Stellate Shape Research Articles

Modulation effects of hexamethylene bisacetamide on growth and differentiation of cultured human malignant glioma cells.

The modulation effects of hexamethylene bisacetamide (HMBA), a differentiation-inducing agent, on growth and differentiation of cells from human malignant glioma cell line SHG-44 were studied. At cytostatic doses (2.5 mM, 5 mM, 7.5 mM, and 10 mM for 15 days), HMBA exerted a marked inhibitory effect on cell proliferation. Exposure to HMBA (5 mM and 10 mM for 12 days) also resulted in an accumulation of cells in G0/G1 phase and a decrease of cells in S phase as analyzed by flow cytometry. The reversible effects of 7.5 mM HMBA and 10 mM HMBA on cell proliferation and 10 mM HMBA on disruption of cell cycle distribution were observed when HMBA was removed from culture media on Day 6 and replaced with HMBA-free media. Colony-forming efficiency (CFE) in soft agar was remarkably decreased by HMBA (2.5 mM, 5 mM, 7.5 mM, and 10 mM for 14 days), and in 7.5 mM HMBA- and 10 nM HMBA treated cells, the CFEs were reduced to 25% and 12.5%, respectively, of that in untreated cells. Cells treated with HMBA (5 mM and 10 mM for 15 days) remained tumorigenic in athymic nude mice, but the growth rates of the xenografts were much slower than those in the control group. The effects of HMBA on cell proliferation, cell cycle distribution, CFE, and growth of xenografts were dose dependent. A more mature phenotype was confirmed by the morphological changes from spindle shape to large polygonal stellate shape and remarkably elevated expression of glial fibrillary acidic protein in cells exposed to HMBA (5 mM, 10 mM for 15 days). Our results showed that a more differentiated phenotype with marked growth arrest was induced in SHG-44 cells by HMBA.

Langhans cells of human arterial intima: uniform by stellate appearance but different by nature

The stellate cells in human arterial intima known as Langhans cells were investigated. Arterial specimens were obtained during carotid endarterectomy and aortic reconstruction and included atherosclerotic lesions as well as areas of the adjacent normal appearing arterial wall. Following immunohistochemical and electron microscopic analysis, most of the stellate cells were found to inhabit the elastic-hyperplastic layer of the intima in the normal arterial wall but in atherosclerotic lesions, stellate cells were distributed throughout all intimal layers. Immunohistochemical examination revealed that different types of intimal cells, including smooth muscle cells (HHF-35; smooth muscle alpha-actin +) and vascular dendritic cells (CD1a +, S-100 +), exhibited a typical stellate appearance but the cell processes of macrophages (HAM56 +, CD68 +) were too short for macrophages to be considered as stellate. No other intimal cells formed processes which could be detected under immunohistochemical examination. In atherosclerotic lesions, some smooth muscle cells transforming to foam cells retained their stellate shape. Smooth muscle cells interacted with each other through gap junctions while other intimal cells including vascular dendritic cells contacted each other without forming any specialized structures. We conclude that Langhans cells comprise two histological types of intimal cells, namely, smooth muscle cells and vascular dendritic cells.

Confocal imaging of the keratocyte network in porcine cornea using the fixable vital dye 5-chloromethylfluorescein diacetate.

This study reports on the combined use of an aldehyde fixable, cell viability fluoroprobe, 5-chloromethylfluorescein diacetate (CMFDA), confocal laser scanning microscopy and digital image reconstruction, to produce high resolution images of corneal keratocyte preparations in situ. The central region of freshly enucleated porcine corneae were removed and stained overnight at 4 degrees C with CMFDA. The tissue was washed, fixed, and frozen for cryosectioning in either a horizontal or antero-posterior orientation. Sections from anterior, central and posterior stroma were examined with a confocal microscope, and the digital images rendered as three-dimensional stereo reconstructions. Fluorescent CMFDA which completely permeated the cell bodies and extremes of the finest ramifying cell processes of all keratocytes provided exceptional high resolution images of the three morphologically distinct cell subpopulations at different levels of the stroma, and enabled improved characterisation of each cell type. Anteriorly was a thin, dense, non-lamellar network of keratocytes subjacent Bowman's membrane. In the central stroma, keratocytes were arranged in layers, the cell bodies had a flattened pyramidal or stellate shape, and the fine cell processes formed extensive distal ramifications. Immediately anterior to Descemet's membrane a small subpopulation of keratocytes with large cell bodies and short branched processes was identified. Extensive and diverse cell-to-cell contacts were orientated in all stromal planes, including ramping cell bridges between keratocyte lamellae in the central stroma. The use of the cell viability dye CMFDA is feasible and valuable for enhancing the visibility of entire keratocyte population in the intact cornea. Diverse multi-directional cell processes and intercellular contacts throughout the keratocyte network suggest a strong capacity for direct communication and cohesion in the maintenance and repair of the stromal matrix. Keratocytes closely related to the epithelium and endothelium have unique morphologies which may relate to specialised functions of these interface cells.

Isolated chick osteocytes stimulate formation and bone-resorbing activity of osteoclast-like cells

Several possible roles have been proposed for osteocytes in bone metabolism, but the precise functions of these cells are still obscure. In this study, we examined the effects of osteocytes on osteoclast formation, using isolated osteocytes from 16-day-old chick embryonic parietal bones and mouse hemopoietic blast cells from spleen cells obtained from 5-fluorouracil-treated mice. Purity of the isolated osteocytes was more than 95%, and their osteocytic phenotypes—such as those having a typical stellate shape and being immunoreactive for osteocyte-specific monoclonal antibody OB 7.3, the nonproliferative phenotype, and those negative for alkaline phosphatase (ALP) activity—were retained during 3 days of culture. When hemopoietic blast cells were cocultured with the isolated osteocytes, the number of osteoclastic tartrate-resistant acid phosphatase (TRAP)-positive multinucleate cells (MNCs) from the blast cells increased even in the absence of any osteotropic factors. This stimulatory effect depended on the number of osteocytes added, and was consistent with that obtained with bone fragments freed of nonosteocytic cells by stripping the periosteum. Conditioned medium (CM) of isolated osteocytes also had stimulatory effects on both formation and bone-resorbing activity of the MNCs, indicating that osteocytes increased osteoclastic development, at least in part, via their production of some soluble factor(s). This osteoclast-inducing activity in the osteocyte CM was observed in the fraction that eluted at around 0.3 M NaCl from a mono-Q anion-exchange column, and this elution profile was similar to that of cell lysates of freshly isolated osteocytes and of stripped bone fragments, indicating the existence of such osteoclastogenesis factor(s) in vivo. Finally, both the osteocyte CM and the active fraction of mono-Q chromatography increased the pit area on dentine slices in the cultures of unfractionated mouse bone cells as well. Thus, osteocytes may play a role in the physiological processes of osteoclastic bone resorption.

Morphological characteristics of neurons in the superficial layers of the rabbit's superior colliculus projecting to the ipsilateral dorsal lateral geniculate nucleus.

The mammalian superior colliculus (SC) may be divided into distinct layers on the basis of cytoarchitectonic criteria. These layers have different functions and patterns of connectivity. The superficial layers (stratum zonale, stratum griseum superficiale (SGS) and stratum opticum) are intimately associated with the processing of visual information. In the present study, we have investigated the morphology of the SC neurons that project ipsilaterally to the dorsal lateral geniculate nucleus (dLGN). Rhodamine latex microspheres were injected into the dLGN of nine adult rabbits. Retrogradely labeled cells were then intracellularly injected with Lucifer Yellow. The somata of the cells from which the projection originated were primarily located in a band occupying the medial third of the SGS and were found to reside at an average depth of 373 microns. Morphological analysis of these neurons revealed that 37% had a stellate shape, 27% were vertical fusiform, 18% were globular/pyriform, 14% were oriented horizontally and 4% were pyramidal in their morphology. Within each morphological class, we have examined the different subtypes with respect to incidence, localisation and characteristics of the dendritic arborizations.

Gonadal hormone regulation of neuronal-glial interactions in the developing neuroendocrine hypothalamus.

Recent evidence indicates that, in addition to their well known effects on neurons, gonadal steroids may exert part of their neural effects through astroglia. In adult female rats astroglia participate in the phasic remodelling of synapses that takes place during the estrous cycle in the arcuate nucleus of the hypothalamus under the influence of estradiol. Astroglia also appear to be involved in the genesis of sex differences in synaptic connectivity. Gonadal steroids influence hypothalamic astroglia differentiation in vitro and in vivo. In monolayer mixed neuronal-glial cultures from fetal rat hypothalami, estradiol induces a progressive differentiation of astrocytes from a flattened epithelioid morphology to bipolar, radial and stellate shapes. This effect of estradiol on astroglia is dependent on the expression of specific molecules on the neuronal surface, such as the polysialic acid-rich form of the neural cell adhesion molecule. In the rat arcuate nucleus in situ, perinatal androgen influences astroglia gene expression and differentiation, resulting in a sex difference in astroglia organization by postnatal day 20. By this day, the amount of neuronal surface covered by astroglial processes is higher in males than in females. This difference in the coverage of neuronal surface by astroglia may be directly related to the reduced number of synaptic contacts that is established on the soma of male neurons compared to females.

Effects of Triiodothyronine on Retention of β-Adrenergic Responsiveness of Voltage-Gated Transmembrane Calcium Current during Culture of Ventricular Myocytes from Neonatal Rabbits

To study the effects of triiodothyronine (T3) on responsiveness of L-type calcium currents to beta-adrenergic stimulation in neonatal hearts, ventricular myocytes were isolated from neonatal rabbits and cultured in medium containing 10% fetal bovine serum to which T3 had been added to achieve either hypothyroid, euthyroid, or hyperthyroid conditions, as assessed by measurement of free T3 concentrations. During a 24-h culture period, the striated rod-shaped myocardial cells progressively assumed a stellate shape with reduced surface area; however, the rate constants for diffusion of Na+ from a microelectrode pipette into the cells remained unchanged. Voltage-dependent characteristics of L-type calcium currents as assessed by whole-cell voltage clamp studies were also unchanged after culture with various concentrations of free T3. By contrast, the stimulation of voltage-gated transmembrane calcium current from baseline by isoproterenol was reduced (p < 0.05) in hypothyroid cells (15 +/- 8%; n = 14) compared with either euthyroid (86 +/- 15%; n = 18), hyperthyroid (54 +/- 16%; n = 12) or freshly isolated (50 +/- 12%; n = 10) myocytes. The differences in beta-adrenergic responsiveness of voltage-gated transmembrane calcium current to isoproterenol between euthyroid, hyperthyroid, and freshly isolated cells were not significant (p > 0.05). These results indicate that retention of beta-adrenergic responsiveness of voltage-gated transmembrane calcium current in neonatal cardiac myocytes depends on physiologic amounts of active thyroid hormone. Our culture method for neonatal cardiac myocytes will be useful for studying physiologic modulation of beta-adrenergic responsiveness.

Heterogeneity of smooth muscle cells in embryonic human aorta

Cellular composition of aortas from 5- to 12-week and 18- to 28-week-old human embryos were investigated using immunocytochemistry, scanning and transmission electron microscopy. The aorta of the 5- to 12-week-old embryos consisted of three sublayers differing in cellular composition. The inner sublayer adjacent to the endothelium contained round and ovoid cells with synthetic phenotype. In the intermediate sublayer, spindle-like cells ultrastructurally similar to smooth muscle cells were found. Cells of the outer sublayer resembled fibroblasts or poorly differentiated mesenchymal cells. There were not definite morphological borders between sublayers. In the 18- to 28-week-old embryo aorta the intima was separated from media by internal elastic lamina. Intimal and innermost medial cells had predominately stellate shape and synthetic phenotype. The outer part of media contained spindle-like cells that had well developed contractile structures. Both the 5- to 12-week-old and the 18- to 28-week-old embryo aortic cells were positively stained for α-actin and myosin and negatively stained for macrophage antigens. Thus, the majority of embryo aortic cells appeared smooth muscle cells, however there was a regional difference in shape and synthetic state of these cells.

Biosynthesis of Type VI Collagen by Glioblastoma Cells and Possible Function in Cell Invasion of Three-Dimensional Matrices

The biosynthesis of type VI collagen was studied in human glioblastoma cell line, U-87 MG. The effects of ascorbic acid on type VI collagen synthesis and secretion were investigated. After ascorbic acid treatment, type VI collagen in cell layers increased from 4.48% in control to 6.63% in the ascorbic acid treated cultures, an increase of 48%. The effect of ascorbic acid on type VI collagen synthesized by glioblastoma cells was lower than that reported for osteosarcoma cells (Engvall et al., 1986). The reason for these differences is still under investigation. The function of type VI collagen in glioblastoma cells is still unknown. We utilized the collagen gel system to elucidate the possible roles of type VI collagen in glioblastoma cells in vitro. Glioblastoma cells in collagen gels showed a stellate shape with long, branched processes in all directions. The strong positive reactivity of type VI collagen detected on cell bodies and cell processes by anti-type VI collagen antibody indicated that this specific collagen was associated with cell surfaces and processes, without releasing or diffusing into the gels. Type VI collagen was directly involved in the cell process extension. When living cells were treated with anti-type VI collagen antibody, a variation of cell morphology was observed. Instead of a stellate shape with processes, cells formed clusters without or with very short processes. These data suggest that type VI collagen, synthesized and secreted by glioblastoma cells, may play a role in tumor cell adhesion and spreading, and enhance cell process extension, penetration, and invasion into collagen gels.

Human wild type p53 inhibits cell proliferation and elicits dramatic morphological changes in human glioma cell lines in vitro

A human pilocytic astrocytoma-derived cell line, a grade III astrocytoma-derived cell line, and a glioblastoma-derived cell line were transfected with the human wild-type p53 gene, in order to demonstrate the possible suppressor role of this gene in low grade as well as in high grade human astrocytomas. p53 exhibited a strong growth suppressor effect on the three cell lines studied, irrespective of the grade of malignancy of the tumours from which they originate. Furthermore, the p53 gene elicited important morphological changes in these cell lines. p53-Transfected cells displayed a flat morphology, a large cell body, and a stellate shape with long processes, characteristic of differentiated astrocytes. In addition, the growth inhibitory effect of p53 was found not to be due to induction of apoptosis. These results indicate that p53 plays a tumour suppressor role in low grade and high grade human astrocytomas and raise the possibility of the involvement of p53 in glioma cell differentiation in vitro.

Spontaneous in vitro differentiation of a myoepithelial cell line (PA 16/23) from a pleomorphic adenoma of the parotid gland is associated with reduced production of the autocrine growth factor interleukin 6.

A myoepithelial cell line (PA 16/23) was derived from a pleomorphic adenoma of the parotid gland. PA 16/23 cells have light microscopic, immunophenotypical and ultrastructural features of immature myoepithelial cells, i.e. they are of fusiform or stellate shape and show keratin and actin cytofilaments located mainly in the perinuclear cytoplasm, desmosomes and tracts of basal lamina. The PA 16/23 cells grew actively and expressed mRNA for and produced interleukin 6 (IL-6) which was released into the culture medium. This cytokine, in turn, acted as an autocrine growth factor on the cells. PA 16/23 cells also expressed high-affinity IL-6 receptors. In these cells, both IL-6 production and proliferation could be modulated by exogenous stimulants, such as IL-6 itself, IL-1, IL-4, tumour necrosis factor alpha, interferon gamma and lipopolysaccharide. From the 40th culture passage onwards, the PA 16/23 cells ceased to grow, either spontaneously or in response to exogenous stimulants. Moreover, they strongly reduced IL-6 production, and underwent morphological differentiation into more mature myoepithelial cells, with an increased amount and a different arrangement of the keratin and actin cytofilaments, which formed thick bundles in the peripheral cytoplasm. These findings suggest a role for IL-6 in modulating the proliferation and, possibly, the differentiation of the PA 16/23 cells.

In vivo phosphorylation in the rat basal nucleus induces PHF-like and APP immunoreactivity

In vivo phosphorylation was stimulated in the rat basal nucleus by stereotaxic injection of the phosphatase inhibitor okadaic acid (OA). Hyperphosphorylation of neurofilaments and of the microtubule-associated proteins tau and MAP2 was associated with their redistribution from the axonal compartment into the cell bodies of large projection neurones where they appeared as paired helical filament (PHF)-like immunoreactivity. Astrocytes showed a dramatic increase in APP immunoreactivity and changed their appearance to a stellate shape with long processes. The results demonstrate that abnormal tau phosphorylation and changes in the expression and/or metabolism of APP can be induced in vivo by altering protein phosphorylation. The present experimental paradigm might, therefore, provide a useful tool to model early steps of the pathomechanism of Alzheimer's disease.

Astroglial differentiation is required for support of neurite outgrowth

Models of astrocyte differentiation stress a lineage program that involves a progressive loss of astroglial support of neuronal differentiation. These models predict that astroglial promotion of neurite extension declines with the "age" of the astrocyte. An alternative view is that astroglial support of neurite growth is regulated by epigenetic factors that induce the cells either to differentiate and support neuronal functions or to undergo cell proliferation and fail to support neurons. To compare the contribution of astroglial cell "age" to astroglial support of neurite extension, mouse cerebellar astroglia were maintained in vitro for 3-90 d, and assayed for their ability to support neurite formation. When cultured in isolation, astroglial support of neurite extension declined with time in vitro, as assayed by quantifying outgrowth from explants of pontine nuclei, falling from a robust level just after the astroglia were harvested to negligible levels 21-90 d later. Since previous studies have shown that neurons can change the state of astroglial cells (Hatten, 1985), we tested the neurite promoting activity of astroglia that were cultured for 21-90 d in vitro and subsequently induced to differentiate by the addition of neurons. When granule neurons were added to aged astroglia and pontine explants plated 2 d later, neurite growth from the explants was exuberant, regardless of the time astroglia spent in vitro prior to the addition of neurons. The state of astroglia that were growth promoting or growth inhibiting was examined by bromodeoxyuridine staining and with antisera to glial filament protein. Aged astroglia cultured alone and thus inhibitory to axon growth, proliferated at high rates and had polygonal shapes. In contrast, aged astroglia to which neurons had been added, proliferated at low rates and developed process-bearing stellate shapes. To test further whether proliferation levels related to the growth-supporting properties of astroglia, astroglia were plated alone in medium without serum, or with the addition of transforming growth factor-beta 1, each treatment known to arrest proliferation. In both cases, promotion of neurite growth was restored in aged astroglia, but the morphology of astroglia did not correlate with the ability to support neurite growth. Finally, the growth-inhibiting properties of aged astroglia do not appear to be mediated by diffusible factors, and require close apposition with living astroglial cells. We conclude that astroglial support of neurite extension depends on the state of differentiation of astroglial cells, and that these properties can be modified by coculture with neurons or conditions that arrest of astroglial proliferation, irrespective of astroglial "age".

Melanotropin receptors in the cartilaginous fish, Potamotrygon reticulatus and in the lungfish, Lepidosiren paradoxa

1. P. reticulatus skins in vitro exhibit punctate melanophores, and darken in response to the melanotropins in a dose-related manner. 2. The relative potencies (Ac-Nle4-alpha-MSH4–13-NH2< alpha-MSH = Ac-Cys4,Cys10-alpha-MSH4–13-NH2) suggest that the supression of the amino-terminal tripeptide may affect the ideal peptide conformation for interaction with the receptor, which is restored with cyclization of the decapeptide fragment. 3. Our data suggest that hormonal control of P. reticulatus pigment cells is mainly exerted by alpha-MSH, as none of the other agonists (isoproterenol, norepinephrine, MCH and melatonin) exhibited pigment dispersing or aggregating activities. 4. L. paradoxa scales possess non-innervated epidermal and dermal melanophores, which exhibit stellate shape in vitro. 5. Norepinephrine is a weak partial agonist in L. paradoxa epidermal melanophores, in the presence or in the absence of the beta-adrenoceptor blocker propranolol, suggesting that catecholamines do not play a role in the control of pigment cells. 6. Both epidermal and dermal melanophore responses to MCH and to its analogue MCH5–15, and to melatonin were also slow, partial and obtained with high doses of the hormones. 7. The relative insensitivity of L. paradoxa melanophores is in accordance with the animal cryptic behavior, since it lives in muddy dark waters, and does not seem to depend on chromatic adaptation for camouflage.

Confocal and Electron Microscopy of cultured cardiac myocytes

Early in heart development cardiac myocytes are spherical in shape, intercellular junctions are distributed at irregular intervals around the periphery of the cell, and myofibrillar organization is essentially random. As myocytes mature, they undergo extensive morphogenesis during which the phenotype changes to a tubular rodlike shape, cell junctions congregate at the distal ends of cells to form intercalated disks, and myofibrils become organized in parallel arrays typical of striated muscle. Although not fully understood, it is known that these changes are a result of interactive processes between intracellular components of the cytoskeleton, integrin membrane receptors, and the extracellular matrix (ECM).In vivo studies on the process of cardiac myocyte maturation and myofibrillogenesis are difficult because of the complex biochemical environment of the intact animal and the many extra- and intracellular interactions which are required for proper development and myofibrillogenesis. Unfortunately, in previously available in vitro modelling systems, isolated myocytes spread out over the culture substratum, assume a stellate nonpolar shape, and myofibril organization remains essentially random.

Myometrial characteristics of the syrian hamster uterine smooth muscle cell line, SHM.

We describe several characteristics of a novel smooth muscle cell line, SHM (Syrian hamster myometrium) derived from a primary uterine leiomyosarcoma which was induced by chronic estrogen plus androgen treatment of a female Syrian (golden) hamster. To determine the usefulness of the SHM cell line as a model for understanding myometrial function and its regulation, we have examined the morphologic and immunocytochemical properties of these cells, and the ability of uterotonic agonists to activate transmembrane signaling via phosphoinositide hydrolysis. The SHM cells exhibited a spindle-shape, smooth musclelike morphology when subconfluent, and a more compact, stellate shape at confluence. Like primary myocytes, SHM cells expressed the intermediate filament desmin and the contractile protein alpha smooth muscle actin, but not the epithelial antigen cytokeratin. Norepinephrine and bradykinin, which stimulate contraction and inositol polyphosphate production in the uterus, also stimulated inositol polyphosphate production in SHM cells. The maximal phosphoinositide signaling responses were lower in SHM cells compared with primary hamster uterine myocytes. We conclude that the SHM cell line exhibits primary uterine myocyte characteristics, and may therefore be a useful system for examining the mechanisms through which myometrial functions are regulated.

Phakomatosis Pigmentovascularis Type IIb With Iris Mammillations

Phakomatosis pigmentovascularis type IIb is a syndrome in which extensive nevus flammeus is associated with persistent aberrant mongolian spots. Herein, we describe a patient with phakomatosis pigmentovascularis who had numerous iris mammillations that were initially mistaken for the Lisch nodules of neurofibromatosis type I. A 5-year-old girl with phakomatosis pigmentovascularis type IIb was found to have bilateral melanosis oculi and numerous iris mammillations. The mammillations differed from Lisch nodules in their smaller size, stellate shape, darker color, increased number, and more regular distribution. Similar iris mammillations have been described in patients with melanosis oculi accompanying nevus of Ota. Patients with phakomatosis pigmentovascularis can present with the clinical manifestations of one or more of the following: Sturge-Weber syndrome, Klippel-Trenaunay syndrome, and melanosis oculi; our patient had clinical features of all three of these entities. In addition, the presence of iris mammillations in this patient can be explained by their known association with melanosis oculi.

Immunohistochemical localization of nerve growth factor in the rat pineal gland

Sympathetic nerve fibers arising from the superior cervical ganglia are the main innervation of the rat pineal gland. Since most organs innervated by these ganglia contain nerve growth factor (NGF), the hypothetical existence of NGF in the pineal gland was investigated. The peroxidase anti-peroxidase technique was applied for the immunohistochemical demonstration of NGF using a polyclonal antiserum on Bouin-fixed, paraffin-embedded pineal glands from adult, young and 6-hydroxydopamine (6-OHDA)-treated rats. Few immunopositive cells were observed in the adult pineal gland. A more conspicuous population of immunoreactive cells displayed a stellate shape resembling the interstitial or glial cells previously described in the rat pineal gland. Since NGF plays a trophic effect on sympathetic neurons during development and adulthood, we postulate that its presence in the pineal gland may exert a trophic role on its sympathetic innervation.

Synthesis of glial fibrillary acidic protein in rat C6 glioma in chemically defined medium: cyclic AMP-dependent transcriptional and translational regulation.

Glial fibrillary acidic protein (GFA) expression was induced in rat C6 glioma in chemically defined medium by the addition of N6, O2'-dibutyryl cyclic AMP (dbcAMP). Induction was dependent on the increase in intracellular cyclic AMP (cAMP), which was linearly correlated with added dbcAMP. Contrary to GFA mRNA synthesis, which can be obtained by cAMP-dependent and -independent pathways, translation of mRNA into GFA was observed only above a cellular cAMP concentration of approximately 0.2 fmol/cell. dbcAMP stimulation did not affect the vimentin concentration, which remained at a low level, but changed the cellular morphology from a bipolar to a stellate shape. A similar morphological change was observed after stimulation of C6 with lipopolysaccharide (LPS). However, LPS did not significantly increase the intracellular concentration of cAMP and the LPS-induced mRNA was not translated into GFA. Our results indicate that GFA synthesis is regulated at the mRNA level and at the translational level and that a cAMP-dependent mechanism determines the ultimate synthesis of GFA by a yet unknown mechanism.

Establishment and characterization of a mouse Schwann cell line which produces myelin in vivo.

A Schwann cell line (MSC 80) was established from purified mouse Schwann cell cultures using large doses of serum. MSC 80 cell line is an aneuploid cell line which has a doubling time of 17 hr and has been maintained through more than 110 passages. Most of MSC 80 cells are of bipolar or stellate (3-5 processes) shape. A few others are irregular in shape, gigantic, and multinucleated. All MSC 80 cells express antigens of myelin-forming Schwann cells such as S-100, 224/58, laminin, and other glycoproteins of the extracellular matrix. However, they also express the non-myelin-forming Schwann cell antigen GFAP. By time-lapse cinematography, MSC 80 cells exhibit the Schwann cell characteristic rhythmical undulations. When induced to form aggregates in agar, they form intercellular junctions and basement membrane-like structures. In addition, after transplantation in or at a distance from a lysolecithin induced lesion, MSC 80 cells form myelin around the host demyelinated axons. MSC 80 cells thus express, when isolated in vitro, some of the normal myelin-forming Schwann cell phenotype. In addition, they present the major advantage of forming myelin when associated with axons in vivo.