Browsing by Author "Xie, Luokun"
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Item Artemisinin Protects Oxidative Stress-induced Neuronal Apoptosis Via Up-Regulation of Akt/Bcl-2 Signaling(2017-03-14) Liu, Ran; Xie, Luokun; Li, Wenjun; Winters, Ali; Chaudhari, Kiran; Prah, Jude; Yang, Shao-Hua; Lin, Shao-PengPurpose: Artemisinin is a powerful anti-malarial drug that has been in use for decades. Recently, the novel biological effects of artemisinin on cancer, inflammation-related disorders, and cardiovascular disease were reported. The aim of this study was to explore the neuroprotective actions of artemisinin. Methods: The model of glutamate-induced oxidative injury in HT22 hippocampal cells was established to simulate cellular ischemic model. We investigated the effect of artemisinin on oxidative stress-induced cell apoptosis death and the activity of Akt/Bcl-2 pathway in HT22 cells. Results: Pretreatment with artemisinin attenuated reactive oxygen species (ROS) generations, preventing the decline of mitochondrial membrane potential and rescued the HT22 cells form glutamate-induced apoptosis death. The Akt/Bcl-2 pathway was activated by artemisinin in time dependent manner. Furthermore, the artemisinin inhibitor MK2206 blocked the neuroprotective effect of artemisinin. Conclusions: Artemisinin protects neuronal HT22 cell from glutamate-induced oxidative injury and apoptosis via Akt/Bcl-signaling, thereby might be applicated for clinical neurological therapy.Item Combining Injectable Plasma Scaffold with Mesenchymal Stem/Stromal Cells for Repairing Infarct Cavity after Ischemic Stroke(JKL International, 2017-04-01) Zhang, Hongxia; Sun, Fen; Wang, Jixian; Xie, Luokun; Yang, Chenqi; Pan, Mengxiong; Shao, Bei; Yang, Guo-Yuan; Yang, Shaohua; Zhuge, Qichuan; Jin, KunlinStroke survivors are typically left with structural brain damage and associated functional impairment in the chronic phase of injury, for which few therapeutic options exist. We reported previously that transplantation of human embryonic stem cell (hESC)-derived neural stem cells together with Matrigel scaffolding into the brains of rats after focal ischemia reduced infarct volume and improved neurobehavioral performance. Matrigel is a gelatinous protein mixture extracted from mouse sarcoma cells, thus would not be approved for use as a scaffold clinically. In this study, we generated a gel-like scaffold from plasma that was controlled by changing the concentration of CaCl2. In vitro study confirmed that 10-20 mM CaCl2 and 10-40% plasma did not affect the viability and proliferation of human and rat bone marrow mesenchymal stem/stromal cells (BMSCs) and neural stem cells (NSCs). We transplanted plasma scaffold in combination of BMSCs into the cystic cavity after focal cerebral ischemia, and found that the atrophy volume was dramatically reduced and motor function was significantly improved in the group transplanted with scaffold/BMSCs compared with the groups treated with vehicle, scaffold or BMSCs only. Our data suggest that plasma-derived scaffold in combination of BMSCs is feasible for tissue engineering approach for the stroke treatment.Item IMPACT OF T-CELLS ON ASTROCYTES IN VIVO & IN VITRO: IMPLICATIONS POST-ISCHEMIC STROKE(2019-03-05) Xie, Luokun; Li, Wenjun; Liu, Ran; Yang, Shaohua; Hersh, JessicaPurpose: Post-ischemic stroke, T-lymphocytes enter the brain. The role of T-cells in the progression of cerebral infarction or repair mechanisms is unclear. We hypothesized that T-cells interact with astrocytes directly leading to an anti-inflammatory response. Methods: In vivo, ischemic stroke was induced by middle cerebral artery occlusion in young adult C57/B6 male mice. Mice were sacrificed at 3 days or 1-month post-ischemic stroke. Paraffin-embedded brain sections demonstrated co-localization of astrocytes and CD4+ and CD8+ T-cells in the ischemic region 1 month after stroke. T-cells were harvested from the brain by digestion; percoll enriched, and incubated with anti-CD3 and CD25 antibodies. T-cells were sorted via flow cytometry. The cytokine profile of brain infiltrated CD4+ and CD8+ T-cells were compared to spleen T-cells using QT-PCR. In vitro, C8-S murine astrocyte type II clone cell line (ATCC® CRL-2535™), and T-cells extracted from the spleens of 3-month-old C57/B6 female mice were placed in co-culture at a 1:1 for 48 and 72 hours and compared to individual cell cultures. Anti-CTLA-4 antibodies were added to each culture condition as another experimental group. Astrocytes and T-cells were collected separately for QT-PCR analysis. Results: In vivo, the following cytokine gene expressions poststroke, were found to be elevated: IFNγ, IL-10, IL-17, TNFα, and perforin. In vitro, IL-10 gene expression was elevated in astrocytes and T-cells individually harvested from 1:1 co-cultures compared to astrocytes and T-cells alone at 48 and 72 hours respectively. IL-10 was produced primarily by T-cells stimulated by direct contact with astrocytes. Anti-CTLA-4 antibodies blocked the direct cell-to-cell interaction by reducing IL-10 gene expression in both astrocytes and T-cells. Conclusions: Our data suggests that T-cells release pro- and anti- inflammatory cytokines while in close proximity to astrocytes after ischemic stroke. In co-cultures, astrocytes directly interact with T-cells increasing their IL-10 gene expression by 72 h., implying a neuroprotective mechanism exists via astrocyte stimulation of T-cell IL-10 production.Item METHYLENE BLUE INHIBITS PROLIFERATION AND MAINTAINS SELF-RENEWAL OF RAT NEURAL STEM/PROGENITOR CELLS(2014-03) Xie, Luokun; Roy Choudhury, Gourav; Park, Yong; Liu, Ran; Zhang, Chun-Li; Yorio, Thomas; Jin, Kunlin; Yang, ShaohuaNeural progenitor cells (NPCs)are important for neurogenesis and brain damage repair. To elucidate the effect of methylene blue on NPCs, we studied the self-renewal and proliferation of NPCs both in vitro and in vivo. We found that methylene blue restrains NPC proliferation and enahnces NPC self-renewal. Purpose (a): Neural stem cell-based treatment holds a new therapeutic opportunity for treating neurodegenerative disorders. While methylene blue has been shown to be neuroprotective in multiple experimental neurodegenerative disease models, its potential effects on neural stem/progenitor cells (NSPCs) has not been addressed. Methylene blue can easily penetrate the blood brain barrier to access the brain parenchyma. Thus, its effects on NSPCs, whether positive or negative, need to be elucidated. Methods (b): We used in both in vitro culture model and in vivo study to test the effects of methylene blue on the proliferation, self-renewal and differentiation of NSPCs. Neurospheres were generated in vitro and were treated with methylene blue. NSPC proliferation was evaluated by Ki67 staining and propidium iodide staining. NSPC self-renewal was determined by serial passage assay. Real-time PCR was applied to test the expression of neural differentiation markers in NSPCs. To address the signal pathway responsible for the methylene blue-induced changes on NSPCs, expression of cyclins and mTORC1 activation were determined by real-time PCR and Western blot, respectively. The effects of methylene blue on NSPC proliferation was also confirmed by intracerebroventricular infusion of methylene blue in rats followed by BrdU and Nestin staining. Results (c): Methylene blue inhibits porliferation of rat NSPCs in the in vitro culture. Methylene blue treatment decreased most cyclin expression. In addition, methylene blue enhanced the self-renewal capacity of NSPCs, demonstrated by more neurosphere growth and inhibited differentiation marker expression in NSPCs. However, methylene blue did not impair committed neuronal differentiation. The change of cyclin expression is associated with the change of mTOR expression in methylene blue-treated NSPCs. Methylene blue repressed transcription of mTOR rather than enhance mTOR degradation. Consistent with in vitro data, methylene blue inhibited neural stem cell division in the subventricular zone, but did not influenced neuronal development in a short term. Conclusions (d): Our findings indicate that methylene blue could delay NSPCs senescence by enhancing NSPCs self-renewal capacity. However, the long-term effects of methylene blue on the in vivo NSPC pool needs further investigation. The impact of methylene blue on NSPCs should be taken into account in future therapy with methylene blue, either for the peripheral diseases or for the CNS disorders.Item Sex Dependent Alteration in Psychomotor and Cognitive Functions After Chronic Metformin Treatment(2017-03-14) Winters, Ali; Shetty, Ritu; Li, Wenjun; Xie, Luokun; Prah, Jude; Liu, Ran; Sumien, Nathalie; Yang, Shaohua; Chaudhari, KiranPurpose: Metformin, the most commonly used anti-hyperglycemic medication has been proposed to have delayed aging and longevity benefits. Without due consideration to gender/sex influence, metformin administration is being tested for non-diabetic benefits. Amid mixed reports on cognition, the purpose of the current study was to identify the influence of sex variation in the psychomotor and cognitive outcomes after long term metformin treatment. Materials and Methods: Young normo-glycemic male and female C57BL/6J mice (aged 4 mo, n=10 each; total n=40 mice) were randomly assigned to either a control group or metformin group (administered 2 mg/ml in drinking water). After 1 month of treatment, a battery of behavioral tests was initiated to assess the psychomotor and cognitive functions. Metformin treatment was continued during behavior assessment. Results: Overall female mice weighed lesser than male mice. Over the experiment time span, metformin neither altered the body weight nor decreased the blood glucose level significantly. There was no variation in muscle strength or reflexes between male and female mice on either treatment. Male mice were more anxious than female mice and metformin treatment decreased anxiety in male mice only. Female mice had better motor learning and maximum coordinated running performance than male mice. Metformin treatment improved motor learning only in male mice. Metformin treatment improved balance function irrespective of sex. Overall male mice had better retention of long term memory which was deteriorated after metformin treatment. Further, metformin impaired the short term memory and cognitive flexibility only in male mice. Conclusions: This study demonstrated that metformin affects psychomotor or cognitive function differently influenced by sex. Our results suggested that chronic metformin was beneficial for psychomotor function and detrimental for short term and long-term memory in male sex. While, in female sex, metformin had beneficial or no effects on brain functions.