Biomedical subjects

V W Yong

Publications and source records attributed to V W Yong.

At least 19 recordsLinked to original sources

PubMed1992

Differential proliferative response of human and mouse astrocytes to gamma-interferon.

We have previously shown that gamma-interferon promoted the proliferation of adult human astrocytes isolated from brain biopsy specimens. In contrast, in the present study, astrocytes derived from neonatal mouse brains and treated with recombinant murine gamma-interferon responded by a decrease (average of 50% at 100 U/ml) in proliferation. The basal rate of proliferation as assessed by bromodeoxyuridine incorporation was markedly increased in neonatal mouse astrocytes when compared to the adult human cells, suggesting that age, and the corresponding metabolic activity of cells, could be important determinants in the mitogenic response of astrocytes to cytokines. However, subsequent examinations of fetal human and adult mouse astrocytes, with comparable basal rate of proliferation to neonatal mouse and adult human cells respectively, showed gamma-interferon to promote DNA synthesis in fetal human astrocytes while inhibiting that of adult mouse astrocytes. The results suggest species differences in the proliferative response of human and mouse astrocytes to the cytokine gamma-interferon.

Adult↗

PubMed1992

Production of soluble autocrine inhibitory factors by human glioma cell lines.

Gliomas in vitro exhibit density-limited growth upon the attainment of confluency, an effect usually attributed to cell-cell contact inhibition. Since gliomas have been demonstrated to secrete an array of soluble factors which can enhance tumor growth, we undertook this study to ascertain whether production of soluble factors by the tumor may also inhibit growth in an autocrine manner, and whether production of such factors is associated with the growth phase of the glioma. We observed that cell-conditioned medium (supernatants) from non-confluent glioma cultures induced growth, while confluent culture supernatants produced pronounced growth suppression. These latter supernatants enhanced proliferation of non-transformed astrocytes. Supernatants derived from all stages of confluency produced inhibition of lymphocyte proliferation. To characterize these factors, dialyzed supernatant was tested and found to continue to produce lymphocyte suppression but no glioma growth limitation. Growth of tumors in indomethacin or in acetylsalicylic acid to abolish prostanoid synthesis abrogated the inhibitory influence on glioma growth but only partially reversed the lymphocyte suppressive capacity. These studies suggest that gliomas do produce a growth phase dependent autocrine inhibitory factor(s), and that the production of these small molecular weight factors is at least partially under control of the cyclooxygenase pathway.

Adult↗

PubMed1992

Proliferation of human and mouse astrocytes in vitro: signalling through the protein kinase C pathway.

While several mitogens for astrocytes have been described, the signal transduction pathway(s) that mediates their proliferative effect remains unclear; in this report, a major role for the protein kinase C (PKC) system is suggested by several lines of evidence. Firstly, biologically active phorbol esters, 4 beta-phorbol-12,13-dibutyrate and phorbol-12-myristate-13-acetate, increase the proliferation of astrocytes as determined by [3H]thymidine incorporation or bromodeoxyuridine immunofluorescence; this effect is not reproduced by a phorbol ester that binds to PKC but does not activate it (4 alpha-phorbol-12,13-didecanoate). Secondly, 2 relatively selective inhibitors of PKC, H7 and staurosporine, attenuate the basal rate of proliferation of astrocytes in concentrations that were not cytotoxic to cells. Thirdly, mitogen-enhanced proliferation of astrocytes can be blocked by PKC inhibitors; this is observed for all astrocyte mitogens tested. Fourthly, measurements of PKC enzyme activity in astrocytes in response to serum-mitogenic factors, or to staurosporine, revealed a statistically significant correlation with proliferation rate. The mediation by PKC is not dependent on species- or age factors, since neonatal mouse or adult human astrocytes gave comparable results. The results have relevance to normal development and reactive gliosis post-injury, 2 conditions where astrocytes undergo proliferation, and to glioma growth.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1992

Malignant glioma-derived soluble factors regulate proliferation of normal adult human astrocytes.

Malignant gliomas are characteristically surrounded by marked gliosis. To assess whether glioma-derived products contribute to the proliferation of astrocytes, a feature of the gliosis response, we evaluated the influence of culture supernatants from malignant human glioma lines and tumor cyst fluids collected from two patients with glioblastoma multiforme on the proliferation of non-transformed adult human astrocytes. Both the culture supernatants and cyst fluids significantly increased DNA synthesis in astrocytes as assessed by a double immunofluorescence glial fibrillary acidic protein-bromodeoxyuridine technique. The net proliferative effect mediated by glioma cell line supernatants was tumor growth phase-dependent, being preferentially expressed during the logarithmic phase of glioma cell growth. Specific growth factor molecules and cytokines known to be secreted by gliomas (epidermal growth factor, fibroblast growth factor, platelet-derived growth factor, transforming growth factor-beta, interleukin-6, and tumor necrosis factor-alpha) could not reproduce the mitogenic effects of the glioma-derived soluble factors. Cytokines which can induce DNA synthesis by adult human astrocytes in vitro, gamma-interferon and interleukin-1, were not detected in the culture supernatant of glioma lines used in this study. In conjunction with the documented effects of glioma products on endothelial and lymphoid cells, the current study suggests that soluble glioma products can contribute to the production of surrounding gliosis observed in vivo.

Astrocytes↗

PubMed1992

Biology of adult human microglia in culture: comparisons with peripheral blood monocytes and astrocytes.

We have compared phenotypic and functional properties of surgically derived adult human microglia to autologous and allogenic peripheral blood-derived monocytes and to astrocytes derived from the same surgical resection. We found that microglia differed from peripheral blood monocytes with respect to adhesion properties and survival rates in vitro. Microglia, similar to resident macrophages in different tissues, expressed many but not all (CD4, Leu-M3, non-specific esterase) monocyte/macrophage associated markers tested, a pattern similar to that of terminally differentiated cells of this lineage. As with other human tissue macrophages, but in contrast to astrocytes, microglia did not undergo DNA synthesis in vitro, assessed using BrdU incorporation. Under basal culture conditions the majority of microglia of all morphologic subtypes (ameboid, bipolar, ramified) expressed MHC class II molecules; by flow cytometric analysis, mean fluorescence intensity of these cells was less than that of blood monocytes (relative to isotype control). In vitro MHC class II antigen expression on microglia, under basal and interferon gamma activating conditions, was greater than on astrocytes. Freshly derived T cells cultured with 1-10% autologous microglia plus Candida albicans underwent active proliferation, indicating the functional capacity of the microglia to serve as antigen-presenting cells.

Astrocytes↗

PubMed1992

Protein kinase C activity correlates with the growth rate of malignant gliomas: Part II. Effects of glioma mitogens and modulators of protein kinase C.

The proliferation rates of gliomas may be modulated by the protein kinase C (PKC) signal transduction system. The present study was undertaken to further examine the role of PKC system in growth regulation of gliomas in vitro by measurement of PKC activity over various phases of tumor growth and by assessing its potential role as a signal transduction system induced by serum mitogens and the known glioma mitogens epidermal growth factor and fibroblast growth factor. All human glioma lines examined, and the rat glioma C6, displayed high PKC activity relative to nonmalignant glial cells, which correlated with their proliferation rates over their respective growth phase. Frozen surgical human malignant glioma specimens also displayed high PKC activity. The relatively selective PKC inhibitor staurosporine (SP) reduced PKC activity and corresponding growth rates in a dose-related manner. Stimulation of PKC with phorbol esters under different concentrations of serum in the growth medium indicated that the high PKC activity, which correlated with their rapid growth rates, is highly susceptible to down-regulation by these agents. Epidermal growth factor and fibroblast growth factor increased both PKC activity and the growth rate of glioma line A172; addition of SP reduced the growth rate to levels observed in SP-treated control tumors, indicating that PKC may be a common signal transduction system induced by these mitogens. These results implicate PKC as an important signal transduction system regulating glioma growth, and offers a potential target for tumor inhibition.

Animals↗

PubMed1991

Gamma-interferon promotes proliferation of adult human astrocytes in vitro and reactive gliosis in the adult mouse brain in vivo.

Reactive gliosis is a characteristic response of astrocytes to inflammation and trauma of the central nervous system. To investigate whether soluble factors (cytokines) from inflammatory mononuclear cells that accumulate at lesion sites can provide the cellular signals to initiate gliosis and to identify such cytokines, we have tested and found that supernatants derived from subsets of activated human T lymphocytes (CD8+ or CD4+) are potent mitogens for cultured human adult astrocytes. This effect is blocked by a neutralizing antibody to gamma-interferon (IFN). Recombinant IFN alone can induce proliferation of human adult astrocytes in vitro and increase the extent of trauma-initiated gliosis in the adult mouse brain. The astrocyte proliferation-inducing activity of supernatants of glial cultures treated with IFN can be completely blocked with IFN-neutralizing antibody, suggesting that the proliferative effect does not require intermediary cytokines or cells. These results implicate IFN as an important mediator of the gliosis observed in pathologic conditions of the adult central nervous system associated with infiltrating lymphocytes.

Animals↗

PubMed1991

Origin of contralateral reactive gliosis in surgically injured rat cerebral cortex.

While reactive gliosis is readily observed close to the site of cerebral injury, astrocyte reactivity can also occur in distant areas either ipsilateral or contralateral to the lesion site. The present experiments were designed to address the origin of contralateral gliosis in adult rats following a cortical stab wound injury. One-month-old rats were subjected to either left cortical stab wound alone, callosotomy alone, callosotomy plus left cortical stab wound, or no surgery; 7 days later, animals were sacrificed. Formalin-fixed, paraffin-embedded sections were obtained and immunostained for GFAP. While untreated controls showed no cortical gliosis, callosotomy alone induced mild bilateral cortical gliosis. Whether or not rats were subjected to a callosotomy, the left cortical stab wound produced identical results: severe ipsilateral cortical gliosis and moderate contralateral gliosis. In all lesion models, both the intensity of GFAP staining and the number of reactive astrocytes were most marked in cortical areas abutting the subarachnoid spaces and decreased gradually into the deeper cortical layers. Our results suggest that the origin of contralateral gliosis in cortical stab injury is more likely due to the release of soluble substance(s) which diffuse to distant areas, rather than the migration of astrocytes through the corpus callosum from the lesion site, or being subsequent to degeneration of neurons which fibers traverse the corpus callosum.

Animals↗

PubMed1991

Protein kinase C activity and intracellular distribution in surgically excised human epileptic neocortex.

Protein kinase C (PKC) activity assayed by phosphorylation of exogenous histone, was measured in neocortex obtained from 32 patients following surgery for focal epilepsy and from 6 non-epileptic patients. PKC activity was not significantly different in either the particulate or cytosolic fraction from epileptic foci (n = 17) versus samples from non-spiking regions (n = 22) or neocortex from non-epileptic patients (n = 6). From 67% to 70% of total PKC activity was present in the cytosolic fraction. Phosphorylation of endogenous cytosolic substrate proteins was also not significantly different in epileptic foci.

Biopsy↗

PubMed1991

Age-dependent decrease of process formation by cultured oligodendrocytes is augmented by protein kinase C stimulation.

The proportion of cultured rat oligodendrocytes (OL) that extended processes of over three soma diameter in length is dependent on the age of the animals from which the brains were derived; up to 70% of neonatal OL attained this criterion within 3 days, and this proportion progressively decreased with advancing ages of the animals (1, 3, and 6 months). The lower extent of process formation from older rat OL could be augmented, and indeed to equal neonatal levels, by treatment of cells with phorbol esters that stimulate protein kinase C: 4 beta-phorbol-12,13-dibutyrate (PDB) and phorbol-12-myristate-13-acetate (PMA). Enhancement of process formation by PDB and PMA was also observed for cultured adult human and bovine OL. For adult OL from all three species, a phorbol ester that binds but that does not activate protein kinase C, 4 alpha-phorbol-12,13-didecanoate, did not result in enhancement of process formation. Selectively to biologically active phorbol esters was shown by the inability of a wide range of growth factors to promote process extension. Immunohistochemical analyses indicate that the type III isozyme of protein kinase C predominates in cultured OL; the apparent intensity of immunoreactive PKC was not different between controls or cultures treated for 12 days with PDB, suggesting that the persistent presence of PDB might not have down-regulated the enzyme, in contrast to other cell types. We propose that stimulation of protein kinase C is critical to the triggering of process formation by cultured OL in vitro.

Aging↗

PubMed1991

Expression and modulation of HLA-DR on cultured human adult astrocytes.

Expression of Class II major histocompatibility complex (MHC) antigens on astrocytes has been implicated as contributing to the immune responses characteristic of chronic autoimmune diseases of the central nervous system. We examined the properties and regulation of HLA-DR on cultured human adult astrocytes. We found that a proportion of human astrocytes from each of fifteen individual donors expressed HLA-DR under basal culture condition; while this proportion differed among the human subjects (range 3-65%), the results for each individual remained relatively constant when analyzed at several time points (up to 125 days in vitro). Attempts to modulate HLA-DR expression by a variety of cytokines likely to be present in inflammatory infiltrates in the brain showed that only gamma-interferon could increase the proportion of human astrocytes that expressed HLA-DR. Whether the variability of HLA-DR expression on astrocytes between different individuals reflects a genetic trait which can influence susceptibility to autoimmune central nervous system diseases remains to be determined.

Adult↗

PubMed1991

Enhanced protein kinase C activity correlates with the growth rate of malignant gliomas in vitro.

Direct measurement of protein kinase C (PKC) activity in vitro revealed a significant increase in the activity of the enzyme in all human malignant glioma lines examined and the rat C6 tumor in comparison with control nonneoplastic astrocyte and mixed glial cultures. The total and particulate PKC activity in these cell types correlated strongly [r = 0.98 (P less than 0.001) and 0.94 (P = 0.002), respectively] with the maximal growth rates as measured by 3H-thymidine incorporation in each of the samples. An alteration in the growth rate of an individual glioma line (A172) by varying the serum concentration in the growth medium produced comparative changes in the measured PKC activity. The addition of the phorbol ester phorbol-12-myristate-13-acetate to this tumor line under high serum conditions produced down-regulation of the enzyme, which was accompanied by a corresponding reduction in thymidine incorporation. The administration of the PKC inhibitor staurosporine produced a dose-related decrease in the basal proliferation rate of glioma lines A172 and C6, as measured by 3H-thymidine uptake and confirmed by flow cytometry, indicating that the high intrinsic PKC activity is amenable to pharmacological manipulation. Cytofluorometric deoxyribonucleic acid cell cycle analysis of the tumors treated with PKC modulators demonstrated that reduced proliferation rates were caused by an inhibition of entrance into the deoxyribonucleic acid synthesis (S) phase (decrease in proliferative index), supporting the evidence that these modulators are not slowing the tumor growth in a nonspecific cytotoxic manner.(ABSTRACT TRUNCATED AT 250 WORDS)

Alkaloids↗

PubMed1990

Identification of GTP-binding proteins in myelin and oligodendrocyte membranes.

Myelin membranes purified from mouse and rat brain are associated with alpha subunits of four signal transducing guanosine triphosphate (GTP)-binding proteins: Go, Gi, Gs, and ras. Four low-molecular-weight (Mr) GTP-binding proteins are also present, as demonstrated by the binding of GTP to proteins immobilized in nitrocellulose. This latter group is more prominent at early stages of myelination and remains associated with isolated myelin membranes despite repetitive cycles of purification. At least one nonmyelin subcellular membrane fraction possesses the same proteins. The total membrane fraction of cultured oligodendrocytes is associated with both groups of GTP-binding proteins. None of the well-known myelin proteins bound GTP by the procedure described.

Animals↗

PubMed1990

Morphologic heterogeneity of human adult astrocytes in culture: correlation with HLA-DR expression.

Human adult astrocytes derived from brain surgical resections showed marked morphologic heterogeneity when cultured in vitro, ranging from a flat, fibroblast-like appearance to process-bearing cells with little soma cytoplasm. The majority of cells were intermediate in morphology, bearing a prominent cytoplasmic cell body with processes radiating from them. The morphologic heterogeneity was more extensive than that of adult rat astrocytes, and was not correlated with the extent of attendant gliosis in the surgical specimens, or the site of the surgical resection. None of the human astrocytes expressed A2B5, thus preventing their classification on basis of lineage into type 1 or type 2 astrocytes. However, functional differences appear to exist between subpopulations of human astrocytes, since the proportion of process-bearing human astrocytes that expressed HLA-DR in vitro was significantly greater than that found for flat astrocytes.

Adolescent↗

PubMed1990

Inhibition of growth of established human glioma cell lines by modulators of the protein kinase-C system.

The protein kinase-C (PKC) second messenger system contributes to regulation of cell growth and differentiation. This study was undertaken to examine the effects of modulators of the PKC enzyme system on the state of differentiation and proliferation rates of human gliomas in vitro. The administration of the PKC-activating phorbol esters 4-beta-phorbol-12,13-dibutyrate (PDB) and phorbol-12-myristate-13-acetate (PMA) resulted in a dose-related inhibition of growth of human glioma cell lines in vitro as measured by 3H-thymidine uptake. The synthetic nonphorbol PKC activator (SC-9) produced an even more pronounced decrease of 3H-thymidine uptake. Diacylglycerol, an endogenous activator of the system, applied externally, transiently decreased the proliferation, in concordance with its short-lived existence in vivo. Conversely, the administration of 4-alpha-phorbol-12,13-didecanoate (alpha-PDD), a phorbol ester that binds but does not activate the enzyme, had no effect on the proliferation rate. At the dosages that maximally decreased proliferation, there was no evidence of direct glioma cell lysis induced by these agents as measured by a chromium-release assay. Immunocytochemical analysis and cytofluorometric measurement of glial fibrillary acidic protein (GFAP) staining in the treated cultures revealed an increase in GFAP staining over control cultures. In contrast to the response of glioma cells, nonmalignant human adult astrocytes treated with the PKC activators responded by increasing their proliferation rate. The authors postulate that the diametrically opposed effects of PKC activators on nonmalignant astrocytes versus glioma growth may be due to a high intrinsic PKC activity in glioma cells, with resultant down-regulation of enzyme activity following the administration of the pharmacological activators.

Astrocytes↗

PubMed1989

Human fetal Schwann cells in culture: phenotypic expressions and proliferative capability.

In this report we examined the phenotypic expressions and the proliferative capability of cultured human fetal Schwann cells. Antigens that were expressed included laminin, nerve growth factor receptor, neural cell adhesion molecule, S-100 protein, and that recognized by the monoclonal antibody HNK-1. In addition, HLA-A,B,C and HLA-DR, respectively, class I and class II antigens of the major histocompatibility complex, were demonstrated on Schwann cells. Mitotic capability was high, with an average of 34% of Schwann cells undergoing proliferation over a 2-day period.

Antigens, Surface↗

PubMed1989

Transplantation of human sympathetic neurons and adrenal chromaffin cells into parkinsonian monkeys: no reversal of clinical symptoms.

Cultured human fetal sympathetic ganglion explants or adrenal chromaffin cell aggregates were implanted into the left striatum of monkeys whose left nigrostriatal pathway had been lesioned with the neurotoxin MPTP. There was no clinical reversal of parkinsonian symptoms and PET scans did not show increased striatal fluorodopa uptake from pre-implant levels. At sacrifice, left striatal contents of dopamine were not statistically different from MPTP-treated but non-implanted controls. Histological examinations revealed pockets of extrinsic cells which were found at the end of needle tracks. There was no evidence of immune rejection. The extrinsic cells did not stain for tyrosine hydroxylase or neurofilament, suggesting that they were not dopaminergic neurons. The failure to reverse clinical parkinsonian symptoms highlights the stage of infancy of neural implantation in Parkinson's disease.

Adrenal Medulla↗

PubMed1988

Growth factors for fetal and adult human astrocytes in culture.

Using a recently developed method for the determination of cell proliferation in culture, we assessed the effects of several growth factors on their capability to promote proliferation of astrocytes derived from human fetuses or adults. Factors tested included fibroblast growth factor, epidermal growth factor, platelet-derived growth factor, glial growth factor from bovine pituitary, nerve growth factor, interleukin-2, dibutyryl cyclic adenosine monophosphate, and 4 beta-phorbol 12,13-dibutyrate. The results show that astrocytes from human adults do not undergo proliferation in vitro even in the presence of these agents and that in the case of fetal astrocytes, only glial growth factor from bovine pituitary, platelet-derived growth factor and the phorbol ester were mitogenic. In addition, the capability of fetal astrocytes to proliferate appears to decrease with the increasing length of time they have been maintained in culture.

Adult↗