Normal Neural Tissues Research Articles

Ret-oncogene Expression Correlates with Neuronal Differentiation of Neuroblastic Tumors

An antibody to the ret proto-oncogene product (RET) was raised and applied to formalin-fixed, paraffin-embedded neuroblastic tumors (NBTs) to investigate its usefulness in diagnosis and evaluation of cell differentiation. In normal neural crest-derived tissues, most ganglion cells were moderately stained while large ganglion cells were weakly stained. In NBTs, the intensity of the staining in moderately differentiated neuroblasts and small ganglion cells was more prominent than in undifferentiated neuroblasts, while the cytoplasm of large ganglionic cells was weakly stained as in normal ganglion cells. The RET immunoreactivity was compared with that of nine neural and neuroendocrine markers. The results revealed a parallelism with the protein gene product 9.5 (PGP9.5), neuron specific enolase (NSE) and NF-150 kD (NF = M). These findings indicated that the RET immunoreactivity was correlated with ganglionic differentiation and maturation. Thus, RET was considered to be a new marker that would be implemented in diagnosis and estimation of neuronal differentiation of NBTs.

Melanocytes in human skin express bcl-2 protein.

The bcl-2 gene was initially identified through its participation in the translocation 14:18 in B-cell lymphomas of follicular center cell origin. This classic translocation juxtaposes the transcriptionally active immunoglobulin heavy-chain locus on chromosome 14 to the bcl-2 gene on chromosome 18, resulting in overexpression of bcl-2 protein. The oncogene bcl-2 is thought to prolong cell survival through interference with programmed cell death. To date, bcl-2 expression has been reported in normal lymphoid, hemopoietic, neural, breast and prostatic tissues. Since melanocytes are neural crest derivatives and have an extended life-span, our objective was to determine whether the bcl-2 protein is expressed in human melanocytes. We analyzed normal skin biopsies for bcl-2 expression using standard immunohistochemistry. As hypothesized, dendritic cells in the basal epidermal layer stained strongly for bcl-2. The distribution and morphology of these cells was typical for melanocytes. Additionally, eccrine sweat glands, lymphocytes and the dermal papilla of hair follicles demonstrated bcl-2 positivity. We believe this to be the first report of bcl-2 expression in normal melanocytes.

The Effect of Lead on the Metabolism of a Nuclear Matrix Protein Which Becomes Prominent in Lead-induced Intranuclear Inclusion Bodies

The influence of lead and calcium on the metabolism of a nuclear matrix protein has been studied in mouse neuroblastoma 2a (Nb2a) cells. This protein, p32/6.3, has an unusual distribution in that it is relatively abundant only in normal neural tissues and in intranuclear inclusion bodies induced in kidney tubule-lining cells of chronically lead-intoxicated animals (Egle, P. M., and Shelton, K. R. (1986) J. Biol. Chem. 261, 2294-2298). The mechanism(s) whereby lead increases p32/6.3 content are of interest, but its slow accumulation in intact animals over a period of weeks to months precludes studies with metabolic inhibitors. However, the enriched levels of p32/6.3 in mouse neuroblastoma 2a (Nb2a) cells permit these studies. The relative abundance of this protein was found to increase in mouse Nb2a cells after 1- and 3-day exposures to lead. This increase could be attributed to a decreased rate of degradation rather than either increased transcription or increased translation. A role for calcium in p32/6.3 regulation was also explored. Although neither increased extracellular calcium nor calcium ionophores had an effect, treatment of the cells with the calcium chelator [ethylenebis(oxyethylenenitrilo)]tetraacetic acid decreased p32/6.3 levels in a concentration-dependent manner, suggesting a role for calcium in the normal metabolism of the protein.

Immunologic recognition of cell surface antigens in normal mouse neural tissues and in neuroepithelial cells of the OTT-6050 mouse teratoma. A radiometric, gel electrophoretic and morphologic (immunofluorescence and immunoperoxidase) study.

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.

Neutral glycosphingolipids and ceramide composition of ethylnitrosourea-induced rat neural tumors: accumulation of ceramide in tumors.

Experimental rat neural tumors in offspring were induced transplacentally by a single injection of a chemical carcinogen, ethylnitrosourea, 20 mg/kg body weight, in the tail vein of the mother. The neutral glycosphingolipid, sulfatide, and ceramide composition of the tumors and the normal tissues from which the tumors originated is described. The content of nonhydroxy fatty acid (NFA) and hydroxy fatty acid (HFA) containing ceramide in all the neural tumors so far examined was significantly increased compared with the corresponding normal neural tissue. Some 8 to 18 mol% of total neutral glycolipids was as ceramide in neurinomas, oligodendrogliomas, and meningiomas. Lactosylceramide in normal neural tissues was about 1 mol% of the total neutral glycosphingolipids. In various neural tumors lactosylceramide increased up to 8 mol%. NFA- and HFA-containing cerebrosides constitute 94-100% of the neutral glycosphingolipids in normal neural tissues. In various neural tumors the mol percent of cerebrosides was significantly reduced. A high performance liquid chromatographic method was modified to analyze simultaneously ceramides, cerebrosides, and higher neutral glycosphingolipids.

An immunocytotoxicity assay for neural cells in monolayer cultures

The immunological analysis of cell surface constituents which may characterize neuronal and glial populations, though still in its infancy, will greatly facilitate the investigation of several important problems in neurobiology. One critical component of such analyses is the way by which a given antiserum can be shown to be active on, and possibly selective for neurons and glial cells from normal neural tissues. This report describes the use of monolayer cultures of normal neural cells for recognition and quantitative titration of antisera directed against them. Sera were collected from rabbits immunized with chick embryo spinal cord cell susptnsions, and found to be reactive to the same cells in the initial cell dissociate as well as in subsequent monolayer cultures of different in vitro ages. A monolayer assay procedure was developed, which ( i) uses small numbers of cells and small volumes of immune reagents, with the possibility of further scaling down; ( ii) applies equally to cultures using different substrata; ( iii) permits differential counts of morphologically different cultured cells; ( iv) allows to recognize cytological damage imposed by the immune serum in the presence, though not in the absence, of complement; and ( v) quantitatively titrates the immune activity with 10- to 20-fold higher sensitivity than other titration procedures. While the study was not intended to investigate the possible specificities of the new antisera, it provided the unexpected observation that non-neuronal cells in these spinal cell cultures were considerably less sensitive than neurons to the complement-dependent action of the antisera.

Acid hydrolase and cytochrome oxidase activities in nitrosourea induced tumors of the nervous system

Nitrosourea induced tumors of the nervous system in rats have proven useful for biochemical studies combined with morphological approaches. The pattern of enzyme activities for acid hydrolases and cytochrome oxidase resemble those previously observed in spontaneous nervous system tumors of man. The activities of 4 acid hydrolases were generally high in the gliomas. This could not be attributed solely to zones of regression or necrosis but was a general characteristic of the neoplasms. The activities were predominantly particulate and most likely lysosomal in localization. In schwannomas a similar increase in hydrolases was found in comparison with normal neural tissues but aryl-sulfatase was not increased. Cytochrome oxidase activities were markedly reduced in all tumors studied. The proportionate reduction with respect to normal brain was comparable to that noted in man. No differences were found with respect to fairly well differentiated gliomas.

Immunological properties of normal neural tissues

The preparation of an antiserum using human brain tissue as antigen is described. The presence in this antiserum of antibodies specific for brain and nerve tissue is demonstrated using double diffusion of reactants in agar gel. The cytotoxic effect of the antiserum in the presence of complement on cultured glial cells in vitro, and localisation of fluorescein-labelled anti-globulin in tissue sections and cells in culture, demonstrate the presence of the organ-specific material in astrocytes, oligodendrocytes and myelin, and its absence from liver, thyroid and kidney-cells, and fibroblasts. A critical evaluation of the techniques used is included.