Lysophosphatidic Acid Receptor 1 Plays a Pathogenic Role in Permanent Brain Ischemic Stroke by Modulating Neuroinflammatory Responses.

Lysophosphatidic acid receptor 1 (LPA1) plays a critical role in brain injury following a transient brain ischemic stroke. However, its role in permanent brain ischemic stroke remains unknown. To address this, we investigated whether LPA1 could contribute to brain injury of mice challenged by permanent middle cerebral artery occlusion (pMCAO). A selective LPA1 antagonist (AM152) was used as a pharmacological tool for this investigation. When AM152 was given to pMCAO-challenged mice one hour after occlusion, pMCAO-induced brain damage such as brain infarction, functional neurological deficits, apoptosis, and blood-brain barrier disruption was significantly attenuated. Histological analyses demonstrated that AM152 administration attenuated microglial activation and proliferation in injured brain after pMCAO challenge. AM152 administration also attenuated abnormal neuroinflammatory responses by decreasing expression levels of pro-inflammatory cytokines while increasing expression levels of anti-inflammatory cytokines in the injured brain. As underlying effector pathways, NF-κB, MAPKs (ERK1/2, p38, and JNKs), and PI3K/Akt were found to be involved in LPA1-dependent pathogenesis. Collectively, these results demonstrate that LPA1 can contribute to brain injury by permanent ischemic stroke, along with relevant pathogenic events in an injured brain.

Tetrahydropiperine, a natural alkaloid with neuroprotective effects in ischemic stroke.

BACKGROUND: Ischemic stroke (IS) is a life-threatening neurological disease with various pathological mechanisms. Tetrahydropiperine (THP) is a natural alkaloid with protective effects against multiple diseases, such as seizure, and pain. This study was to examine the impact of THP on IS and investigate its potential mechanism. MATERIAL AND METHODS: We employed network pharmacology and molecular docking techniques to identify the target proteins of THP for intervention in IS. Adult male Sprague-Dawley rats were used to create a permanent middle cerebral artery occlusion model. PC-12 cells were chosen to establish an oxygen-glucose deprivation (OGD) cell model. Disease modeling followed by nimodipine (NIMO); 3-methyladenine (3-MA) and rapamycin (RAP) interventions. Open field test, Longa score, balance beam test, and forelimb grip test were used to measure motor and neurological functions. The degree of neurological damage recovery was assessed through behavioral analysis, and cerebral infarction volume was determined using TTC staining. Morphological changes were examined through HE and Nissl staining, and ultrastructural changes in neurons were observed using transmission electron microscopy. The protein expression of autophagy and related pathways was analyzed through Western blot (WB). The appropriate hypoxia time and drug concentration were determined using CCK-8 assay, which also measured cell survival rate. RESULTS: The network pharmacology findings indicated that the impact of THP on IS was enhanced in the PI3K/Akt signaling pathway. THP demonstrated robust docking capability with proteins associated with the autophagy and PI3K/Akt/mTOR, as indicated by the molecular docking outcomes. THP significantly improved behavioral damage, reduced the area of cerebral infarction, ameliorated histopathological damage from ischemia, increase neuronal survival, and alleviated ultrastructural damage in neurons (P < 0.05). THP enhanced the survival of PC-12 cells induced by OGD and ameliorated the morphological harm to the cells (P < 0.05). THP was found to elevate the quantities of P62, LC3-Ⅰ, PI3K, P-AKt/Akt, and P-mTOR/mTOR proteins while reducing the levels of Atg7 and Beclin1 proteins. The results of transmission electron microscopy showed no autophagosomes in the THP, 3-MA, and 3-MA + THP groups. CONCLUSION: The activation of the PI3K/Akt/mTOR signaling pathway by THP inhibits autophagy and provides relief from neurological damage in IS.

Stem-Cell-Based Therapy: The Celestial Weapon against Neurological Disorders.

Stem cells are a versatile source for cell therapy. Their use is particularly significant for the treatment of neurological disorders for which no definitive conventional medical treatment is available. Neurological disorders are of diverse etiology and pathogenesis. Alzheimer's disease (AD) is caused by abnormal protein deposits, leading to progressive dementia. Parkinson's disease (PD) is due to the specific degeneration of the dopaminergic neurons causing motor and sensory impairment. Huntington's disease (HD) includes a transmittable gene mutation, and any treatment should involve gene modulation of the transplanted cells. Multiple sclerosis (MS) is an autoimmune disorder affecting multiple neurons sporadically but induces progressive neuronal dysfunction. Amyotrophic lateral sclerosis (ALS) impacts upper and lower motor neurons, leading to progressive muscle degeneration. This shows the need to try to tailor different types of cells to repair the specific defect characteristic of each disease. In recent years, several types of stem cells were used in different animal models, including transgenic animals of various neurologic disorders. Based on some of the successful animal studies, some clinical trials were designed and approved. Some studies were successful, others were terminated and, still, a few are ongoing. In this manuscript, we aim to review the current information on both the experimental and clinical trials of stem cell therapy in neurological disorders of various disease mechanisms. The different types of cells used, their mode of transplantation and the molecular and physiologic effects are discussed. Recommendations for future use and hopes are highlighted.

Piperine ameliorates ischemic stroke-induced brain injury in rats by regulating the PI3K/AKT/mTOR pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Piperine (PIP), a main active component isolated from Piper nigrum L., exerts neuroprotective effects in a rat model of ischemic stroke (IS). However, studies on the effects of PIP on neuroprotection and autophagy after IS are limited. AIM OF THE STUDY: This study aimed to prove the protective effects of PIP against brain IS and elucidate its underlying mechanisms. MATERIALS AND METHODS: Specific pathogen-free male Sprague-Dawley rats were selected to establish a permanent middle cerebral artery occlusion model. The experiment was randomly divided into six groups: sham group, model group, PIP intervention group (10, 20, and 30 mg/kg group), and nimodipine group (Nimo group, 12 mg/kg). Neurological function score, postural reflex score, body swing score, balance beam test, and grip strength test were used to detect behavioral changes of rats. The area of cerebral infarction was detected by TTC staining, and the number and morphological changes of neurons were observed by Nissl and HE staining. In addition, the ultrastructure of hippocampal dentate gyrus neurons was observed using a transmission electron microscope. Western blot was used to detect the expression of PI3K/AKT/mTOR signaling pathway proteins and autophagy-related proteins, namely, Beclin1 and LC3, in the hippocampus and cortex. Cell experiments established an in vitro model of oxygen-glucose deprivation (OGD) with the HT22 cell line to verify the mechanism. The experiment was divided into five groups: control group, OGD group, OGD + PIP 20 µg/mL group, OGD + PIP 30 µg/mL group, and OGD + PIP 40 µg/mL group. CCK-8 was used to measure cell activity, and Western blot was used to measure the expression of PI3K/AKT/mTOR signaling pathway proteins and autophagy-related proteins (Beclin1 and LC3). RESULTS: Compared with the model group, the neurological function scores, body swing scores, and postural reflex scores of rats in the 10, 20, and 30 mg/kg PIP intervention groups and Nimo groups decreased, whereas the balance beam score and grip test scores increased (all p < 0.05). After 10, 20, and 30 mg/kg PIP and Nimo intervention, the cerebral infarction area of pMCAO rats was reduced (p < 0.01), and Nissl and HE staining results showed that the number of neurons survived in the 30 mg/kg PIP and Nimo intervention groups increased. Cell morphology and structure were significantly improved (p < 0.05). Most of the hippocampal dentate gyrus neurons and their organelles gradually returned to normal in the 30 mg/kg PIP and Nimo intervention groups, with less neuronal damage. The expression levels of p-mTOR, p-AKT, and p-PI3K in the hippocampus and cortex of the 30 mg/kg PIP and Nimo intervention groups decreased, whereas the expression level of PI3K increased (all p < 0.05). In addition, the expression level of autophagy-related proteins, namely, Beclin1 and LC3-II, in the 30 mg/kg PIP and Nimo intervention groups decreased (all p < 0.05). Results of CCK-8 showed that after 1 h of OGD, the 30 and 40 µg/mL PIP intervention groups had higher cell viability than the OGD group (p < 0.01). Western blot results showed that compared with the OGD group, the expression level of p-mTOR, p-AKT, and p-PI3K in the 30 and 40 µg/mL PIP intervention groups decreased, and the expression level of PI3K increased (all p < 0.05). Moreover, the expression level of autophagy-related proteins, namely, Beclin1 and LC3-II, in the 30 and 40 µg/mL PIP intervention groups decreased (all p < 0.05). CONCLUSIONS: This study shows that PIP is a potential compound with neuroprotective effects. PIP can inhibit the PI3K/AKT/mTOR pathway and autophagy. Its inhibition of autophagy is possibly related to modulating the PI3K/AKT/mTOR pathway. These findings provide new insights into the use of PIP for the treatment of IS and its underlying mechanism.

Atrial Natriuretic Peptide Antibody-Functionalised, PEGylated Multiwalled Carbon Nanotubes for Targeted Ischemic Stroke Intervention.

Stroke is one of the major causes of disability and the second major cause of death around the globe. There is a dire need for an ultrasensitive detection tool and an effective and efficient therapeutic system for both detection and treatment of stroke at its infancy stage. Carbon nanotubes are promising nanomaterials for tackling these challenges. The loading of dexamethasone and decoration of PEGylated multiwalled carbon nanotube with atrial natriuretic peptide (ANP) antibody and fluorescein isothiocyanate for targeting ischemic site in the rat stroke model is presented here. Functionalisation of carbon nanotubes with dexamethasone (DEX), polyethylene glycol (PEG), fluorescein isothiocyanate (FITC), and ANP antibody caused a 63-fold increase in the D band intensity as illustrated by Raman. The characteristic band intensity increase was observed at 1636 nm following functionalisation of carbon nanotubes with polyethylene glycol and dexamethasone as confirmed by Fourier Transform Infrared. These findings have demonstrated the coupling capability of atrial natriuretic peptide antibody to DEX-PEG-CNTs. The baseline plasma atrial natriuretic peptide levels were ranging from 118 to 135.70 pg/mL prior to surgery and from 522.09 to 552.37 following common carotid artery occlusion. A decrease in atrial natriuretic peptide levels to 307.77 was observed when the rats were treated with FITC-DEX-PEG-ANP-CNTs, PEG-CNTs and DEX with a significant drop in the FITC-DEX-PEG-ANP-CNTs treated group. Fluorescence was detected in FITC-DEX-PEG-CNTs and FITC-DEX-PEG-ANP-CNTs treated ischemic stroke rats. The highest fluorescence intensity was reported in plasma (2179) followed by the kidney (1563) and liver (1507). These findings suggest a beneficial role that is played by the FITC-DEX-PEG-ANP-CNTs in the reduction of inflammation in the ischemic stroke induced rats that could induce a successful treatment of ischemic stroke.

Receptor for Advanced Glycation End Products Is Involved in LPA5-Mediated Brain Damage after a Transient Ischemic Stroke.

Lysophosphatidic acid receptor 5 (LPA5) has been recently identified as a novel pathogenic factor for brain ischemic stroke. However, its underlying mechanisms remain unclear. Here, we determined whether the receptor for advanced glycation end products (RAGE) could be involved in LPA5-mediated brain injuries after ischemic challenge using a mouse model of transient middle cerebral artery occlusion (tMCAO). RAGE was upregulated in the penumbra and ischemic core regions after tMCAO challenge. RAGE upregulation was greater at 3 days than that at 1 day after tMCAO challenge. It was mostly observed in Iba1-immunopositive cells of a post-ischemic brain. Suppressing LPA5 activity with its antagonist, TCLPA5, attenuated RAGE upregulation in the penumbra and ischemic core regions, particularly on Iba1-immunopositive cells, of injured brains after tMCAO challenge. It also attenuated blood-brain barrier disruption, one of the core pathogenesis upon RAGE activation, after tMCAO challenge. As an underlying signaling pathways, LPA5 could contribute to the activation of ERK1/2 and NF-κB in injured brains after tMCAO challenge. Collectively, the current study suggests that RAGE is a possible mediator for LPA5-dependent ischemic brain injury.

Hesperidin reduces adverse symptomatic intracerebral hemorrhage by promoting TGF-ß1 for treating ischemic stroke using tissue plasminogen activator.

Treatment with recombinant tissue plasminogen activator (rt-PA) is the most effective therapeutic option against brain ischemic stroke at the present time. However, elevated incidence of symptomatic intracerebral hemorrhage (SIH) greatly hinders ideal treatment outcome of rt-PA. We sought to assess the impacts of hesperidin on SIH following rt-PA therapies. Patients with ischemic stroke were assigned into two groups in a random fashion, to receive either rt-PA + placebo (Pc) or rt-PA + hesperidin. Treatment outcome was evaluated 24 h after the initial reperfusion using the transcranial Doppler ultrasonography (TCD) and the NIH Stroke Scale (NIHSS). Further, serum concentrations of transforming growth factor (TGF)-ß1, matrix metalloproteinase (MMP)-2, and MMP-9 were examined. Following the initial administration, stroke patients continued to receive either daily Pc or daily hesperidin, and the treatment outcome after 7 days was examined using the TCD, NIHSS, Glasgow Outcome Scale (GOS), and the Modified Rankin Scale (MRS). Combined treatment of rt-PA with hesperidin yielded significant improvement of outcomes, as revealed by better TCD and NIHSS scores as well as decreased SIH incidences, which could be attributable to elevation of TGF-ß1 and reduction in serum levels of both MMP-2 and MMP-9 caused by hesperidin. Follow-up hesperidin treatment for 7 consecutive days also markedly enhanced the recovery of stroke patients, as indicated by TCD, MRS, GOS, and NIHSS. Findings of the present study strongly suggested potential clinical application of hesperidin supplement in rt-PA therapies to reduce SIH and thereby improve the treatment outcomes of rt-PA in patients with ischemic stroke.

Echinochrome A Attenuates Cerebral Ischemic Injury through Regulation of Cell Survival after Middle Cerebral Artery Occlusion in Rat.

Of late, researchers have taken interest in alternative medicines for the treatment of brain ischemic stroke, where full recovery is rarely seen despite advanced medical technologies. Due to its antioxidant activity, Echinochrome A (Ech A), a natural compound found in sea urchins, has acquired attention as an alternative clinical trial source for the treatment of ischemic stroke. The current study demonstrates considerable potential of Ech A as a medication for cerebral ischemic injury. To confirm the effects of Ech A on the recovery of the injured region and behavioral decline, Ech A was administered through the external carotid artery in a rat middle cerebral artery occlusion model after reperfusion. The expression level of cell viability-related factors was also examined to confirm the mechanism of brain physiological restoration. Based on the results obtained, we propose that Ech A ameliorates the physiological deterioration by its antioxidant effect which plays a protective role against cell death, subsequent to post cerebral ischemic stroke.

Multiple Combination of Angelica gigas Extract and Mesenchymal Stem Cells Enhances Therapeutic Effect.

Alternative medicines attract attention because stroke is rarely expected to make a full recovery with the most advanced medical technology. Angelica gigas (AG) is a well-known herbal medicine as a neuroprotective agent. The present study introduced mesenchymal stem cells (MSCs) to identify for the advanced treatment of the cerebrovascular disease. The objective of this research is validation of the enhanced effects of multiple combined treatment of AG extract with MSCs on stroke through angiogenesis. Our results confirmed that AG extract with MSCs improved the neovascularization increasing expression of angiogenesis-regulated molecules. The changes of brain and the behavioral ability showed the increased effects of AG extract with MSCs. As a result, AG extract and MSCs may synergistically increase the therapeutic potential by enhancing neovascularization. This mixed approach provides a new experimental protocol of herbal medicine therapy for the treatment of a variety of diseases including stroke, trauma, and spinal cord injury.

Computational Fluid Dynamics as an Engineering Tool for the Reconstruction of Hemodynamics after Carotid Artery Stenosis Operation: A Case Study.

Background and objectives: Brain ischemic stroke is caused by impaired or absolutely blocked blood flow into the brain regions. Despite the large number of possible origins, there is no general strategy for preventive treatment. In this paper, we aimed to predict the hemodynamics in a patient who experienced a critical stenosis operation in the carotid artery. This is a unique study where we used medical data together with the computational fluid (CFD) technique not to plan the surgery, but to predict its outcome. Materials and Methods: AngioCT data and blood perfusion of brain tissue (CT-perfusion) together with CFD technique were applied for stroke formation reconstruction in different clinical conditions. With the use of self-made semiautomatic algorithm for image processing and 3DDoctror software, 3D-vascular geometries before and after surgical intervention were reconstructed. As the paper is focused on the analysis of stroke appearance, apparent stroke was simulated as higher and lower pressure values in the cranial part due to different outcomes of the surgical intervention. This allowed to investigate the influence of spatial configuration and pressure values on blood perfusion in the analyzed circulatory system. Results: Application of CFD simulations for blood flow reconstruction for clinical conditions in the circulatory system accomplished on average 98.5% and 98.7% accuracy for CFD results compared to US-Doppler before and after surgical intervention, respectively. Meanwhile, CFD results compared to CT-perfusion indicated an average 89.7% and 92.8% accuracy before and after surgical intervention, respectively. Thus, the CFD is a reliable approach for predicting the patient hemodynamics, as it was confirmed by postoperative data. Conclusions: Our study indicated that the application of CFD simulations for blood flow reconstruction for clinical conditions in circulatory system reached 98% and 90% accuracy for US-Doppler and CT-perfusion, respectively. Therefore, the proposed method might be used as a tool for reconstruction of specific patients' hemodynamics after operation of critical stenosis in the carotid artery. However, further studies are necessary to confirm its usefulness in clinical practice.

Salvia miltiorrhiza enhances the survival of mesenchymal stem cells under ischemic conditions.

OBJECTIVES: To validate the enhanced therapeutic effect of Salvia miltiorrhiza Bunge (SM) for brain ischemic stroke through the anti-apoptotic and survival ability of mesenchymal stem cells (MSCs). METHODS: The viability and the expression level of cell apoptotic and survival-related proteins in MSCs by treatment of SM were assessed in vitro. In addition, the infarcted brain region and the behavioural changes after treatment of MSCs with SM were confirmed in rat middle cerebral artery occlusion (MCAo) models. KEY FINDINGS: We demonstrated that SM attenuates apoptosis and improves the cell viability of MSCs. In the rat MCAo model, the recovery of the infarcted region and positive changes of behaviour are observed after treatment of MSCs with SM. CONCLUSIONS: The therapy using SM enhances the therapeutic effect for brain ischemic stroke by promoting the survival of MSCs. This synergetic effect thereby proposes a new experimental approach of traditional Chinese medicine and stem cell-based therapies for patients suffering from a variety of diseases.

Tissue plasminogen activator deficiency preserves neurological function and protects against murine acute ischemic stroke.

BACKGROUND: We tested the hypothesis that tissue plasminogen activator (tPA) deficiency protected against acute ischemic stroke (AIS)-induced brain injury. METHODS AND RESULTS: Wild-type mice (n=54) were categorized into group 1 (sham control, n=18) and group 3 [AIS by permanent ligation of left common carotid artery (CCA) and cramping right CCA for 1h and then reperfusion followed by hypoxia (11% of oxygen supply for 2h), n=36]. Similarly, tPA knockout (tPA(-/-)) mice (n=54) were randomized into group 2 (sham control, n=18) and group 4 (AIS, n=36). By day 28 after AIS procedure, mortality rate was higher in group 3 (77.8%) than in group 4 (38.9%) and lowest in groups 1 (0%) and 2 (0%) (p<0.001). By days 3 and 28, MRI demonstrated a pattern of changes in brain-infarct volume identical to that of mortality among four groups (p<0.001). By day 28, protein expressions of inflammatory (MMP-9, TNF-α, NF-κB, iNOS, PAI-1, RANTES), oxidative (NOX-1, NOX-2, oxidized protein), apoptotic (cleaved caspase-3 & PARP, Bax), and fibrotic (Smad3, TGF-ß) biomarkers and cellular expressions of inflammation (CD11, F4/80, GFAP), DNA-damage (γ-H2AX) and brain-edema (AQP4) markers exhibited an identical pattern compared to that of mortality (all p<0.001), whereas protein expressions of endothelial (eNOS, CD31), anti-fibrotic (Smad1/5, BMP-2) biomarkers, and number of small vessels displayed an opposite pattern (all p<0.001) among four groups. Expressions of protein and cellular angiogenesis markers (VEGF, SDF-1α, CXCR4) were progressively increased from groups 1 and 2 to group 4 (all p<0.0001). CONCLUSION: tPA deficiency protected the brain from AIS injury.

Ginsenoside Represses Symptomatic Intracerebral Hemorrhage after Recombinant Tissue Plasminogen Activator Therapy by Promoting Transforming Growth Factor-ß1.

BACKGROUND: Currently, the most effective treatment for brain ischemic stroke is recombinant tissue plasminogen activator (rt-PA); however, increased incidence of symptomatic intracerebral hemorrhage severely reduced its favorable treatment outcome. METHODS: We aimed to investigate the effect of ginsenoside (Gs) on symptomatic intracerebral hemorrhage after rt-PA treatment. Stroke patients were randomly divided into 2 treatment groups, one receiving rt-PA + placebo (Pc) and the other rt-PA + Gs. Twenty-four hours after the treatment, outcomes were assessed with transcranial Doppler (TCD) ultrasonography and National Institutes of Health Stroke Scale (NIHSS), and plasma levels of transforming growth factor-ß1 (TGF-ß1), matrix metalloproteinase (MMP)-2, and MMP-9 were also measured. After initial cotreatment, the patients were continuously administered with either Pc or Gs, and the treatment outcomes at 7 days were assessed with TCD, NIHSS, modified Rankin scale (MRS), and Glasgow outcome scale (GOS). RESULTS: Cotreatment of rt-PA with Gs significantly improved outcomes in patients compared to the Pc group, as indicated by improved TCD and NIHSS scores and reduced incidence of symptomatic intracerebral hemorrhage, which could be attributed to a Gs-induced increase in TGF-ß1 and a decrease in both MMP-2 and MMP-9 serum levels. Seven days of Gs treatment also significantly improved outcomes in patients compared to the Pc group, assessed by TCD, NIHSS, MRS, and GOS. CONCLUSION: Our study supports the clinical use of Gs as a potential supplement with rt-PA treatment, which reduces symptomatic intracerebral hemorrhage, therefore improving the treatment outcome of stroke patients.

Study on the correlation of duration of reproductive period and prognosis of acute ischemic stroke / 中华神经科杂志

Objective To study a possible correlation between the duration of reproductive period ( from puberty to menopause) and the prognosis of ischemic stroke. Methods Female in-patients with acute ischemic stroke confirmed by CT/MRI in the Neurology Department of the First Affiliated Hospital of Zhengzhou University from 09/03/2006 to 08/30/2008 were enrolled in this study. The probable risk factors of prognosis were analyzed and recovery was assessed by modified Rankin score (MRS) at 6 months followup. Multivariate Logistic regression was used for statistic analysis. Results 371 female patients were enrolled. The average age was (66. 2 ± 10. 0) years; average menopause age was (48. 5 ± 3.9 ) years and average duration of reproductive period was (33.3 ± 4. 3) years. There is a negative correlation between the duration and MRS (OR =0. 285, 95% CI: 0. 095-0. 850, P =0.024). There is no correlation between menopause age and prognosis of stroke. Conclusions Duration of reproductive period is a predictor for prognosis of ischemic stroke. Patients with longer reproductive period have better prognosis.