Application and advances of biomimetic membrane materials in central nervous system disorders.

Central nervous system (CNS) diseases encompass spinal cord injuries, brain tumors, neurodegenerative diseases, and ischemic strokes. Recently, there has been a growing global recognition of CNS disorders as a leading cause of disability and death in humans and the second most common cause of death worldwide. The global burdens and treatment challenges posed by CNS disorders are particularly significant in the context of a rapidly expanding global population and aging demographics. The blood-brain barrier (BBB) presents a challenge for effective drug delivery in CNS disorders, as conventional drugs often have limited penetration into the brain. Advances in biomimetic membrane nanomaterials technology have shown promise in enhancing drug delivery for various CNS disorders, leveraging properties such as natural biological surfaces, high biocompatibility and biosafety. This review discusses recent developments in biomimetic membrane materials, summarizes the types and preparation methods of these materials, analyzes their applications in treating CNS injuries, and provides insights into the future prospects and limitations of biomimetic membrane materials.

Astrocyte-derived Interleukin-31 causes poor prognosis in elderly patients with intracerebral hemorrhage.

The incidence of intracerebral hemorrhage (ICH) is increasing every year, with very high rates of mortality and disability. The prognosis of elderly ICH patients is extremely unfavorable. Interleukin, as an important participant in building the inflammatory microenvironment of the central nervous system after ICH, has long been the focus of neuroimmunology research. However, there are no studies on the role IL31 play in the pathologic process of ICH. We collected para-lesion tissue for immunofluorescence and flow cytometry from the elderly and young ICH patients who underwent surgery. Here, we found that IL31 expression in the lesion of elderly ICH patients was significantly higher than that of young patients. The activation of astrocytes after ICH releases a large amount of IL31, which binds to microglia through IL31R, causing a large number of microglia to converge to the hematoma area, leading to the spread of neuroinflammation, apoptosis of neurons, and ultimately resulting in poorer recovery of nerve function. Interfering with IL31 expression suppresses neuroinflammation and promotes the recovery of neurological function. Our study demonstrated that elderly patients release more IL31 after ICH than young patients. IL31 promotes the progression of neuroinflammation, leading to neuronal apoptosis as well as neurological decline. Suppression of high IL31 concentrations in the brain after ICH may be a promising therapeutic strategy for ICH.

Indole-3-carbinol (I3C) reduces apoptosis and improves neurological function after cerebral ischemia-reperfusion injury by modulating microglia inflammation.

Indole-3-carbinol(I3C) is a tumor chemopreventive substance that can be extracted from cruciferous vegetables. Indole-3-carbinol (I3C) has been shown to have antioxidant and anti-inflammatory effects. In this study, we investigated the cerebral protective effects of I3C in an in vivo rats model of middle cerebral artery occlusion (MCAO). 8-10 Week-Old male SD rat received I3C (150 mg/kg, once daily) for 3 days and underwent 3 h of middle cerebral artery occlusion (MCAO) followed by reperfusion. The results showed that I3C pretreatment (150 mg/kg, once daily) prevented CIRI-induced cerebral infarction in rats. I3C pretreatment also decreased the mRNA expression levels of several apoptotic proteins, including Bax, caspase-3 and caspase-9, by increasing the mRNA expression levels of the anti-apoptotic protein Bcl-2. Inhibited apoptosis in the brain cells of MCAO rats. In addition, we found that I3C pretreatment reduced neuronal loss, promoted neurological recovery after ischemia-reperfusion injury and increased seven-day survival in MCAO rats. I3C pretreatment also significantly reduced the expression of inducible nitric oxide synthase (INOS), interleukin-1ß (IL-1ß) and interleukin-6 (IL-6) mRNA in ischemic brain tissue; Increased expression of interleukin-4 (IL-4) and interleukin-10 (IL-10) mRNA. At the same time, I3C pretreatment significantly decreased the expression of the M1 microglial marker IBA1 after cerebral ischemia-reperfusion injury and increased the expression of these results in the M2 microglial marker CD206. I3C pretreatment also significantly decreased apoptosis and death of HAPI microglial cells after hypoxia induction, decreased interleukin-1ß (IL-1ß) and interleukin-6 (IL-6) mRNA The expression of interleukin-4 (IL-4) and interleukin-10 (IL-10) mRNAs was increased. These results suggest that I3C protects the brain from CIRI by regulating the anti-inflammatory and anti-apoptotic effects of microglia.

Targeting the AKT-P53/CREB pathway with epicatechin for improved prognosis of traumatic brain injury.

AIMS: The aim of this study was to evaluate the effect of epicatechin, on neurological recovery and neuroinflammation after traumatic brain injury (TBI) to investigate its potential value in clinical practice. METHODS: TBI model was established in adult rats by CCI method. The effect of epicatechin was evaluated after intraperitoneal injection. Neurological recovery after TBI was assessed by Morris Water Maze, mNSS score, Rotarod test and Adhesive removal test. Protein and gene expression was assessed by Western blot, ELISA, PCR and immunofluorescence. Furthermore, the use of AKT pathway inhibitors blocked the therapeutic effects of epicatechin clarifying AKT-P53/CREB as a potential pathway for the effects of epicatechin. RESULTS: Administering epicatechin after TBI prevented neuronal death, reduced neuroinflammation, and promoted neurological function restoration in TBI rats. Network pharmacology study suggested that epicatechin may exert its therapeutic benefits through the AKT-P53/CREB pathway CONCLUSION: These results indicate that epicatechin, a monomeric compound derived from tea polyphenols, possesses potent antioxidant and anti-inflammatory properties after TBI. The mechanism may be related to the regulation of the AKT-P53/CREB signal pathway.

Myricetin suppresses traumatic brain injury-induced inflammatory response via EGFR/AKT/STAT pathway.

Traumatic brain injury (TBI) is a common disease in neurosurgery with a high fatality and disability rate which imposes a huge burden on society and patient's family. Inhibition of neuroinflammation caused by microglia activation is a reasonable strategy to promote neurological recovery after TBI. Myricetin is a natural flavonoid that has shown good therapeutic effects in a variety of neurological disease models, but its therapeutic effect on TBI is not clear. We demonstrated that intraperitoneal injection of appropriate doses of myricetin significantly improved recovery of neurological function after TBI in Sprague Dawley rats and inhibited excessive inflammatory responses around the lesion site. Myricetin dramatically reduced the expression of toxic microglia markers generated by TBI and LPS, according to the outcomes of in vivo and in vitro tests. In particular, the expression of inducible nitric oxide synthase, cyclooxygenase 2, and some pro-inflammatory cytokines was reduced, which protected learning and memory functions in TBI rats. Through network pharmacological analysis, we found that myricetin may inhibit microglia hyperactivation through the EGFR-AKT/STAT pathway. These findings imply that myricetin is a promising treatment option for the management of neuroinflammation following TBI.

COMMD4 is a novel prognostic biomarker and relates to potential drug resistance mechanism in glioma.

Background: Glioma as the most frequently discovered tumor affecting the brain shows significant morbidity and fatality rates with unfavorable prognosis. There is an urgent need to find novel therapeutic targets to overcome the low chemotherapeutic efficacy of glioma. This research examined whether the copper-metabolism-domain protein, COMMD4, had predictive and therapeutic significance in glioma. Methods: Using the freely accessible CGGA (The Chinese Glioma Atlas) and TCGA (The Cancer Genome Atlas) databases, we examined the function of COMMD4 in GBM and LGG. CIBERSORT and TIMER were utilized to assess the associations between COMMD4 and immune cells. The Gene Set Enrichment Analysis (GSEA) was employed to examine the functional data. Furthermore, the link between COMMD4 expression and predicted treatment response was evaluated via CellMiner Cross-Database. Meanwhile, qRT-PCR was conducted to examine COMMD4 expression in human glioma. Finally, Migration and invasion of glioma cells (U-87, U-251) were assessed using transwell assays. R was used to analyze the statistical data. Results: According to our findings, COMMD4 expression level was higher in patients having grade-dependent glioma who also showed an unfavorable prognosis. Furthermore, qRT-PCR confirmed the high expression of COMMD4 in glioma tissues and cells. Additionally, using integrated correlation analysis, we acquired significant prognostic findings between isocitrate dehydrogenase 1(IDH1) and COMMD4. Meanwhile, a link between COMMD4 and many tumor-infiltrating immune cells was observed. GSEA and drug response analysis revealed the potential mechanism of COMMD4 in drug resistance of glioma. Conclusion: The current findings validated COMMD4 as a novel biological marker, which might offer insights into the possible drug resistance mechanisms and the impact of the immune microenvironment on glioma. COMMD4 might be used to predict glioma prognosis.

The cuproptosis-related signature associated with the tumor environment and prognosis of patients with glioma.

Background: Copper ions are essential for cellular physiology. Cuproptosis is a novel method of copper-dependent cell death, and the cuproptosis-based signature for glioma remains less studied. Methods: Several glioma datasets with clinicopathological information were collected from TCGA, GEO and CGGA. Robust Multichip Average (RMA) algorithm was used for background correction and normalization, cuproptosis-related genes (CRGs) were then collected. The TCGA-glioma cohort was clustered using ConsensusClusterPlus. Univariate Cox regression analysis and the Random Survival Forest model were performed on the differentially expressed genes to identify prognostic genes. The cuproptosis-signature was constructed by calculating CuproptosisScore using Multivariate Cox regression analysis. Differences in terms of genomic mutation, tumor microenvironment, and enrichment pathways were evaluated between high- or low-CuproptosisScore. Furthermore, drug response prediction was carried out utilizing pRRophetic. Results: Two subclusters based on CRGs were identified. Patients in cluster2 had better clinical outcomes. The cuproptosis-signature was constructed based on CuproptosisScore. Patients with higher CuproptosisScore had higher WHO grades and worse prognosis, while patients with lower grades were more likely to develop IDH mutations or MGMT methylation. Univariate and Multivariate Cox regression analysis demonstrated CuproptosisScore was an independent prognostic factor. The accuracy of the signature in prognostic prediction was further confirmed in 11 external validation datasets. In groups with high-CuproptosisScore, PIK3CA, MUC16, NF1, TTN, TP53, PTEN, and EGFR showed high mutation frequency. IDH1, TP53, ATRX, CIC, and FUBP1 demonstrated high mutation frequency in low-CuproptosisScore group. The level of immune infiltration increased as CuproptosisScore increased. SubMap analysis revealed patients with high-CuproptosisScore may respond to anti-PD-1 therapy. The IC50 values of Bexarotene, Bicalutamide, Bortezomib, and Cytarabine were lower in the high-CuproptosisScore group than those in the low-CuproptosisScore group. Finally, the importance of IGFBP2 in TCGA-glioma cohort was confirmed. Conclusion: The current study revealed the novel cuproptosis-based signature might help predict the prognosis, biological features, and appropriate treatment for patients with glioma.

Prognostic Biomarker ZNF311 and Its Correlation With Tumor Progression and Immune Infiltrates in Glioma.

BACKGROUND: Gliomas, particularly high-grade gliomas, are the most common primary brain tumors. From the Chinese Glioma Genome Atlas (CGGA) database, the relationships between the altered molecular pathways and gliomas could be easily observed. A close connection in the occurrence of the pathogenesis exists between the microenvironment, the glioma, and the associated genes. METHODS: Validation of the role of ZNF311 oncogene was confirmed by data from the CGGA dataset on glioblastoma and low-grade glioma. Furthermore, we used CIBERSORT to analyze the correlation between ZNF311 and cancer immune infiltrates. RESULTS: According to our analysis, ZNF311 was expressed higher in patients with grade-depended glioma with poor prognosis. In addition, we obtained valuable prognostic results between isocitrate dehydrogenase 1 (IDH1) and ZNF311 through the analysis of integrated correlations. Similarly, we simultaneously revealed the prognostic results between 1p/19q and ZNF311. In addition, we found that ZNF311 is correlated with a large number of tumor-infiltrating immune cells. CONCLUSIONS: Based on the study findings, we conclude that ZNF311 is potentially a novel biomarker for assessing prognosis and immune infiltration in glioblastoma and diffuse glioma cases.

Novel Insights into MSK1 Phosphorylation by MRKß in Intracerebral Hemorrhage-Induced Neuronal Apoptosis.

Neuronal apoptosis is regarded as one of the most important pathophysiological changes of intracerebral hemorrhagic (ICH) stroke-a major public health problem that leads to high mortality rates and functional dependency. Mitogen-and stress-activated kinase (MSK) 1 is implicated in various biological functions in different cell types, including proliferation, tumorigenesis and responses to stress. Our previous study showed that MSK1 phosphorylation (p-MSK1) is related to the regulation of LPS-induced astrocytic inflammation, and possibly acts as a negative regulator of inflammation. In this study, we identified a specific interaction between MSK1 and MRKß (MLK-related kinase)-a member of the MAPK pathway-during neuronal apoptosis. In an ICH rat model, western blotting and immunohistochemical analysis revealed that both MRKß and phosphorylation of MSK1 (p-MSK1 Ser376) were significantly upregulated in cells surrounding the hematoma. Triple-immunofluorescent labeling demonstrated the co-localization of MRKß and p-MSK1 in neurons, but not astrocytes. Furthermore, MRKß was partially transported into the nucleus, and interacted with p-MSK1 in hemin-treated neurons. Immunoprecipitation showed that MRKß and p-MSK1 exhibited an enhanced interaction during the pathophysiology process. Utilizing small interfering RNAs to knockdown MRKß or MSK1, we verified that MSK1 Ser376 is a phosphorylation site targeted by MRKß. We also observed that the phosphorylation of NF-κB p65 at Ser276 was mediated by the MRKß-p-MSK1 complex. Furthermore, it was found that the neuronal apoptosis marker, active caspase-3, was co-localized with MRKß and p-MSK1. In addition, flow cytometry analysis revealed that knockdown of MRKß or MSK1 specifically resulted in increased neuronal apoptosis, which suggested that the MRKß-p-MSK1 complex might exert a neuroprotective function against ICH-induced neuronal apoptosis. Taken together, our data suggest that MRKß translocated into the nucleus and phosphorylated MSK1 to protect neurons via phosphorylation of p65-a subunit of nuclear factor κB.

Combination Glioma Therapy Mediated by a Dual-Targeted Delivery System Constructed Using OMCN-PEG-Pep22/DOX.

Accumulating studies have investigated the efficacy of receptor-mediated delivery of hydrophobic drugs in glioma chemotherapy. Here, a delivery vehicle comprising polyethylene glycol (PEG) and oxidized nanocrystalline mesoporous carbon particles (OMCN) linked to the Pep22 polypeptide targeting the low-density lipoprotein receptor (LDLR) is designed to generate a novel drug-loaded system, designated as OMCN-PEG-Pep22/DOX (OPPD). This system effectively targets glioma cells and the blood-brain barrier and exerts therapeutic efficacy through both near-infrared (NIR) photothermal and chemotherapeutic effects of loaded doxycycline (DOX). Pathological tissue microarrays show an association of LDLR overexpression in human glioma tissue with patient survival.NIR irradiation treatment and magnetic resonance imaging results show that OPPD reaches the effective glioma-killing temperature in a glioma-bearing rat with a skull bone removal model and considerably reduces glioma sizes relative to the drug-loaded system without the Pep22 peptide modification and the control respectively. Thus, OPPD not only effectively targets LDLR-overexpressing glioma but also exerts a dual therapeutic effect by transporting DOX into the glioma and generating thermal effects with near-infrared irradiation to kill tumor cells. These collective findings support the utility of the novel OPPD drug-loaded system as a promising drug delivery vehicle for clinical application in glioma therapy.

Up-regulation of FOS-like antigen 1 contributes to neuronal apoptosis in the cortex of rat following traumatic brain injury.

Neuronal apoptosis is an important process of secondary brain injury which is induced by neurochemical signaling cascades after traumatic brain injury (TBI). Present study was designed to investigate whether FOS-like antigen 1 (Fra-1) is involved in the neuronal apoptosis. Western blot analysis and immunohistochemistry in a rat TBI model revealed a significant increase in the expression of Fra-1 in the ipsilateral brain cortex, which was in parallel with increase in the expression of active caspase-3. With immunofluorescence double-labeling, Fra-1 was colocalized with active caspase-3 and with NeuN, a neuronal marker. In an in vitro cell injury model, H2O2 exposure induced cell apoptosis and reduced cell viability and at the same time, a similar increased expression of active caspase-3, p53 and Fra-1 was found in PC12 cells. Down-regulation of Fra-1 through transfection with Fra-1 siRNA remarkably elevated cell viability, reduced the expression of active caspase-3 and p53, and decreased apoptosis of PC12 cells after H2O2 exposure. Taken together, present findings suggest that Fra-1 may be involved in the induction of neuronal apoptosis through up-regulating p53 signaling pathway and that this action may contribute to the secondary neuropathological process after TBI.

O-GlcNAc Glycosylation of nNOS Promotes Neuronal Apoptosis Following Glutamate Excitotoxicity.

Ischemic stroke is a dominant health problem with extremely high rates of mortality and disability. The main mechanism of neuronal injury after stroke is excitotoxicity, during which the activation of neuronal nitric oxide synthase (nNOS) exerts a vital role. However, directly blocking N-methyl-D-aspartate receptors or nNOS can lead to severe undesirable effects since they have crucial physiological functions in the central nervous system. Here, we report that nNOS undergoes O-linked-ß-N-acetylglucosamine (O-GlcNAc) modification via interacting with O-GlcNAc transferase, and the O-GlcNAcylation of nNOS remarkably increases during glutamate-induced excitotoxicity. In addition, eliminating the O-GlcNAcylation of nNOS protects neurons from apoptosis during glutamate stimulation by decreasing the formation of nNOS-postsynaptic density protein 95 complexes. Taken together, our data suggest a novel function of the O-GlcNAcylation of nNOS in neuronal apoptosis during glutamate excitotoxicity, suggesting a novel therapy strategy for ischemic stroke.

Upregulated Expression of SSTR3 is Involved in Neuronal Apoptosis After Intracerebral Hemorrhage in Adult Rats.

Somatostatin which is a multifunctional growth hormone inhibitory neuropeptide shows diverse physiological effects, such as neurotransmission, cell growth, apoptosis, and endocrine signaling as well as exerts inhibitory effects on hormonal products and other secretory proteins. SSTR3 is a member of superfamily of somatostatin receptors (SSTR), which are G-protein-coupled plasma membrane receptors. Previous studies proved that SSTR3 regulates antiproliferative signaling and apoptosis in several cells. Here, we explored a potential role of SSTR3 in the regulation of neuronal apoptosis in the course of intracerebral hemorrhage (ICH). An ICH rat model was established and assessed by behavioral tests. We found SSTR3 was significantly upregulated surrounding the hematoma after ICH by Western blot and immunohistochemistry. Double immunofluorescence-manifested SSTR3 was strikingly increased in neurons, not astrocytes or microglia. Moreover, increasing SSTR3 level was found to be accompanied by the upregulation of p53, Bax, and active caspase-3 in vivo and in vitro studies. Furthermore, we detected that neuronal apoptosis marker active caspase-3 was co-localized with SSTR3 suggesting its potential role in neuronal apoptosis. In addition, in vitro study, revealed that SSTR3 knockdown specifically resulted in reducing neuronal apoptosis in PC12 cells, which further indicated that SSTR3 might exert its pro-apoptotic function on neuronal apoptosis. All our findings suggested that upregulated SSTR3 may be involved in neuronal apoptosis after ICH.

Spatiotemporal Patterns of RING1 Expression after Rat Spinal Cord Injury.

Ring finger protein 1 (RING1) is a RING domain characterized protein belonging to the RING finger family. It is an E3 ubiquitin-protein ligase that mediated monoubiquitination of histone H2A and the core component of PRC1 complex, which is the repressive multiprotein complex of Polycomb group (PcG). Previous studies showed the important tumorigenic role of RING1 via promoting cell proliferation and the crucial function in maintaining transcriptional program stability during development. However, its mechanism for spinal cord injury (SCI) is still unknown. In our research, we established an acute SCI model in adult rats and studied the expression and function profiles of RING1. RING1 protein level detected by western blot peaked at day 3 after trauma and then decreased gradually. Immunohistochemistry showed the increase of RING1 expression displayed in the white matter more obviously than in the gray matter. Furthermore, increased co-expression of RING1 and GFAP confirmed activated astrocytes in injured spinal cord via double immunofluorescence staining. Meanwhile, we also found the co-localization of PCNA, a famous marker of proliferative cells, with RING1 and GFAP, which indicated RING1 might play a role in astrocyte proliferation after SCI. In vitro studies, RING1 protein level in C6 cells increased after LPS challenge and RING1 was required for astrocyte proliferation and activation induced by LPS. In summary, we took a new insight into the function of RING1 in the cellular and molecular mechanism underlying the pathophysiology of SCI.

Inhibition of PTTG1 expression by microRNA suppresses proliferation and induces apoptosis of malignant glioma cells.

The present study aimed to investigate the role of pituitary tumor-transforming gene 1 (PTTG1) in the proliferation, invasion and apoptosis of human malignant glioma U251 cells. Firstly, 2 microRNAs (miRNAs) targeting PTTG1 messenger (m)RNA were ligated into a pcDNA6.2-GW/EmGFP-miR expression vector. The recombinant plasmids, miRNA-1 and miRNA-2 (miR-2), were transfected into U251 cells using the liposome method. PTTG1 mRNA and protein levels were evaluated using quantitative polymerase chain reaction and western blot analysis. The proliferation and invasion abilities of U251 cells were determined using methylthiazol tetrazolium and Matrigel assays. Flow cytometry analysis with Annexin V/propidium iodide double staining was used to determine the percentage of apoptotic cells. PTTG1 expression was effectively suppressed by miR-2. U251 cell growth was inhibited between 10.7 and 34.7% in the miR-2 group compared with the blank group. The Matrigel assay demonstrated that the percentage of infiltrating U251 cells was significantly lower in the miR-2 group (12.3±1.0%) compared to the blank group (24.7±1.4%; P<0.001) and the negative control group (24.0±2.0%; P<0.05). A higher percentage of apoptotic U251 cells were observed in the miR-2 group compared with the blank group (53.6 vs. 32.4%) using flow cytometry due to cycle arrests at the G2/M phase. The miR-2-transfected U251 cells were subcutaneously injected into nude mice, and these mice possessed a decreased tumor tissue growth rate and higher percentage of apoptotic cells compared with the blank and negative control groups. In conclusion, PTTG1 gene expression in human malignant glioma U251 cells was effectively suppressed by exogenous miR-2. The downregulation of PTTG1 induced glioma cell apoptosis and cell cycle arrest at the G2/M phase, which inhibited cell proliferation, reverse invasion and infiltration of glioma cells.

An IDH1 mutation inhibits growth of glioma cells via GSH depletion and ROS generation.

The isocitrate dehydrogenase 1 (IDH1) gene mutation occurs frequently in glioma. While some studies have demonstrated that IDH1 mutations are associated with prolonged survival, the mechanism remains unclear. In this study, we found that growth was significantly inhibited in glioma cells overexpressing the mutated IDH1 gene. Furthermore, these cells were characterized by decreased intracellular NADPH levels accompanied by glutathione (GSH) depletion and reactive oxygen species (ROS) generation. Moreover, the increased apoptosis and the decreased proliferation were found in the glioma cells overexpressing the mutant IDH1 gene. Accordingly, our study demonstrates that using H2O2-regulated mutant IDH1 glioma cells could obviously increase the inhibition of cell growth; nevertheless, GSH had the opposite result. Our study provides direct evidence that mutation of IDH1 profoundly inhibits the growth of glioma cells, and we speculate that this is the major factor behind its association with prolonged survival in glioma. Finally, our study indicates that depletion of GSH and generation of ROS are the primary cellular events associated with this mutation.

Phosphorylation of mitogen- and stress-activated protein kinase-1 in astrocytic inflammation: a possible role in inhibiting production of inflammatory cytokines.

PURPOSE: It is generally accepted that inflammation has a role in the progression of many central nervous system (CNS) diseases, although the mechanisms through which this occurs remain unclear. Among mitogen-activated protein kinase (MAPK) targets, mitogen- and stress-activated protein kinase (MSK1) has been thought to be involved in the pathology of inflammatory gene expression. In this study, the roles of MSK1 activation in neuroinflammation were investigated. METHODS: The bacterial lipopolysaccharide (LPS)-induced brain injury model was performed on Sprague-Dawley rats. The dynamic expression changes and the cellular location of p-MSK1 in the brain cortex were detected by Western blot and immunofluorescence staining. The synthesis of inflammatory cytokines in astrocytes was detected by enzyme-linked immunosorbent assay (ELISA). RESULTS: Phosphorylated MSK1 (p-MSK1 Thr-581) was induced significantly after intracerebral injection of LPS into the lateral ventricles of the rat brain. Specific upregulation of p-MSK1 in astrocytes was also observed in inflamed cerebral cortex. At 1 day after LPS stimulation, iNOS, TNFα expression, and the astrocyte marker glial fibrillary acidic protein (GFAP) were increased significantly. Also, in vitro studies indicated that the upregulation of p-MSK1 (Thr-581) may be involved in the subsequent astrocyte inflammatory process, following LPS challenge. Using an enzyme-linked immunosorbent assay (ELISA), it was confirmed that treatment with LPS in primary astrocytes stimulated the synthesis of inflammatory cytokines, through MAPKs signaling pathways. In cultured primary astrocytes, both knock-down of total MSK1 by small interfering RNAs (siRNA) or specific mutation of Thr-581 resulted in higher production of certain cytokines, such as TNFα and IL-6. CONCLUSIONS: Collectively, these results suggest that MSK1 phosphorylation is associated with the regulation of LPS-induced brain injury and possibly acts as a negative regulator of inflammation.

PAX3 is overexpressed in human glioblastomas and critically regulates the tumorigenicity of glioma cells.

Paired box 3 (PAX3) is overexpressed in glioma tissues compared to normal brain tissues, however, the pathogenic role of PAX3 in human glioma cells remains to be elucidated. In this study, we selected the human glioma cell lines U251, U87, SHG-44, and the normal human astrocytes, 1800, which have differential PAX3 expression depending upon the person. SiRNA targeting PAX3 and PAX3 overexpression vectors were transfected into U87 and SHG-44 glioma cell lines, and cell proliferation, invasion, apoptosis, and differentiation were examined by CCK-8 assays, transwell chamber assays, tunnel staining, Annexin V/PI analysis, and Western blotting, respectively. In addition, we used subcutaneous tumor models to study the effect of PAX3 on the growth of glioma cells in vivo. We found that PAX3 was upregulated in the three glioma cell lines. PAX3 knockdown inhibited cell proliferation and invasion, and induced apoptosis in the U87MG glioblastoma cell line, whereas PAX3 upregulation promoted proliferation, inhibited apoptosis, and increased invasion in the SHG-44 glioma cell line. Moreover, we found that targeting PAX3 expression in glioma cell lines together with chemotherapeutic treatment could increase glioma cell susceptibility to the drug. In subcutaneous tumor models in nude mice using glioma cell lines U-87MG and SHG-44, inhibition of PAX3 expression in glioblastoma U-87MG cells suppressed tumorigenicity, and upregulation of PAX3 expression in glioma SHG-44 cells promoted tumor formation in vivo. These results indicate that PAX3 in glioma is essential for gliomagenesis; thus, targeting PAX3 or its downstream targets may lead to novel therapies for this disease.

Cyclic AMP response element modulator-1 (CREM-1) involves in neuronal apoptosis after traumatic brain injury.

The cyclic AMP response element-binding protein (CREB) family can regulate biological functions of various types of cells by forming homo- or heterodimers to bind the target DNA sequences; it plays an essential role in individual neuronal function and entire neuronal circuits. One attractive activity of the CREB family is regulating the transcription of apoptosis-suppressor gene bcl-2. Cyclic AMP response element modulator-1 (CREM-1) is one member of the family with limited acquaintance. To investigate whether CREM-1 is involved in central nervous system injury and repair, we performed an acute traumatic brain injury (TBI) model in adult rats. Western blot analysis and immunohistochemistry showed a significant upregulation of CREM-1 in ipsilateral peritrauma cortex. Immunofluorescent labeling indicated that CREM-1 was localized mainly in the nuclei of neurons; co-localization of CREM-1 and active-caspase-3 in the ipsilateral cortex suggested that CREM-1 might participate in neuronal apoptosis. To further investigate the function of CREM-1, a neuronal cell line PC12 was employed to establish an apoptosis model. We analyzed the association of CREM-1 with p-CREB on PC12 cells by Western blot, immunofluorescent labeling, and co-immunoprecipitation. The result implied that the association of CREM-1 with p-CREB was enhanced in apoptotic cells. Additionally, knocking CREM-1 down with siRNA demonstrated the probable pro-apoptotic role played by CREM-1 in neuronal apoptosis. Together with our data, we hypothesized that CREM-1 might play an important role in regulating neuronal death after TBI by interacting with CREB.

Expression of Spy1 protein in human non-Hodgkin's lymphomas is correlated with phosphorylation of p27 Kip1 on Thr187 and cell proliferation.

Aberrations in cell cycle control are often observed in tumors and might even be necessary in tumor development. Spy1, a novel cell cycle regulatory protein, can control cell progression and survival through the atypical activation of cyclin-dependent kinases (CDKs). In this progression, the phosphorylation of p27(Kip1) at Thr187 by CDK2 was shown to be a chief role. In this study, we studied 183 human specimens including reactive lymphoid and Non-Hodgkin's Lymphomas (NHLs) tissues. Immunohistochemistry (IHC) analysis suggested that Spy1 and pThr187-p27 were overexpressed in NHLs. The expression of Spy1 was positively related to pThr187-p27 and proliferation marker Ki-67 expression. In a multivariate analysis, high Spy1 and pThr187-p27 expressions were showed to be associated with poor prognosis in NHLs. While in vitro, following release of Jurkat cells from serum starvation, the expression of Spy1 was upregulated, as well as pThr187-p27 and CDK2. And an increased interaction between Spy1 and pThr187-p27 was demonstrated at 4 h after serum stimulation. Additionally, transfecting cells with Spy1-siRNA could diminish the expression of pThr187-p27 and arrest cell growth. Our results suggest that Spy1 may be a possible prognostic indicator in NHLs, and it was correlated with phosphorylation of p27(Kip1) on Thr187. These findings provide a rational framework for further development of Spy1 inhibitors as a novel class of anti-tumor agents.