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Notes: Summary A rabbit antiserum against mouse neonatal brain cell surface membranes labeled by immunoperoxidase (PAP) the cells of the central and peripheral nervous systems of adult and neonatal mice and their processes, as well as the differentiating neuroepithelial cells of three OTT-6050 mouse teratoma-derived tumors. Indirect immunofluorescence on living 14-day-old monolayer cultures of neonatal mouse brain demonstrated reaction of the immune serum with external surface membrane antigens of neuroblasts and of primitive and mature glial cells. Radioimmune assays (RIA) showed almost complete loss of antiserum binding to neonatal mouse brain plasma membranes after absorption with adult or neonatal mouse brain membranes, and no loss of binding after absorption by liver, spleen, kidney, and heart membranes. Cross-reactivity of the immune serum to several non-neural cell types was demonstrated by immunoperoxidase on sperm and sperm-precursors, on moderate numbers of epithelial cells in the medulla of adult mouse thymus, and, in the neonate, on a range of mesenchymal cells. This cross-reactivity was reflected in the RIA by a moderate reduction of immune serum binding to neonatal mouse brain plasma membranes after absorption with testis pellets and with thymus membranes. PAP staining showed loss of crossreactivity after testis or thymus absorption, without climination of neural cell recognition. Absorption with adult or neonatal mouse brain eliminated cross-reactivity. In the teratoma-derived tumors, absorption of the antiserum with testis or thymus eliminated or markedly reduced the PAP staining of primitive neuroepithelial cells, and only moderately reduced, but did not remove, that of neural cells in the mature neuropil. Among the proteins of neonatal mouse brain plasma membranes separated by polyacrylamide gel electrophoresis, there were six distinct bands indicating major proteins ranging from 26,000–54,000 daltons. Autoradiography of the antigen-antibody complexes with125I protein A on the same gels demonstrated three discrete bands of activity at 10,000–12,000, 76,000, and 97,000 daltons, and one greater than 130,000 daltons, suggesting that the immune serum recognizes only minor protein components of the mouse brain plasma membranes. The application of the PAP method to the recognition of neural cell surface antigens considerably enhances the potential of this antiserum as a tool for the early identification of primitive neural cells in the experimental mouse teratoma.

Notes: Abstract— Non-histone chromosomal proteins (NHCP) from mouse brain at different stages of development and from adult liver and kidney of strain related mice were analyzed by SDS-polyacrylamide gel electrophoresis and were compared with the mouse teratoma, OTT-6050. The fetal, neonatal and adult brains were qualitatively similar in their NHCP profiles but had quantitative differences. The NHCP composition of the adult brain was clearly distinct from that of the liver and kidney and was dissimilar from that of the teratoma.

Notes: Abstract The cellular nature of the giant eosinophilic cells of tuber and of the cells comprising subependymal giant cell astrocytoma (SEGA) in tuberous sclerosis (TS) remains unclear. To assess the characteristics of these lesions, 13 tubers and 6 SEGA were immunohistochemically studied with glial and neuron-associated antigens. In addition to conventional ultrastructure, 6 tubers and 8 SEGA were fibrillary acidic protein (GFAP) and somatostatin. Eosinophilic giant cells of tubers were positive for vimentin (100%), GFAP (77%) and S-100 protein (92%); such cells were also found to a various extent to be reactive for neuron-associated antigens, including neurofilament (NF) proteins (38%) or class III β-tubulin (77%). SEGA also showed variable immunoreactivity for GFAP (50%) or for S-100 protein (100%); NF epitopes, class III β-tubulin, and calbindin 28-kD were expressed in 2 (33%), 5 (83%) and 4 (67%) cases, respectively. Cytoplasmic staining for somatostatin (50%), met-enkephalin (50%), 6-hydroxytryptamine (33%), β-endorphin (33%) and neuropeptide Y (17%) was noted in SEGA, but not in tubers. Ultrastructurally, the giant cells of tubers and the cells of SEGA contained numerous intermediate filaments, frequent lysosomes and occasional rectangular or rhomboid membrane-bound crystalloids exhibiting lamellar periodicity and structural transition to lysosomes. Some SEGA cells showed features suggestive of neuronal differentiation, including stacks of rough endoplasmic reticulum, occasional microtubules and a few dense-core granules. Furthermore, in one case of tuber, a process of a single large cell was seen to be engaged in synapse formation. Intermediate filaments within a few cells of both lesions were decorated by gold particle-labeled GFAP antiserum. Within the tumor cells of SEGA, irregular, non-membrane-bound, electron-lucent areas often contained somatostatin-immunoreactive particles, whereas the latter could not be detected in tuber. The present study provides further evidence of divergent glioneuronal differentiation, both in the giant cells of tubers and the cells of SEGA. The findings of similar cells at different sites, including the subependymal zone, white matter (“heterotopias”), and cortex indirectly supports the idea that these lesions of TS result from a migration abnormality.

Notes: Abstract The cellular nature of the giant eosinophilic cells of tuber and of the cells comprising subependymal giant cell astrocytoma (SEGA) in tuberous sclerosis (TS) remains unclear. To assess the characteristics of these lesions, 13 tubers and 6 SEGA were immunohistochemically studied with glial and neuron-associated antigens. In addition to conventional ultrastructure, 6 tubers and 8 SEGA were subjected to immunoelectron microscopic study for glial fibrillary acidic protein (GFAP) and somatostatin. Eosinophilic giant cells of tubers were positive for vimentin (100%), GFAP (77%) and S-100 protein (92%); such cells were also found to a various extent to be reactive for neuron-associated antigens, including neurofilament (NF) proteins (38%) or class III β-tubulin (77%). SEGA also showed variable immunoreactivity for GFAP (50%) or for S-100 protein (100%); NF epitopes, class III b-tubulin, and calbindin 28-kD were expressed in 2 (33%), 5 (83%) and 4 (67%) cases, respectively. Cytoplasmic staining for somatostatin (50%), met-enkephalin (50%), 5-hydroxytryptamine (33%), β-endorphin (33%) and neuropeptide Y (17%) was noted in SEGA, but not in tubers. Ultrastructurally, the giant cells of tubers and the cells of SEGA contained numerous intermediate filaments, frequent lysosomes and occasional rectangular or rhomboid membrane-bound crystalloids exhibiting lamellar periodicity and structural transition to lysosomes. Some SEGA cells showed features suggestive of neuronal differentiation, including stacks of rough endoplasmic reticulum, occasional microtubules and a few dense-core granules. Furthermore, in one case of tuber, a process of a single large cell was seen to be engaged in synapse formation. Intermediate filaments within a few cells of both lesions were decorated by gold particle-labeled GFAP antiserum. Within the tumor cells of SEGA, irregular, non-membrane-bound, electron-lucent areas often contained somatostatin-immunoreactive particles, whereas the latter could not be detected in tuber. The present study provides further evidence of divergent glioneuronal differentiation, both in the giant cells of tubers and the cells of SEGA. The findings of similar cells at different sites, including the subependymal zone, white matter ("heterotopias"), and cortex indirectly supports the idea that these lesions of TS result from a migration abnormality.

Notes: Abstract Subependymal giant cell astrocytoma (SEGA) is the most common neoplastic process involving the brain in patients with tuberous sclerosis complex (TSC). Morphologically, these tumors exhibit a wide range of cytoarchitecture with spindle and epithelioid cells resembling astrocytes, and also large, occasionally giant cells, some of which have a distinctly ganglion-like appearance. Unresolved questions regarding SEGAs center on: (a) their cytogenesis, i.e., whether they are derived from single or multiple precursors; and (b) their differentiating capacity along glial or neuronal lines. We sought to determine whether SEGAs represent truly mixed tumors or whether they consist of a single population of cells with a capacity for divergent differentiation. Twenty SEGAs were assessed for immunophenotypic features of either neuronal or glial differentiation or both. Only tumors from patients with a clinically confirmed diagnosis of TSC were included. Immunoreactivity for glial fibrillary acidic protein (GFAP) and/or S-100 protein was considered indicative of a glial phenotype, whereas the presence of neuronal differentiation was assessed by staining for cytoskeletal proteins [neurofilament epitopes, class III β-tubulin, microtubule-associated protein 2 (MAP2), synaptophysin], neurosecretory substances [serotonin, cholecystokinin, β-endorphin, substance P, somatostatin, met-enkephalin, neuropeptide Y, vasoactive intestinal polypeptide (VIP)], and for the 28-kDa neuron-associated calcium binding protein calbindin. Of the tumors examined, 18 exhibited both glial and neuronal epitopes, the staining pattern being variable. In 19 tumors, the constituent spindle, polygonal and giant or ganglion-like cells showed variable immunoreactivity for GFAP and S-100 protein, both within the cell body and processes. Neuron-associated cytoskeletal proteins were present in 18 cases. Class III β-tubulin immunoreactivity was demonstrated in 17 tumors, both within the bodies of all three cell types and to varying degrees within their processes. Neurofilament protein and calbindin staining was present in 8 tumors, with reactivity for the former being distributed in a phosphorylation-dependent manner. MAP2 was detected in a few cells of two tumors. Immunoreactivity for neuropeptides was observed in 17 lesions. Somatostatin and met-enkephalin staining was noted in 10 tumors (50%), being present particularly within polygonal cells. Neuropeptide Y, serotonin and β-endorphin reactivity was found in 6 (30%), 5 (25%), and 4 tumors (20%), respectively; β-endorphin was lacking in giant cells, whereas neuropeptide Y and serotonin were seen within their cell bodies. Substance P and VIP were evident in only occasional polygonal cells of 2 (10%) and 1 tumor (5%), respectively. Stains for cholecystokinin were negative. The observation of immunoreactivity for both glial- and neuron-associated epitopes within tumor cells of the same morphology suggests that SEGAs represent proliferations of cell lineages with the capacity to undergo divergent glioneuronal as well as neuroendocrine differentiation to a greater extent than do other mixed glial-neuronal neoplasms.

Notes: Summary While the embryonal central neuroepithelial tumors present complex conceptual and clinical problems, advances in cell type identification by special neurohistological, immunohisto- and immunocytochemical techniques have permitted discrimination of distinct cytomorphogenetic entities. These are based in part on their resemblance to the normal phases of neurocytogenesis. Four of these tumors, medulloepithelioma, desmoplastic infantile ganglioglioma, pineoblastoma and medulloblastoma, are designated as multipotential in light of their capacity to undergo divergent differentiation. Cytomorphogenetic, clinical and experimental data implicate fetal neural cell targets for transformation and raise the possibility that aberrant developmental regulatory mechanisms may contribute to the biologic behavior of these tumors. Growth factors and some neuroregulatory neurotransmitters (such as serotonin) are known to act as modulators of normal neuromorphogenesis. They could play a regulatory role in central neuroepithelial tumors on the hypothesis that the aberrant behavior of the embryonal neoplasms could either be modified by fuctional receptor responses or result from abnormal receptor responses to these substances. Future challenges include 1) the definition of new cytomorphogenetic entities and subgroups of the currently defined forms of embryonal CNS tumors based on the presence of specific growth factors and neuroregulatory neurotransmitters, or their receptors, 2) the characterization of neoplastic receptor responses mediating any modulatory role of the presently known growth factors or neuroregulatory neurotransmitters on the growth and maturation potential of the embryonal central neuroepithelial tumors and 3) the further definition of developmental, stage-specific modulators that might be operative in these tumors.