Glutamate leads to neuronal cell damage by generating neurotoxicity during brain development. The objective of this study is to identify proteins that differently expressed by glutamate treatment in neonatal cerebral cortex. Sprague-Dawley rat pups (post-natal day 7) were intraperitoneally injected with vehicle or glutamate (10 mg/kg). Brain tissues were isolated 4 h after drug treatment and fixed for morphological study. Moreover, cerebral cortices were collected for protein study. Two-dimensional gel electrophoresis and mass spectrometry were carried out to identify specific proteins. We observed severe histopathological changes in glutamate-exposed cerebral cortex. We identified various proteins that differentially expressed by glutamate exposure. Identified proteins were thioredoxin, peroxiredoxin 5, ubiquitin carboxy-terminal hydrolase L1, proteasome subunit alpha proteins, isocitrate dehydrogenase, and heat shock protein 60. Heat shock protein 60 was increased in glutamate exposed condition. However, other proteins were decreased in glutamate-treated animals. These proteins are related to anti-oxidant, protein degradation, metabolism, signal transduction, and anti-apoptotic function. Thus, our findings can suggest that glutamate leads to neonatal cerebral cortex damage by regulation of specific proteins that mediated with various functions.
Animals ; Brain ; Cerebral Cortex ; Chaperonin 60 ; Electrophoresis, Gel, Two-Dimensional ; Glutamic Acid ; Humans ; Infant, Newborn ; Isocitrate Dehydrogenase ; Mass Spectrometry ; Metabolism ; Neurons ; Peroxiredoxins ; Proteasome Endopeptidase Complex ; Proteolysis ; Proteomics ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; Thioredoxins ; Ubiquitin Thiolesterase

Catalpol, a major bioactive component from Rehmannia glutinosa, which has been used to treat diabetes. The present study was designed to elucidate the anti-diabetic effect and mechanism of action for catalpol in db/db mice. The db/db mice were randomly divided into six groups (10/group) according to their blood glucose levels: db/db control, metformin (positive control), and four dose levels of catalpol treatment (25, 50, 100, and 200 mg·kg), and 10 db/m mice were used as the normal control. All the groups were administered orally for 8 weeks. The levels of fasting blood glucose (FBG), random blood glucose (RBG), glucose tolerance, insulin tolerance, and glycated serum protein (GSP) and the globe gene expression in liver tissues were analyzed. Our results showed that catalpol treatment obviously reduced water intake and food intake in a dose-dependent manner. Catalpol treatment also remarkably reduce fasting blood glucose (FBG) and random blood glucose (RBG) in a dose-dependent manner. The RBG-lowering effect of catalpol was better than that of metformin. Furthermore, catalpol significantly improved glucose tolerance and insulin tolerance via increasing insulin sensitivity. Catalpol treatment significantly decreased GSP level. The comparisons of gene expression in liver tissues among normal control mice, db/db mice and catalpol treated mice (200 and 100 mg·kg) indicated that there were significant increases in the expressions of 287 genes, whichwere mainly involved in lipid metabolism, response to stress, energy metabolism, and cellular processes, and significant decreases in the expressions of 520 genes, which were mainly involved in cell growth, death, immune system, and response to stress. Four genes expressed differentially were linked to glucose metabolism or insulin signaling pathways, including Irs1 (insulin receptor substrate 1), Idh2 (isocitrate dehydrogenase 2 (NADP), mitochondrial), G6pd2 (glucose-6-phosphate dehydrogenase 2), and SOCS3 (suppressor of cytokine signaling 3). In conclusion, catalpol ecerted significant hypoglycemic effect and remarkable therapeutic effect in db/db mice via modulating various gene expressions.
Animals ; Blood Glucose ; metabolism ; Diabetes Mellitus, Experimental ; drug therapy ; genetics ; metabolism ; Disease Models, Animal ; Dose-Response Relationship, Drug ; Drugs, Chinese Herbal ; administration & dosage ; analysis ; Gene Expression ; drug effects ; Glucosephosphate Dehydrogenase ; genetics ; metabolism ; Humans ; Hypoglycemic Agents ; administration & dosage ; Insulin ; metabolism ; Insulin Receptor Substrate Proteins ; genetics ; metabolism ; Iridoid Glucosides ; administration & dosage ; analysis ; Isocitrate Dehydrogenase ; genetics ; metabolism ; Liver ; drug effects ; metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Rehmannia ; chemistry ; Suppressor of Cytokine Signaling 3 Protein ; genetics ; metabolism

Glioblastoma (GBM) can be classified into molecular subgroups, on the basis of biomarker expression. Here, we classified our cohort of 163 adult GBMs into molecular subgroups according to the expression of proteins encoded by genes of alpha thalassemia/mental retardation syndrome X-linked (ATRX), isocitrate dehydrogenase (IDH) and TP53. We focused on the survival rate of molecular subgroups, depending on each and various combination of these biomarkers. ATRX, IDH1 and p53 protein expression were evaluated immunohistochemically and Kaplan-Meier analysis were carried out in each group. A total of 15.3% of enrolled GBMs demonstrated loss of ATRX expression (ATRX-), 10.4% expressed an aberrant IDH1 R132H protein (IDH1+), and 48.4% exhibited p53 overexpression (p53+). Survival differences were statistically significant when single protein expression or different combinations of expression of these proteins were analyzed. In conclusion, in the case of single protein expression, the patients with each IDH1+, or ATRX-, or p53- GBMs showed better survival than patients with counterparts protein expressed GBMs. In the case of double protein pairs, the patients with ATRX-/p53-, ATRX-/IDH1+, and IDH1+/p53- GBMs revealed better survival than the patients with GBMs with the remained pairs. In the case of triple protein combinations, the patients with ATRX-/p53-/IDH+ showed statistically significant survival gain than the patients with remained combination of proteins-expression status. Therefore, these three biomarkers, individually and as a combination, can stratify GBMs into prognostically relevant subgroups and have strong prognostic values in adult GBMs.
Adult ; Aged ; Biomarkers, Tumor/metabolism ; Brain Neoplasms/*diagnosis/metabolism/mortality ; DNA Helicases/*metabolism ; Disease-Free Survival ; Glioblastoma/*diagnosis/metabolism/mortality ; Humans ; Immunohistochemistry ; Isocitrate Dehydrogenase/*metabolism ; Kaplan-Meier Estimate ; Middle Aged ; Nuclear Proteins/*metabolism ; Retrospective Studies ; Tumor Suppressor Protein p53/genetics/*metabolism ; Young Adult

OBJECTIVETo evaluate the incidence rate of IDH1 in acute myeloid leukemia and analyze its effect on clinical characteristics and prognosis.

METHODSMononuclear cells in bone marrow samples were collected from 192 adult patients with newly diagnosed AML. Polymerase chain reaction (PCR) and direct sequencing were used to amplify exon 4 of IDH1 gene, the gene sequencing was used to analyze the gene mutations, at same time, the detection of NPM1, FLT3-TKD, FLT3-ITD, C-KIT, CEPBA, TET2 and JAK2V617F and MLL mutations were carried out, the follow-up was used to determine its therapeutic efficacy and outcomes of patients. The clinical and laboratory data of these cases were collected, and their clinical characteristics and prognosis were then analyzed.

RESULTSAmong the 192 AML patients, 13 cases were detected with IDH1 gene mutation, the mutation rate was 6.77% [95% CI (5.70%-13.38%)]. The sequencing chart of IDH1 gene showed double peaks, the mutations were heterozygous, out of them c.G395A (p.R132H) was found in 8 cases, c.C394T was found in 4 cases (p.R132C), c.C394A (p.R132S) was found in 1 cases, R132H and R132C are common, 13 cases showed missense mutation. The median age in mutation group was 52 years old, the median age in unnutration group was 40 years, there was significant difference between them (P = 0.010). Mutation rate of IDH1 gene in M1 and M2 was significantly higher than that in other FAB subtypes. There were no significant difference in sex, newly diagnosed peripheral white blood cell count, hemoglobin, platelet count, peripheral blood and bone marrow original cell proportion of primitive cells between them. Mutation of IDH1 gene had certain correlation with NPM1 gene mutation, but no correlation with FLT3-TKD, FLT3-ITD, C-KIT, TET2 and JAK2V617F and MLL natations was found. In addition, the IDH1 mutation easily occurred in patients with normal karyotype or in patients with middle prognostic risk karyotype, IDH1 mutation occurred in 11 cases with normal karyotype, the mutation rate was 10.28%, IDH1 mutation were observed in 2 cases with abnormal karyotype, the mutation rate was 3.50%, there was significant difference. In AML patients with middle prognostic risk karyotype. The complete remission (CR) and the 3 year survival (OS) rate of IDH1 mut patients were less than that in IDH1 wt, there was significant difference (P < 0.05).

CONCLUSIONSThe IDH1 mutation more easily occurr in older AML patients and mutations effect of IDH1 on clinical characteristics may represent a molecular marker for poor prognosis in AML.

Abnormal Karyotype ; Adult ; Exons ; Heterozygote ; Humans ; Isocitrate Dehydrogenase ; metabolism ; Leukemia, Myeloid, Acute ; enzymology ; Leukocyte Count ; Mutation ; Mutation, Missense ; Platelet Count ; Polymerase Chain Reaction ; Prognosis ; Remission Induction ; Survival Rate

OBJECTIVETo study the correlation between IDH1 mutation, MGMT expression, clinicopathologic features and post-radiotherapy prognosis in patients with astrocytoma.

METHODSDetection of IDH1 mutation and MGMT expression was carried out in 48 cases of astrocytoma (WHO grade II to III) by EnVision method with immunohistochemical staining. Follow-up data, including treatment response and overall survival time, were analyzed.

RESULTSThe rates of IDH1 mutation and MGMT expression in astrocytomas were 62.7% (30/48) and 47.9% (23/48), respectively. There was a negative correlation between IDH1 mutation and MGMT expression (r = -0.641, P < 0.01). The age of patients with IDH1 mutation was younger at disease onset. The IDH1 mutation rate in patients with WHO grade II astrocytoma was higher than that in patients with WHO grade III tumor (P < 0.05). The age at onset was an independent factor affecting the expression of mutant IDH1. After radiotherapy, patients with IDH1 mutation+/MGMT- tumor carried a longer overall survival time than patients with IDH1 mutation-/MGMT+ tumor (P < 0.05).

CONCLUSIONSThere is a correlation between IDH1 mutation and MGMT expression in WHO grade II to III astrocytoma. Age at onset is an independent factor affecting the expression of mutant IDH1. Tumors with IDH1+/MGMT- pattern show better response to radiotherapy than tumors with IDH1-/MGMT+ pattern. Detection of IDH1 mutation and MGMT protein expression can provide some guidance in choice of treatment modalities in patients with astrocytoma.

Adult ; Age Factors ; Age of Onset ; Aged ; Astrocytoma ; genetics ; metabolism ; mortality ; pathology ; radiotherapy ; Brain Neoplasms ; genetics ; metabolism ; mortality ; pathology ; radiotherapy ; DNA Modification Methylases ; metabolism ; DNA Repair Enzymes ; metabolism ; Female ; Humans ; Isocitrate Dehydrogenase ; genetics ; Male ; Middle Aged ; Mutant Proteins ; genetics ; Mutation ; Prognosis ; Tumor Suppressor Proteins ; metabolism

OBJECTIVETo correlate the presence of chromosome 1p/19q deletion with the expression of R132H mutant IDH1 status in oligodendroglial tumors, and to explore molecular markers for predicting chemosensitivity of oligodendroglial tumors.

METHODSThe study included 75 oligodendroglial tumors (38 oligodendrogliomas and 37 oligoastrocytomas). Immunohistochemistry was used to detect the expression of R132H mutant IDH1 protein, and fluorescence in situ hybridization (FISH) was employed to detect 1p/19q deletion.

RESULTSDeletion of chromosome 1p and/or 19q was detected in 37 cases (37/75, 49.3%), among which co-deletion of 1p and 19q was seen in 34 cases (closely correlated, P < 0.01). Oligodendrogliomas WHOIIhad a slightly higher deletion rate than oligodendrogliomas WHO III, although without statistical significance. Oligodendrogliomas WHO IIand WHO III had a significantly higher deletion rate of chromosome 1p/19q than oligoastrocytomas WHO II and WHO III (P < 0.05). While combined loss of 1p/19q was always detected in oligodendrogliomas when FISH was positive, isolated 1p or 19q deletion was only found in oligoastrocytomas. The expression of R132H mutant IDH1 was detected in 51 of 75 cases (68.0%), in which oligodendrogliomas had a higher positive rate than oligoastrocytomas. Statistical analysis demonstrated a significant correlation between the expression of R132H mutant IDH1 protein and the presence of combined 1p/19q deletion in oligodendrogliomas (P < 0.05).

CONCLUSIONSA significant correlation was observed between the expression of R132H mutant protein and 1p/19q LOH.Expression of 132H mutant IDH1 protein is the potential biomarker for predicating the presence of 1p/19q deletion in oligodendrogliomas.

Aged ; Brain Neoplasms ; genetics ; metabolism ; Chromosome Deletion ; Chromosomes, Human, 19-20 ; genetics ; Chromosomes, Human, Pair 1 ; Humans ; Immunohistochemistry ; In Situ Hybridization, Fluorescence ; Isocitrate Dehydrogenase ; genetics ; metabolism ; Middle Aged ; Mutant Proteins ; metabolism ; Neoplasm Proteins ; genetics ; metabolism ; Oligodendroglioma ; genetics ; metabolism

The current World Health Organization classification system of primary brain tumors is solely based on morphologic criteria. However, there is accumulating evidence that tumors with similar histology have distinct molecular signatures that significantly impact treatment response and survival. Recent practice-changing clinical trials have defined a role for routine assessment of O-6-methylguanine-DNA methyltransferase (MGMT) promoter methylation in glioblastoma patients, especially in the elderly, and 1p and 19q codeletions in patients with anaplastic glial tumors. Recently discovered molecular alterations including mutations in IDH-1/2, epidermal growth factor receptor (EGFR), and BRAF also have the potential to become targets for future drug development. This article aims to summarize current knowledge on the molecular biology of high-grade gliomas relevant to daily practice.
Aged ; Brain Neoplasms ; genetics ; metabolism ; pathology ; Chromosome Deletion ; DNA Methylation ; DNA Modification Methylases ; genetics ; metabolism ; DNA Repair Enzymes ; genetics ; metabolism ; Glioblastoma ; genetics ; metabolism ; pathology ; Glioma ; genetics ; metabolism ; pathology ; Humans ; Isocitrate Dehydrogenase ; genetics ; metabolism ; Neoplasm Grading ; Oligodendroglioma ; genetics ; metabolism ; pathology ; Point Mutation ; Promoter Regions, Genetic ; Receptor, Epidermal Growth Factor ; metabolism ; Tumor Suppressor Proteins ; genetics ; metabolism

OBJECTIVESTo analyze the long-term survivors of glioblastoma and to identify any prognostic factors that potentially contribute to survival.

METHODSFifteen glioblastomas patients underwent surgery from June 2007 to April 2009 who survived longer than 3 years were enrolled in. Clinical characteristics such as age, location of tumor, extent of resection, and radiotherapy or chemotherapy were analyzed. The expressions of epidermal growth factor receptor (EGFR), tumor protein 53 (P53), phosphatase and tensin homolog (PTEN), O6-methylguanine-DNA methyltransferase (MGMT), isocitrate dehydrogenase 1 gene (IDH1), and neurofibromatosis type 1 (NF-1) in tumor samples were measured by immunohistochemical method, and the status of P53 and IDH1 were detected by direct DNA sequencing as well. And the patients who survived less than 1 year were set as control. Kaplan-Meier analysis was used to evaluate the prognostic factors.

RESULTSThe average age of patients at diagnosis was 45.6 years. And the overall survival time was 3-6 years (median survival time 3.5 years). Thirteen patients underwent a total resection, and 14 patients took orally temozolomide. The occurrence frequency of these molecular markers in long-term survivors was PTEN (13/15), IDH1 (13/15), IDH1 mutation (12/15), P53 (8/15), P53 mutation (7/15), EGFR (6/15), MGMT (4/15) and NF-1 (3/15). There was a good correlation between IDH1 protein expression and IDHI mutation, and between P53 protein expression and P53 mutation. And the survival analysis showed that age above 50 years at diagnosis (OR = 0.262, 95%CI: 0.102 - 0.672), total resection (OR = 0.372, 95%CI: 0.149 - 0.931) and combined oral temozolomide (OR = 0.131, 95%CI: 0.044 - 0.390) were favorable clinical prognostic factors. PTEN (OR = 0.201, 95%CI: 0.074 - 0.549) and IDH1 (OR = 0.151, 95%CI: 0.050 - 0.454) expression, IDH1 mutation (OR = 0.276, 95%CI: 0.108 - 0.709) in tumor cells contributed to a favorable prognosis.

CONCLUSIONSThere is probably no single molecular marker that is responsible for long-term survival of patients with glioblastoma, may be a balance between all these molecular events result in a favorable outcome.

Adult ; Brain Neoplasms ; diagnosis ; metabolism ; Female ; Glioblastoma ; diagnosis ; metabolism ; Humans ; Isocitrate Dehydrogenase ; metabolism ; Kaplan-Meier Estimate ; Male ; Middle Aged ; Mutation ; PTEN Phosphohydrolase ; metabolism ; Prognosis ; Survivors

OBJECTIVETo investigate mutation status of isocitrate dehydrogenase (IDH) 1 and IDH2 genes in Chinese patients with gliomas in correlation with clinicopathological characteristics.

METHODSFormalin-fixed and paraffin-embedded (FFPE) tissue samples of 234 gliomas were collected including the matched blood samples in 30 patients. DNA was extracted, followed by PCR-Sanger sequencing to detect IDH1 and IDH2 gene mutations. Immunohistochemistry was performed using mutation-specific antibody recognizing IDH1R132H mutation. Immunostains for p53 and epidermal growth factor receptor (EGFR) were also performed. Oligodendroglial tumors with IDH mutation were double stained with IDH1R132H and GFAP by immunofluorescence to investigate the location of IDH1R132H expression.

RESULTS(1) By IDH1 heterozygous somatic mutation analysis, Arg132His (c: G395A) was found in 31.6% (74 of 234) of the cases. IDH mutations were more frequent in oligoastrocytomas (9/13), anaplastic oligoastrocytomas (7/11), oligodendrogliomas(18/26, 69.2%), anaplastic oligodendrogliomas (8/10), and less frequent in diffuse astrocytomas (17/47, 36.2%), anaplastic astrocytomas (5/18), and glioblastomas (10/69, 14.5%). The mutation rate inversely correlated with the tumor grade in a linear fashion in astrocytic tumors (P = 0.007). Primary glioblastomas were characterized by a lower frequency of mutations than secondary glioblastomas (5/55 vs. 5/14, P = 0.036); IDH mutation was not detected in pilocytic astrocytoma and ependymoma. No IDH2 mutation was identified in this study cohort. (2) Immunohistochemistry of IDH1R132H demonstrated a strong cytoplasmic staining in 80 cases, which was highly correlated with IDH mutation status (P = 0.001). IDH1R132H was highly specific to tumor cells. (3) p53 immunostain was significantly correlated the IDH mutation in diffuse astrocytoma, anaplastic astrocytoma and secondary glioblastomas (P = 0.007, 0.026, 0.038 respectively). (4) No correlation between EGFR and IDH mutation was found.

CONCLUSIONSHigh prevalence of IDH heterozygous somatic mutation occurs in the earlier stage of gliomas, which can be detected by mutation-specific antibody IDH1R132H. Furthermore, evaluation of p53 and EGFR expression combined with IDH mutation analysis may significantly aid in the diagnosis and differential diagnoses of gliomas in Chinese patients.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Astrocytoma ; genetics ; metabolism ; Brain Neoplasms ; genetics ; metabolism ; Child ; Ependymoma ; genetics ; metabolism ; Female ; Glioblastoma ; genetics ; metabolism ; Glioma ; genetics ; metabolism ; Humans ; Isocitrate Dehydrogenase ; genetics ; metabolism ; Male ; Middle Aged ; Oligodendroglioma ; genetics ; metabolism ; Point Mutation ; Receptor, Epidermal Growth Factor ; metabolism ; Tumor Suppressor Protein p53 ; metabolism ; Young Adult

Adenosine Triphosphate ; metabolism ; Colorectal Neoplasms ; diagnosis ; genetics ; DNA Methylation ; Diphosphates ; metabolism ; Glioma ; diagnosis ; genetics ; Humans ; Isocitrate Dehydrogenase ; genetics ; Janus Kinase 2 ; genetics ; Mutation ; Myeloproliferative Disorders ; diagnosis ; genetics ; Polymorphism, Single Nucleotide ; Proto-Oncogene Proteins ; genetics ; Proto-Oncogene Proteins p21(ras) ; Sequence Analysis, DNA ; methods ; Templates, Genetic ; ras Proteins ; genetics