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L F Eng

Publications and source records attributed to L F Eng.

At least 73 records · Page 4Linked to original sources
PubMed1987

The cytoskeleton of the human cerebellar cortex: an immunohistochemical study of normal and pathological material.

Monoclonal antibodies to non-phosphorylated and phosphorylated neurofilaments, as well as monoclonal and polyclonal antibodies to other cytoskeletal elements, were applied to the study of the cerebellar cortex of normal and pathological human material. The methods proved to be applicable to formalin fixed paraffin embedded tissue, provided the period of formalin fixation was short. The main difference between normal and pathological material was found in Purkinje cells and their dendrites. While normal Purkinje perikarya and dendrites expressed only non-phosphorylated neurofilaments, reactive dendrites stained more intensely with antibodies to phosphorylated neurofilaments. Similar observations were made on the abnormal dendritic ramifications of the partially deafferented, hypertrophic, inferior olive. The significance of the appearance of phosphorylated neurofilament epitopes in abnormal dendrites remains unknown and requires further investigation.

Adult↗
PubMed1987

Glial fibrillary acidic protein in chronic relapsing experimental allergic encephalomyelitis in SJL/J mice.

The gliotic scar in the demyelinated plaque is a prominent feature of the pathology of multiple sclerosis. Chronic relapsing experimental allergic encephalomyelitis (EAE) in the SJL/J mouse has many characteristics in common with multiple sclerosis in the human, including the development of intense gliosis during the course of the demyelinating disease. With the use of antibody to the astrocyte marker glial fibrillary acidic protein (GFAP) we have measured the increase of GFAP over an 11-month course of chronic EAE. Intense staining of astrocyte fibers was seen around EAE lesions, which were most frequently observed in the cerebellum, periventricular areas, and in the spinal cord. The relative amount of GFAP was estimated by preparation of cytoskeletal proteins from the affected CNS areas, separation of proteins by polyacrylamide gel electrophoresis, and quantitation of GFAP in relation to the 70-kD neurofilament protein (NF) in gel scans. The ratio GFAP/70-kD NF protein in control animals did not change significantly over 11 months, whereas this ratio gradually increased to 2.54 in animals with chronic relapsing EAE 6 months after immunization. Although some decrease of neural fibers may have contributed partially to this change in ratio, the amounts of GFAP were greatly increased. These results indicate that the SJL/J mouse with chronic relapsing EAE provides an excellent model with which to investigate the formation and development of the gliotic plaque analogous to that seen in demyelinated areas in multiple sclerosis tissue.

Animals↗
PubMed1987

Molecular differentiation of neurons from ependyma-derived cells in tissue cultures of regenerating teleost spinal cord.

Cells cultured from the caudalmost area of regenerating teleost spinal cord differ both morphologically and in terms of molecular architecture from those of more rostral (more fully differentiated) areas of the cord. The caudalmost regenerating cord consists of an ependymal tube. Cells from this region have flattened or partially flattened cell somas and short spike projections in vitro; they do not exhibit the rounded cell somas nor the long, thin, branching neurites typical of differentiated neurons. A series of cultures taken from different areas along the length of regenerating spinal cord were examined for molecular differentiation by staining with a monoclonal antibody against non-phosphorylated neurofilament protein (SMI 32). None of the cells from the caudalmost culture of the regenerating spinal cord stained with antibody SMI 32. In cultures of more rostral regenerated cord, the cells with neuronal morphology do stain positively with the anti-neurofilament antibody. This result suggests that the cells in the cultures of caudalmost cord represent relatively undifferentiated ependymal cells, or ependyma-derived cells, which later differentiate into neurons and glia in the regenerating spinal cord.

Animals↗
PubMed1987

Immunohistopathologic spectrum of the glial fibrillary acidic protein in neurooncology.

To conventional histologic and special staining techniques as well as electron microscopy, immunohistological methods have recently been added to characterize neuroectodermal and non-neuroectodermal tumors of both the central and the peripheral nervous system. An ever increasing panel of these morphologic techniques enables us today, to characterize precisely such tumors in or around the central and peripheral nervous system, sometimes only hampered by preservation of tissue inadequate for the respective methods.

Astrocytes↗
PubMed1986

Reaction of Lewy bodies with antibodies to phosphorylated and non-phosphorylated neurofilaments.

The reaction of Lewy bodies in the substantia nigra and the sympathetic ganglia was studied in 10 cases of Parkinson's disease and other Lewy body-related conditions, using monoclonal antibodies to non-phosphorylated (02-40) and phosphorylated (03-44 and 07-5) neurofilaments. Although many Lewy bodies were unstained, a peripheral ring of positive immunoreactivity was frequently observed. This reaction was more common and more intense with antibodies to phosphorylated neurofilaments, suggesting a posttranslational phosphorylation of elements in the Lewy body. Although Lewy bodies appear to have some antigenic sites in common with neurofilaments, certain differences between neurofilaments and Lewy body filaments are emphasized.

Antibodies, Monoclonal↗
PubMed1986

Astrocyte culture on nitrocellulose membranes and plastic: detection of cytoskeletal proteins and mRNAs by immunocytochemistry and in situ hybridization.

Neonatal rat brain astrocyte secondary cultures were established on nitrocellulose membrane filters (13-mm diameter Millipore disk) and on plastic coverslips in serum-supplemented medium. On these substrata, cultured astrocytes changed their shape from flat and polygonal to stellate in the absence of hormones or growth factor supplements. Cultures became confluent after 4 days, and astrocytes on nitrocellulose filters continued to differentiate morphologically and biochemically, as evidenced by extensive cytoplasmic process formation and glial fibrillary acid protein (GFAP) accumulation. Cultures were immunostained for GFAP and vimentin. mRNAs to GFAP, vimentin, alpha and beta tubulin, and actin also were detected by in situ hybridization with biotinylated cDNA probes. The astrocyte culture method on nitrocellulose provides a simple, versatile means of comparing undifferentiated, morphologically mature, reactive, and neoplastic astrocytes in vitro.

Animals↗
PubMed1986

Specific chromosomal abnormalities characterize four established cell lines derived from malignant human gliomas.

The four permanent human glioma-derived cell lines reported here are the first such lines for which the karyotypes have been followed from the original biopsies through the establishment of the lines in culture. Although ploidy changes were seen, each line retained either distinctive marker chromosomes or the overall original chromosomal distribution allowing the origin of each line to be established with certainty. D-263 MG expresses glial fibrillary acidic protein, all lines except D-245 MG are tumorigenic in athymic mice, and each line displays a unique pattern with respect to in vitro growth parameters and expression of biochemically defined markers, oncofetal antigens and lymphoid-associated markers. D-245 MG and D-259 MG are able to grow in the absence of supplemental glutamine; glutamine synthetase was detected in these cell lines both by immunocytochemistry and by direct assay. Thus, the four permanent human glioma-derived cell lines described here are representative of glioma lines in their general characteristics. D-259 MG retains numerous double minute chromosomes (DMs), D-263 MG contains two marker chromosomes with breaks in 9p, and D-247 MG and D-245 MG with stemlines containing 96 and 89 chromosomes contain eight and six normal copies (respectively) of chromosome No. 7. The retention in these four cell lines of the most common chromosomal abnormalities seen in biopsies of malignant human gliomas provides the opportunity to investigate the meaning of these specific chromosomal changes.

Animals↗
PubMed1986

Alpha tocopherol decreases lipid peroxidation, neuronal necrosis, and reactive gliosis in reaggregate cultures of fetal rat brain.

This study explored the effects of the lipid-soluble free radical scavenger, alpha tocopherol (vitamin E), on neuronal injury and glial protein accumulation in a well-characterized, three-dimensional, mixed neuronal and glial culture system derived from fetal rat prosencephalon. As these reaggregated spheroidal cultures grew and enlarged, they developed small central areas of necrosis (presumably due to nutritional compromise) which we used as a model for central nervous system injury. Treatment with vitamin E (delivered in egg phosphatidylcholine liposomes beginning on day 8 in vitro) did not alter the appearance of the central necrotic areas, but it strongly suppressed the reaction of adjacent astroglia and microglia. Histological examination showed that by day 35 in vitro control cultures which received liposomes without tocopherol were nearly devoid of neurons and contained many glial and microglial cells. In contrast, tocopherol-treated cultures contained many viable-appearing neurons and did not exhibit an overgrowth of glia. Both glial fibrillary acidic protein (as measured by immunoassay) and lipid peroxidation (as estimated by malondialdehyde levels) were markedly reduced in the tocopherol-treated cultures. We speculate that the vitamin exerts its protective effect on injured nervous tissues by scavenging free radicals, stabilizing cellular membranes, and quenching the cascade of biochemical events that follows necrosis in brain. This work suggests that the signal(s) to initiate gliosis are mediated, at least indirectly, by free radical formation.

Animals↗
PubMed1986

The immunohistochemical application of three anti-GFAP monoclonal antibodies to formalin-fixed, paraffin-embedded, normal and neoplastic brain tissues.

The intermediate filament, glial fibrillary acidic protein (GFAP) has proven to be an important glial marker in diagnostic neuropathology. We report the histochemical application of three monoclonal antibodies (Mab) produced in this laboratory, 1B4, 2E1, and 4A11, which are monospecific to GFAP by radioimmunoassay, immunoblot electrophoresis, and immunoperoxidase histochemistry. The goal of this study was to compare the specificity and sensitivity of these Mab to GFAP on surgical brain biopsy specimens which had been routinely processed for diagnostic neuropathology with that of a high titer, highly specific, reference polyvalent anti-GFAP antiserum. The Mab stained astrocytes specifically in normal brain. When combined in a "cocktail" preparation, the quality of the immunoperoxidase detection of GFAP by these Mab closely approached that of the reference serum in 71 intracranial and intraspinal neoplasms. As these three Mab represent a continuous supply of a well defined, monospecific reagent, the monoclonal "cocktail" represents a standard reagent for large multi-institutional studies and for studies extending over a period of time.

Antibodies, Monoclonal↗
PubMed1986

A glial fibrillary acidic protein-expressing and tumorigenic cell line derived from an avian sarcoma virus-induced rat astrocytoma.

A permanent cell line, S635c15, was derived from an anaplastic astrocytoma induced by the Schmidt-Ruppin strain of avian sarcoma virus (ASV) in a female F-344 rat. Persistent expression of the astrocytic differentiation protein, glial fibrillary acidic protein (GFAP), was detected both in cultured cells after 100 passages in vitro and in transplanted tumors. Subcutaneous and intracerebral transplantation of S635c15 cells in syngeneic rats resulted in a 100% tumor incidence and a reproducible mortality distribution. S635c15 cells formed discrete masses after subcutaneous injection but grew intracranially as infiltrative lesions. Tumor blood flow and blood-to-tissue transport studies yield comparable values to other rat glioma models; S635c15 intracranial tumors proved to be a homogeneous model with little variation within and between tumors with respect to morphology, GFAP expression, blood flow, and permeability. This cell line provides a GFAP-expressing brain tumor model that extends the use of autochthonous ASV-induced astrocytomas by allowing in vitro and in vivo studies. It may be useful for further studies in neurobiology and brain tumor biology, diagnosis, and therapy.

Animals↗
PubMed1986

Turnover of glial filaments in mouse spinal cord.

Twenty-day-old mice received a single tail vein injection of [guanido-14C]arginine. The cytoskeleton was extracted from the spinal cords at varying lengths of time thereafter. Glial fibrillary acidic protein (GFAP) formed a distinct, broad band that was widely separated from other protein bands in one-dimensional polyacrylamide gels. The purity of the GFAP band was verified by Western blot analysis of one- and two-dimensional electrophoretic patterns. In addition, enzyme-linked immunosorbent assay and quantitative Western blot analysis indicated that 95% of the total spinal cord GFAP was extracted in the cytoskeletal preparation. The specific activity of GFAP was obtained by eluting the protein from the cytoskeletal GFAP band in preparative one-dimensional gels. Specific activity reached a peak 2 h after injection with [14C]arginine. Forty percent of the incorporated radioactivity was still present in cytoskeletal GFAP at 9 weeks, indicating that a significant proportion of glial filaments turns over relatively slowly in vivo.

Animals↗
PubMed1985

Immunocytochemical localization of glial fibrillary acidic protein (GFAP) in the area postrema of the cat. Light and electron microscopic study.

Glial fibrillary acidic protein (GFAP) was demonstrated in the cytoplasm and processes of ependymal cells and astroglial components of the area postrema of the cat. These observations differ from the findings in the ependyma of the ventricular cavities which are consistently negative for the protein. Since some studies have suggested sensory functions of the glial cells in this emetic chemoreceptor trigger zone, a careful consideration of morphological and biochemical attributes of these cells seems appropriate.

Animals↗
PubMed1985

Hormones and growth factors induce the synthesis of glial fibrillary acidic protein in rat brain astrocytes.

Glial fibrillary acidic protein (GFAP) is the major constituent of glial filaments and is restricted within the CNS to astrocytes. As with other classes of intermediate filament proteins, the regulation of GFAP expression is poorly understood. Utilizing highly purified cultures of astrocytes and a chemically defined (CD) medium, we have demonstrated that the expression of GFAP is subject to regulation by hormones and growth factors. The concentration of GFAP/mg protein was induced 2-4-fold in the presence of hydrocortisone, putrescine, prostaglandin F-2 alpha (PGF2 alpha), and pituitary fibroblast growth factor (FGF). Augmentation of the levels of GFAP continued for up to 3 weeks after conversion to CD medium and paralleled the morphological maturation of astrocytes. The accumulation of GFAP resulted from an increase in its specific rate of synthesis. Conversion of astrocytes from serum-supplemented (SS) to CD medium did not alter its rate of degradation. GFAP appeared quite stable under both sets of conditions, exhibiting a half-life of approximately 7.5 days. The data demonstrate that GFAP expression in astrocytes is subject to hormonal regulation, which may have implications for gliosis.

Animals↗
PubMed1985

Immunohistochemical study of glial fibrillary acidic protein in avian sarcoma virus-induced gliomas in dogs.

Anaplastic astrocytomas, gliosarcomas and sarcomas were induced in 33 neonatal dogs by intracerebral injection of purified Schmidt-Ruppin strain of avian sarcoma virus (ASV). A total of 11 anaplastic gliomas, 7 gliosarcomas and 18 sarcomas were induced. Ten of the 11 anaplastic astrocytomas and all 7 gliosarcomas were positive for GFAP using the peroxidase-antiperoxidase method on formalin-fixed, paraffin embedded tissue. None of the sarcomas were positive for GFAP. One anaplastic astrocytoma was analyzed by gel electrophoresis and immunoblot and the amount of GFAP in the tumor was compared with comparable samples from normal white and gray matter. Densitometric analysis of the GFAP stained gels showed that 12-13% of tumor protein was GFAP compared with 2-3% for the non-neoplastic white and gray matter. The results of this study add further evidence to the astrocytic origin of many of the induced anaplastic primary brain tumors by ASV.

Animals↗
PubMed1985

Monoclonal antibodies reactive with epitopes restricted to glial fibrillary acidic proteins of several species.

The highly reproducible histochemical localization of glial fibrillary acidic protein (GFAP)+ qualifies it as an important marker of astrocytes in both research and clinical applications. The primary objective of this study was to produce monoclonal antibodies having the advantage of invariant specificity, affinity, and titer to GFAP-specific epitopes of wide species distribution. We report here the characterization of four monoclonal antibodies that recognize the same or spatially close epitopes specific to GFAP. The epitope(s) detected has been phylogenetically conserved; human, bovine, ovine, canine, porcine, rabbit, guinea pig, rat, murine, and chicken brain homogenates all specifically absorb monoclonal antibody activity. Of importance to the routine application of these new anti-GFAP monoclonal antibodies is the demonstration here of the stability of the antigen-antibody interaction in normal, reactive, and neoplastic astrocytes of both rat and human origin following various methods of fixation.

Animals↗
PubMed1985

A serially transplantable human giant cell glioblastoma that maintains a near-haploid stem line.

We have karyotyped a human giant cell glioblastoma removed from an 11-year-old girl and have established from it a subcutaneously transplantable line in athymic nude mice. The original tumor contained near-haploid cells with 25 or 26 chromosomes, including two copies of #1, (7 or 7p+) and #18. There were also hyperdiploid (49-52) cells that were tetraploid for these same three chromosome types; doubled versions of the hyperdiploid population were also seen. The stemline of the mouse-grown tumor was 26,X, +1, +7p+, +18 in the first passage and has remained consistently near-haploid through ten serial in vivo passages. Growth stabilization has occurred with an average latency of less than 3 months. This transplantable line is available for evaluating chemotherapeutic responsiveness of human giant cell glioblastoma and for studying near-haploidy in solid human tumors.

Animals↗
PubMed1985

Glial fibrillary acidic protein (GFAP): the major protein of glial intermediate filaments in differentiated astrocytes.

The glial fibrillary acidic protein (GFA protein or GFAP) is the major protein constituent of glial intermediate filaments in differentiated fibrous and protoplasmic astrocytes of the central nervous system. Proteins having similar molecular weights, isoelectric points, and immunoreactivity with GFAP have been found in cells of neural crest and ectodermal origin. A putative function ascribed to glial filaments is its role as a component of the cytoskeleton in defining and maintaining the shape of the astrocyte. Since 1980, over 350 reports have utilized antisera to GFAP for immunochemical and immunocytochemical studies.

Animals↗