| Abstract |
In the in vitro establishment initiated in vivo by carcinogens
(pre)neoplastic liver cells showed similarities and differences to the
processes of hepatocarcinogenesis in test animals. In vivo and in vitro
only some of the initiated cells are able to survive in the long term,
only a few are immortalized or neoplaslically transformed in vitro. The
tumor promoter phenobarbital has an inhibiting effect on diethyl
nitrosamine-induced, established liver cell lines. Epidermal growth
factors such as hepatocyte-stimulating substances can promote DNA
synthesis or migration when the receptors are expressed. Depending on the
liver cell type and degree of differentiation, growth factors act
differently and synergistically with a large number of other components.
They are only partially cell-specific and often have a paracrine effect,
which results in different effects in primary, mixed-cell cultures than in
pure hepatocyte cultures.
The ten examined liver cell lines showed partly good differentiation and
stable morphology (CL 38). In lines that were difficult to establish (CL
52), the in vitro progression correlated with the occurrence of genetic
instability. The chromosome analyzes did not reveal any aberrations
specifically attributable to immortalization or neoplastic transformation.
A 3p or 11p deletion was often found, but with the progression and
occurrence of a mitotic rate sufficient for karyotyping, several
structural aberrations, karyotype heterogeneity and, not only after in
vivo transplantation, variability were detectable. Regardless of the
degree of differentiation or a morphology that remained stable for years,
all cell lines showed constant and increasing karyotype alterations. Thus,
newly occurring chromosome aberrations were only associated with changes
in growth or morphology in exceptional cases. A comparison with liver cell
lines from the literature and two spontaneous tumors, however, showed that
some regions of the RNO # 1, 3, 10 and # 11, 4, 6 and 7 not only in liver
cells, but generally in carcinomas of the rat are involved. The CL and FL
induced in vivo by continuous diethyl nitrosamine administration did not
show any carcinogen-specific chromosome alteration. In contrast, a single
dose of methylnitrosourea in vivo resulted in # 4 aberrations in all three
examined NT cell lines, as were also described for ethylnitrosourea-
induced neurogenic tumors.
Numerical chromosome losses associated with the progression were only
partially due to translocations, preferably to centromeres and dead cells.
When hypodiploidy was reached, the cell lines only remained if they were
able to (hypo) tetraploidize. Endoreduplication of individual chromosomes
or non-disjunction preserved For the limited survival time of already
propagated subcultures, as well as the lines CL 44 and CL 49-VII, a
telomere loss or a lack of telomerase is possible. The cell line CL 50
shows considerable cell division disorders with its mononuclear cells,
multipolar mitoses, micronuclei and PCCs. In relation to the latency
period until tumor formation, the degree of malignancy of the lines
correlates neither with the heterogeneity of the cell population, nor does
the aneuploidy correlate with the number of cylogenetically recognizable
aberrations. The essentially adherent line CL 49-VIII shows high cell
death in mitosis in the event of hyperconfluence. Nevertheless, it shows
short latency times and high metastasis in vivo. Oligonucleosomes typical
of apoptosis could be determined for cell loss, which is dependent on cell
density. The established lines CL 50 and CL 49 are a suitable in vitro
system for investigating the complex mechanisms of karyo-and cytokinesis,
or factors that lead to cell death through apoptosis.
As a sensitive in vitro system, the establishment of the liver cell lines
can provide generally valid information for primary epilhelic cell
cultures from parenchymal tissues. The classic karyotype analysis is not
very suitable for the detection of genetic aberrations in the case of
insufficient proliferation and lack of mitoses in initiated or
preneoplastic cells. For the cell type-specific investigation of early
stages of the cancerogenesis process, in situ hybridization on interphase
nuclei is the current method of choice. However, chromosome and gene
specific probes are not commercially available for Rattus norvegicus.
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