Qianxue Chen

Hemorrhagic Transformation after Tissue Plasminogen Activator Reperfusion Therapy for Ischemic Stroke: Mechanisms, Models, and Biomarkers.

Intracerebral hemorrhagic transformation (HT) is well recognized as a common cause of hemorrhage in patients with ischemic stroke. HT after acute ischemic stroke contributes to early mortality and adversely affects functional recovery. The risk of HT is especially high when patients receive thrombolytic reperfusion therapy with tissue plasminogen activator, the only available treatment for ischemic stroke. Although many important publications address preclinical models of ischemic stroke, there are no current recommendations regarding the conduct of research aimed at understanding the mechanisms and prediction of HT. In this review, we discuss the underlying mechanisms for HT after ischemic stroke, provide an overview of the models commonly used for the study of HT, and discuss biomarkers that might be used for the early detection of this challenging clinical problem.

Glioma Stem Cell-Targeted Dendritic Cells as a Tumor Vaccine Against Malignant Glioma

Purpose Cancer stem cells have recently been thought to be closely related to tumor development and reoccurrence. It may be a promising way to cure malignant glioma by using glioma stem cell-targeted dendritic cells as a tumor vaccine. In this study, we explored whether pulsing dendritic cells with antigens of glioma stem cells was a potent way to induce specific cytotoxic T lymphocytes and anti-tumor immunity. Materials and Methods Cancer stem cells were cultured from glioma cell line U251. Lysate of glioma stem cells was obtained by the repeated freezing and thawing method. Dendritic cells (DCs) were induced and cultured from the murine bone marrow cells, the biological characteristics were detected by electron microscope and flow cytometry. The DC vaccine was obtained by mixing DCs with lysate of glioma stem cells. The DC vaccine was charactirizated through the mixed lymphocyte responses and cell killing experiment in vitro. Level of interferon-γ (IFN-γ) in the supernatant was checked by ELISA. Results After stimulation of lysate of glioma stem cell, expression of surface molecules of DC was up-regulated, including CD80, CD86, CD11C and MHC-II. DCs pulsed with lysate of glioma stem cells were more effective than the control group in stimulating original glioma cells-specific cytotoxic T lymphocytes responses, killing glioma cells and boosting the secretion of IFN-γ in vitro. Conclusion The results demonstrated DCs loaded with antigens derived from glioma stem cells can effectively stimulate naive T cells to form specific cytotoxic T cells, kill glioma cells cultured in vitro.

Clinical and Imaging Characteristics of Cerebral Schistosomiasis

In recent years, there has been a trend for increased incidence of cerebral schistosomiasis. It is often misdiagnosed because of the diversity of clinical symptoms. We wished to explore clinical characteristics and imaging findings in cerebral schistosomiasis. We retrospectively analyzed clinical data, laboratory tests, CT, and MRI results in 11 patients with cerebral schistosomiasis. All patients had chronic cerebral schistosomiasis (five with epilepsy type, five with brain tumor type, and one patient with stroke type). All patients with brain tumor type were misdiagnosed as having gliomas. There were typical findings on CT and MRI. In conclusion, clinical manifestations of cerebral schistosomiasis are variable, and the rate of misdiagnosis is high. For more precise diagnosis, a combination of laboratory and imaging data is required.

Methazolamide improves neurological behavior by inhibition of neuron apoptosis in subarachnoid hemorrhage mice

Subarachnoid hemorrhage (SAH) results in significant nerve dysfunction, such as hemiplegia, mood disorders, cognitive and memory impairment. Currently, no clear measures can reduce brain nerve damage. The study of brain nerve protection after SAH is of great significance. We aim to evaluate the protective effects and the possible mechanism of methazolamide in C57BL/6J SAH animal model in vivo and in blood-induced primary cortical neuron (PCNs) cellular model of SAH in vitro. We demonstrate that methazolamide accelerates the recovery of neurological damage, effectively relieves cerebral edema, and improves cognitive function in SAH mice as well as offers neuroprotection in blood- or hemoglobin-treated PCNs and partially restores normal neuronal morphology. In addition, western blot analyses show obviously decreased expression of active caspase-3 in methazolamide-treated SAH mice comparing with vehicle-treated SAH animals. Furthermore, methazolamide effectively inhibits ROS production in PCNs induced by blood exposure or hemoglobin insult. However, methazolamide has no protective effects in morality, fluctuation of cerebral blood flow, SAH grade, and cerebral vasospasm of SAH mice. Given methazolamide, a potent carbonic anhydrase inhibitor, can penetrate the blood–brain barrier and has been used in clinic in the treatment of ocular conditions, it provides potential as a novel therapy for SAH.

Daphnetin Protects against Cerebral Ischemia/Reperfusion Injury in Mice via Inhibition of TLR4/NF-κB Signaling Pathway

Growing evidences indicate that immune-mediated mechanisms contribute to the development of cerebral ischemia/reperfusion (I/R) injury. Daphnetin (DAP) is a coumarin derivative extracted from Daphne odora var., which displays anti-inflammatory properties. However, the effect of DAP on cerebral I/R injury is not yet clear. Recent studies have demonstrated that TLR4/NF-κB signaling pathway takes part in the damaging inflammatory process of cerebral I/R injury. The present study aimed to investigate the effect of DAP on cerebral I/R injury in vivo and its possible mechanisms. DAP was administered before middle cerebral artery occlusion and reperfusion in mice. The neurological scores, cerebral infarct sizes, the levels of inflammatory cytokines, apoptotic neural cells, and the levels of TLR4, NF-κB p65, and IκBα were estimated. The results showed that an obvious improvement of neurological scores and infarct sizes was observed in DAP-treated mice after MCAO/R. DAP treatment decreased the overexpression of TNF-α, IL-1β, and IL-6 and attenuated neural cells apoptosis. Moreover, DAP treatment decreased the TLR4 expression, IκB-α degradation, and nuclear translocation of NF-κB. Taken together, our results suggested that DAP exerted neuroprotective and anti-inflammatory effects on cerebral I/R injury. The potential mechanism was involved in the inhibition of TLR4/NF-κB mediated inflammatory signaling pathway.

BMP4 reverses multidrug resistance through modulation of BCL-2 and GDNF in glioblastoma.

Patients with glioblastoma are commonly treated with chemotherapy. But a significant proportion of patients develop disease progression after an initial response to chemotherapy. Presently, there is no standard of care for such patients. The bone morphogenetic protein 4 (BMP4) has been reported to play a tumor-suppressing role in glioblastoma, but its role in glioblastoma multidrug resistance (MDR) is not clear. We reported that BMP4 can reverse MDR of glioblastoma through the inhibition of B-cell lymphoma 2(BCL-2) and glial cell derived neurotrophic factor (GDNF). We showed that the expression level of BMP4 was lower in glioblastoma compared to normal brain tissue, and also showed that BMP4 expression decreased in multidrug resistance cell line U251/TMZ compared to U251 cells. Our research demonstrated that over-expression of BMP4 can reverse the multidrug resistance. BCL-2 and GDNF were inhibited when BMP4 was over-expressed, and this data were consistent with the negative relationship in human samples; analysis of 40 patients glioblastoma and brain samples revealed a significant negative correlation between BMP4 and BCL-2, GDNF. When BCL-2 and GDNF were knocked down, the effect of BMP4 in regulating MDR was partially lost. This novel result showed, for the first time, that BMP4 can reverse MDR in glioblastoma, which involved negative inhibition of BCL-2 and GDNF.

Ellagic acid inhibits proliferation and induces apoptosis in human glioblastoma cells.

PURPOSE To investigate the anticancer activity of ellagic acid (EA) in U251 human glioblastoma cells and its possible molecular mechanism. METHODS The cells were treated with EA at various concentrations for different time periods. Cell viability and cell proliferation were detected by cell counting kit-8(CCK-8) assay and live/dead assay respectively. Cell apoptosis were measured with Annexin V-FITC/PI double staining method by flow cytometry and Mitochondrial membrane potential assay separately. Cell cycle was measured with PI staining method by flow cytometry. The expressions of Bcl-2, Survivin, XIAP, Caspase-3, Bax, JNK, p-JNK, ERK1/2, p-ERK1/2, p38, p-p38, DR4, DR5, CHOP and GRP78-related proteins were detected by western blot after EA treatment. RESULTS Cell viability and proliferation of glioblastoma cells treated with EA were significantly lower than the control group. EA caused robust apoptosis of the glioblastoma cells compared to the control group. EA significantly decreased the proportion at G0/G1 phases of cell cycling accompanied by increased populations at S phase in U251 cell lines. And the expressions of anti-apoptotic proteins were dramatically down-regulated. CONCLUSION Ellagic acid potentially up-regulated DR4, DR5 and MAP kinases (JNK, ERK1/2 and p38). EA also caused significant increase in the expressions of CHOP and GRP78. Our findings suggest that EA would be beneficial for the treatment of glioblastoma.

LRIG1 enhances chemosensitivity by modulating BCL-2 expression and receptor tyrosine kinase signaling in glioma cells.

Purpose Leucine-rich repeats and immunoglobulin-like domains 1 (LRIG1) are an inhibitor of receptor tyrosine kinases (RTKs) that was discovered in recent years, and many studies showed that LRIG1 is a tumor suppressor gene and may be related to tumor drug resistance. In this study, we explored whether LRIG1 protein expression can improve the chemosensitivity of glioma cells and what was its mechanism. Materials and Methods We collected 93 cases of glioma tissues and detected the expression of LRIG1 and BCL-2. We constructed a multidrug resistance cell line U251/multidrug resistance (MDR) and examined the change of LRIG1 and BCL-2 at mRNA and protein expression levels. LRIG1 expression was upregulated in U251/MDR cells and we detected the change of multidrug resistance. Meanwhile, we changed the expression of LRIG1 and BCL-2 and explored the relationship between LRIG1 and BCL-2. Finally, we also explored the relationship between LRIG1 and RTKs. Results LRIG1 was negatively correlated with BCL-2 expression in glioma tissue and U251/MDR cells, and upregulation of LRIG1 can enhance chemosensitivity and inhibit BCL-2 expression. Furthermore, LRIG1 was negatively correlated with RTKs in U251/MDR cells. Conclusion These results demonstrated that LRIG1 can improve chemosensitivity by modulating BCL-2 expression and RTK signaling in glioma cells.

GSK-3β as a target for protection against transient cerebral ischemia

Stroke remains the leading cause of death and disability worldwide. This fact highlights the need to search for potential drug targets that can reduce stroke-related brain damage. We showed recently that a glycogen synthase kinase-3β (GSK-3β) inhibitor attenuates tissue plasminogen activator-induced hemorrhagic transformation after permanent focal cerebral ischemia. Here, we examined whether GSK-3β inhibition mitigates early ischemia-reperfusion stroke injury and investigated its potential mechanism of action. We used the rat middle cerebral artery occlusion (MCAO) model to mimic transient cerebral ischemia. At 3.5 h after MCAO, cerebral blood flow was restored, and rats were administered DMSO (vehicle, 1% in saline) or GSK-3β inhibitor TWS119 (30 mg/kg) by intraperitoneal injection. Animals were sacrificed 24 h after MCAO. TWS119 treatment reduced neurologic deficits, brain edema, infarct volume, and blood-brain barrier permeability compared with those in the vehicle group. TWS119 treatment also increased the protein expression of β-catenin and zonula occludens-1 but decreased β-catenin phosphorylation while suppressing the expression of GSK-3β. These results indicate that GSK-3β inhibition protects the blood-brain barrier and attenuates early ischemia-reperfusion stroke injury. This protection may be related to early activation of the Wnt/β-catenin signaling pathway.

Novel microcatheter-based intracarotid delivery approach for MCAO/R mice.

The intra-arterial (IA) model by microcatheter administration was an effective way to deliver drugs or cells to the brain. All of these models were carried out introduced in rat rather than mice for the difficult and technically challenging due to their small caliber. In 2014, Alejandro Santillan first introduced this model in mice and we found that most of the operational steps were similar with the middle cerebral artery occlusion and reperfusion (MCAO/R) model. We attempted to combine these two techniques into a single model in mice and discovered that this technique was indeed possible. In our work, 12C57Bl/6J male mice were carried on middle cerebral artery occlusion for 60min and then the intra-arterial microcatheter was placed into the internal carotid artery (ICA) from the external carotid artery (ECA). GFP-Luc-Pro labeled mNSCs were infused through the microcatheter and then the blood flow perfusion was reestablished subsequently. The results showed that all 12 mice were carried on successfully the model of middle cerebral artery occlusion, and the placement of the microcatheter and the mNSCs perfusion were completed smoothly without exception. Which means that it is logical to combine the two models into one in order to facilitate studying of stroke. Meanwhile, during the dissection, we found the variation of occipital artery (OA) was noticeable and we classified first time this variation into four categories to attempt to protect the OA.