Neuroprotective Effects of Lycium barbarum Polysaccharide on Focal Cerebral Ischemic Injury in Mice.

Increasing evidence demonstrates inflammation contributes to neuronal death following cerebral ischemia. Lycium barbarum polysaccharide (LBP) has been reported to prevent scopolamine-induced cognitive and memory deficits. We recently indicated that LBP exerts neuroprotective effect against focal cerebral ischemic injury in mice via attenuating the mitochondrial apoptosis pathway. The aim of this study was to investigate the neuroprotective effects of LBP against the behavioral dysfunction induced by focal cerebral ischemia injury in mice. Following 7 successive days of pretreatment with LBP (10, 20 and 40 mg/kg) and nimodipine (4 mg/kg) by intragastric gavage, mice were subjected to middle cerebral artery occlusion (MCAO). Following reperfusion, cerebral blood flows, the total power of the spontaneous EEG, and morphological changes were estimated. Learning and memory ability, and motor coordination were determined by Morris water maze task, rotarod and grip test. Western blot analysis, Real-Time fluorogenic PCR assays, and immunofluorescence staining were used to examine the expression of proinflammatory mediators and activation of microglia. The present study showed that LBP pretreatment significantly enhanced regional cortical blood flow and the total power of the spontaneous EEG, improved memory and motor coordination impairments, and inhibited over-activation of microglia and astrocytes after MCAO. Further study demonstrated LBP suppressed MCAO-induced activations of P65 NF-κB and P38 MAPK, and prevented up-regulations of proinflammatory mediators in hippocampus. Our data suggest that LBP can exert functional recovery of memory and motor coordination deficits and neuroprotective effect against cerebral ischemic injury in mice.

Ligustilide prevents cognitive impairment and attenuates neurotoxicity in D-galactose induced aging mice brain.

Ligustilide (LIG) is a principal active ingredient of traditional Chinese medicine, Radix Angelica sinensis, which has versatile pharmacological activities including neuroprotection. Previous studies have demonstrated that LIG has beneficial effects on cognition deficits associated with cerebral damage or neurodegenerative disorders. In present study, we investigated the neuroprotective effect of LIG on cognitive impairment and neurotoxicity in the brain of aging mouse induced by d-galactose (d-gal). The aging model mice were induced by subcutaneous (S.C.) injection of d-gal once daily for 8 weeks and LIG (80 mg/kg) was simultaneously administered orally. The Morris water maze (MWM) test was used to assess the spatial learning and memory abilities. The activity of Na(+)-K(+)-ATPase and the content of lipid peroxidation product malondialdehyde (MDA) in brain were examined. The levels of glial fibrillary acidic protein (GFAP), growth-associated protein GAP-43, and cleaved caspase-3 in brain were also determined by immunohistochemistry. The MWM test showed that LIG administration markedly improved behavioral performance of d-gal treated mice. This action could be partly explained by the results that LIG reduced the level of MDA as well as increased the activity of Na(+)-K(+)-ATPase in the brain of d-gal induced aging mice. Moreover, LIG significantly raised the expression of GAP-43 and reduced cleaved caspase-3 and GFAP levels in the brain of d-gal treated mice. These results demonstrated that LIG improves d-gal-induced cognitive dysfunction and brain toxicity, which suggests that LIG may be developed as a new medicine for the treatment of aged-related conditions.

Reducing iron in the brain: a novel pharmacologic mechanism of huperzine A in the treatment of Alzheimer's disease.

Huperzine A (HupA), a natural inhibitor of acetylcholinesterase derived from a plant, is a licensed anti-Alzheimer's disease (AD) drug in China and a nutraceutical in the United States. In addition to acting as an acetylcholinesterase inhibitor, HupA possesses neuroprotective properties. However, the relevant mechanism is unknown. Here, we showed that the neuroprotective effect of HupA was derived from a novel action on brain iron regulation. HupA treatment reduced insoluble and soluble beta amyloid levels, ameliorated amyloid plaques formation, and hyperphosphorylated tau in the cortex and hippocampus of APPswe/PS1dE9 transgenic AD mice. Also, HupA decreased beta amyloid oligomers and amyloid precursor protein levels, and increased A Disintegrin And Metalloprotease Domain 10 (ADAM10) expression in these treated AD mice. However, these beneficial effects of HupA were largely abolished by feeding the animals with a high iron diet. In parallel, we found that HupA decreased iron content in the brain and demonstrated that HupA also has a role to reduce the expression of transferrin-receptor 1 as well as the transferrin-bound iron uptake in cultured neurons. The findings implied that reducing iron in the brain is a novel mechanism of HupA in the treatment of Alzheimer's disease.

Ligustilide alleviates brain damage and improves cognitive function in rats of chronic cerebral hypoperfusion.

ETHNOPHARMACOLOGICAL RELEVANCE: Ligustilide (LIG), a main lipophilic component of Danggui (Chinese Angelica root, Radix Angelica sinensis) which is a popular used herb to treat menstrual disorders in traditional chinese medicine, has been reported to possess some neuroprotective effects on permanent focal ischemia and transient forebrain ischemia. AIM OF THE STUDY: Based on previous work, we intended to investigate the protective effects of LIG on parietal cortex and hippocampus of rats in chronic cerebral hypoperfusion model. MATERIALS AND METHODS: Chronic cerebral hypoperfusion was induced by permanent, bilateral common carotid artery's occlusion (2VO). The rats were treated with LIG (80mg/kg, by oral) from the eighth day after surgery for seven consecutive days. Their spatial learning and memory abilities were assessed using the Morris water maze. After six days for maze test, rats were sacrificed. Coronal sections in cortex and hippocampus were stained with cresyl violet or labeled with NeuN (Neuronal Nuclei), MAP-2 (Microtubule-Associated Protein-2), Caspase-3 and GFAP (Glial Fibrillary Acidic Protein) antibodies. RESULTS: LIG treatment for seven days decreased escape latency and swimming distance of 2VO rats from the third day in maze tests, and increased percent time in the target quadrant. LIG prevented neuronal loss, dendrites damage and neuronal apoptosis in both parietal cortex and hippocampus of 2VO rats; and it also inhibited astrocytic activation and proliferation stimulated by hypoperfusion. CONCLUSIONS: These results demonstrate that LIG show obvious neuroprotective potential for treating chronic cerebral hypoperfusion injury, which may be attributed to its anti-apoptosis of neuron and anti-proliferation of astrocyte both in cortex and in hippocampus of 2VO rats. We suggest that LIG can be developed as an effective drugs for the prevention of vascular dementia (VD).

Myrica rubra Extracts Protect the Liver from CCl(4)-Induced Damage.

The relationship between the expression of mitochondrial voltage-dependent anion channels (VDACs) and the protective effects of Myrica rubra Sieb. Et Zucc fruit extract (MCE) against carbon tetrachloride (CCl(4))-induced liver damage was investigated. Pretreatment with 50 mg kg(-1), 150 mg kg(-1) or 450 mg kg(-1) MCE significantly blocked the CCl(4)-induced increase in both serum aspartate aminotransferase (sAST) and serum alanine aminotransferase (sALT) levels in mice (P < .05 or .01 versus CCl(4) group). Ultrastructural observations of decreased nuclear condensation, ameliorated mitochondrial fragmentation of the cristae and less lipid deposition by an electron microscope confirmed the hepatoprotection. The mitochondrial membrane potential dropped from -191.94 ± 8.84 mV to -132.06 ± 12.26 mV (P < .01) after the mice had been treated with CCl(4). MCE attenuated CCl(4)-induced mitochondrial membrane potential dissipation in a dose-dependent manner. At a dose of 150 or 450 mg kg(-1) of MCE, the mitochondrial membrane potentials were restored (P < .05). Pretreatment with MCE also prevented the elevation of intra-mitochondrial free calcium as observed in the liver of the CCl(4)-insulted mice (P < .01 versus CCl(4) group). In addition, MCE treatment (50-450 mg kg(-1)) significantly increased both transcription and translation of VDAC inhibited by CCl(4). The above data suggest that MCE mitigates the damage to liver mitochondria induced by CCl(4), possibly through the regulation of mitochondrial VDAC, one of the most important proteins in the mitochondrial outer membrane.

[Determination of butylidenephthalide in Ligusticum chuanxiong by HPLC].

OBJECTIVE: To determine butylidenephthalide in Ligusticum Chuanxiong with RP-HPLC. METHOD: The sample was extracted with methanol using sonication. The ESTD was used to quantify butylidenephthalide. HPLC separation was carried out in a Hypersil ODS columm (4.6 mm x 150 mm, 5 microm) , eluted at 1 mL x min(-1) with methanol-5% isopropyl alcohol (60: 40) at 25 degrees C. The detection wavelength was 230 nm. RESULT: The linear range was 0.07-0.7 microg for butylidenephthalide. The average recovery was 95.3%, and RSD was 2.3% (n =6). CONCLUSION: This method was simple and could be used to determine butylidenephthalide with satisfactory accuracy and reproducibility.

Ligustilide reduces phenylephrine induced-aortic tension in vitro but has no effect on systolic pressure in spontaneously hypertensive rats.

Radix Angelica sinensis, known as Danggui in Chinese, has been used to treat cardiovascular diseases in traditional Chinese medicine for a long time. Experimental evidence showed that the essential oil of Danggui could reduce blood pressure in rabbits, cats or hypertensive dogs when given intravenously. In this study, we investigated the effects of Z-ligustilide, the main lipophilic component of the essential oil of Danggui on aortic tension induced by phenylephrine, an alpha-adrenergic agonist, in vitro and the systolic blood pressure in SHR rats. We demonstrated for the first time that ligustilide can significantly reduce the phenylephrine-induced aortic tension in vitro with IC(50) about 64 mug/ml, but has no in vivo effect on systolic blood pressure in SHR rats when administrated orally. The data on transport of ligustilide across Caco-2 monolayer suggested an efficient intestinal absorption of ligustilide in vivo, implying that the non-effectiveness of ligustilide in vivo is not due to the poor absorption in the gastrointestinal tract. Further studies on whether ligustilide is one of the main anti-hypertensive components of the essential oil are needed.

Identification of transcriptionally regulated genes in response to cellular iron availability in rat hippocampus.

The present study was attempted to identify transcriptionally regulated genes of the normal neurocytes responsive to iron availability. Postnatal rat hippocampus cells were primarily cultured either under the iron-loaded or depleted conditions. These cultured cells were applied for the generation of subtracted complementary DNA libraries by the suppression subtraction hybridization (SSH) and for the subsequent identification of differentially expressed transcripts by reverse Northern blot. The differentially expressed genes were chosen to perform sequencing, and then some of them were performed by Northern blot analysis for observation of their expression in the hippocampus of rats with the different iron status. The results indicated that five unique transcripts were strong candidates for differential expression in cellular iron repletion, one of them is a novel sequence (GenBank No. AF 433878), while 26 unique transcripts were strong candidates for differential expression in cellular iron deprivation, one of them is a novel sequence (GenBank No. AY 912101). The revealed known genes responsive to iron availability were previously unknown to respond to iron availability, or have not been determined in the brain, have not even been currently determined in their physiological and biological functions. Interestingly, the proteins encoded by most of the known genes are either directly pointed to or indirectly associated with the molecules that play important, even key roles in cellular signal transduction and the cell cycle. These findings lead to the important suggestion that the cellular responses to iron availability involve extensive transcriptional regulation and cellular signal transduction. Therefore, iron may serve as a signal, which directly and/or indirectly regulates or modulates cell functions.

[Experimental study of effect of tanshinone on artery restenosis in rat carotid injury model].

OBJECTIVE: To observe the preventive and therapeutic effect of tanshinone (TA) on artery restenosis in the rat carotid injury model and explor the mechanism. METHOD: Male SD rats were randomly divided into model control group, and low dose, moderate dose and high dose TA groups. Each group had 10 rats. The rats in the high, moderate and low dose groups were respectively fed with TA 120, 40,13.3 mg x kg(-1) x d(-1) by gast rogavage; the rats in the model control group were fed with the same volume solvent. Two days later, the rat's right carotid artery was injuried by balloon dilatation to induce intimal thickening for establishing the restenosis model. After 2 weeks of treatment, the artery was harvested and stained by hematoxylin-elsin (HE) and immunohistochemistry of PCNA, NF-kappaB and iNOS. The morphological changes were checked under microscope. The area of the intimal and medial layer of the vessels, and their ratios were analyzed with image analysis software. The expression level of PCNA, NF-kappaB and iNOS were used as the positive index. RESULT: The intimal area and intima-to-media ratio of the injuried artery increased obviously, suggesting the model was successful. Compared with the model group, TA significantly decreased the intimal area and intima-to-media ratio (P < 0.05), and also decreased the positive index of PCNA and the positive ratio of NF-kappaB and iNOS (P < 0.05). CONCLUSION: TA can effectively inhibit intimal thickening and inflammation. This result suggestes that TA may play a positive role in the prevention of restenosis after PTCA.

[The new progress of the study about volatile oil of the angelica].

To summarize the new progress of the study about volatile oil of the angelica, including the distillable methods, the analysis of the chemical components, the pharmacological effects and the clinical applications. We tracked and searched the correlative references and study reports about volatile oil of the angelica in CNKI data base(1994-2004) and Medline data base (1997-2004). We summarized and compared the different distillable methods of volatile oil of the angelica, meanwhile we summarized many study reports about the analysis of the chemical components of volatile oil of the angelica and it's pharmacological effects, including the toxicity of the volatile oil and it's effects on the uterus smooth muscle, cardiovascular system, respiratory system, central nerve system and immune system. Finally we summarized the clinical application of the volatile oil of the angelica. There are three distillable methods of volatile oil of the angelica . The harvest efficiency of volatile oil is different with different distillable methods. The chemical components are very complicated and the new chemical components are separated and identified. The volatile oil has bidirectional effects on the uterus smooth muscle. It can inhibit the contraction of the uterus smooth muscle induced by different mechanisms. Meanwhile it can depress the blood pressure and ameliorate the cardiac ischemia. The volatile oil can resist the arrhythmia and asthma, restrain the central system, improve the immune function. Nowadays the volatile oil of the angelica is applied to therapy the dysmenorrhea and disorder of the catamenia. The chemical components of the volatile oil of the angelica are very complicated, moreover the pharmacological effects of the volatile oil are comprehensive. People make the new progress of the study about volatile oil of the angelica.

Tanshinone inhibits intimal hyperplasia in the ligated carotid artery in mice.

Vascular smooth muscle cell (VSMC) proliferation is considered to play a central role in the development of intimal hyperplasia with pathological artery healing. Danshen, the Salvia miltiorrhiza Bge., has long been regarded as an effective traditional Chinese medicine for cardiovascular diseases. In this paper, the effects of tanshinone (TA), the lipid-soluble pharmacological constituents of danshen, on the intima hyperplasia and proliferating state of VSMC were described in a mouse carotid artery injured by complete cessation of blood flow. This study showed that oral administration of TA could significantly decrease the intimal thickening of injured vessels and proliferating cell nuclear antigen (PCNA)-positive VSMC in intimal area. These results suggested that the suppressive effects of TA on intimal hyperplasia might partly result from its inhibitory effect against VSMC proliferation.

[The experimental study of the effect of tanshinone on artery restenosis in mouse].

OBJECTIVE: To observe the preventive and therapeutic effect of tanshinone (TA) on artery restenosis in mice and primarily explore the mechanism. METHOD: Female KM mice were randomly divided into model control, low dose and high dose TA groups. Each group had 12 mice. The low and high dose drug groups were respectively given TA 3 and 6 g x kg(-1) x d(-1) by ig; the model control group was given the same volume solvent. The controlateral carotid of ligated artery of model control group was regarded as normal control. 2 days later, the mice' s left common carotid artery was dissected and ligated near the carotid bifurcation, leading to intima hyperplasia and then establishing restenosis model. 4 weeks later, the artery was harvested and stained by hematoxylin-elsin (HE) and immunohistochemistry of PCNA. The morphological changes were checked under microscope; the area of the intimal and medial layer of the vessels, and their ratios were analyzed with image analysis software. The expression level of PCNA was expressed as the positive index. RESULT: Compared with those of normal artery, the intimal area, media area and intima-to-media ratio of ligated artery increased obviously (P < 0.01). But TA could significantly decrease all of these parameters (P < 0.01), and also decrease the positive index of PCNA (P < 0.01). CONCLUSION: TA effectively inhibits intima hyperplasia, which is mainly characterized with the proliferation and migration of smooth muscle cell induced by abnormal hemodynamic changes. This result suggestes that TA may play a positive role in the prevention of restenosis after PTCA.