Stanley A. Vinores

Electron-immunocytochemical localization of neuron-specific enolase in cytoplasm and on membranes of primary and metastatic cerebral tumours and on glial filaments of glioma cells

A series of primary and metastatic human brain tumours was evaluated immunocy‐tochemically for the electron microscopic localization of neuron‐specific enolase (NSE). All contained cells which, regardless of the cell type, demonstrated an irregular distribution of NSE in their cytoplasm and on membranes. This was in contrast to the staining pattern in normal central nervous system (CNS) cells which, as previously reported (Vinores et al. 1984b), show only diffuse cytoplasmic staining usually not associated with membranes. In the tumours, the interior of nuclei and the cristae and matrices of mitochondria were consistently negative, as in normal CNS cells. Except in one low‐grade fibrillary astrocytoma, the cytoplasmic filaments in neoplastic astrocytes were often, but not invariably, stained for NSE. The fine structural localization of NSE in neoplastic cells suggests that the conversion of 2‐D‐glycerophosphate to phosphoenolpyruvate by enolase may occur on the membrane and, in the case of astrocytic tumours, on the cytoplasmic filaments as well as in the cytoplasm. When cells which contain only the non‐neuronal form of enolase (NNE) transform to neoplastic cells, they may acquire the ability to produce NSE. This presumably enables them to accommodate the increased metabolic demands of neoplasia by allowing them to elude the regulatory controls that are specific for NNE.

Progressive ultrastructural damage and thickening of the basement membrane of the retinal pigment epithelium in spontaneously diabetic BB rats.

Ultrastructural examination of the neural retina and retinal pigment epithelium was performed in spontaneously diabetic BB rats and non-diabetic littermate controls at various time points during the course of diabetes. Four types of progressive changes occur in diabetic rats: (1) There is a progressive thickening of the basal laminae of the retinal pigment epithelium and the retinal capillaries. (2) In addition to this thickening, there is a progressive, often focal, thickening of the rest of Bruchs membrane, and by 11 1/2 months it contains numerous randomly oriented collagen fibrils. (3) There are progressive ultrastructural changes in retinal pigment epithelial cells consisting of marked dilation of the basal infoldings, the formation of large vacuoles and cell swelling. (4) Rare degenerative changes were noted in retinal capillary pericytes. Since non-diabetic littermates do not show comparable changes, it is likely that they are diabetes-related, suggesting that progressive damage to the retinal pigment epithelium may occur during the course of diabetes.

Prevention or moderation of some ultrastructural changes in the RPE and retina of galactosemic rats by aldose reductase inhibition

Rats were maintained on a 50% galactose diet with and without the aldose reductase inhibitor, Sorbinil, or on a normal diet of rat chow, and the retinas and retinal pigment epithelium were examined ultrastructurally at time points ranging from 4 weeks to 20 months. Several ultrastructural changes were observed in the retinal pigment epithelium and retinal capillaries of galactosemic rats that were not seen in rats on a normal diet. Only some of these changes are similar to those that have been previously noted in diabetic (glucosemic) rats. As in diabetics, galactosemic rats demonstrated thickening of the basal laminae of the retinal pigment epithelium and retinal capillaries. They also manifested vacuolization and degenerative foci in the retinal pigment epithelium that were similar, but not identical, to changes seen in diabetic rats. Each of these changes was significantly inhibited by Sorbinil. Unlike diabetics, galactosemic rats did not have dilated basal infoldings, but did have outer retinal folds and a significant increase in large-lipofuscin-like aggregates in the retinal pigment epithelium, both of which were partly prevented by Sorbinil. These data suggest that multiple mechanisms may be involved in retinal and retinal pigment epithelium changes in diabetic and galactosemic rats, and that enhanced polyol metabolism is likely to be involved in some of the changes in both.

A morphological and immunohistochemical study of human retinal pigment epthelial cells, retinal glia, and fibroblasts grown on Gelfoam matrix in an organ culture system - A comparison of structural and nonstructural proteins and their application to cell type identification

Retinal pigment epithelial (RPE) cells, retinal glia, and fibroblasts undergo marked phenotypical change when outside their usual microenvironment, as occurs in epiretinal membrane formation. To explore their phenotypic potential without the influence of other cell types, each was cultured on Gelfoam matrix and assessed immunohistochemically and ultrastructurally. All cell types demonstrated vimentin and a universalβ-tubulin epitope, TU27. RPE and retinal glial cells were positive for cytokeratin, Len 7, and neuron-specific (γγ) enolase, as were glia and ibroblasts for S-100 protein and RPE cells and fibroblasts for glutamine synthetase. RPE cells alone showed positivity for class III β-tubulin and retinal S-antigen (monolayer cultures only) ; occasional retinal glia, which immunohistochemical findings suggest are Willer cell derived, demonstrated GFA protein. Therefore, class III β-tubulin may be useful in distinguishing RPE cells from retinal glia and ibroblasts, and Leu-7 may help to identify RPE cells and fibroblasts; these cell types are difficult to distinguish in clinical material using more traditional morphological criteria.

Localization of neuron-specific (γγ) enolase in proliferating (supportive and neoplastic) Schwann cells. An immunohisto- and electron-immunocyto-chemical study of ganglioneuroblastoma and schwannomas

SummaryNeuron-specific (γγ) enolase, a glycolytic enzyme used as a relatively specific marker for normal neurons and neuroendocrine cells, has recently been found in a variety of neoplastic cells and in reactive astrocytes. Its localization was investigated by immunohisto- and electron-immunocyto-chemistry, in the proliferating supportive Schwann cells of a peripheral ganglioneuroblastoma and in the neoplastic Schwann cells of four acoustic tumours. By light microscopy, the neoplastic Schwann cells showed moderate uneven diffuse immunopositivity for enolase. By electron-immunocy-tochemistry, both types of Schwann cells demonstrated immunopositivity discretely limited to their cell surface membranes. The neoplastic ganglion cells and axons of the ganglioneuroblastoma and the normal neurons and axons included in the schwannomas were, as expected, intensely immunopositive. The visualization of γγ enolase on the cell surface membranes of both neoplastic and non-neoplastic proliferating Schwann cells suggests that increased glycolytic activity may occur on the surface of these proliferating cells irrespective of the nature of the proliferation.

Nerve growth factor modification of the ethylnitrosourea model for multiple schwannomas.

The administration of ethylnitrosourea (ENU) to pregnant rats late in gestation or to neonatal rats results in the induction of Schwann cell tumors in a high percentage of perinatally exposed animals. Exogenous administration of nerve growth factor (NGF) significantly reduces the number of Schwann cell tumors and other neurogenic tumors developing in ENU-treated rats. Administration of antibodies directed against NGF prior to neonatal ENU exposure results in a substantial increase in the incidence of Schwann cell tumors, particularly in the trigeminal nerves of both rats and mice. Transplacental ENU treatment causes early neoplastic proliferation (ENP) at 90 days of age in the Schwann cell population of trigeminal nerves in nearly all exposed rats. A variety of NGF treatment protocols (single or multiple inoculations or microinfusion prior to or following ENU exposure) resulted in a significant reduction in ENU-induced ENP in trigeminal nerves. These results indicate that NGF may convey protection either directly or indirectly, by an unknown mechanism, to Schwann cells and other supportive neural cells by reducing their sensitivity to ENU-induced neoplastic transformation.

The growth of two murine hemangioendotheliomas intracranially, subcutaneously, and in culture, and their comparison with human cerebellar hemangioblastomas: morphological and immunohistochemical studies

SummaryTwo thorium dioxide-induced murine hemangioendotheliomas, 42021 TCT and 44347 TST, were grown subcutaneously (for up to 22 and 15 passages respectively) or intracranially (single passage) and were adapted to culture as a monolayer and, in a limited fashion, in an organ culture system or in rotary suspension. They remained viable and malignant following 20–21 years of storage in liquid nitrogen, and had ultrastructural similarities to human hemangioblastomas. The murine tumors were positive for Griffonia (Bandeiraea) simplicofolia isolectin B4 binding, establishing their endothelial nature; however, unlike human hemangioblastic tumors, they did not cross-react with antisera to human factor VIII or fibronectin and they did not demonstrate Ulex europaeus type I lectin (UEA I) binding (as is also the case for non-neoplastic murine vascular endothelial cells). A variety of morphological cell types in cultures derived from the tumors were also positive for Griffonia (Bandeiraea) simplicifolia isolectin B4 binding. Both murine hemangioendotheliomas, when implanted in the cerebrum, were potent inducers of reactive gliosis, but there was no evidence of uptake of glial fibrillary acidic protein. Unlike the human cerebellar hemangioblastomas, murine tumors were malignant and invasive and did not contain stromal cells, nor did they demonstrate Weibel-Palade bodies or extensive pinocytotic activity. Thus, the murine tumors appear to more closely resemble angiosarcomas or epitheloid hemangioblastomas than the cerebellar hemangioblastomas.