1-Bromopropane is used as a cleaning agent or adhesive solvent in the workplace. The present study investigated the long-term effects of exposure to 1-bromopropane on biochemical components in the ...central nervous system (CNS) of rats. Four groups, each of nine male Wistar rats, were exposed to 200, 400, or 800
ppm 1-bromopropane or fresh air only, 8
h per day, 7 days a week for 12 weeks. We measured the levels of neuron-specific γ-enolase, glia-specific β-S100 protein, creatine kinase (CK) subunits B and M, heat shock protein Hsp27 (by enzyme immunoassay), enzymatic activity of CK and levels of glutathione (GSH), oxidized glutathione (GSSG) and sulfhydrul (SH) base in the cerebrum, cerebellum, brainstem and spinal cord. γ-Enolase decreased dose-dependently in the cerebrum, which showed a decrease in wet weight, at 400
ppm or over, but no change was noted in β-S100 protein in any brain region or spinal cord. Hsp27 decreased in the cerebellum, brainstem and spinal cord. Protein-bound SH base, non-protein SH base and total glutathione decreased in every brain region. CK activity decreased dose-dependently at 200
ppm or over, and the ratio of CK activity to CK-B concentration tended to decrease in all regions. The decrease in γ-enolase in the cerebrum suggests the involvement of biochemical changes in neurons with decrease in the wet weight of the cerebrum. Glutathione depletion and changes in proteins containing SH base as a critical site might be the underlying neurotoxic mechanism of 1-bromopropane. The biochemical changes in the cerebrum indicate that long-term exposure to 1-bromopropane has effects on the CNS.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
1-Bromopropane is used widely as an alternative to ozone-depleting solvents. The neurotoxic effects of this agent have been described in humans and experimental animals. Here we investigated the ...underlying mechanisms of the neurotoxic effects of 1-bromopropane by examining the initial biochemical changes in the central nervous system. Four groups of 9 Wistar male rats each were exposed to 200, 400, or 800 ppm 1-bromopropane or only fresh air, 8 h per day for 7 days. At the end of the experiment, the cerebrum, cerebellum, brain stem and lumbar enlargement of the spinal cord were dissected out from each rat (n = 8) for biochemical analyses. Morphological examinations of the nervous system were performed in the remaining rat of each group. 1-Bromopropane dose-dependently decreased neurospecific γ-enolase, total glutathione, and nonprotein sulfhydryl groups in the cerebrum and cerebellum. Creatine kinase activity decreased dose-dependently in the brain and spinal cord. Histopathological examination showed swelling of preterminal axons in gracile nucleus and degeneration of myelin in peripheral nerves. Our results of low levels of γ-enolase suggested that 1-bromopropane might primarily cause functional or cellular loss of neurons in the cerebrum and cerebellum. Glutathione depletion or modification to functional proteins containing a sulfhydryl base as a critical site might be the underlying mechanism of 1-bromopropane neurotoxicity.
: We have examined the regulation of neuron‐specific γ‐enolase gene (NSE) expression in oligodendrocytes at various steps of their differentiation/maturation. We have demonstrated for the first time ...that NSE is expressed in oligodendroglial cells in vitro and in vivo, and only at a certain stage of differentiation. A heterogeneity of the γ subunit was observed in cultured oligodendrocytes and the same one was found in adult rat brain. The level of γ mRNA increased when precursor cells differentiated into oligodendrocytes. By contrast, no significant change in α‐enolase gene expression was observed. High NSE (γγ and αγ) enolase activity was detected in cultured oligodendrocytes. Treatment with basic fibroblast growth factor, which stimulates the proliferation of oligodendrocyte precursor cells and reversibly blocks their differentiation, resulted in lower αγ‐ and γγ‐enolase activities in these cells, but it enhanced αα‐enolase activity slightly. These data indicate that γ‐enolase gene expression is associated with the differentiation of the oligodendrocytes and that it is repressed in adult fully mature cells.
Background. Increased levels of gamma‐enolase (γ‐enolase) have been observed in the sera of patients with renal cell carcinoma. To evaluate the prognostic information of γ‐enolase in this disease, ...161 consecutive patients were assessed before initiation of therapy.
Methods. γ‐Enolase was analyzed in serum using an immunoradiometric assay. The patients were clinically staged and followed up for a median time of 36 months (range, 5–104 months). Actuarial survival was calculated using the Kaplan‐Meier method.
Results. Elevated levels of γ‐enolase was found in 28 of 61 (46%) patients with distant metastases, compared with 8 of 56 (14%) when the tumor was confined to the kidney. A correlation also was observed between γ‐enolase and tumor grade, with poorly differentiated tumors having the highest levels. In 28 patients with distant metastases and elevated γ‐enolase, the survival time was significantly shorter than that of 31 patients with normal γ‐enolase levels (P < 0.001). The median survival time was 5 and 11 months, respectively. Using Cox proportional hazard model, clinical stage, serum γ‐enolase, and tumor grade were identified as independent prognostic factors.
Conclusion. Serum γ‐enolase can be useful as an adjunct in the staging of renal cell carcinoma. It also gives predictive information and might be of value as a marker in adjuvant therapy. Cancer 1993; 72:1324‐8.
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BFBNIB, FZAB, GIS, IJS, KILJ, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
The isoforms of gamma-enolase were characterized in serum from patients with small-cell lung cancer (SCLC) and in extracts from SCLC cell lines and malignant melanoma tumor tissue. Large variations ...in the expression of the 3 gamma-isoforms of enolase were observed. These forms probably represent the homodimeric gamma gamma-enolase, the heterodimeric alpha gamma-enolase and the monomeric forms of gamma-enolase. Only the dimeric forms are enzymatically active. The predominant gamma-enolase in the cell lines is the heterodimeric alpha gamma-enolase. The SCLC cell lines can be divided into two groups: one with negligible gamma gamma-enolase expression and considerable amounts of the nonneuronal alpha alpha-enolase and a second group with a large fraction of gamma gamma-enolase concomitant with a low expression of alpha-enolase. Similar patterns are observed in tissue extracts from malignant melanoma. When changing buffer conditions by increasing the ionic strength and decreasing the Mg2+ concentration, interconversions between the isozymes occur. In contrast to the predominant alpha gamma-enolase in extracts from cell lines, the multiple forms of gamma-enolase in serum might be caused by a subunit exchange facilitated by the low Mg2+ concentration in plasma. However, there seems to be a stable equilibrium between the isoforms in undiluted patient serum. The induction of subunit exchange by perturbation in ionic strength and/or Mg2+ concentration indicates a need for caution when choosing diluents for use in assays for neuron-specific enolase.
The behavior of marker proteins of neurons (gamma-enolase) and glial cells (alpha-enolase, beta-S100 protein and creatine kinase-B) was investigated quantitatively by using enzyme immunoassay systems ...in toluene-exposed rat brains. Three groups of animals were exposed to toluene vapor at 300 ppm, 1000 ppm, and 3000 ppm, respectively, 8 h/day, 6 days/week, for 2 weeks. After subacute repeated solvent exposure, both neuron-specific gamma-enolase and glial marker proteins displayed an overall concentration-dependent increase tendency in separate brain regions. In cerebrum, only the 3000 ppm group showed a significant increase in alpha-enolase by 27% and creatine kinase-B (CK-B) by 26%. alpha-Enolase and gamma-enolase exhibited a pronounced elevation in cerebellum relative to other brain regions, while beta-S100 protein appeared to be the most markedly altered marker in brainstem. The development of gliosis, which is a frequent phenomenon following CNS damage, is presumed to be responsible for the elevation of glial marker content. Energy metabolism disruption in brain tissues may also bring about the compensatory oversynthesis of glycolytic enzymes such as gamma-enolase, alpha-enolase and CK-B. The dose-dependent alteration patterns following toluene exposure suggest the feasibility of using these brain specific markers to evaluate solvent-induced CNS effects.
Chicken γ-enolase cDNA was cloned and its sequence and tissue-specific expression were analyzed. The cDNA, consisting of 2273
bp of nucleotides, was composed of 86
bp of the 5′-noncoding region, 1305
...bp of an open reading frame encoding a protein of 434 amino acids and 882
bp of the 3′-noncoding region. The deduced amino acid sequence showed higher homology (more than 90%) to those of mammalian γ-enolases than to those of chicken α- or β-enolases. Northern blot analysis has revealed that the mRNA for γ-enolase (2.3
kb) is expressed in the brain and, to much less but significant extents, in the pituitary and adrenal gland of the chicken.
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IJS, IMTLJ, KILJ, KISLJ, NUK, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
We have isolated and sequenced a cDNA clone encoding the human gamma enolase. Comparison of our cDNA sequence and the rat gamma enolase sequence revealed 97% homology at the level of amino acid ...sequence. The two coding regions were 91% homologous on the nucleotide level, whereas the 3' noncoding regions were much less homologous (32%). Further comparison of our cDNA sequence with the human alpha enolase revealed an 82% homology at the amino acid level and a 75% homology at the nucleotide level for the two coding regions, whereas the 3' nontranslated regions were only 30% homologous. Using a portion of the 3' nontranslated region of our cDNA, shown to be specific for human gamma enolase, a single 2.5 kb mRNA was detected in human brain tissue. This same gamma enolase message was also found in a number of human normal nonneuronal tissues, and in several human tumor-derived cell lines. Expression of the mRNA for the gamma enolase subunit should thus be used with caution when identifying the cells of neuronal or neuroendocrine origin.
Three cases of cerebellar hemangioblastoma were studied using the immunoperoxidase technique to localize gamma-enolase, also known as neuron-specific enolase. The stromal cells demonstrated positive ...staining for gamma-enolase, while endothelial cells and pericytes showed no reactivity. Two vascular lesions, an angiosarcoma and a cutaneous angioma, were studied and found to be nonreactive for gamma-enolase. All tumors were also tested for factor VIII/von Willebrand factor, glial fibrillary acidic protein, and the S-100 protein. The lack of expression of gamma-enolase in endothelial cells of hemangioblastomas demonstrates a clear antigenic distinction from neighboring gamma-enolase-positive stromal cells. The significance of this finding and its implications for stromal cell histogenesis are discussed.
Human gamma-enolase cDNA prepared by reverse transcriptase-polymerase chain reaction was cloned into the Escherichia coli expression vector pKK223-3. The resulting plasmid, pHTK503, expressed human ...gamma-enolase as a 46-kDa protein in SDS-PAGE, and in the cells as the active gamma gamma form (designated as recombinant human NSE; R-NSE). R-NSE was purified from E. coli by several chromatographic elutions. Finally, 6.0 mg of R-NSE from 8.1 g cells was purified with a specific activity of 86 units/mg protein. The structural properties of R-NSE were compared with the NSE purified from human brain tissue (B-NSE). The biochemical and enzymatic characteristics were essentially the same, except for the isoelectric point (4.5 for B-NSE and 4.7 for R-NSE). In an NSE immunoassay system, R-NSE and standard NSE were almost equal in reactivity to the anti-NSE antibody. These results indicate that R-NSE can be used as standard assay material.
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