Interrelationship between differentiation and malignancy-associated properties in glioma

Frame, Margaret C.; Freshney, R. Ian; Vaughan, Peter F. T.; Graham, David I.; Shaw, Roosevelt

doi:10.1038/bjc.1984.44

Cited by 51 publications

(17 citation statements)

References 26 publications

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“…22 The CB 109 23 and the U251MG cell lines (American Type Culture Collection, Rockville, MD) were derived from human glioblastomas multiforme. All other cell cultures were derived from brain tumor biopsy specimens from the Regional Neurosurgery Unit of the Royal Victoria Hospital and were used experimentally within 5 to 10 passages.…”

Section: Cell Culturementioning

confidence: 99%

The Clinical Significance of Cathepsin S Expression in Human Astrocytomas

Flannery¹,

Gibson²,

Mirakhur³

et al. 2003

The American Journal of Pathology

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Early local invasion by astrocytoma cells results in tumor recurrence even after apparent total surgical resection, leading to the poor prognosis associated with malignant astrocytomas. Proteolytic enzymes have been implicated in facilitating tumor cell invasion and the current study was designed to characterize the expression of the cysteine proteinase cathepsin S (CatS) in astrocytomas and examine its potential role in invasion. Immunohistochemical analysis of biopsies demonstrated that CatS was expressed in astrocytoma cells but absent from normal astrocytes, oligodendrocytes, neurones and endothelial cells. Microglial cells and macrophages were also positive. Assays of specific activity in 59 astrocytoma biopsies confirmed CatS expression and in addition demonstrated that the highest levels of activity were expressed in grade IV tumors. CatS activity was also present in astrocytoma cells in vitro and the extracellular levels of activity were highest in cultures derived from grade IV tumors. In vitro invasion assays were carried out using the U251MG cell line and the invasion rate was reduced by up to 61% in the presence of the selective CatS inhibitor 4-Morpholineurea-LeuHomoPhe-vinylsulphone. We conclude that CatS expression is up-regulated in astrocytoma cells and provide evidence for a potential role for CatS in invasion. Astrocytomas are the commonest primary brain tumors and within this group the malignant grade III and IV tumors predominate (anaplastic astrocytomas and glioblastomas). Despite their non-metastatic nature, prognosis is poor because tumor infiltration of surrounding brain leads to recurrence even after apparently radical surgery. Multiple stereotactic biopsies of astrocytomas have demonstrated isolated tumor cells at considerable distances from the main tumor mass, 1 and recent investigations 2 have shown that tumor cells may be cultured from histologically normal brain at a distance greater than 4 cm from the gross tumor. Astrocytoma invasion is an active process that involves interactions with the host extracellular matrix (ECM), proteolytic modification of the ECM, and migration of the tumor cells into the modified matrix. 3 Members of the metalloproteinase, serine proteinase, and cysteine proteinase superfamilies have been linked to glioma invasion. 4 The cysteine proteinase cathepsin B (CatB) is expressed in gliomas in vitro 5 and immunohistochemical studies have demonstrated its presence in astrocytomas and glioblastomas in contrast to its absence from astrocytes in normal brain. 6,7 The expression and activity of CatB is higher in anaplastic astrocytomas and glioblastomas compared with low-grade tumors and normal brain 8 and high levels of CatB are positively correlated with poor survival. 9 Human glioblastoma cell lines also exhibit higher levels of CatB expression than lower grade astrocytomas 10 and the functional relevance of the enzyme is confirmed by a marked reduction in in vitro invasiveness of a glioblastoma cell line transfected with antisense CatB cDNA. 11 There is evide...

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Section: Cell Culturementioning

confidence: 99%

The Clinical Significance of Cathepsin S Expression in Human Astrocytomas

Flannery¹,

Gibson²,

Mirakhur³

et al. 2003

The American Journal of Pathology

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“…This process extension can be enhanced by the synthetic steroid testololactone (13). Expression of glialcell-specific properties can also be induced by cAMP or Bt2cAMP (14)(15)(16). In this study the maturation of the RT4-B, RT4-E, and RT4-D cell types in response to Bt2cAMP and testololactone is examined.…”

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confidence: 99%

Cell-type-specific responses of RT4 neural cell lines to dibutyryl-cAMP: branch determination versus maturation.

Droms¹,

Sueoka²

1987

Proc. Natl. Acad. Sci. U.S.A.

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This report describes the induction of celltype-specific maturation, by dibutyryl-cAMP and testololactone, of neuronal and glial properties in a family of cell lines derived from a rat peripheral neurotumor, RT4. This maturation allows further understanding of the process of determination because of the close lineage relationship between the cell types of the RT4 family. The RT4 family is characterized by the spontaneous conversion of one of the cell types, RT4-AC (stem-cell type), to any of three derivative cell types, RT4-B, RT4-D, or RT4-E, with a frequency of about i0-5. The RT4-AC cells express some properties characteristic of both neuronal and glial cells. Of these neural properties expressed by RT4-AC cells, only the neuronal properties are expressed by the RT4-B and RT4-E cells, and only the glial properties are expressed by the RT4-D cells. This in vitro cell-type conversion of RT4-AC to three derivative cell types is a branch point for the coordinate regulation of several properties and seems to resemble determination in vivo. In our standard culture conditions, several other neuronal and glial properties are not expressed by these cell types. However, addition of dibutyrylcAMP induces expression of additional properties, in a celltype-specific manner: formation of long cellular processes in the RT4-B8 and RT4-E5 cell lines and expression of highaffinity uptake of y-aminobutyric acid, by a glial-cell-specific mechanism, in the RT4-D6-2 cell line. These new properties are maximally expressed 2-3 days after addition of dibutyrylcAMP. This indicates that conversion of RT4-AC to the derivative cell types is also a branch point for the regulation of cell-type-specific properties whose expression is responsive to cAMP. Thus, the potential for maturation in response to increased cAMP is a property that segregates in a cell-typespecific manner and is activated at the determinational level in this system. Differentiation can be considered to involve two distinct processes: determination and maturation. Determination is the process by which multipotential stem cells segregate genetic potential to give rise to cells with a more restricted fate or range of fates. This process will subsequently be referred to as branching. In contrast, maturation is the process by which cells come to express more of the celltype-specific properties characteristic of that particular cell type. Several lines of evidence indicate that maturation is a result of the activation of genes that were preprogrammed to be functional in response to extracellular signals. The adipose precursor cell line (3T3) (1), a mammary epithelial precursor cell line (Rama 25) (2), and the pheochromocytoma cell line (PC12) (3) exemplify this.This report describes the cell-type-specific maturation, in response to N6,02'-dibutyryladenosine 3',5'-cyclic monophosphate (Bt2cAMP) and testololactone, of the four mor- RT4-AC(Stem cell type) + SloP., ± GFAP') + Na+-influx + K+-efflux Tumorigenic J

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“…We report here that the glial fibrillary acidic protein (GFAP), an intermediate filament protein almost exclusively confined to glial cell types in the central nervous system (CNS) and the PNS in vertebrates (9)(10)(11), is expressed under certain conditions in the RT4-AC and RT4-D cell lines. Previously, only a few immortalized cell lines that express GFAP, all of them derived from the CNS, have been reported; clonal propagation of cell lines from GFAP-positive astrocytic tumors generally results in the irreversible loss of the GFAP marker (12)(13)(14)(15). GFAP expression in the RT4 cell lines is shown here to segregate from the RT4-AC line to the derivative cell lines in a similar fashion to S-100, including at the RNA level.…”

Section: Introductionmentioning

confidence: 68%

“…Additionally, we have shown that GFAP expression in the GFAP-responsive RT4 cell lines is cell-density dependent in a manner already described for astrocytes in primary culture and for one continuous brain-derived cell line (the rat glioma line C6) (27,30,31 Primary astrocyte cultures and the rat C6 line have also been reported to enhance GFAP levels in response to Bt2cAMP addition (13,25,27,31). As observed for the above cell types, GFAP induction in response to Bt2cAMP addition occurs along with a significant morphological differentiation in the GFAP-positive RT4 cell lines.…”

Section: Resultsmentioning

confidence: 98%

Induction and segregation of glial intermediate filament expression in the RT4 family of peripheral nervous system cell lines.

Freeman¹,

Sueoka²

1987

Proc. Natl. Acad. Sci. U.S.A.

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We have found glial fibrillary acidic protein (GFAP), the major component of astrocyte intermediate fiaments, to be expressed in cell lines of the RT4 peripheral neurotumor family. The RT4 family is a "stem-cell-like" cell line, RT4-AC, that spontaneously undergoes differentiation in culture to three derivative cell types. This process, termed cell-type conversion, results in a segregation among the derivative cell types of parental cell phenotypes that have been described as neuronal-like or glial-like. We have identified a 50-kDa GFAP-immunoreactive cytoskeletal protein and GFAP mRNA in continuous RT4-AC and RT4-D (glial-type derivative) cell lines, but not in two presumptive neuronal-type cell lines. This result suggests that GFAP gene expression is coordinately coupled with the expression of other glial properties during cell-type conversion. In addition, the RT4-AC and RT4-D sublines were found to significantly express GFAP only at high cell densities and not during logarithmic growth and to express GFAP precociously during morphological differentiation following treatment with 1 mM N6,02'-dibutyryladenosine 3',5'-cyclic monophosphate. These observations closely reflect reports of glial filament expression in astrocyte cultures, suggesting that a common regulatory mechanism is employed by central and peripheral nervous system glia.Our laboratory has described the isolation and characterization of clonal cell lines from the peripheral nervous system (PNS) tumor RT4 (1-8). The RT4 tumor was induced by ethylnitrosourea injection of a newborn BDIX rat, a procedure that predominantly gives rise to PNS tumors (1). An unstable cell line, termed RT4-AC, has been described (1, 3) that expresses the glial marker protein S-100 and that also displays voltage-sensitive Na' and K+ channels. The RT4-AC cell line was described as a "stem-cell-type" line because of its ability to repeatedly give rise to three distinct progeny cell types in culture (Fig. 1). These derivative cell types have been described as glial-like (the RT4-D cell type), based on the expression of S-100 and on the loss of the excitable membrane phenotype, or neuronal-like (the RT4-B and RT4-E cell types), based on the loss of S-100 expression and on the retention and enhancement of the excitable membrane phenotype. This differentiation phenomenon has been termed cell-type conversion, and the apparent coordinate segregation of "glial" or "neuronal" properties with conversion has been termed conversion coupling. Clonal RT4 cell lines corresponding to these four cell types have been passaged continuously in culture for over 5 years.Droms and Sueoka (8) have shown that a normally unexpressed high-affinity y-aminobutyric acid uptake mechanism, generally considered to be a glial property in the PNS, can be induced in the RT4-D cell line by a combination treatment with N6,02'-dibutyryladenosine 3',5'-cyclic monophosphate (Bt2cAMP) and testololactone, a synthetic androgen. Under the conditions described, the high-affinity y-aminobutyric acid uptake mecha...

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Interrelationship between differentiation and malignancy-associated properties in glioma

Cited by 51 publications

References 26 publications

The Clinical Significance of Cathepsin S Expression in Human Astrocytomas

The Clinical Significance of Cathepsin S Expression in Human Astrocytomas

Cell-type-specific responses of RT4 neural cell lines to dibutyryl-cAMP: branch determination versus maturation.

Induction and segregation of glial intermediate filament expression in the RT4 family of peripheral nervous system cell lines.

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