ABSTRACT
Xenogeneic mouse models are widely used for the study of human tumor growth and metastasis. To date, few methods have been developed to track and quantitate the colonization of mouse organs with transplanted human cells. In this paper, a family of nonradioisotopic DNA oligonucleotide probes that are complementary to sequences within the human Alu element are characterized. These probes can be used in Southern hybridization reactions to quantitate the colonization of mouse organs with human derived cells. One oligonucleotide probe, the Alu-C probe, was identified as the most sensitive and specific in the family of probes synthesized for the distinction of human genomic DNA in a mouse genomic DNA background. The Alu-C probe can identify 0.05 ng human diploid DNA in a mouse background of 500 ng of genomic DNA. This represents 7.5 human diploid cells admixed with 75,000 mouse diploid cells. The Alu-C probe can therefore be employed to assess human colonization in xenograft models for a variety of human tumors and non-neoplastic tissues.
Subject(s)
Colonic Neoplasms/genetics , Colonic Neoplasms/pathology , DNA/blood , Lymphocyte Transfusion , Repetitive Sequences, Nucleic Acid , Transplantation, Heterologous , Animals , Base Sequence , Blotting, Southern , Cell Line , DNA/genetics , Humans , Mice , Mice, SCID , Molecular Sequence Data , Oligonucleotide Probes , Sensitivity and Specificity , Tumor Cells, CulturedABSTRACT
In order to determine the biological effects of activation of the c-fos protooncogene on growth and differentiation of BALB/MK mouse keratinocytes, these cells were transfected with the plasmid pMAN-fos, which encompasses the human c-fos coding region under transcriptional control of a mouse mammary tumor virus promoter, as well as the gene encoding neomycin phosphotransferase. Of approximately 70 individual clones obtained by selection in Geneticin, 5 clones that constitutively expressed c-fos mRNA as well as p55fos protein were selected for phenotypic analysis. Each of these clones displayed density-dependent growth arrest at confluency when deprived of serum and mitogens. DNA synthesis could be reinitiated in quiescent cultures by treatment with epidermal growth factor to a similar extent as in the parental line. In four of five fos transfectants, insulin-like growth factor 1 potentiated the mitogenic response to epidermal growth factor more than 10-fold, compared to only 2.5-fold in the parental cells. The enhanced response to insulin-like growth 1 could not be attributed to changes in the number or the affinity of receptors for the growth factor. Finally, the constitutive expression of c-fos did not interfere with the induction of terminal differentiation by calcium.
