F. Anders

Tumour formation in platyfish-swordtail hybrids as a problem of gene regulation

Bei bestimmten Zahnkarpfen-Bastarden treten stets erbbedingte Melanome auf. Diese entstehen dadurch, dass bestimmte Gene, die für die Differenzierung von Farbzellen verantwortlich sind, enthemmt und gleichzeitig zu einer gesteigerten Aktivität angeregt werden. Die Enthemmung beruht auf einem Verlust bestimmter Repressionsgene und die Aktivitätssteigerung auf einer Einführung polyfaktorieller Systeme von Induktionsgene. Es bestehen schwerwiegende Indizien dafür, dass die Induktorsubstanzen mit Aminosäuren identisch sind.

Xiphophorus as an in vivo model for studies on normal and defective control of oncogenes.

Publisher Summary Oncogenes are genes that code for neoplastic transformation and possibly for the maintenance of the neoplastic state of a cell. This chapter unifies populational, morphological, developmental, and cell biological findings obtained during research on the biology of the oncogene Tu of Xiphophorus. In addition, it shows that neoplasia of multicellular animals including humans results from the elimination, deletion, impairment, or insufficicncy of regulatory genes that normally control the oncogene, or from the introduction of uncontrolled accessory oncogenes into the genome. The Xiphophorus tumor system has provided the opportunity to reduce the enormous complexity of cancer etiology to a few biological elements basically involved in neoplasia. The development of a tumor requires an oncogene that after impairment, deletion, or elimination of its regulatory genes is permitted to mediate neoplastic transformation. Emphasis is being placed today in cancer research on the actual oncogenes themselves, but, the most important genes involved in neoplasia are these regulatory genes. However, although detected by classical genetics in the Xiphophorus system, these genes are not at present open to a more finely detailed molecular biological analysis. Their actual mode of action is, therefore, still far from being understood.

The involvement of reactive oxygen species in oral cancers of betel quid/tobacco chewers

Most biological reactions, including carcinogenesis, are complex processes involving thousands of compounds, their metabolites and intermediates. The separation of events which form part of a direct chain leading to neoplastic transformation from those which are mere by-products is a herculean task. In this study, we focused on the pros and cons of reactive oxygen species (ROS) being involved in the development of oral cancer among chewers of tobacco and areca nuts. The results revealed that bursts of ROS generation occur at different stages of carcinogenesis, and are caused by different mechanisms. This observation may have considerable practical implications. Different strategies will be required in the administration of chemopreventive agents in order to trap ROS formed in the alkaline (due to the addition of slaked lime) chewing mixture within the saliva of a chewer, to scavenge ROS within mucosal cells exposed to an array of tobacco- or areca nut-related carcinogens or tumour promoters, and to inhibit the action of ROS released from ROS-generating white cells during lymphocytic infiltration of the oral mucosa at a precancerous stage. The remission of oral leukoplakias following the administration of vitamin A (200,000 IU/week) or vitamin A (100,000 IU/week) plus beta-carotene (180 mg/week) for 6 months, the inhibition of new leukoplakias during this trial period, and the reduction of micronucleated oral mucosal cells in chewers treated with beta-carotene or vitamin A are indeed promising results. However, a better understanding of the role of ROS in various stages of carcinogenesis will provide the basis for selection of the proper chemopreventive agents and the design of a treatment regime which may either prevent the formation of precancerous lesions, induce their remission, or inhibit the progression of precancerous lesions into malignant cancers.

Regulation of Gene Expression in the Gordon-Kosswig Melanoma System

The Xiphophorin fish, including platyfish and swordtails, live in brooks, rivers, ponds and pools in Central America, where they have developed many species, subspecies, and populations. In some populations nearly all individuals have certain spot patterns composed of macromelanophores, in others only a certain percentage of the individuals are spotted, then again others exhibit no spots whatsoever (Gordon, 1942, 1947; Gordon and Gordon, 1950, 1957; Kallman, personal communication). Such spot patterns are, without a doubt, phylogenetic late acquirements which succeed in overcoming ecological and geographical barriers from population to population, and species to species.

Cell cultures derived from embryos and melanoma of poeciliid fish.

SummaryIn poeciliid fish, melanoma of different degrees of malignancy can be produced by crossing specific genotypes. For a detailed investigation of the processes leading to proliferation or differentiation of the melanoma cells, it is necessary to establish cell cultures. The aim of the present study was to find out the optimal conditions for initiating and culturing poeciliid fish cells for the purpose of establishing cell cultures of melanoma. The optimal method was developed by using small pieces of late embryos as starting material and includes: (a) dispersion of tissue by mild stepwise treatment with a trypsin-EDTA mixture at low temperature; (b) culture of cells in the complex medium 199; (c) supplementation of medium with high percentage (20%) of fetal bovine serum; and (d) stabilization of pH by buffering the medium with HEPES. Under these conditions, primary and secondary cultures of embryonic cells have been initiated. An epithelial-like cell line has been subcultured for more than 80 passages. The method developed for embryonic tissues was used to start cell cultures from melanoma of platyfish-swordtail hybrids. Until now, only cells of rapidly growing malignant albino melanoma could be maintained in primary cultures. Secondary cultures could not be initiated since the melanoma cells tended to differentiate and stopped growing before a confluent monolayer was formed.

Genetic basis of susceptibility for development of neoplasms following treatment with N-methyl-N-nitrosourea (MNU) or X-rays in the platyfish/swordtail system

Specific genotypes of the xiphophorine fish develop neoplasms following treatment with N-methyl-N-nitrosourea or X-rays. Several of these neoplasms can be related to the presence of specific chromosomes. The implications of these findings are discussed.

Sex chromosome aberrations involving loss and translocation of tumor-inducing loci inXiphophorus

Karyotypic changes involving a deletion and a translocation of certain sex-linked tumor-inducing loci of the platyfish,Platypoecilus (Xiphophorus) maculatus, have been investigated. The effects of these chromosome aberrations on tumor formation and viability of the fish are discussed.

Genetics of susceptibility in the platyfish/swordtail tumor system to develop fibrosarcoma and rhabdomyosarcoma following treatment with N-methyl-N-nitrosourea (MNU)

About 7000 animals of 65 different genotypes of the xiphophorine fish were treated with the direct acting chemical carcinogen N-methyl-N-nitrosourea (MNU; 10−3 M; four times for 1 hour in two week intervals), in order to find out, whether the susceptibility for development of fibrosarcomas and rhabdomyosarcomas is directly related to the genotype. A genotype specific susceptibility was found, ranging from zero to about nine percent. The highest susceptibilities were found in certain backcross hybrids involving P.variatus/X.helleri-hybrids and X.helleri as the recurrent parent. These genotypes were further analysed. Both P.variatus and X.helleri, as well as their F 1 proved to be insusceptible; while from the three backcrosses, which were tested, namely the BC 1, BC 4 and BC 15, both the BC 1 and the BC 4, were susceptible, but the BC 15 was insusceptible. The results are interpreted on the basis of the assumption that the differential susceptibility is a function of the type of control of a tumor gene (Tu-Fi-Rh) endogenous to P.variatus and involved in development of fibrosarcomas and rhabdomyosarcomas. Accordingly, in P.variatus and in the F1 the Tu-Fi-Rh is controlled by repressing genes (R-genes) linked as well as non-linked to Tu-Fi-Rh; because simultaneous mutation of both R-genes following treatment with MNU is an extremely unlikely event, these genotypes have an extremely low susceptibility. By contrast, in the BC 1 and the BC 4 the non-linked R-genes become eliminated and only the linked R-gene remains for repression of Tu-Fi-Rh; this condition confers a high degree of susceptibility, because one single mutation may lead to impairment of the R-gene and to Tu-Fi-Rh-mediated formation of fibrosarcomas and rhabdomyosarcomas. In the BC 15, furthermore, also the Tu-Fi-Rh has become eliminated, resulting in a loss of the susceptibility. The results suggest that in the xiphophophorine fish the susceptibility for responding to MNU-treatment with the development of fibrosarcomas and rhabdomyosarcomas is related directly to the genotype.

The induction of a specific pigment cell type by total genomic DNA injected into the neural crest region of fish embryos of the genus Xiphophorus

SummaryWe report genetic transformation in an intact higher organism, i.e., in xiphophorine fish. The gene to be transferred (Tu) is responsible for the formation of T-melanophores in the platyfish and is involved in the formation of melanomas in platyfish-swordtail hybrids. After injection of Tu-donor DNA into the neural crest region of embryos from Tu-free fish, some of the recipients developed T-melanophores. In a few cases, one or two single T-melanophores were formed during late embryogenesis. In most cases, many T-melanophores developed in young fish and were arranged in several colonies or in a pattern. DNase-degraded Tu-donor DNA, Tu-free fish DNA, as well as DNA from E. coli and adenovirus-2, did not induce T-melanophores. When using DNA from different strains of Tu-donor fish which differed in a regulating gene linked to Tu, the percentages of fish showing T-melanophores paralleled the degree of phenotypic expression of the Tu gene in the DNA donor. The results suggest that the Tu gene has been successfully transferred together with the linked regulating gene.

Virus-like particles induced by bromodeoxyuridine in melanoma and neuroblastoma of xiphophorus

Summary5-Bromodeoxyuridine (BrdUrd) injected into the muscular tissue of fish bearing melanoma or neuroblastoma induces the production of virus-like particles in these tumors. The particles in the melanoma are morphologically similar to papovaviruses of polyoma-type, those in the neuroblastoma resemble oncornavirouses of B- and C-type.