/L at d₀, d₂₈, d₅₆ and d₈₄, respectively. Existed iNOS was located at hepatocyte at d₀, stellate cells at d₂₈, stellate cells and macrophages at d₅₆, and macrophages in portal triads at d₈₄. Macrophages infiltrated more into portal triads and expressed more iNOS along with PHT development. AUC values of Gly were positively correlated with existed iNOS levels in portal triads. CONCLUSION Gly reduces indirectly PHT in IPPRL with CCl₄-induced chronic hepatitis. The underlying mechanisms may relate to rescue NO bioavailability from macrophage-derived peroxynitrite in portal triads.

Ethnopharmacological relevance: Fuzi-Lizhong pill (FLZ) is a traditional Chinese medicine for treating patients with Spleen Yang deficient syndrome. Ghrelin, a peptide with 28 amino acid residues, plays multiple roles in thermogenesis. This study aims to explore FLZ regulating ghrelin to compensate hypothermia in rats with hypothyroid and indigestion.Materials and methods: In litter-matched rats, hypothermia was developed with both thyroidectomy at d(1) and interscapular brown adipose (IBA) removal at d(42), indigestion was induced with both high fat diet and fasting-feeding cycle from d(56); the littermates with hypothermia and indigestion were administrated with FLZ from d(70). Adaptive thermogenesis, thyroid hormones, metabolites, ghrelin dynamics were measured at d(98).Results: The results showed that plasma ghrelin levels were inversely correlated with the gastric ghrelin levels and adaptive thermogenesis in rats undergone both thyroidectomy and IBA removal. Fatty diet and FLZ enhanced the increase of plasma ghrelin of hypothyroid rats. These were supported by the changes of plasma thyroid related hormones, plasma metabolites, gastric ghrelin mRNA and protein, and the effects of fatty diet or FLZ.Conclusions: Our results suggest that more ghrelin release compensate chronic hypothermia in rats with both hypothyroidism and indigestion. It could explain the mechanisms of FLZ in relieving chronic hypothermia. (C) 2013 Elsevier Ireland Ltd. All rights reserved.

Objective To compare the curative effect of Fritillariae Cirrhosae Bulbus(FCB) in six species from five origins on recurrent asthma in mice.Methods Eighty one female Kunming mice were randomly divided into nine groups with equal weights,such as model,control,Dexamethasone(DEX,positive control),and six FCB groups.The mice in the model,DEX,and FCB groups were sc injected with ovalbumin(OVA) on days 1,7,and 14,then challenged with the aerosols of normal saline contained OVA on days 22-28.The mice in DEX and FCB groups were ig administered with DEX(0.6 mg/kg) and FCB(830 mg/kg) respectively for 28 d,and the mice in the control group were given 0.1% carboxymethyl cellulose.The surface tension of bronchoalveolar lavage was measured,the lung tissues were taken for hematoxylin-eosin(HE) staining,and the tracheal stenosis and volume difference of pulmonary alveolus were analyzed by the microphotograph.Results Compared with the control group,the tracheal stenosis,surface tension,and volume difference of pulmonary alveolus were increased obviously in the model group(P 0.01).The pathological section revealed vessel wall thickening in bronchiole,inflammatory cell infiltrating,gland hyperplasia,and mucus hypersecretion,but FCB could reverse these pathological changes(P 0.01).Each FCB had its feature to cure asthma.F.unibracteata and F.delavayi had the most potency on increasing alveolar surfactant;F.unibracteata and F.taipaiensis had the most potency on attenuating tracheal stenosis;F.unibracteata and F.przewalskii had the most potency on relieving the inflammation;F.unibracteata and F.taipaiensis had the most potency on reducing glandular hyperplasia or intimal thickening.Each FCB had its feature to cure asthma,but F.unibracteata was the best.Conclusion FCB has the best potency to prevent and cure the recurrent asthma in mice,but each FCB has its feature.The diversity of FCB species should be protected.

This study is aimed to investigate the effects of Sal B on portal hypertension (PH). PH with chronic hepatitis was induced by carbon tetrachloride (CCl4) in rats. The model was confirmed with elevated portal pressures and increased serum CD163 levels. The inducible nitric oxide synthase (iNOS) or heme oxygenase-1 (HO-1) in portal triads was assessed. The isolated portal perfused rat liver (IPPRL) was performed at d0, d28, d56 , and d84 in the progression of chronic hepatitis. After constricting with phenylephrine, the portal veins were relaxed with Sal B. The EC50 of Sal B for relaxing portal veins was −2.04×10−9, 7.28×10−11, 1.52×10−11, and 8.44×10−11 mol/L at d0, d28, d56, and d84, respectively. More macrophages infiltrated in portal triads and expressed more iNOS or HO-1 as PH advanced. The areas under the curve (AUCs) of Sal B for reducing PH were positively correlated with the levels of iNOS or HO-1 in portal triads, and so did with serum CD163 levels. Sal B reduces PH in IPPRL with chronic hepatitis, via promoting portal relaxation due to macrophage-originated NO or CO in portal triads, partly at least.

Including herbal medicine, complementary and alternative medicine (CAM) is popular worldwide. The traditional Chinese medicine xinkeshu has been widely used to treat coronary heart disease in China. This study was designed to investigate the protective effect and probable mechanism of xinkeshu tablet to atherosclerotic myocardial ischemia rabbit. Rabbits were divided into four groups (n = 12 each) and fed with different diet for 12 weeks: Control (standard diet), Model (high-cholesterol diet), XKS (high-cholesterol diet with 184.8 mg/kg/d xinkeshu), and Atorvastatin (high-cholesterol diet with 5.0 mg/kg/d atorvastatin). Plasma lipoprotein, ECG, endothelium-dependent vessel relaxation, histomorphological study, and expressions of eNOS and VCAM-1 on coronary arteries were assessed. The findings showed that, similar to atorvastatin, xinkeshu presented significant effects on rescuing endothelium-dependent vessel relaxation, inhibiting atherosclerotic progress, preventing myocardial ischemia, and changing eNOS and VCAM-1 expression. However, xinkeshu showed no lipoprotein lowering effect in hypercholesterolemia rabbits. The results of the present study indicated that xinkeshu exerted potent antiatherogenic and anti-ischemic properties on atherosclerotic myocardial ischemia rabbit. An endothelial protecting effect may be involved in the mechanism other than antihyperlipidemic effect.

This study is aimed to investigate the Aconitum Lizhong pill (ALZ) pharmacological actions on hypothermia with indigestion, especially the ghrelin roles. The littermate-matched rats were randomly divided into four groups. Control did sham operation or standard diet, Model carried out interscapular brown adipose (IBA) removal with standard diet, Fat-diet did IBA removal with fat-diet, and ALZ did IBA removal and fat-diet with 4.536 g/kg/d ALZ. The potency of adaptive thermogenesis, ghrelin levels in plasma or gastric mucosa, thyroid hormones and metabolite in sera, expression of ghrelin mRNA, and protein in gastric mucous membrane were determined. ALZ relieved the hypothermia processes with indigestion, via inhibiting ghrelin expression and increasing ghrelin secretion; the dynamics from the therapy is supported with the energy changes as less body weight loss, less plasma lipid decrease, more plasma T3 or T4 increase with TSH decrease, and more compensation of thermogenic AUC decrease. Ghrelin played key roles in the actions of ALZ on the hypothermia with indigestion. The pharmacological mechanisms of ALZ involved the homeostasis of ghrelin expression and secretion.