Heterogeneous response to differentiation induction with different polar compounds in a clonal rat rhabdomyosarcoma cell line (BA-HAN-1C)

Summary Differentiation-inducing ability of y-radiation, N,N-dimethylformamide and their combination has been tested on human rhabdomyosarcoma RMZ-RC2 clone cells. Ionising radiation at 2-5 Gy doses induced a more differentiated morphology, with the appearance of an increased proportion of multinuclear myotube-like cells, and a significant increase in myosin-positive and multinuclear cells. Radiation appeared to act by inducing de novo differentiated elements. N,N-dimethylformamide was able to induce an increased myosin expression, but did not affect multinuclear cell proportion. The combined treatment (ionising radiation and N,N-dimethylformamide) resulted in an additive increase in the proportion of myosin-positive cells, approaching 25-35%, but de novo differentiated elements were not increased above the levels obtained with irradiation alone.Differentiation induction therapy has been mainly investigated in leukaemic cells and in rodent solid tumours. In the last decade a few suitable human model systems have been developed (Reiss et al., 1986;Waxman et al., 1988).Induction of differentiation has been recently studied in human rhabdomyosarcoma cells. In vitro treatment with retinoic acid (Garvin et al., 1986) and phorbol esters (Aguanno et al., 1990) can induce myogenic differentiation; moreover in vivo growth and metastasisation in nude mice were impaired by differentiation induction (Lollini et al., 1991). We have previously shown that also some (but not all) antineoplastic drugs can induce myogenic differentiation of human rhabdomyosarcoma cells (Lollini et al., 1989). However in our model system, as in other solid tumours, a complete differentiation was not obtained; the possibility that combined differentiation therapy regimens might be more effective is now being investigated (Wiemann et al., 1988).In rodent rhabdomyosarcoma models, differentiation could be induced also by different polar compounds, e.g. N,Ndimethylformamide (Dexter, 1977) and N-methylformamide (Gerharz et al., 1989). Moreover, these compounds have been reported to enhance radiosensitivity of some human tumour cell cultures (Leith et al., 1982;Leith et al., 1985;Arundel et al., 1987). The aims of the present work are (a) to analyse the effect in vitro of a common therapeutic approach, radiation therapy, on the differentiation of human rhabdomyosarcoma cells, which are known to be radiosensitive (Kelland et al., 1989), and (b) to assess whether the combination of ionising radiation with an inducer of differentiation, N,N-dimethylformamide, may lead to an enhancement of myogenic differentiation. Materials and methodsCells and standard culture conditions Clone RMZ-RC2, derived from a human alveolar rhabdomyosarcoma and previously characterised (Nanni et al., 1986), was used between the 20th and the 30th in vitro passages. Cells were routinely maintained in Dulbecco's modified Eagle medium supplemented with 100 U ml-I penicillin, 100i,gml-' streptomycin (hereafter referred to as DMEM) and with 10% foetal calf serum (FCS). All media constitu...

Section: Discussionmentioning

confidence: 48%

Section: Discussionmentioning

confidence: 99%

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Induction of myogenic differentiation in human rhabdomyosarcoma cells by ionising radiation, N,N-dimethylformamide and their combination

Nicoletti

Giovanni²,

Landuzzi

et al. 1992

“…The ability to form myotube-like structures has been reported also in rat rhabdomyosarcomas induced by nickel (PotDeprun et al, 1983) or by dimethylbenzathracene (Gerharz et al, 1989).…”

Section: Discussionmentioning

confidence: 90%

In vitro differentiation of rhabdomyosarcomas induced by nickel or by Moloney murine sarcoma virus

Nanni

Azzarello²,

Tessarollo³

et al. 1991

Summary In vitro cultures and clonal derivatives have been established from rat rhabdomyosarcomas induced by Moloney-Murine Sarcoma Virus (MSV) or by nickel sulfide; differentiation ability has been studied as expression of desmin, embryonic and adult myosin isoforms, a-actin isoforms and cellular fusion. The two rhabdomyosarcoma models showed different levels of myogenic differentiation. Multinucleated myotube-like structures were frequently observed in cultures derived from nickel-induced tumours. Desmin was present in 50-80% of cells and embryonic myosin in up to 10%. In MSV-tumour-derived cultures and in their metastases or clonal derivatives two cell types are present in different ratios: spindle-shaped cells, adherent to plastic surfaces, and rounded cells, loosely attached or floating free in the medium. These cultures showed features of myogenic differentiation (10-80% desmin-positive cells), but embryonic myosin expression and production of multinucleated myotube-like structures were very rare events. Cultures from autochthonous lymph node and lung metastatic cells showed similar patterns of differentiation. Retinoic acid increased differentiated features (myotube formation and embryonic myosin expression) only in nickel-induced rhabdomyosarcoma cells. The two models described here mimic the heterogeneity in differentiation pattern found among human rhabdomyosarcomas. Myogenic differentiation ability was retained at a good level by nickelinduced tumours, whereas it was strongly impaired in MSV-induced tumours.

“…Cloning procedures were performed as previously described (Gerharz et al, 1989;Engers et al, 1994). Briefly, a single-cell suspension of tumour cells was diluted to a concentration of 3 cells ml-', and 0.1 ml of this cell suspension was inoculated into each well of a micro-well plate 96 (Gibco, Eggenstein, Germany).…”

Section: Isolation Of Clonal Subpopulationsmentioning

confidence: 99%

Chromophilic renal cell carcinoma: cytomorphological and cytogenetic characterisation of four permanent cell lines

Gerharz

Hildebrandt²,

Moll³

et al. 1996

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Summary Chromophilic renal cell carcinoma is a distinct type of human renal cancer, only recently recognised and defined by its characteristic histomorphological aspect and cytogenetic aberrations. We are the first to report on the establishment and cytogenetic characterisation of a panel of four permanent cell lines, i.e. chromphi-l, -2, -3 and -4, derived from strictly defined renal cell carcinomas (RCCs) of the chromophilic type and kept in continuous culture for up to 5 years. Immunohistochemistry revealed coexpression of vimentin and cytokeratins in all cell lines -the cytokeratin polypeptide patterns, however, varying between the different cell lines. By light and transmission electron microscopy, various amounts of cytoplasmatic glycogen deposition were observed, being most pronounced in chromphi-3 and -4. The mean population doubling time ranged from 24 h (chromphi-1) to 51 h (chromphi-4). Chromphi-1 tumour cells produced slowly growing tumours in nude mice using the subrenal capsule assay. In all cell lines, cytogenetic analysis revealed numerical chromosomal aberrations known to be characteristic for chromophilic RCCs, i.e. loss of the Y chromosome, tri-or tetrasomy of chromosomes 7 and 17 as well as various combinations of additional structural and numerical chromosomal aberrations. Karyological aberrations were least pronounced in chromphi-2 and most complex in chromphi-1.Chromosomal aberrations typically affecting the short arm of chromosome 3 in clear cell RCCs were not observed in any of our cell lines.