Activation of STAT3, MAPK, and AKT in malignant astrocytic gliomas: correlation with EGFR status, tumor grade, and survival

Diffuse astrocytic gliomas are the most common human glial tumors with glioblastoma being the most malignant form. Epidermal growth factor receptor (EGFR) gene amplification is one of the most common genetic changes in glioblastoma and can lead to the activation of various downstream signaling molecules, including STAT3, MAPK, and AKT. In this study, we investigated the activation status of these 3 signaling molecules as well as wild-type (EGFRwt) and mutant (EGFRvIII) EGFR in 82 malignant astrocytic gliomas (55 glioblastomas and 27 anaplastic astrocytomas) using immunohistochemistry. The presence of EGFRwt, but not EGFRvIII, immunopositivity correlated significantly with prevalent EGFR gene amplification in glioblastomas. STAT3 and AKT activation correlated significantly with EGFR status, although the correlation for p-STAT3 was attributed exclusively to EGFRvIII. The distribution of these 3 activated molecules varied significantly with tumor grade; although activation of STAT3 was essentially identical between anaplastic astrocytomas and glioblastomas, an increase in the activation of MAPK and AKT appeared to correlate with the progression of anaplastic astrocytoma to glioblastoma. Finally, activated STAT3 and AKT were marginally predictive of improved and worse prognosis, respectively. Taken together, these findings begin to elucidate the interrelationship between these signaling pathways in astrocytic gliomas in vivo.     

胶质瘤中EGFR扩增、过表达及其与肿瘤增殖关系的研究

目的探讨表皮生长因子受体（EGFR）在胶质瘤中扩增、过表达及其在肿瘤增殖中的作用机制。方法应用SP免疫组化法、RT—PCR、Southern印迹法检测6例正常脑组织、50例不同级别胶质瘤及2个体外胶质瘤细胞系中在EGFR蛋白水平、mRNA水平、DNA水平的表达及扩增情况，以及与肿瘤增殖活性的相关性。结果不同级别的胶质瘤中，表达与扩增程度不同，Ⅱ级与Ⅲ、Ⅳ级肿瘤之间比较具有显著性差异：EGFR过表达与Ki-67在胶质瘤中的表达呈正相关．而EGFR过表达与扩增并不一致。结论EGFR过表达与胶质瘤的增殖活性增高有密切相关性，推测EGFR过表达是肿瘤进展中的早期事件而非进展期事件。     

Differential BDNF signaling in dentate gyrus and perirhinal cortex during consolidation of recognition memory in the rat

Consolidation of long‐term memory is dependent on synthesis of new proteins in the hippocampus and associated cortical regions. The neurotrophin brain‐derived neurotrophic factor (BDNF) is tightly regulated by activity‐dependent cellular processes and is strongly linked with mechanisms underlying learning and memory. BDNF activation of tyrosine receptor kinase (TrkB) stimulates intracellular signaling cascades implicated in plasticity, including the extracellular‐signal related kinase (ERK)/mitogen‐activated protein kinase (MAPK) pathway and the phosphatidylinositide‐3‐kinase (PI3K)/Akt pathway. Here, we investigate the role of BDNF, ERK/MAPK, and PI3K/AKT signaling cascade in recognition memory in the rat. We report that recognition memory was associated with increased release of BDNF in the dentate gyrus and perirhinal cortex. This was associated with significant increases in p44ERK activation and c‐fos expression in the dentate gyrus and PI3K activation and c‐fos expression in the perirhinal cortex. Furthermore, both recognition memory and the associated cell signaling events in dentate gyrus and perirhinal cortex were blocked by intraperitoneal injection of the Trk receptor inhibitor tyrphostin AG879. These data are consistent with the hypothesis that BDNF‐stimulated intracellular signaling plays a role in consolidation of recognition memory in the rat. © 2012 Wiley Periodicals, Inc.     

ERK activation and expression of neuronal cell cycle markers in the hippocampus after entorhinal cortex lesion

Current findings suggest that neuronal cell death is frequently associated with the aberrant expression of cell cycle‐regulatory proteins in postmitotic neurons. Aberrant cell cycle reentry has been implicated in diverse neurodegenerative conditions, including Alzheimer's disease (AD). Previously we reported that the appearance of cell cycle markers in postmitotic neurons of the entorhinal cortex (EC) after excitotoxic hippocampal damage is associated with the expression of phospho‐tau and amyloid precursor protein (APP). However, the question of the signaling pathway involved in this cell cycle reentry remains unresolved. Differentiated neurons use the molecular mechanisms initially acquired to direct cell proliferation, such as the Ras–extracellular signal‐regulated kinase (ERK1/2) pathway, to regulate synaptic plasticity. In this work we explored whether ERK1/2‐related signaling might contribute to the cell cycle reentry in hippocampal neurons after a unilateral EC lesion. We showed that, within the first 24 hr after hippocampal deafferentation, numerous neurons expressed phospho‐ERK1/2, concomitantly with the gradual increases in cyclin D1 and cyclin B immunoreactivity in the dentate gyrus and hilus. Several of these immunopositive cells to phospho‐ERK1/2 and cyclin B in hippocampus are postmitotic neurons, insofar as they are positive to NeuN. The intracisternal administration of U0126 (an MEK inhibitor), previous to the excitotoxic lesion, decreased the activation of ERK1/2 and the expression of cyclin D1 and cyclin B in the hippocampus. The present findings support the notion that ERK1/2 plays a role in cell cycle reactivation in mature neurons efferently connected to the lesion site. © 2012 Wiley Periodicals, Inc.     

Implications of mitogen‐activated protein kinase signaling in glioma

Gliomas are the most common primary central nervous system tumors. Gliomas originate from astrocytes, oligodendrocytes, and neural stem cells or their precursors. According to WHO classification, gliomas are classified into four different malignant grades ranging from grade I to grade IV based on histopathological features and related molecular aberrations. The induction and maintenance of these tumors can be attributed largely to aberrant signaling networks. In this regard, the mitogen‐activated protein kinase (MAPK) network has been widely studied and is reported to be severely altered in glial tumors. Mutations in MAPK pathways most frequently affect RAS and B‐RAF in the ERK, c‐Jun N‐terminal kinase (JNK), and p38 pathways leading to malignant transformation. Also, it is linked to both inherited and sequential accumulations of mutations that control receptor tyrosine kinase (RTK)‐activated signal transduction pathways, cell cycle growth arrest pathways, and nonresponsive cell death pathways. Genetic alterations that modulate RTK signaling can also alter several downstream pathways, including RAS‐mediated MAP kinases along with JNK pathways, which ultimately regulate cell proliferation and cell death. The present review focuses on recent literature regarding important deregulations in the RTK‐activated MAPK pathway during gliomagenesis and progression. © 2015 Wiley Periodicals, Inc.     

Comparative status of activated ERK1/2 and PARP cleavage in human gliomas

Gliomas are the most common form of cerebral tumors. Understanding molecular features of glioma will eventually allow for targeted intervention and more promising approaches for treating gliomas. The present study is therefore carried out to check the levels of activated ERK1/2 with respect to phospho‐tyrosine and cleavage of poly ADP‐ribose polymerase (PARP). Recent experiments support that extracellular signal regulated kinase (ERK), a mitogen activated protein (MAP) kinase might have a critical role in cell proliferation. PARP is a DNA‐repair enzyme activated by DNA strand breaks. Overactivation of PARP after cellular insult lead to cell death caused by rapid depletion of cellular ATP. Three glioblastoma multiforme (GBM) and two astrocytoma biopsies (core tumor) and peripheral tissues were analyzed for the expression of p‐ERK1/2 and PARP. Results indicate higher p‐ERK1/2 in GBM. Cleaved fragments of PARP (89 kDa) were found to be more in core tumor tissue samples as compared to peripheral tumor tissues of both astrocytoma and GBM.     

Extracellular signal‐regulated kinase (ERK) is involved in LPS‐induced disturbances in intestinal motility

Background Lipopolysaccharide (LPS) is a causative agent of sepsis. A relationship has been described between LPS, free radicals, and cyclooxygenase‐2 (COX‐2). Here, we investigate the role of extracellular signal‐regulated kinase (ERK) mitogen‐activated protein kinases (MAPK) in the effect of LPS on intestinal motility, oxidative stress status, and COX‐2 expression. Methods Rabbits were injected with (i) saline, (ii) LPS, (iii) U0126, an ERK MAPK inhibitor, or (iv) U0126+LPS. Duodenal contractility was studied in an organ bath with acetylcholine, prostaglandin E₂, and KCl added. Neuromuscular function was assessed by electrical field stimulation (EFS). Neurotransmitter blockers were used to study the EFS‐elicited contractile response. The formation of products of oxidative damage to proteins (carbonyls), lipids, [malondialdehyde (MDA), and 4‐hydroxyalkenals (4‐HDA)] was quantified in plasma and intestine. The protein expression of phospho‐ERK (p‐ERK), total ERK, and COX‐2 in the intestine was measured by western blot, and p‐ERK was localized by immunohistochemistry. Key Results Acetylcholine, prostaglandin E₂, and KCl‐induced contractions decreased with LPS. Electrical field stimulation induced a neurogenic contraction that was reduced by LPS. Lipopolysaccharide increased p‐ERK and COX‐2 expression and the levels of carbonyls and MDA+4‐HDA. U0126 blocked the effect of LPS on acetylcholine, prostaglandin E₂, KCl, and EFS‐induced contractions, the levels of carbonyls and MDA+4‐HDA and p‐ERK and COX‐2 expression. Phospho‐ERK was detected mostly in the neurons of the myenteric and submucosal ganglia. Conclusions & Inferences We can suggest that ERK is involved in the mechanism of action of LPS in the intestine.     

ITGA4基因在脑胶质瘤中的表达及临床意义

目的 探讨整合素α4(ITGA4)基因在脑胶质瘤中的表达与临床意义。方法 计算机检索CGGA数据库获取mRNAseq-325数据集中325例脑胶质瘤的转录组测序（RNA-seq）数据及临床资料,并用m RNAseq-693数据集中693例脑胶质瘤进行验证。结果 胶质瘤ITGA4基因呈高表达（P<0.05）,与胶质瘤IDH基因型、1p/19q共缺失状态、WHO病理分级有关（P<0.05）,与胶质瘤MGMT启动子甲基化状态无明显关系（P>0.05）。多因素Cox回归分析显示,ITGA4高表达是胶质瘤生存预后不良的独立危险因素（P<0.05）。生存曲线分析显示,ITGA4高表达胶质瘤病人生存期明显缩短（P<0.05）。GO分析和KEGG分析显示,ITGA4基因调控的生物过程包括细胞粘附、血管生成及细胞迁移等,主要调控PI3K/AKT信号通路。结论 我们的结果提示胶质瘤ITGA4呈高表达,与胶质瘤不良生存预后密切相关,主要调控PI3K/AKT信号通路。     

PI3K/AKT pathway alterations are associated with clinically aggressive and histologically anaplastic subsets of pilocytic astrocytoma

Pilocytic astrocytomas (PA) are well-differentiated gliomas having a favorable prognosis when compared with other diffuse or infiltrative astrocytomas. Molecular genetic abnormalities and activation of signaling pathways associated with clinically aggressive PA and histologically anaplastic PA have not been adequately studied. We performed molecular genetic, gene expression, and immunohistochemical studies using three PA subsets, including conventional PA (n?=?43), clinically aggressive/recurrent PA (n?=?24), and histologically anaplastic PA (n?=?25). A clinical diagnosis of NF1 was present in 28% of anaplastic PA. Molecular cytogenetic studies demonstrated heterozygous PTEN/10q and homozygous p16 deletions in 6/19 (32%) and 3/15 (20%) cases of anaplastic PA, respectively, but in neither of the two other groups. BRAF duplication was identified in 33% of sporadic anaplastic PA and 63% of cerebellar examples. BRAF (V600E) mutation was absent in four (of 4) sporadic cases lacking duplication. IDH1(R132H) immunohistochemistry was negative in 16 (of 16) cases. Neither PDGFRA nor EGFR amplifications were present. pERK staining levels were similar among the three PA subsets, but a stepwise increase in cytoplasmic pAKT and to a lesser extent pS6 immunoreactivity was noted by immunohistochemistry in aggressive PA groups. This was particularly true in histologically anaplastic PA when compared with conventional PA (p?相似文献   

Repression of tau hyperphosphorylation by chronic endurance exercise in aged transgenic mouse model of tauopathies

The present study was undertaken to investigate whether chronic endurance exercise affects tau phosphorylation levels in the brain with Alzheimer's disease (AD)‐like pathology. To address this, the transgenic (Tg) mouse model of tauopathies, Tg‐NSE/htau23, which expresses human tau23 in the brain, was chosen. Animals were subjected to chronic exercise for 3 months from 16 months of age. The exercised Tg mouse groups were treadmill run at speeds of 12 m/min (intermediate exercise group) or 19 m/min (high exercise group) for 1 hr/day, 5 days/week, during the 3‐month period. Chronic endurance exercise in Tg mice increased the expression of Cu/Zn‐superoxide dismutase (SOD) and catalase, and also their enzymatic activities in the brain. In parallel, chronic exercise in Tg mice up‐regulated the expression of phospho‐PKCα, phospho‐AKT, and phospho‐PI3K, and down‐regulated the expressions of phospho‐PKA, phosphor‐p38, phospho‐JNK, and phospho‐ERK. Moreover, chronic exercise up‐regulated both cytosolic and nuclear levels of β‐catenin, and the expression of T‐cell factor‐4 (Tcf‐4) and cyclin D1 in the brain. As a consequence of such changes, the levels of phospho‐tau in the brain of Tg mice were markedly decreased after exercise. Immunohistochemical analysis showed an exercised‐induced decrease of the phospho‐tau levels in the CA3 subregion of the hippocampus. These results suggest that chronic endurance exercise may provide a therapeutic potential to alleviate the tau pathology. © 2009 Wiley‐Liss, Inc.     

MYB upregulation and genetic aberrations in a subset of pediatric low-grade gliomas

Recent studies of genetic abnormalities in pediatric low-grade gliomas (LGGs) have focused on activation of the ERK/MAPK pathway by KIAA1549-BRAF gene fusions in the majority of pilocytic astrocytomas (PAs) and by rare mutations in elements of the pathway across histopathologically diverse LGGs. This study reports that MYB, an oncogene not previously implicated in gliomagenesis, is activated in a diverse subset of pediatric LGGs. The study cohort comprised 57 pediatric LGGs and a comparative cohort of 59 pediatric high-grade gliomas (HGGs). The LGG cohort included 34 PAs and 23 diffuse gliomas; fibrillary astrocytomas (n = 14), oligodendroglial tumors (n = 7), and angiocentric gliomas (n = 2). MYB copy number abnormalities were disclosed using Affymetrix 6.0 SNP arrays and confirmed using interphase fluorescence in situ hybridization. Novel MYB amplifications that upregulate MYB RNA and protein expression were demonstrated in 2/14 diffuse astrocytomas. In addition, focal deletion of the terminal region of MYB was seen in 1 of 2 angiocentric gliomas (AGs). Increased expression of MYB was demonstrated by quantitative RT-PCR and immunohistochemistry. MYB upregulation at the protein level was demonstrated in a proportion of diffuse LGGs (60%), pilocytic astrocytomas (41%), and HGGs (19%), but abnormalities at the genomic level were only a feature of diffuse gliomas. Our data suggest that MYB may have a role in a subset of pediatric gliomas, through a variety of mechanisms in addition to MYB amplification and deletion.     

Circadian expression and functional characterization of PEA‐15 within the mouse suprachiasmatic nucleus

The circadian timing system influences the functional properties of most, if not all, physiological processes. Central to the mammalian timing system is the suprachiasmatic nucleus (SCN) of the hypothalamus. The SCN functions as a ‘master clock’ that sets the phasing of ancillary circadian oscillator populations found throughout the body. Further, via an entraining input from the retina, the SCN ensures that the clock oscillators are synchronized to the daily light/dark cycle. A critical component of the SCN timing and entrainment systems is the p44/42 mitogen‐activated protein kinase (ERK/MAPK) pathway. Here, we examined the expression and function of phosphoprotein‐enriched in astrocytes (PEA‐15), an ERK scaffold protein that serves as a key regulator of MAPK signaling. A combination of immunolabeling and Western blotting approaches revealed high levels of PEA‐15 within the SCN. PEA‐15 expression was enriched in distinct subpopulations of SCN neurons, including arginine vasopressin (AVP)‐positive neurons of the SCN shell region. Further, expression profiling detected a significant circadian oscillation in PEA‐15 expression within the SCN. Brief photic stimulation during the early subjective night led to a significant increase in PEA‐15 phosphorylation, an event that can trigger ERK/PEA‐15 dissociation. Consistent with this, co‐immunoprecipitation assays revealed that PEA‐15 is directly bound to ERK in the SCN and that photic stimulation leads to their dissociation. Finally, we show that PEA‐15 regulates ERK/MAPK‐dependent activation of the core clock gene period1. Together, these data raise the prospect that PEA‐15 functions as a key regulator of the SCN timing system.     

STK40在脑胶质瘤中的表达及其临床意义的生信分析

目的 探讨丝氨酸/酸酸蛋白激酶40（STK40）在脑胶质瘤中的表达及其临床意义。方法 计算机检索GEPIA数据库,运用生物信息学方法分析STK40在胶质瘤组织及正常脑组织中的表达差异;同时,检索CGGA数据库中693例脑胶质瘤的基因信息及临床资料,运用Kaplan-Meier生存曲线分析STK40表达水平与脑胶质瘤生存预后的关系;采用多因素Cox比例回归风险模型分析脑胶质瘤生存预后的影响因素。结果 GEPIA数据库分析显示,胶质母细胞瘤组织STK40表达水平较正常脑组织明显增高（P<0.05）。STK40表达水平与胶质瘤病理级别呈正相关,随胶质瘤病理级别增高,STK40表达水平明显增高（P<0.0001）。多因素Cox风险比例回归模型分析结果显示,STK40高表达是脑胶质瘤生存预后不良的独立危险因素（P<0.05）。Kaplan-Meier生存曲线分析显示,STK40高表达组中位总生存期较低表达组明显缩短（P<0.0001）;而且STK40高表达明显降低胶质瘤放/化疗的效果（P<0.0001）。Pearson相关性分析显示,ERK下游产物SRF与STK40呈明显正相关（r=0.45;P<0.0001）,JNK下游产物ATF2与STK40表达水平呈明显负相关（r=-0.167;P<0.0001）。结论 胶质瘤STK40呈高表达,与病人生存预后不良有关,其作用机制可能与ERK/MAPK及JNK/MAPK信号通路有关。     

MMP‐9 controls Schwann cell proliferation and phenotypic remodeling via IGF‐1 and ErbB receptor‐mediated activation of MEK/ERK pathway

Phenotypic remodeling of Schwann cells is required to ensure successful regeneration of damaged peripheral axons. After nerve damage, Schwann cells produce an over 100‐fold increase in metalloproteinase‐9 (MMP‐9), and therapy with an MMP inhibitor increases the number of resident (but not infiltrating) cells in injured nerve. Here, we demonstrate that MMP‐9 regulates proliferation and trophic signaling of Schwann cells. Using in vivo BrdU incorporation studies of axotomized sciatic nerves of MMP‐9?/? mice, we found increased Schwann cell mitosis in regenerating (proximal) stump relative to wild‐type mice. Treatment of cultured primary Schwann cells with recombinant MMP‐9 suppressed their growth, mitogenic activity, and produced a dose‐dependent, biphasic, and selective activation of ERK1/2, but not JNK and p38 MAPK. MMP‐9 induced ERK1/2 signaling in both undifferentiated and differentiated (using dbcAMP) Schwann cells. Using inhibitors to MEK and trophic tyrosine kinase receptors, we established that MMP‐9 regulates Ras/Raf/MEK—ERK pathways through IGF‐1, ErbB, and PDGF receptors. We also report on the early changes of MMP‐9 mRNA expression (within 24 h) after axotomy. These studies establish that MMP‐9 controls critical trophic signal transduction pathways and phenotypic remodeling of Schwann cells. © 2009 Wiley‐Liss, Inc.     

Dystroglycan modulates the ability of insulin‐like growth factor‐1 to promote oligodendrocyte differentiation

The adhesion receptor dystroglycan positively regulates terminal differentiation of oligodendrocytes, but the mechanism by which this occurs remains unclear. Using primary oligodendrocyte cultures, we identified and examined a connection between dystroglycan and the ability of insulin‐like growth factor‐1 (IGF‐1) to promote oligodendrocyte differentiation. Consistent with previous reports, treatment with exogenous IGF‐1 caused an increase in MBP protein that was preceded by activation of PI3K (AKT) and MAPK (ERK) signaling pathways. The extracellular matrix protein laminin was further shown to potentiate the effect of IGF‐1 on oligodendrocyte differentiation. Depletion of the laminin receptor dystroglycan using siRNA, however, blocked the ability of IGF‐1 to promote oligodendrocyte differentiation of cells grown on laminin, suggesting a role for dystroglycan in IGF‐1‐mediated differentiation. Indeed, loss of dystroglycan led to a reduction in the ability of IGF‐1 to activate MAPK, but not PI3K, signaling pathways. Pharmacological inhibition of MAPK signaling also prevented IGF‐1‐induced increases in myelin basic protein (MBP), indicating that MAPK signaling was necessary to drive IGF‐1‐mediated enhancement of oligodendrocyte differentiation. Using immunoprecipitation, we found that dystroglycan, the adaptor protein Grb2, and insulin receptor substrate‐1 (IRS‐1), were associated in a protein complex. Taken together, our results suggest that the positive regulatory effect of laminin on oligodendrocyte differentiation may be attributed, at least in part, to dystroglycan's ability to promote IGF‐1‐induced differentiation. © 2010 Wiley‐Liss, Inc.