Naoxintong Capsule for Secondary Prevention of Ischemic Stroke: A Multicenter, Randomized, and Placebo-Controlled Trial.

OBJECTIVE: To examine whether the combination of Naoxintong Capsule with standard care could further reduce the recurrence of ischemic stroke without increasing the risk of severe bleeding. METHODS: A total of 23 Chinese medical centers participated in this trial. Adult patients with a history of ischemic stroke were randomly assigned in a 1:1 ratio using a block design to receive either Naoxintong Capsule (1.2 g orally, twice a day) or placebo in addition to standard care. The primary endpoint was recurrence of ischemic stroke within 2 years. Secondary outcomes included myocardial infarction, death due to recurrent ischemic stroke, and all-cause mortality. The safety of drugs was monitored. Results were analyzed using the intention-to-treat principle. RESULTS: A total of 2,200 patients were enrolled from March 2015 to March 2016, of whom 143 and 158 in the Naoxintong and placebo groups were lost to follow-up, respectively. Compared with the placebo group, the recurrence rate of ischemic stroke within 2 years was significantly lower in the Naoxintong group [6.5% vs. 9.5%, hazard ratio (HR): 0.665, 95% confidence interval (CI): 0.492-0.899, P=0.008]. The two groups showed no significant differences in the secondary outcomes and safety, including rates of severe hemorrhage, cerebral hemorrhage and subarachnoid hemorrhage (P>0.05). CONCLUSION: The combination of Naoxintong Capsule with standard care reduced the 2-year stroke recurrence rate in patients with ischemic stroke without increasing the risk of severe hemorrhage in high-risk patients. (Trial registration No. NCT02334969).

Bilobalide inhibits inflammation and promotes the expression of Aß degrading enzymes in astrocytes to rescue neuronal deficiency in AD models.

The pathogenesis of Alzheimer's disease (AD) involves multiple cell types including endothelial cells, glia, and neurons. It suggests that therapy against single target in single cell type may not be sufficient to treat AD and therapies with protective effects in multiple cell types may be more effective. Here, we comprehensively investigated the effects of bilobalide on neuroinflammation and Aß degrading enzymes in AD cell model and mouse model. We find that bilobalide inhibits Aß-induced and STAT3-dependent expression of TNF-α, IL-1ß, and IL-6 in primary astrocyte culture. Bilobalide also induces robust expression of Aß degrading enzymes like NEP, IDE, and MMP2 to facilitate astrocyte-mediated Aß clearance. Moreover, bilobalide treatment of astrocyte rescues neuronal deficiency in co-cultured APP/PS1 neurons. Most importantly, bilobalide reduces amyloid and inflammation in AD mouse brain. Taken together, the protective effects of bilobalide in in vitro cultures were fully recapitulated in in vivo AD mouse model. Our study supports that bilobalide has therapeutic potential for AD treatment.

Correction: Melatonin-induced ApoE expression in mouse astrocytes protects endothelial cells from OGD-R induced injuries.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Melatonin-induced ApoE expression in mouse astrocytes protects endothelial cells from OGD-R induced injuries.

Stroke is a leading reason of death and long-term disability, and most studies mainly focus on efforts to protect neurons. However, failed clinical trials suggest that therapies against single target in neurons may not be sufficient and the involvement of endothelial cells and glial cells have been underestimated. Astrocytes are the major source of ApoE in the brain and endothelial cells express high level of ApoE receptors. Thus, ApoE may mediate the interaction between astrocytes and endothelial cells. To address whether and how ApoE-mediated astrocytes-endothelial cells interaction contributes to the pathogenesis of stroke, we used oxygen and glucose deprivation-reoxygenation (OGD-R) as a stroke model and investigated the effects of OGD-R on astrocytes-endothelial cell co-cultures in the current study. We find that OGD-R leads to various damages to endothelial cells, including compromised cell viability, increased ROS level, enhanced caspase activity, and higher apoptotic rate. Meanwhile, mouse astrocytes could secrete ApoE to activate PI3K/eNOS signaling in endothelial cells to prevent OGD-R induced injuries. In addition, OGD-R induces down-regulation of ApoE in astrocyte-endothelial cell co-cultures while melatonin restores astrocytic ApoE expression via pCREB pathway and protects endothelial cell in OGD-R treated co-cultures. Our study provides evidence that astrocytes could protect endothelial cells via ApoE in OGD-R condition and Melatonin could induce ApoE expression to protect endothelial cells.

DNA Aptamers Targeting BACE1 Reduce Amyloid Levels and Rescue Neuronal Deficiency in Cultured Cells.

ß-amyloid (Aß) plays an essential role in the pathogenesis of Alzheimer's disease (AD). Beta-site amyloid precursor protein cleaving enzyme 1 (BACE1) is indispensable for Aß production, and knockout of BACE1 has no overt phenotypes in mouse. Thus, fine modulation of BACE1 may be a safe and effective treatment for AD patients. However, the large active site of BACE1 makes it challenging to target BACE1 with classical small-molecule inhibitors. DNA aptamer can have high affinity and specificity against diverse targets, and it provides an alternative strategy to target BACE1. In this study, we used a novel cell-systematic evolution of ligands by exponential enrichment (SELEX) strategy to select specific DNA aptamers optimized to target BACE1 under physiological status. After 17 rounds of selection, we identified two DNA aptamers against BACE1: BI1 and BI2. The identified aptamers interacted with BACE1 in pull-down assay, inhibited BACE1 activity in in vitro fluorescence resonance energy transfer (FRET) assay and HEK293-APP stable cell line, reduced Aß in the culture medium of HEK293-amyloid protein precursor (APP) stable cell line and APP-PS1 primary cultured neurons, and rescued Aß-induced neuronal deficiency in APP-PS1 primary cultured neurons. In contrast, the identified aptamers had no effect on α- or γ-secretase. In addition, cholesteryl tetraetylene glycol (TEG) modification further improved the potency of the identified aptamers. Our study suggests that it is feasible and effective to target BACE1 with DNA aptamers, and the therapeutic potential of the identified aptamers deserves further investigation.

Observational Study of Chinese Medicine Syndrome Distribution in Patients with Acute Myocardial Infarction and Its Impact on Prognosis.

OBJECTIVE: To investigate the distribution of Chinese medicine (CM) syndrome in patients with acute myocardial infarction (AMI) on admission and its impact on prognosis. METHODS: A total of 525 AMI patients were prospectively recruited and classifified into 4 groups based on their clinical characteristics: excess-heat, excess-cold, deficiency-heat and deficiency-cold syndromes. Major adverse cardiovascular events (MACEs) were followed up. RESULTS: The excess syndrome was more common than deficiency syndrome (72.95% vs. 27.05%; P<0.05). Totally 495 (94.29%) of 525 AMI patients were followed up (median 277 days). There were 59 (11.92%) MACEs. After adjusted with confounding factors in Cox regression models, the hazard ratio (95% confifidence interval) of excess-heat, excess-cold, defificiency-heat and defificiency-cold syndrome groups were 1, 1.25 (0.63, 2.49; P<0.05), 2.37 (1.14, 4.94; P<0.05), 3.76 (1.71, 8.28; P<0.05), respectively. CONCLUSIONS: Excess syndrome was more common in AMI patients and had better prognosis, while defificiency-cold syndrome had the poorest prognosis. CM syndrome was of value in predicting long-term outcomes in AMI patients.

Subcutaneous rotenone rat model of Parkinson's disease: Dose exploration study.

Subcutaneous administration of rotenone has recently attracted attention because of its convenience, simplicity and efficacy in replicating features of Parkinson's disease (PD) in animal models. However, the wide range of doses reported in the literature makes it difficult to evaluate the effectiveness of this technique objectively. The aim of the present study was to identify the optimum dose of subcutaneous rotenone for establishing a model of PD. We injected male Wistar rats subcutaneously with one of three doses of rotenone (1.5, 2, or 2.5mg/kg) daily for 5 weeks. Rotenone caused a dose-dependent increase in α-synuclein in the substantia nigra. Furthermore, at 2 and 2.5mg/kg, rotenone caused a significant decrease in the number of tyrosine hydroxylase-immunoreactive neurons in the substantia nigra, and dopamine in the striatum. However, mortality at 2.5mg/kg was 46.7%, compared with just 6.7% at 2mg/kg; the high mortality observed at 2.5mg/kg would limit its application. The 2mg/kg dose showed no detrimental effect on body weight after 5 weeks of daily injections. Furthermore, rats in the 2mg/kg group showed a longer latency to descend from a horizontal bar and a grid wall, decreased rearing, and shorter latency to fall from a rotarod than rats that received vehicle or saline. Mitochondrial damage, observed by transmission electron microscopy, was also evident at this dose. Together, our data indicate that daily subcutaneous injection of 2mg/kg rotenone in rats facilitates the formation of α-synuclein and reproduces the typical features of PD, while maintaining low mortality.

MicroRNA-454 regulates stromal cell derived factor-1 in the control of the growth of pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is a highly malignant carcinoma with an extremely high lethality. We recently reported that hypoxia-inducible factor 1 (HIF-1) targets quiescin sulfhydryl oxidase 1 to facilitate PDAC cell growth and invasion. Here, we analyzed the control of another HIF-1 target, stromal cell derived factor-1 (SDF-1), in PDAC cells. We detected significantly more CD68+ macrophages in the PDAC, compared to normal human pancreas (NT). Since macrophages are recruited to the tissue through their expression of CXCR4 in response to SDF-1, we thus examined the SDF-1 levels in the PDAC specimens. Surprisingly, the SDF-1 protein but not mRNA significantly increased in PDAC, compared to NT. Moreover, a SDF-1-targeting microRNA, miR-454, was found to decrease in PDAC. Promoter luciferase assay confirmed that bindings of miR-454 to 3'-UTR of SDF-1 mRNAs inhibited SDF-1 protein translation. Co-culture of bone marrow derived macrophages and miR-454-modified PDAC cells in a transwell migration experiment showed that macrophages migrated less towards miR-454-overexpressing PDAC cells, and migrated more towards miR-454-depleted cells. Implanted miR-454-depleted PDAC cells grew significantly faster than control, while implanted miR-454-overexpressing PDAC cells grew significantly slower than control. Together, our data suggest that miR-454 may regulate SDF-1 in the control of the growth of PDAC.

PI3K/Akt pathway contributes to neuroprotective effect of Tongxinluo against focal cerebral ischemia and reperfusion injury in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Tongxinluo (TXL), a compound prescription, is formulated according to the collateral disease doctrine of traditional Chinese medicine, and is widely used for the treatment of cardio-cerebrovascular diseases in China. AIM OF THE STUDY: We aimed to investigate the neuroprotective effect of TXL on focal cerebral ischemia and reperfusion injury in rats by attenuating its brain damage and neuronal apoptosis, and to assess the potential role of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) pathway in this protection. MATERIALS AND METHODS: Adult Male Sprague-Dawley rats (n=120) were randomly divided into 5 groups: sham, cerebral ischemia and reperfusion (I/R), cerebral ischemia and reperfusion plus TXL (1.6g/kg/day) (TXL1.6), TXL1.6 plus LY294002 and dimethyl sulfoxide (DMSO) (TXL1.6+LY294002), TXL1.6 plus DMSO (TXL1.6+vehicle). Prior to the grouping, TXL1.6 was selected to be the optimal dose of TXL by evaluating the neurological deficits score of five group rats (Sham, I/R, TXL0.4, TXL0.8 and TXL1.6, n=30) at 0, 1, 3, 5, and 7 days after reperfusion. Rats, being subjected to middle cerebral artery occlusion (MCAO) for 90min followed by 24h reperfusion, were the cerebral ischemia/reperfusion models. At 24h after reperfusion, cerebral infarct area was measured via tetrazolium staining and neuronal damage was showed by Nissl staining. The double staining of Terminal deoxynucleotidyl transferase-mediated deoxyuridine 5-triphosphate nick end labeling (TUNEL) staining and immunofluorescence labeling with NeuN, was performed to evaluate neuronal apoptosis. Proteins involved in PI3K/Akt pathway were detected by Western blot. RESULTS: The results showed that TXL markedly improved neurological function, reduced cerebral infarct area, decreased neuronal damage, and significantly attenuated neuronal apoptosis, while these effects were eliminated by inhibition of PI3K/Akt with LY294002. We also found that TXL up-regulated the expression levels of p-PDK1, p-Akt, p-c-Raf, p-BAD and down-regulated Cleaved caspase 3 expression notably, which were partially reversed by LY294002. Additionally, the increment of p-PTEN level on which LY294002 had little effect was also detected in response to TXL treatment. CONCLUSIONS: These findings demonstrated that TXL provided neuroprotection against cerebral ischemia/reperfusion injury and neuronal apoptosis, and this effect was mediated partly by activation of the PI3K/Akt pathway.

Pachymic acid modified carbon nanoparticles reduced angiogenesis via inhibition of MMP-3.

Angiogenesis is a process of new blood vessel generation, which is consistently and robustly correlated with tumor formation, growth, and metastasis. The disruption of angiogenesis, and the imbalanced endothelial remodeling and regression, are the main pathogenesis of malignant tumor. Recently, multi-walled nanotubes (MWNTs) have been proposed as a new tool for drug delivery in cancer treatment, which also displayed anti-angiogenic property. In the present study, we modified MWNTs with pachymic acid (PA) extracted from Heterosmilax chinensis, a traditional Chinese medicine used for cancer treatment, and compared their effects on blood vessel development. MWNTs and PA/MWNTs were evaluated for their influences on chorioallantoic membrane (CAM) vessel morphology and extracellular matrix metalloproteinase-3 (MMP-3) expression, a crucial proteinase associated with tumor metastasis. MWNTs functioned as an inhibitor of forming branch while PA was not able to promote this inhibition. Subsequently, MWNTs suppressed the endothelial cell maturation, accounting for the ceased elongation of CAM blood vessel, while PA/MWNTs increased the suppressive effect, indicating the potential roles of PA in preventing angiogenesis. PA/MWNTs also showed greater anti-angiogenic property as MMP-3 expression in CAM tissue was significantly decreased by PA/MWNTs compared to MWNTs. These results emphasize the anti-angiogenic activities of PA, supporting a new promising therapy for cancer from the perspective of traditional Chinese medicine.

An-Gong-Niu-Huang Wan protects against cerebral ischemia induced apoptosis in rats: up-regulation of Bcl-2 and down-regulation of Bax and caspase-3.

ETHNOPHARMACOLOGICAL RELEVANCE: The An-Gong-Niu-Huang Wan (AGNH), a Chinese traditional medicine, has been used for treatment of cerebral diseases for centuries in China and other Asian countries, and is approved by the State Food and Drug Administration of China for the treatment of stroke. The aim of present study is to test the neuroprotective effects of AGNH on cerebral ischemia in rats and to explore the underlying mechanisms. MATERIALS AND METHODS: 75 Male Sprague-Dawley rats were randomly divided into 5 groups: sham, ischemia-reperfusion (I/R), and I/R plus 0.065 g/kg/d AGNH, 0.125 g/kg/d AGNH and 0.25 g/kg/d AGNH. Cerebral ischemia was induced by 1.5h of middle cerebral artery occlusion (MCAO). Neurological functional deficits were evaluated according to Zea longa׳s score, cerebral infarct area was measured by tetrazolium staining. Cell injury and apoptosis were assessed by Nissl staining and DNA fragmentation assay. The expression of Bax, Bcl-2 and caspase-3 were analyzed by Western blot. RESULTS: Rats subjected to MCAO exhibited worsened neurological score, infarct area, cell damage and apoptosis. These were all attenuated by AGNH (0.125 and 0.25 g/kg/d). Moreover, AGNH reversed cerebral ischemia induced decreases in Bcl-2 expression and increases in Bax and caspase-3 expression. CONCLUSIONS: These results suggest that AGNH exerts neuroprotective effects, and the neuroprotection is likely to relate to depressed Bax/Bcl-2 ratio and caspase-3 level, leading to inhibition of apoptotic cell death.

Xiao-Xu-Ming decoction preserves mitochondrial integrity and reduces apoptosis after focal cerebral ischemia and reperfusion via the mitochondrial p53 pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Xiao-Xu-Ming decoction (XXMD) has been used to treat stroke and other neurological diseases for more than 1000 years. The purpose of this study was to investigate the effects of XXMD on mitochondrial damage and apoptosis after cerebral ischemia and reperfusion. MATERIALS AND METHODS: Male Sprague-Dawley rats were randomly divided into 3 groups: sham, cerebral ischemia and reperfusion (I/R), and cerebral ischemia and reperfusion plus XXMD (60 g/kg/day) (XXMD60). Focal cerebral ischemia and reperfusion models were induced by middle cerebral artery occlusion. Cerebral ischemic injury was evaluated by hematoxylin and eosin staining. Ultrastructural features of mitochondria in the penumbra of the ischemic cortex were analyzed by transmission electron microscopy. Apoptosis was evaluated by terminal deoxynucleotidyl transferase-mediated deoxyuridine 5-triphosphate nick end labeling (TUNEL) staining and cleaved caspase 3 immunohistochemistry. Proteins in the mitochondrial p53 pathway were detected by western blot and immunofluorescence. RESULTS: The results showed that XXMD treatment markedly attenuated ischemic changes, preserved mitochondrial integrity, and significantly reduced apoptosis. In addition, we found that XXMD treatment reduced p53 and Bax levels and increased Bcl-2 levels in mitochondrial fractions. XXMD significantly blocked the release of cytochrome c and Smac/Diablo from mitochondria, and inhibited activation of caspase 9 and caspase 3 in cytoplasmic fractions. Increased expression of c-IAP1 was observed in the XXMD60 group. CONCLUSIONS: The findings demonstrated that XXMD protected mitochondria from ischemic injury and inhibited apoptosis. The mitochondrial p53 pathway could be partially involved in the protective effects.

PI3K/Akt Pathway Contributes to Neurovascular Unit Protection of Xiao-Xu-Ming Decoction against Focal Cerebral Ischemia and Reperfusion Injury in Rats.

In the present study, we used a focal cerebral ischemia and reperfusion rat model to investigate the protective effects of Xiao-Xu-Ming decoction (XXMD) on neurovascular unit and to examine the role of PI3K (phosphatidylinositol 3-kinase)/Akt pathway in this protection. The cerebral ischemia was induced by 90 min of middle cerebral artery occlusion. Cerebral infarct area was measured by tetrazolium staining, and neurological function was observed at 24 h after reperfusion. DNA fragmentation assay, combined with immunofluorescence, was performed to evaluate apoptosis of neuron, astrocyte, and vascular endothelial cell which constitute neurovascular unit. The expression levels of proteins involved in PI3K/Akt pathway were detected by Western blot. The results showed that XXMD improved neurological function, decreased cerebral infarct area and neuronal damage, and attenuated cellular apoptosis in neurovascular unit, while these effects were abolished by inhibition of PI3K/Akt with LY294002. We also found that XXMD upregulated p-PDKl, p-Akt, and p-GSK3 ß expression levels, which were partly reversed by LY294002. In addition, the increases of p-PTEN and p-c-Raf expression levels on which LY294002 had no effect were also observed in response to XXMD treatment. The data indicated the protective effects of XXMD on neurovascular unit partly through the activation of PI3K/Akt pathway.

Xiao-Xu-Ming Decoction Protects against Blood-Brain Barrier Disruption and Neurological Injury Induced by Cerebral Ischemia and Reperfusion in Rats.

Xiao-Xu-Ming decoction (XXMD) is an effective prescription in the treatment of ischemic stroke, but the mechanisms involved are not well known. In the present study, 120 male Sprague-Dawley rats were randomly divided into 5 groups: sham control (sham), ischemia and reperfusion (IR), and IR plus 15, 30, and 60 g/kg/day XXMD. The stroke model was induced by 90 min of middle cerebral artery occlusion followed by reperfusion. The brain lesion areas were evaluated by 2,3,5-triphenyltetrazolium chloride staining, and neurological deficits were observed at different time points after reperfusion. Blood-brain barrier (BBB) disruption was evaluated by assessing brain water content and Evans blue content. Pathological changes in BBB ultrastructure were observed with transmission electron microscopy. MMP-9, -2, and VEGF expression levels were quantitatively determined by western blotting and immunohistochemistry. We found that XXMD (60 g/kg/day) treatment reduced cerebral infarct area, improved behavioral function, and attenuated ultrastructure damage and permeability of BBB following ischemia and reperfusion. Moreover, XXMD downregulated the expression levels of MMP-9, -2, and VEGF. These findings indicate that XXMD alleviates BBB disruption and cerebral ischemic injury, which may be achieved by inhibiting the expression of MMP-9, -2, and VEGF.

Quantitative evaluation of severity of behavioral and psychological symptoms of dementia in patients with vascular dementia.

To quantitatively evaluate severity of behavioral and psychological symptoms of dementia (BPSD) for vascular dementia (VD). Changes of 51 patients with VD in BPSD between the first and 24th week were assessed using the Neuropsychiatric Inventory (NPI) and the behavioral pathology in Alzheimer's disease (BEHAVE-AD) rating scale, in detrended fluctuation analysis (DFA) represented by diurnal activity (DA), evening activity (EA), and nocturnal activity (NA), and the relationships were analyzed. The subscores of activity disturbances, diurnal rhythm disturbances, and anxieties and phobias in the BEHAVE-AD score, and that of agitation, irritability, and sleep disorder in the NPI score were significantly increased compared with the first week, as was for the changes for EA in the DFA value. A linear correlation was observed between the changes of activity disturbances plus anxieties and phobias, and those of DA, and between the development of diurnal rhythm and those of EA, the vehement and autism scores and those of DA, and the difference in sleep disorder scores and those of EA, respectively. Analysis of DA, NA, and EA may reflect the fluctuational degrees of VD-BPSD, can provide a useful assessment of VD-BPSD accompanied by clinical scores for VD.

Associations of GSTM1 and GSTT1 polymorphisms with pancreatic cancer risk: evidence from a meta-analysis.

Glutathione S-transferases (GSTs), including glutathione S-transferase M1 (GSTM1) and glutathione S-transferase T1 (GSTT1), are multifunctional enzymes which play vital roles in the detoxification of a variety of carcinogens. The genetic polymorphisms of GSTM1 and GSTT1 have been implicated in pancreatic cancer risk, but the results of published studies remain conflicting. Thus, a meta-analysis was conducted to estimate the effect of GSTM1 and GSTT1 polymorphisms on the risk of developing pancreatic cancer. A comprehensive search was performed in the PubMed, Embase, Web of Science, and Wanfang databases to identify the available studies on the associations of GSTM1 and GSTT1 polymorphisms with pancreatic cancer risk. The pooled odds ratio (OR) with its corresponding 95 % confidence interval (95 % CI) was used to estimate the associations. Stratified analyses by ethnicity and sensitivity analyses were performed to further identify the relationships. Overall, the null genotype of GSTT1 was associated with an increased risk of pancreatic cancer (OR = 1.61, 95 % CI 1.06-2.44, P OR = 0.025), but similar association was not found between the null genotype of GSTM1 and pancreatic cancer risk. Besides, a significant association of GSTT1 polymorphism with pancreatic cancer risk was identified in Asians (OR = 2.58, 95 % CI 1.67-3.98, P OR < 0.001), but not in Caucasians (OR = 1.16, 95 % CI 0.94-1.43, P OR = 0.170). Sensitivity analyses by sequential omission of individual study confirmed the stability of our results. Meta-analysis of available data thus far shows that the null genotype of GSTT1 is a risk factor for pancreatic cancer, particularly in the Asian population. The currently available data are not sufficient enough to identify the association between the GSTM1 polymorphism and pancreatic cancer risk.

[Protective effects of Chinese herbal medicine Naoshuantong on neurovascular unit in rats with cerebral ischemia/reperfusion injury].

OBJECTIVE: To investigate the protective effects of Naoshuantong, a compound traditional Chinese herbal medicine, on the main components of neurovascular unit in rats with cerebral ischemia/reperfusion injury. METHODS: A total of 30 male Sprague-Dawley rats were randomly divided into sham-operated, ischemia/reperfusion, and ischemia/reperfusion plus Naoshuantong groups. The cerebral ischemia was induced by 1.5 h of middle cerebral artery occlusion, followed by unlocking the thread to induce reperfusion injury. After 24 h of reperfusion, neurological functional deficits were assessed, apoptosis of main components of neurovascular unit was determined by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end-labeling combined with neuronal nuclei antigen, glial fibrillary acidic protein or CD31 immunofluorescence double staining. RESULTS: Naoshuantong significantly reduced neurological functional deficits, and reduced neuron, astrocyte and vascular endothelium cell apoptosis induced by ischemia/reperfusion injury in the border zone of ischemic cortex. CONCLUSION: Naoshuantong is effective in protecting the neurovascular unit including neurons, astrocytes and vascular endothelium cells against cerebral ischemia/reperfusion-induced apoptosis.

Apocynum venetum leaf extract attenuates disruption of the blood-brain barrier and upregulation of matrix metalloproteinase-9/-2 in a rat model of cerebral ischemia-reperfusion injury.

We investigated the neuroprotective effects of Apocynum venetum leaf extract (AVLE) on a rat model of cerebral ischemia-reperfusion injury and explored the underlying mechanisms. Rats were randomly divided into five groups: sham, ischemia-reperfusion, AVLE125, AVLE250, and AVLE500. Cerebral ischemia was induced by 1.5 h of occlusion of the middle cerebral artery. Cerebral infarct area was measured by tetrazolium staining at 24 and 72 h after reperfusion, and neurological function was evaluated at 24, 48 and 72 h after reperfusion. Pathological changes on the ultrastructure of the blood-brain barrier (BBB) were observed by transmission electron microscopy. BBB permeability was assessed by detecting leakage of Evan's blue (EB) dye in brain tissue. The expression and activities of matrix metalloproteinase (MMP)-9/-2 were measured by western blot analyses and gelatin zymography at 24 h after reperfusion. AVLE (500 mg/kg/day) significantly reduced cerebral infarct area, improved recovery of neurological function, relieved morphological damage to the BBB, reduced water content and EB leakage in the brain, and downregulated the expression and activities of MMP-9/-2. These findings suggest that AVLE protects against cerebral ischemia-reperfusion-induced injury by alleviating BBB disruption. This action may be due to its inhibitory effects on the expression and activities of MMP-9/-2.

Apocynum venetum leaf extract protects rat cortical neurons from injury induced by oxygen and glucose deprivation in vitro.

This study aimed to investigate the protective effect of Apocynum venetum leaf extract (AVLE) on an in vitro model of ischemia-reperfusion induced by oxygen and glucose deprivation (OGD) and further explored the possible mechanisms underlying protection. Cell injury was assessed by morphological examination using phase-contrast microscopy and quantified by measuring the amount of lactate dehydrogenase (LDH) leakage; cell viability was measured by XTT reduction. Neuronal apoptosis was determined by flow cytometry, and electron microscopy was used to study morphological changes of neurons. Caspase-3, -8, and -9 activation and Bcl-2/Bax protein expression were determined by Western blot analysis. We report that treatment with AVLE (5 and 50 µg/mL) effectively reduced neuronal cell death and relieved cell injury induced by OGD. Moreover, AVLE decreased the percentage of apoptotic neurons, relieved neuronal morphological damage, suppressed overexpression of active caspase-3 and -8 and Bax, and inhibited the reduction of Bcl-2 expression. These findings indicate that AVLE protects against OGD-induced injury by inhibiting apoptosis in rat cortical neurons by down-regulating caspase-3 activation and modulating the Bcl-2/Bax ratio.