Medulloblastoma cell line secretes platelet-derived growth factor

Medulloblastoma is the most common malignant childhood brain tumor in which aggressive growth produces recurrence in approximately 50% of appropriately treated cases and metastases along the neuraxis in 30%. To date, no studies exist concerning the production of autocrine growth factors by this brain tumor type. Malignant brain tumors in adults often produce platelet-derived growth factor (PDGF). A medulloblastoma cell line, TE-671, has been used for many years in pediatric neuro-oncologic studies. We assayed this medulloblastoma cell line for the production of PDGF. PDGF is produced by medulloblastoma cells grown in monolayer tissue culture and stimulates PDGF-sensitive 3T3 fibroblasts to incorporate tritiated thymidine in a dose-dependent fashion. This biologic activity is blocked by PDGF antibodies in a dose-dependent relationship. We postulate that PDGF produced by medulloblastoma cells plays a role in the growth of this tumor by stimulating mitogenic activity.     

Intracerebral transplantation of the A7 immortalized astrocytic cell line

The A7 cell line is an astrocyte-like cell immortalized by SV40 large T antigen, using retroviral-mediated gene transfer. These cells were transplanted into rat brains, and the graft-host interaction was investigated immunohistochemically. The A7 cells survived focally 2, 6 and 8 weeks after transplantation and retained the immunocytochemical properties observed in vitro. No immunological response was observed. GAP-43 and N-cadherin immunoreactivities were not expressed by A7 cells, but were seen in the matrix within the area of the graft and in the surrounding brain tissue. This indicates that A7 cells may stimulate expression of GAP-43 and N-cadherin immunoreactivity by host tissue. Expression of Thy 1.1 was not observed within the graft site after 2 weeks of survival, but 6 and 8 weeks after transplantation Thy 1.1 was observed within the graft area, indicating the possible co-existence of grafted cells and host tissue. Although indirect, these observations suggest that the A7 cells induce changes in host brain, including possible growth or regeneration of host tissue into the graft area.     

An immortalized cell line expresses properties of activated microglial cells.

Murine cultured microglial cells were immortalized after infection with a v-raf/v-myc recombinant retrovirus. This immortalized cell line (BV-2) shares properties with body macrophages with respect to the antigen profile, their phagocytic capacity and antimicrobial activity. BV-2 cells are not constitutively able to kill tumor cells in vitro, but acquire antitumor activity following an increase in [Ca++]i. BV-2 cells, like microglial cells, are however, distinct from peripheral macrophages by their expression of inwardly rectifying K+ channels in concert with a lack in outwardly rectifying K+ channels and the formation of spineous processes. The BV-2 cell line thus represents a suitable model for in vitro studies of activated microglial cells.     

Effects of dopaminergic cell degeneration on electrophysiological characteristics and GAD65/GAD67 expression in the substantia nigra: different action on GABA cell subpopulations.

The motor disturbances occurring in Parkinson's disease have been partially attributed to a hyperactivity of gamma-aminobutyric acid (GABA)-ergic nigral cells largely in the substantia nigra pars reticulata (SNr) secondary to the degeneration of dopaminergic nigrostriatal neurons. However, some aspects of this response remain unclear. In this work, different electrophysiological and neurochemical parameters were studied in GABAergic cells of the SN after unilateral nigrostriatal dopaminergic lesion using 6-hydroxydopamine injection in rats. Our data showed that 1) the SN under normal conditions contains different subsets of GABAergic cells according to their firing pattern and glutamic acid decarboxylase mRNA levels, and 2) the response of these GABAergic cell subgroups was different after the ipsi- and contralateral dopaminergic cell degeneration. These findings indicate a complex regulation of nigral GABAergic activity after nigrostriatal dopaminergic degeneration that probably involves local mechanisms, the nigro-striato-nigral loop, as well as interhemispheric mechanisms whose anatomical basis remains unstudied.     

Distribution and ultrastructure of neurons in opossum piriform cortex displaying immunoreactivity to GABA and GAD and high-affinity tritiated GABA uptake

GABAergic neurons have been identified in the piriform cortex of the opossum at light and electron microscopic levels by immunocytochemical localization of GABA and the GABA-synthesizing enzyme glutamic acid decarboxylase and by autoradiographic visualization of high-affinity 3H-GABA uptake. Four major neuron populations have been distinguished on the basis of soma size, shape, and segregation at specific depths and locations: large horizontal cells in layer Ia of the anterior piriform cortex, small globular cells with thin dendrites concentrated in layers Ib and II of the posterior piriform cortex, and multipolar and fusiform cells concentrated in the deep part of layer III in anterior and posterior parts of the piriform cortex and the subjacent endopiriform nucleus. All four populations were well visualized with both antisera, but the large layer Ia horizontal cells displayed only very light 3H-GABA uptake, thus suggesting a lack of local axon collaterals or lack of high-affinity GABA uptake sites. The large, ultrastructurally distinctive somata of layer Ia horizontal cells receive a very small number of symmetrical synapses; the thin, axonlike dendrites of small globular cells are exclusively postsynaptic and receive large numbers of both symmetrical and asymmetrical synapses, in contrast to somata which receive a small number of both types; and the deep multipolar and fusiform cells receive a highly variable number of symmetrical and asymmetrical synapses on somata and proximal dendrites. Labeled puncta of axon terminal dimensions were found in large numbers in the neuropil surrounding pyramidal cell somata in layer II and in the endopiriform nucleus. Moderately large numbers of labeled puncta were found in layer I at the depth of pyramidal cell apical dendrites with greater numbers in layer Ia at the depth of distal apical segments than in layer Ib. High-affinity GABA uptake was demonstrated in the termination zone of the projection from the anterior olfactory nucleus to the anterior piriform cortex. Cell bodies of origin of this projection displayed heavy retrograde labeling with 3H-GABA. Matching neuropil and cellular labeling was demonstrated with the GABA-BSA antiserum but not with the GAD antiserum, thus suggesting that GABA is normally present in these cells but is taken up from the neuropil rather than synthesized. No comparable high-affinity GABA uptake was demonstrated in the association fiber systems that originate in the piriform cortex.(ABSTRACT TRUNCATED AT 400 WORDS)     

Bovine anterior pituitary progenitor cell line expresses interleukin (IL)-18 and IL-18 receptor

In the anterior pituitary gland, inflammatory mediators regulate cell function through an immuno-endocrine pathway. Recent studies have shown that undifferentiated stem cells act as immunomodulators. These studies prompted us to establish a progenitor cell line from the bovine anterior pituitary gland and to detail its function. First, we localised interleukin (IL)-18 by immunohistochemistry to the marginal cell layer of Rathke's pouch that is assumed to embody a stem/progenitor cell compartment of the postnatal pituitary gland. A cloned anterior pituitary-derived cell line from the bovine anterior pituitary gland was established from single cell clone by the limiting dilution method and was designated as bovine anterior pituitary-derived cell line (BAPC)-1. BAPC-1 cells constantly expressed mRNAs for IL-18 and IL-18 receptor, and grew steadily and rapidly in the medium containing epidermal growth factor and basic fibroblast growth factor. The cell line also expressed the mRNAs for the stem/progenitor cell- related factors such as Nanog, Oct-4, Ptch1, Nestin, Notch1, Hes1, Lrp and Fzd4, and the mRNAs for embryonic pituitary-related factors, such as Lhx3, PitX1 and Pit-1. The nuclei of BAPC-1 were immunostained positively for Pit-1, Hes1 and beta-catenin antibodies. Furthermore, BAPC-1 cells expressed mRNAs for cytokine such as IL-1alpha, IL-6, IL-7, IL-12 and IL-15. Stimulation of BAPC-1 cells with IL-18 increased expression of mRNAs for IL-1alpha, IL-6, IL-1beta and IL-8. At day 6 in culture, BAPC-1 cells also express growth hormone mRNA. These results strongly suggest that BAPC-1 is a stem/progenitor cell line and modulates the immuno-endocrine function of the anterior pituitary cells through its cytokine production.     

实验性癫痫与γ-氨基丁酸和谷氨酸脱羧酶的关系

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gamma-Aminobutyric acid (GABA) enhances glutamate cytotoxicity in a cerebellar cell line

A temperature-sensitive rat cerebellar cell line SC9 has been used to study the role of the inhibitory neurotransmitter gamma-aminobutyric acid (GABA) in glutamate cytotoxicity. GABA increases glutamate toxicity in a dose-dependent fashion, but NMDA and kainic acid were not toxic in the presence or absence of GABA. The specificity of this cytotoxicity was further indicated by the NMDA-selective antagonist 2-amino-7-phosphonoheptanoic acid (APV), which does not block glutamate effect. These observations, as well as binding experiments with 3H-glutamate, suggest that glutamate cytotoxicity in these cells depends on quisqualate-selective uptake sites of the amino acid. The study may open therefore a novel pathway for understanding the cytotoxic effect of excitatory amino acids in brain structures that are enriched with GABA and glutamate uptake sites.     

Non-correspondence of [3H]GABA uptake and GAD localization in goldfish amacrine cells

The effects induced by penicillin (PEN) upon the synaptic responses of CA1 hippocampal pyramidal cells (HPCs) were studied in the 'in vitro' slice. Low concentrations of PEN (0.17-0.34 mM) evoked an increase in amplitude and duration of the orthodromic excitatory post-synaptic potential induced by stratum radiatum, while stimuli which were subthreshold in control conditions became effective in eliciting action potentials. These changes were not paralleled by any decrease of the recurrent inhibitory post-synaptic potential due to inhibitory interneurons located at or near the soma. However, the latter decreased and then disappeared as PEN concentrations were brought to levels higher than 0.68 mM. Since low concentrations of PEN increase CA1 HPCs responsiveness without decreasing somatic inhibition, it is concluded that this action is probably due to a reduced dendritic inhibitory mechanism.     

Dark-rearing-induced reduction of GABA and GAD and prevention of the effect by BDNF in the mouse retina

Gamma‐aminobutyric acid (GABA) is an important retinal neurotransmitter. We studied the expression of GABA, glutamate decarboxylase 65 (GAD65) and GAD67 by immunocytochemistry and Western blot, in the retinas of control and dark‐reared C57BL/6J black mice. This study asked three questions. First, is visual input necessary for the normal expression of GABA, GAD65 and GAD67? Second, can the retina recover from the effects of dark‐rearing if returned to a normal light–dark cycle? Third, does BDNF prevent the influence of dark‐rearing on the expression of GABA and GAD? At postnatal day 10 (P10), before eye opening, GABA immunoreactivity was present in the ganglion cell layer (GCL), in the innermost rows of the inner nuclear layer (INL) and throughout the inner plexiform layer (IPL) of control and dark‐reared retinas. In P30 control retinas, GABA immunoreactivity showed similar patterns to those at P10. However, in P30 dark‐reared retinas, the density of GABA‐immunoreactive cells was lower in both the INL and GCL than in control retinas. In addition, visual deprivation retarded GABA immunoreactivity in the IPL. Western blot analysis showed corresponding differences in the levels of GAD65 but not of GAD67 expression between control and dark‐rearing conditions. In our study, dark‐rearing effects were reversed when the mice were put in normal cyclic light–dark conditions for 2 weeks. Moreover, dark‐reared retinas treated with BDNF showed normal expression of both GABA and GAD65. Our data indicate that normal expression of GABA and GAD65 is dependent on visual input. Furthermore, the data suggest that BDNF controls this dependence.     

A novel N18TG2 x mesencephalon cell hybrid expresses properties that suggest a dopaminergic cell line of substantia nigra origin.

A dopaminergic neuroblastoma was derived using somatic cell fusion of rat embryonic mesencephalon cells and the murine neuroblastoma-glioma cell line N18TG2. The resulting interspecies hybrid, named MES23.5, has retained a stable phenotype and karyotype for a continuous culture period of 1 year. The hybrid exhibits several properties that suggest that the parent primary neurons originated in the substantia nigra. The cell line contains tyrosine hydroxylase, which is identifiable both by biochemical and immunological methods and synthesizes dopamine, but no other catecholamine. Additionally, the cell line expresses apparent voltage-gated CA2+ channels as measured by high-affinity omega-conotoxin binding. The MES23.5 omega-conotoxin receptors are of similar affinity class to those found in adult rat mesencephalon. No dihydropyridine receptors, as measured by PN200-100 ligand binding, are present. None of these properties are found in the N18TG2 parent. At least three neuronal features, namely, tyrosine hydroxylase, dopamine synthesis, and omega-conotoxin receptor expression, are quantitatively elevated after sustained treatment with cAMP analogs. The cell line expresses a complex range of neural properties found in the dopaminergic neurons of the substantia nigra, and may therefore be useful elucidating further details of their cell biology.     

GABA levels and GAD immunoreactivity in the deep cerebellar nuclei of rats with altered olivo-cerebellar function.

Immunocytochemistry was used to examine the distribution, size, and density of glutamic acid decarboxylase immunoreactive (GAD+) puncta in two animal models with movement disorders, the genetically dystonic (dt) rat and rats with 3-acetylpyridine (3AP) lesions of the inferior olive. In both models, GAD activity is increased in the deep cerebellar nuclei (DCN) where the enzyme is localized primarily in the terminals of Purkinje cells. GABA levels were also measured in the DCN. The general distribution of GAD+ puncta in the DCN was similar in all groups. Immediately after the 3AP lesions, however, GABA levels were elevated in 3AP rats in comparison with both normal rats and age-matched dt rats. GAD+ puncta were also larger than normal in the 3AP group at this time, although the magnitude of this effect declined over a 2-week recovery period. Puncta density was decreased in the medial nucleus only in 25-day-old dt rats in comparisons with normal littermates. These findings are discussed in the context of previously reported differences in the firing rate of Purkinje cells in dt and 3AP-treated rats.     

Ethinylestradiol and d-norgestrel regulation of plasminogen activator in a human melanoma cell line

The influence of ethinylestradiol (EE₂) and d-norgestrel (d-Ng) was studied in a melanoma cell line producing a tissue-type plasminogen activator (t-PA) very similar to or identical with the t-PA isolated in extracts from human uterus. The cell cultures were exposed to the two contraceptive steroids by addition of EE₂ or d-Ng dissolved in a week alcoholic solution to the culture media, in which the released t-PA was assayed by an immunoradiometric method. A strongly stimulating effect of ethanol (0.76% w/v) on the t-PA production was demonstrated. Whereas, EE₂ in the concentration of 1.7 × 10^-7M and d-Ng in the concentration of 8.6 × 10^-7M both caused a significantly reduced secretion of t-PA, and this effect was independent of whether the cell cultures were grown to confluency in the presence of the two synthetic steroids or not. It was concluded, that the two contraceptive steroids had an inhibitory effect on the production of t-PA in melanoma cell culture.