Extracellular vesicle encapsulated Homer1a as novel nanotherapeutics against intracerebral hemorrhage in a mouse model.

Homer1a and A2 astrocytes are involved in the regulation of inflammation induced by intracerebral hemorrhage (ICH). However, there is no anticipated treatment strategy based on the anti-inflammatory effect of Homer1a and A2 astrocytes. Here, we successfully induced A2 astrocytes in vitro, and then we report an efficient method to prepare Homer1a+ EVs derived from A2 astrocytes which making it more stable, safe, and targetable to injured neurons. Homer1a+ EVs promotes the conversion of A1 to A2 astrocytes in ICH mice. Homer1a+ EVs inhibits activation and nuclear translocation of NF-κB, thereby regulating transcription of IL-17A in neurons. Homer1a+ EVs inhibits the RAGE/NF-κB/IL-17 signaling pathway and the binding ability of IL-17A: IL17-AR and RAGE: DIAPH1. In addition, Homer1a+ EVs ameliorates the pathology, behavior, and survival rate in GFAPCreHomer1fl/-Homer1a± and NestinCreRAGEfl/fl ICH mice. Our study provides a novel insight and potential for the clinical translation of Homer1a+ EVs in the treatment of ICH.

D-allose Inhibits TLR4/PI3K/AKT Signaling to Attenuate Neuroinflammation and Neuronal Apoptosis by Inhibiting Gal-3 Following Ischemic Stroke.

BACKGROUND: Ischemic stroke (IS) occurs when a blood vessel supplying the brain becomes obstructed, resulting in cerebral ischemia. This type of stroke accounts for approximately 87% of all strokes. Globally, IS leads to high mortality and poor prognosis and is associated with neuroinflammation and neuronal apoptosis. D-allose is a bio-substrate of glucose that is widely expressed in many plants. Our previous study showed that D-allose exerted neuroprotective effects against acute cerebral ischemic/reperfusion (I/R) injury by reducing neuroinflammation. Here, we aimed to clarify the beneficial effects D-allose in suppressing IS-induced neuroinflammation damage, cytotoxicity, neuronal apoptosis and neurological deficits and the underlying mechanism in vitro and in vivo. METHODS: In vivo, an I/R model was induced by middle cerebral artery occlusion and reperfusion (MCAO/R) in C57BL/6 N mice, and D-allose was given by intraperitoneal injection within 5 min after reperfusion. In vitro, mouse hippocampal neuronal cells (HT-22) with oxygen-glucose deprivation and reperfusion (OGD/R) were established as a cell model of IS. Neurological scores, some cytokines, cytotoxicity and apoptosis in the brain and cell lines were measured. Moreover, Gal-3 short hairpin RNAs, lentiviruses and adeno-associated viruses were used to modulate Gal-3 expression in neurons in vitro and in vivo to reveal the molecular mechanism. RESULTS: D-allose alleviated cytotoxicity, including cell viability, LDH release and apoptosis, in HT-22 cells after OGD/R, which also alleviated brain injury, as indicated by lesion volume, brain edema, neuronal apoptosis, and neurological functional deficits, in a mouse model of I/R. Moreover, D-allose decreased the release of inflammatory factors, such as IL-1ß, IL-6 and TNF-α. Furthermore, the expression of Gal-3 was increased by I/R in wild-type mice and HT-22 cells, and this factor further bound to TLR4, as confirmed by three-dimensional structure prediction and Co-IP. Silencing the Gal-3 gene with shRNAs decreased the activation of TLR4 signaling and alleviated IS-induced neuroinflammation, apoptosis and brain injury. Importantly, the loss of Gal-3 enhanced the D-allose-mediated protection against I/R-induced HT-22 cell injury, inflammatory insults and apoptosis, whereas activation of TLR4 by the selective agonist LPS increased the degree of neuronal injury and abolished the protective effects of D-allose. CONCLUSIONS: In summary, D-allose plays a crucial role in inhibiting inflammation after IS by suppressing Gal-3/TLR4/PI3K/AKT signaling pathway in vitro and in vivo.

d-allose protects brain microvascular endothelial cells from hypoxic/reoxygenated injury by inhibiting endoplasmic reticulum stress.

Ischemic stroke is an acute brain disease with a high mortality rate. Currently, the only effective method is to restore the blood supply. But the inflammation and oxidative stress induced by this approach can damage the integrity of the endothelial system, which hampers the patient's outcome. d-allose has the biological activity to protect against ischemia-reperfusion injury, however, the underlying mechanism remains unclear. Here, brain microvascular endothelial cells (RBMECs) were used as the study material to establish an IR-injury model. Cell viability of RBMECs was suppressed after hypoxia/reoxygenation (H/R) treatment and significantly increased after d-allose supplementation. RNAseq results showed 180 differentially expressed genes (DEGs) between the therapy group (H/R + Dal) and the model group (H/R), of which 151 DEGs were restored to control levels by d-allose. Enrichment analysis revealed that DEGs were mainly involved in protein processing in endoplasmic reticulum. 6 DEGs in the unfolded protein response (UPR) pathway were verified by qRT-PCR. All of them were significantly down-regulated by d-allose, indicating that endoplasmic reticulum stress (ERS) was relieved. In addition, d-allose significantly inhibited the phosphorylation level of eIF2α, a marker of ERS. The downstream molecules of Phosphorylation of eIF2α, Gadd45a and Chac1, which trigger cycle arrest and apoptosis, respectively, were also significantly inhibited by d-allose. Thus, we conclude that d-allose inhibits the UPR pathway, attenuates eIF2α phosphorylation and ERS, restores the cell cycle, inhibits apoptosis, and thus enhances endothelial cell tolerance to H/R injury.

Extended endoscopic endonasal approach for resecting anterior intrinsic third ventricular craniopharyngioma.

Background: The surgical treatment of the extended endoscopic endonasal approach (EEEA) is a safe and effective treatment for suprasellar craniopharyngiomas. However, due to damage to the hypothalamus and third ventricle floor (TVF), EEEA is generally regarded as unsuitable in treating intrinsic third ventricle craniopharyngioma (ITVC) that is entirely within the third ventricle. Until now, there have been only a small number of reports using EEEA to treat TVC via a supra-infrachiasmatic approach. Given that the translamina terminalis (TLT) corridor was used in the transcranial subfrontal approach, EEEA via a suprachiasmatic approach may be feasible and practical to treat ITVC. In the current study, we accumulated experience applying the suprachiasmatic translamina terminalis (STLT) corridor for anterior treatment of ITVC. Methods: From March 2016 to December 2020, 14 patients with ITVC in our center were analyzed retrospectively. All patients underwent surgery by EEEA via an STLT corridor. The multilayer reconstruction technique was adopted to achieve skull base reconstruction. Data concerning the patient's tumor resection, vision, hypophyseal hormone, and complications were collected. Results: Gross-total resection was achieved in 13 (92.8%) of14 patients, with achievement of near-total (90%) resection in the remaining 1 patient. Nine cases (64.3%) were papillary craniopharyngiomas, and the other 5 cases were adamantinomatous subtypes. Postoperatively, 3 patients with pituitary insufficiency received hormone replacement therapy. No permanent diabetes insipidus or hypothalamic obesity was found. All pairs showed significant improvement or stability in vision except 1 patient who encountered visual deterioration. No other neurological deficit occurred postoperatively. Observation results for the exudation of nasal tissue and the length of hospitalization were satisfactory. After a mean follow-up period of 26.2 months, tumor recurrence was not observed. Conclusion: TLT is a minimally invasive corridor used in EEEA for treating anterior ITVC without increasing risks of visual and hormonal deficits. The multilayered reconstruction technique we used is a safe and effective method for achieving watertight closure and avoiding cerebrospinal fluid leaks and infection. The endonasal approach via STLT provides a new, safe and efficacious operative strategy that should be considered a surgical alternative in treating ITVC.

Clinical Efficacy of the Multilayered Skull Base Reconstruction Using In Situ Bone Flap in Endoscopic Endonasal Approach for Craniopharyngioma.

Objective The aim of the study is to summarize and analyze the efficacy of the multilayered skull base reconstruction using in situ bone flap in endoscopic endonasal approach (EEA) for craniopharyngiomas. Methods A retrospective review of 65 patients who underwent resection of their histopathology confirmed craniopharyngiomas performed at a single institution. Based on the team's understanding and mastery of skull base reconstruction techniques, patients were divided into two groups according to the methods of reconstruction in two periods. First (March 2015 through August 2016), osseous reconstruction was not adopted and served as the control group (34 cases). Second (September 2016 through July 2019), in situ bone flap repair of the skull base (complete osseous reconstruction) served as observation group (31 cases). The length of hospitalization and nasal exudation, bed rest time of hospital discharge, the incidence of cerebrospinal fluid leaks, lumbar drainage, and intracranial/pulmonary infections were collected and compared. Results Compared with the control group, patients in the observation group had obviously less lumbar drainage and CSF leakage ( p < 0.05), but had no significant difference in cases of re-operation, meningitis, and pulmonary infection. At the meantime, cases of nasal exudation, bed rest, and hospitalization of the observation group were significantly reduced ( p < 0.05) in the observation group. Conclusion The multilayered reconstruction technique (especially using in situ bone flap, combined with vascularized pedicled nasoseptal flap) is a safe and effective method in achieving watertight closure after EEEA, and can significantly reduce the incidence of cerebrospinal fluid leaks, and facilitate rehabilitation in skull base reconstruction of craniopharyngiomas.

Effect of PITX2 genetic variants on the susceptibility to stroke in the Chinese Han population.

PURPOSE: Stroke is a multifactorial and complex disease caused by the obstruction or rupture of cerebrovascular. To explore the influence of genetic factors on stroke susceptibility, we investigated the association between four single nucleotide polymorphisms (SNPs) in the paired-like homeodomain transcription factor 2 (PITX2) gene and stroke risk. METHODS: A total of 977 volunteers including 476 stroke patients and 501 control individuals were recruited. The association between PITX2 polymorphisms and stroke risk was evaluated using genetic models and haplotype analyses. The strength of the association between each studied polymorphisms and stroke risk was evaluated by calculating odds ratios (ORs) and 95% confidence intervals (CIs). What's more, multifactor dimensionality reduction (MDR) was used to predict the interaction between SNPs. RESULTS: Our study showed that rs6817105 in PITX2 was related to a significant increase in stroke susceptibility (OR = 1.42, 95% CI = 1.04-1.94, p = 0.028). Stratified analyses based on gender indicated that rs6817105, rs13143308, and rs6843082 polymorphisms were significantly associated with an increased risk of stroke in male (OR = 0.68, 95% CI = 0.47-0.99, p = 0.042; OR = 0.53, 95% CI = 0.30-0.96, p = 0.035; and OR = 0.55, 95% CI = 0.30-0.99, p = 0.047). Besides, SNP rs6817105 was significantly increased the risk of stroke in people at age over 65 years (OR = 1.87, 95% CI =1.12-3.11, p = 0.016). MDR showed that the interaction model of rs6817105 and rs3853445 emerged as the best predictor between the PITX2 gene and stroke susceptibility. CONCLUSIONS: This study indicated that there was a significant association between the PITX2 gene and stroke risk, and provided some data as far as possible to support the prevention of stroke.

Pseudocapsule-Based Resection for Pituitary Adenomas via the Endoscopic Endonasal Approach.

INTRODUCTION: The endoscopic endonasal approach (EEA) is a safe and effective treatment for pituitary adenomas (PAs). Since extracapsular resection (ER) of PAs improves tumor resection and endocrine remission rates, the interface between the pseudocapsule and gland draws increasing attention. However, it is difficult to precisely dissect the tumor along the exact boundary, and complete removal of the tumor increases the risks of normal tissue damage and cerebrospinal fluid (CSF) leakage. In this study, we investigated the extracapsular resection as well as the pseudocapsule histology to evaluate the effectiveness and safety of pseudocapsule-related surgical interventions. METHODS: From December 2017 to December 2019, 189 patients of PAs via EEA in our single center were analyzed retrospectively. The images, operative details, and clinical follow-up of patients were collected. Sixty-four patients underwent pseudocapsule-based ER, and 125 patients also underwent traditional intracapsular resection (IR) with or without intensive excision for FPAs. The clinical characteristics, tumor resection, endocrinological outcomes, and postoperative morbidities of the two groups were compared. Informed consent for publication of our article was obtained from each patient. Histological examination of pseudocapsule was performed using hematoxylin and eosin and reticulin staining. RESULTS: The gross total recession was 62 (96.9%) in the ER group and 107 (85.6%) cases in the IR group, whereas the endocrine remission rate was 29/31 (93.5%) and 40/53 (75.5%) cases, respectively. Anterior pituitary functions were not aggravated postoperatively in any patient, but transient diabetes insipidus (DI) occurred more in the IR group (64.0%) than in ER (48.4%). Pseudocapsule specimens were obtained in 93 patients, and clusters of small cell aggregation were detected in 11 pseudocapsule specimens (11.8%) whereas other patients showed no remarkable developed pseudocapsule. Intraoperative CSF leak occurred more in the ER group (28.1%) than in the IR group (13.6%), but no difference was seen between two groups postoperatively. No case of intracranial hematoma or pituitary crisis occurred in both groups. After a mean follow-up of 22.8 months, tumor recurrence was observed in 4 (2.1%) cases. CONCLUSION: Pseudocapsule-based extracapsular resection of PAs via EEA is an effective and safe procedure to achieve complete resection with high and sustained endocrine remission and without deteriorating pituitary function.

Metastatic large cell neuroendocrine lung cancer to the foramen magnum: A case report.

RATIONABLE: Large cell neuroendocrine carcinoma of the lung is rare, especially in the area of the foramen magnum. No previous studies have reported metastatic large cell neuroendocrine lung cancer to the foramen magnum. This paper will be the first time to report this special case. PATIENT CONCERNS: A case of a 37-year-old woman presented with headache that had developed 20 days previously. Imaging examination revealed a circular abnormal signal at the posterior margin of the foramen magnum. DIAGNOSES: The patient we report was diagnosed with a metastatic intracranial tumor. INTERVENTIONS: The patient underwent occipital craniotomy. Pathological results showed metastatic neuroendocrine carcinoma of the brain. Whole body PET-CT examination showed that fusiform soft tissue shadows could be seen near the hilum of the lower lobe of the left lung. OUTCOMES: The final bronchoscopy pathological results showed the large cell neuroendocrine carcinoma of the lung. The patient underwent further chemotherapy and radiotherapy in the oncology department. LESSONS: Diagnosis and treatment of large cell neuroendocrine carcinoma of the lung are difficult. The prognosis is poorer, and effective treatment is urgently needed.

High-Mobility Group Box 1 Neutralization Prevents Chronic Cerebral Hypoperfusion-Induced Optic Tract Injuries in the White Matter Associated with Down-regulation of Inflammatory Responses.

Chronic cerebral hypoperfusion (CCH)-induced white matter lesions (WMLs) are region-specific with the optic tract (OT) displaying the most severe damages and leading to visual-based behavioral impairment. Previously we have demonstrated that anti-high-mobility group box 1 (HMGB1) neutralizing antibody (Ab) prevents CCH-induced hippocampal damages via inhibition of neuroinflammation. Here we tested the protective role of the Ab on CCH-induced OT injuries. Rats were treated with permanent occlusion of common carotid arteries (2-VO) or a sham surgery, and then administered with PBS, anti-HMGB1 Ab, or paired control Ab. Pupillary light reflex examination, visual water maze, and tapered beam-walking were performed 28 days post-surgery to investigate the behavioral deficits. Meanwhile, WMLs were measured by Klüver-Barrera (KB) and H&E staining, and glial activation was further assessed to evaluate inflammatory responses in OT. Results revealed that anti-HMGB1 Ab ameliorated the morphological damages (grade scores, vacuoles, and thickness) in OT area and preserved visual abilities. Additionally, the increased levels of inflammatory responses and expressions of TLR4 and NF-κB p65 and phosphorylated NF-κB p65 (p-p65) in OT area were partly down-regulated after anti-HMGB1 treatment. Taken together, these findings suggested that HMGB1 neutralization could ease OT injuries and visual-guided behavioral deficits via suppressing inflammatory responses.

HMGB1 Neutralization Attenuates Hippocampal Neuronal Death and Cognitive Impairment in Rats with Chronic Cerebral Hypoperfusion via Suppressing Inflammatory Responses and Oxidative Stress.

High-mobility group box-1 (HMGB1) acts as a proinflammatory molecule once released into the extracellular space and inhibition of HMGB1 signaling has been reported be neuroprotective in neurodegenerative diseases. Besides, chronic cerebral hypoperfusion (CCH) causes cognitive impairment in neurodegenerative diseases. Here we tested the protective role of HMGB1 inhibition using anti-HMGB1 neutralizing antibody (Ab) against CCH in rats after bilateral common carotid artery occlusion (2VO). 169 male Sprague-Dawley rats underwent 2VO or sham operation. PBS, anti-HMGB1 Ab (1 mg/kg), or control IgG Ab (1 mg/kg) was intravenously administered post-operation. HMGB1 translocation, blood-brain barrier (BBB) permeability and glial activation were evaluated at 3 d, as well as the levels of inflammatory cytokines and oxidative stress. NeuN immunostaining and Morris Water Maze (MWM) were performed at 3 d, 4 w and 12 w. We found that anti-HMGB1 neutralizing Ab inhibited HMGB1 translocation in hippocampal CA1 subarea and improved hippocampal HMGB1 level. Besides, anti-HMGB1 Ab preserved BBB integrity and reduced glial activation, in association with the related changes in oxidative stress (increased activities of superoxide dismutase (SOD) and catalase (CAT), and decreased malondialdehyde (MDA) production) and inflammatory cytokines (increased gene expression of IL-1ß, IL-6 and TNF) at 3 d. Additionally, anti-HMGB1 neutralizing Ab improved hippocampal CA1 neuronal survival and behavioral outcomes in the chronic phase (4 w and 12 w). Taken together, these findings suggest that HMGB1 neutralization suppresses hippocampal inflammatory responses and oxidative stress in the acute phase, and these changes exert long-lasting beneficial effects in the chronic phase of CCH.

PRL-3 is a potential glioblastoma prognostic marker and promotes glioblastoma progression by enhancing MMP7 through the ERK and JNK pathways.

Purpose: Glioblastoma is the most common and aggressive type of primary brain malignancy and is associated with a poor prognosis. Previously, we found that phosphatase of regenerating liver-3 (PRL-3) was significantly up-regulated in glioblastoma as determined by a microarray analysis. However, the function of PRL-3 in glioblastoma remains unknown. We aimed to investigate the clinical relationship between PRL-3 and glioblastoma, and uncover the mechanisms of PRL-3 in the process of glioblastoma. Methods: PRL-3 expression was evaluated in 61 glioblastoma samples and 4 cell lines by RT-qPCR and immunohistochemistry. Kaplan-Meier analysis was performed to evaluate the prognostic value of PRL-3 for overall survival (OS) and progression-free survival (PFS) for glioblastoma patients. Proliferation was evaluated by Cell Counting Kit-8 (CCK-8) assay and EdU proliferation assay, migration and invasion by wound-closure/Transwell assays, and qRT-PCR/immunoblotting/IHC were used for both in vivo and in vitro investigations. Result: A high PRL-3 expression level was closely correlated with unfavorable OS and PFS for glioblastoma patients, and was also significantly correlated with Ki-67 expression. Down-regulation of PRL-3 inhibited glioma cell proliferation, invasion and migration through ERK/JNK/matrix metalloproteinase 7 (MMP7) in vitro and in vivo. Conclusions: PRL-3 expression enhances the invasion and proliferation of glioma cells, highlighting this phosphatase as a novel prognostic candidate and an attractive target for future therapy in glioblastoma.

Induction of antigen-specific cytotoxic T-cell response by dendritic cells generated from ecto-mesenchymal stem cells infected with an adenovirus containing the MAGE-D4a gene.

The present study aimed to investigate the feasibility of using ecto-mesenchymal stem cell (EMSC)-derived dendritic cells (DCs) for glioma immunotherapy following infection by a recombinant adenovirus containing the melanoma-associated antigen D4a (MAGE-D4a) gene. The ex vivo cultured EMSCs were infected by the adenoviral plasmid containing MAGE-D4a (pAd/MAGE-D4a). Efficiency of transfection was evaluated through the detection of green fluorescent protein-marked MAGE-D4a. The MAGE-EMSCs were induced to differentiate into DCs, termed as MAGE-EMSCs-DCs. The morphology was subsequently analyzed under a microscope, and methyl thiazolyl tetrazolium (MTT) and interferon-γ (IFN-γ) assays were performed to analyze the cytotoxicity of the MAGE-EMSC-DCs on the human glioma U251 cell line. Following purification by magnetic-activated cell sorting, the EMSCs grew into swirls, with a long spindle shape and were fibroblast-like. The gene transfected with recombinant adenovirus vectors maintained high and stable expression levels of MAGE-D4a, and its efficiency was increased in a multiplicity of infection-dependent manner. The results of the MTT assay indicated that the T cells, primed by the recombinant MAGE-D4a-infected EMSC-DCs in vitro, recognized MAGE-D4a-expressing tumor cell lines in a human leukocyte antigen class I-restricted manner, and evoked a higher cytotoxic T cell (CTL) response. The CTL response induced by the MAGE-EMSC-DCs, co-cultured with the U251 cells for 24 h, produced 765.0 pg/ml IFN-γ, which was significantly greater when compared to the control wells. T lymphocytes stimulated by MAGE-EMSC-DCs evoke a higher CTL response to human glioma cell lines, and may serve as a promising therapeutic modality for the treatment of MAGE-D4a-expressing glioma.

D-allose protects the blood brain barrier through PPARγ-mediated anti-inflammatory pathway in the mice model of ischemia reperfusion injury.

Our early experiments confirmed that D-allose was closely involved in the blood brain barrier (BBB) protection from ischemia reperfusion (IR) injury, but the regulatory mechanism is not fully defined. In this study, we aimed to investigate the role of D-allose in the protection of BBB integrity and the relevant mechanisms involved in the mice model of middle cerebral artery occlusion and reperfusion (MCAO/Rep). D-allose was intravenously injected via a tail vein (0.2mg/g and 0.4mg/g, 1h before ischemia), GW9662 was intraperitoneal injected to the mice (4mg/kg) before inducing ischemia 24h. Pretreatment with D-allose ameliorated the neurological deficits, infarct volume and brain edema in brains of MCAO/Rep mice. D-allose inhibited cell apoptosis in the mice model of MCAO/Rep. We observed that D-allose remarkably decreased BBB permeability and prevented the reduction of ZO-1, Occludin and Claudin-5 in mice brains with MCAO/Rep injury. D-allose also repressed the levels of TNF-α, NF-κB, interleukin (IL)-1ß and IL-8 in inflammatory responses. The increases of intercellular adhesion molecular-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and CD11b/CD18 were significantly inhibited by D-allose during the MCAO/Rep injury. And D-allose decreased the L-selectin and P-selectin levels after MCAO/Rep. Moreover, D-allose induced up-regulation of peroxisome proliferator-activated receptor γ (PPARγ), and down-regulation of TNF-α and NF-κB after MCAO/Rep, which were abolished by utilization of GW9662. In conclusion, we provided evidences that D-allose may has therapeutic potential against brain IR injury through attenuating BBB disruption and the inflammatory response via PPARγ-dependent regulation of NF-κB.

[Alarming effect of intraoperative neuroelectrophysiological monitoring in microvascular decompression for primary trigeminal neuralgia].

OBJECTIVE: To explore the alarming effect of intraoperative neuroelectrophysiological monitoring in microvascular decompression (MVD) for primary trigeminal neuralgia. METHODS: In 2014, a total of 44 patients with an initial diagnosis of primary trigeminal neuralgia were consecutively recruited for surgery. And 41 of them with an intraoperative confirmation of primary trigeminal neuralgia underwent MVD. Intraoperative neuroelectrophysiological monitoring was employed for brainstem auditory evoked potentials (BAEPs), spontaneous electromyogram for obicularis oculi, obicularisoris and masseter muscles. The real-time alarming report was offered to the operator who adjusted operations accordingly. RESULTS: There were abnormal changes in 23 cases (56.10%) with a total of 77 instances (BAEPs 27, trigeminal nerve 32, facial nerve 18). The outcomes were no facial pain (n=26), pain relief (n=15) and facial numbness (n=6, two with concurrent hearing disturbance). And the rates of facial pain disappearance and sequela occurrence were much better than those in controls without monitoring. CONCLUSION: Intraoperative neuroelectrophysiological monitoring helps enhance the MVD effect and decrease operative squela through alarming reporting.

Resveratrol inhibits oxygen-glucose deprivation-induced MMP-3 expression and cell apoptosis in primary cortical cells via the NF-κB pathway.

Resveratrol (Res) or trans-3,4',5-trihydroxystilbene, has been proven to exert neuroprotective effects in cerebral ischemia. The aim of the present study was to investigate whether Res has neuroprotective effects in primary cortical neurons subjected to transient oxygen-glucose deprivation (OGD) via inhibiting the expression of the gene encoding stromelysin-1, also known as matrix metalloproteinase-3 (MMP-3), and via inhibiting cell apoptosis. Primary cortical cells were exposed to OGD, followed by reoxygenation to induce transient ischemia. Res (50 µM) was added into the culture medium during transient ischemia in the presence or absence of the nuclear factor (NF)-κB inhibitor pyrrolidine dithiocarbamate (PDTC; 10 µM) or 500 µM of the nitric oxide (NO) donor NOC-18. Cell viability was assessed using the tetrazolium reduction (MTT) assay. Cell apoptosis was evaluated by flow cytometry. MMP-3 expression was analyzed by western blot and reverse transcription-polymerase chain reaction (RT-PCR), while the levels of inducible NO synthase (iNOS), NF-κB, caspase-3, cleaved caspase-3, B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax) were assayed by western blot. NO was detected using a spectrophotometric method. We found that the cellular viability was significantly reduced by transient OGD and that this effect was reversed by Res treatment. In addition, OGD was shown to induce cell apoptosis, the expression of Bax and the activation of caspase-3, and inhibit the expression of Bcl-2, and these effects were also reversed by Res treatment. Res treatment significantly reduced the level of MMP-3 that was induced by transient OGD, via inhibition of NF-κB expression. In addition, Res inhibited iNOS expression and NO synthesis that were induced by OGD. MMP-3 expression induced by NO was attenuated by Res treatment and was partially restored by exogenous NO using NOC-18. Taken together, these findings indicate that OGD induces apoptosis through canonical apoptosis signaling and by modulating the expression of MMP-3; Res can reverse the OGD-induced MMP-3 expression and cell apoptosis via the NF-κB-iNOS/NO pathway. Therefore, Res may be a promising agent for the treatment of neuronal injury associated with stroke.

Resveratrol protects primary cortical neuron cultures from transient oxygen-glucose deprivation by inhibiting MMP-9.

It was recently shown that resveratrol exerts neuroprotective effects against cerebral ischemia in mice. The aim of the present study was to further confirm these effects in in vitro primary cortical neuron cultures with transient oxygen-glucose deprivation (OGD), and to investigate whether these effects are due to the inhibition of matrix metalloproteinase-9 (MMP-9) and of cell apoptosis. Neuronal primary cultures of cerebral cortex were prepared from BALB/c mice embryos (13-15 days). Cells from 14- to 16-day cultures were subjected to OGD for 3 h, followed by 21 h of reoxygenation to simulate transient ischemia. Different doses of resveratrol were added into the culture medium during the simulation of transient ischemia. The effect of the extracellular signal-regulated kinase (ERK) inhibitor U0126 was studied by adding U0126 (5 µg/µl, 4 µl) into the culture medium during transient ischemia; as a control, we used treatment of cells with 50 µM of resveratrol. Cell viability was investigated using the 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) reduction assay. Cell apoptosis was assessed by flow cytometry. The effects of resveratrol on the expression of MMP-9 were analyzed by western blotting and reverse transcription-polymerase chain reaction (RT-PCR), while the levels of ERK, phosphorylated (p)-ERK, cleaved caspase-3, Bax and Bcl-2 were measured by western blotting. The results of the MTT assay showed that cell viability is significantly reduced by transient OGD. OGD induced cell apoptosis, the expression of Bax and the activation of caspase-3 and ERK, inhibited the expression of Bcl-2 and increased the expression of MMP-9, while these effects were reversed by treatment with resveratrol. The therapeutic efficacy of resveratrol was shown to be dose-dependent, with the most suitable dose range determined at 50-100 µM. Treatment with U0126 inhibited MMP-9 and Bax expression and caspase-3 activation, while it further promoted the expression of the anti-apoptotic molecule Bcl-2, suggesting that resveratrol inhibits MMP-9 expression and cell apoptosis by attenuating the activation of ERK1/2. In conclusion, OGD can induce apoptosis through canonical apoptotic signals and by regulating the expression of MMP-9; the anti-apoptotic activity of resveratrol and its inhibitory effect on MMP-9 expression contribute in the reduced activation of ERK.

Allicin protects rat cortical neurons against mechanical trauma injury by regulating nitric oxide synthase pathways.

Allicin, a small molecule that is responsible for the typical smell and most of the functions of garlic, possesses a broad spectrum of pharmacological activities and is considered to have therapeutic potential in many pathologic conditions. In the present study, we investigated the potential protective effect of allicin in an in vitro model of traumatic brain injury (TBI) using primary cultured rat cortical neurons. We found that allicin treatment significantly reduced mechanical trauma-induced lactate dehydrogenase (LDH) release and inhibited apoptotic neuronal death in a dose-dependent manner. These protective effects were observed even if allicin treatment was delayed to 2h after injury. Allicin significantly decreased the expression of inducible nitric oxide synthase (iNOS) and increased the phosphorylation of endothelial NOS (eNOS) but had no effect on neuronal NOS (nNOS) expression. Allicin-induced protection in cortical neurons was augmented by iNOS and nNOS antagonists and was partly reversed by blocking eNOS phosphorylation. In addition, allicin treatment inhibited the TBI-induced activation of ERK and further enhanced the phosphorylation of Akt in TBI-injured neurons. The Akt inhibitor LY294002 attenuated the allicin-induced increase in eNOS expression and phosphorylation, whereas the ERK inhibitor PD98059 had opposite effects on the expression of iNOS and eNOS. Pretreatment with LY294002 or PD98059 partly prevented or further enhanced allicin-induced neuroprotection, respectively. Collectively, these data demonstrate that allicin treatment may be an effective therapeutic strategy for traumatic neuronal injury and that the potential underlying mechanism involves Akt- and ERK-mediated regulation of NOS pathways.

Osthole attenuates spinal cord ischemia-reperfusion injury through mitochondrial biogenesis-independent inhibition of mitochondrial dysfunction in rats.

BACKGROUND: Osthole, the main bioactive compounds isolated from the traditional Chinese medical herb broad Cnidium monnieri (L.) cusson, has been shown to exert spectrum of pharmacologic activities. The aim of this study was to investigate the potential neuroprotective effects of osthole against spinal cord ischemia-reperfusion injury in rats. MATERIALS AND METHODS: Osthole was administrated at the concentration of 0.1, 1, 10, 50, or 200 mg/kg (intraperitoneally) 1 h before spinal cord ischemia. The effects on spinal cord injury were measured by spinal cord water content, infarct volume, hematoxylin and eosin staining, and neurologic assessment. Mitochondria were purified from injured spinal cord tissue to determine mitochondrial function. RESULTS: We found that treatment with osthole (10 and 50 mg/kg) significantly decreased spinal cord water content and infarct volume, preserved normal motor neurons, and improved neurologic functions. These protective effects can be also observed even if the treatment was delayed to 4 h after reperfusion. Osthole treatment preserved mitochondrial membrane potential level, reduced reactive oxygen species production, increased adenosine triphosphate generation, and inhibited cytochrome c release in mitochondrial samples. Moreover, osthole increased mitochondria respiratory chain complex activities in spinal cord tissue, with no effect on mitochondrial DNA content and the expression of mitochondrial-specific transcription factors. CONCLUSIONS: All these findings demonstrate the neuroprotective effect of osthole in spinal cord ischemia-reperfusion injury model and suggest that oshtole-induced neuroprotection was mediated by mitochondrial biogenesis-independent inhibition of mitochondrial dysfunction.

Anti-inflammatory effect of D-allose in cerebral ischemia/reperfusion injury in rats.

D-allose, a type of rare sugar, can produce inhibitory effects on activated leukocytes in various organs, including immunosuppressive effects and anti-inflammatory effects, as well as anti-oxyradical effects. The present experiment was performed to investigate the potential anti-inflammatory effects of D-allose in acute cerebral ischemia/reperfusion (I/R) injury. Transient middle cerebral artery occlusion model was applied in rats. D-allose was administered two times via a tail vein (300 mg/kg, 1 hour before ischemia and 10 hours after reperfusion). After 22 hours of reperfusion following 2 hours of ischemia, brain damage was evaluated by cerebral infarct volume. Myeloperoxidase (MPO) activity assay by enzyme-linked immunosorbent assay, and protein expression of MPO and cyclooxygenase-2 (COX-2) by immunohistochemistry were evaluated to investigate the potential mechanisms of D-allose. The experimental results showed that D-allose exhibited significant neuroprotective effects against acute cerebral I/R injury. The infarct volume in D-allose-treated rats (90.9 ± 13.5 mm(3)) was significantly smaller than that in vehicle rats (114.9 ± 15.3 mm(3), p < 0.01). D-allose treatment significantly suppressed the MPO activity and the number of MPO-positive cells compared with those in the vehicle group, suggesting that treatment with D-allose can reduce the infiltration of leukocytes into the ischemic tissue. Treatment of D-allose also significantly decreased the number of COX-2-positive cells and microglial activation in the ischemic tissue. The present results demonstrate that D-allose exerts potent neuroprotective effects against acute cerebral I/R injury, and constitute the first evidence of anti-inflammatory effects of D-allose which considerably contributes to the beneficial effects. Treatment with D-allose might provide a new strategy and clinically beneficial outcome for acute ischemic stroke.

Protective effects of hydrogen sulfide in a rat model of traumatic brain injury via activation of mitochondrial adenosine triphosphate-sensitive potassium channels and reduction of oxidative stress.

BACKGROUND: Hydrogen sulfide (H2S) is considered an important neuromodulator in the central nervous system. We designed the present study to investigate the effects of exogenous H2S in a rat model of traumatic brain injury (TBI) and the mechanism(s) that underlie this effect. METHODS: We induced a TBI model by controlled cortical impact injury. We intraperitoneally administered sodium hydrosulfide (NaHS) (an H2S donor) (3 mg/kg) or vehicle alone at 5 min after a TBI operation. We then measured the H2S level, brain edema, blood-brain barrier integrity, neurologic dysfunction, and lesion volume in all animals. Moreover, we assessed the role of mitochondrial adenosine triphosphate-sensitive potassium (mitoKATP)channels by intraperitoneal injection of the selective blocker 5-hydroxydecanoate before NaHS administration. In addition, we detected the levels of oxidative products and the activities of antioxidant enzymes in brain tissue. RESULTS: Administration of NaHS significantly increased the H2S level of brain tissue in TBI-challenged rats. The TBI-challenged animals exhibited significant brain injuries, characterized by an increase of blood-brain barrier permeability, brain edema, and lesion volume, as well as neurologic dysfunction, which were significantly ameliorated by NaHS treatment. However, the protective effects of H2S in TBI could be abolished by the mitoK(ATP) channel blocker 5-hydroxydecanoate. Moreover, we found that NaHS treatment increased endogenous antioxidant enzymatic activities and decreased oxidative product levels in brain tissue of TBI-challenged rats. CONCLUSIONS: Exogenous H2S administered at an appropriate dose can exert a protective effect against TBI via activation of mitoK(ATP) channels and reduction of oxidative stress.