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Biomedical subjects

D Hanahan

Publications and source records attributed to D Hanahan.

At least 91 records · Page 5Linked to original sources
PubMed1991

Switch to the angiogenic phenotype during tumorigenesis.

Tumor growth and metastasis are angiogenesis-dependent. Virtually all solid tumors are neovascularized by the time they are detected. However, there is a prevascular phase during early tumor development where few or no tumor cells are angiogenic and expansion of the tumor is restricted to a few mm3. When enough tumor cells become angiogenic, the tumor can expand progressively and shed metastatic cells. This angiogenic switch has recently been quantitated for human breast cancer, as well as for prostate cancer. We have studied the problem of how tumors switch to the angiogenic phenotype by using transgenic mice in which tumors develop at a predictable time and in discrete prevascular and vascular stages. When the transgene is the bovine papilloma virus (BPV) genome, angiogenic fibrosarcomas develop from non-angiogenic precursors called fibromatoses. The fibrosarcomas secrete growth factors for capillary endothelial cells. In contrast, the fibromatoses do not secrete endothelial cell growth factors. When the transgene consists of the large "T" antigen of SV40 under the control of the rat insulin promoter, 70% of pancreatic islets become hyperplastic and 4-10% of these become angiogenic at 6-7 weeks. Tumors arise from these neovascularized hyperplastic islets and reach > 1000 x the volume of the preangiogenic islets. The onset of angiogenic activity coincides with the secretion of acidic fibroblast growth factor (aFGF) and other growth factors not fully identified at this writing. These studies help to explain the switch to the angiogenic phenotype during tumorigenesis and provide models to discover antiangiogenic therapies directed at the source of angiogenic activity. Such therapy, when developed, may be co-administered with currently available angiogenesis inhibitors which are directed at the target of angiogenic activity, vascular endothelial cells.

Animals↗
PubMed1990

Differential plasmid rescue from transgenic mouse DNAs into Escherichia coli methylation-restriction mutants.

Plasmids comprising transgene insertions in four lines of transgenic mice have been retrieved by plasmid rescue into a set of Escherichia coli strains with mutations in different members of the methylation-dependent restriction system (MDRS). Statistical analysis of plasmid rescue frequencies has revealed that the MDRS loci detect differential modifications of the transgene insertions among mouse lines that show distinctive patterns of transgene expression. Plasmids in mice that express hybrid insulin transgenes during development can be readily cloned into E. coli strains carrying mutations in two of the MDRS loci, mcrA and mcrB. In mice in which transgene expression is inappropriately delayed into adulthood, plasmids can only be cloned into E. coli that carry mutations in all known MDRS activities. Differential cloning frequencies in the presence or absence of the various methylation-dependent restriction genes represent a further way to distinguish regions of mammalian chromosomes. These multiply deficient E. coli strains will also facilitate the molecular cloning of modified chromosomal DNA.

Animals↗
PubMed1990

Determinants of the B-cell response against a transgenic autoantigen.

The failure to induce self-tolerance of simian virus 40 large tumor antigen (T antigen) expressed in the pancreatic beta cells of transgenic mice results in an autoimmune response against this protein and the cells that synthesize it. In every transgenic mouse with delayed onset of T-antigen expression and consequent nontolerance, B cells, T cells, and macrophages are attracted to and infiltrate the pancreatic islets. In contrast, the incidence, onset, and intensity of the B-cell response to produce anti-T-antigen autoantibodies vary considerably with genetic background. Thus the initial attraction of lymphocytes to the cells synthesizing a non-self antigen can be separated from the activation of a B-cell response against it. Haplotypes of the major histocompatibility complex (MHC) differentially influence the character of the autoimmune response, with H-2d and H-2k conferring a high incidence of humoral autoimmunity. Additional non-MHC linked genes are also implicated in control of the B-cell response.

Animals↗
PubMed1990

Diabetes induced in male transgenic mice by expression of human H-ras oncoprotein in pancreatic beta cells.

Transgenic mice expressing an insulin-promoted H-ras hybrid gene in pancreatic beta cells developed beta-cell degeneration and diabetes. The disease was manifested in male mice by hyperglycemia, glycosuria, and reduced plasma insulin levels, which appeared around 5 months of age and led to premature death. Histological analyses revealed large holes within the islets of Langerhans and a reduced number of beta cells. The destruction of the islets was not associated with an obvious inflammatory activity. Ultrastructural analysis showed extensive engorgement in the endoplasmic reticulum of the residual beta cells from diabetic males. The females carrying the insulin-promoted ras gene did not manifest any of the physiological abnormalities observed in males and showed only minor histological and ultrastructural changes, even at much greater ages.

Animals↗
PubMed1990

Proglucagon processing similar to normal islets in pancreatic alpha-like cell line derived from transgenic mouse tumor.

A pancreatic alpha-like cell line has been established from a glucagonoma arising in transgenic mice expressing a hybrid gene consisting of the rat glucagon-promoter sequence fused to the sequence encoding the SV40 T-antigen oncoprotein. The alpha-tumor cell 1 (alpha TC1) line maintained many characteristics of differentiated alpha-cells for greater than 40 passages in culture and expressed levels of glucagon mRNA 5- to 10-fold higher than those reported previously in rat and hamster islet cell lines. By radioimmunoassay, the cells synthesized considerable amounts of glucagon, glucagonlike peptide I (GLP-I), the major proglucagon fragment, and small amounts of unprocessed proglucagon but no free GLP-II. This distribution of peptides is similar to that found in extracts of rodent pancreases and is distinct from that seen with other islet cell lines, which process proglucagon in patterns more characteristic of intestinal cells. The GLP-I peptide in the alpha TC1 cell line was in the form of GLP-I-(1-37), which is inactive as a stimulator of insulin secretion, and not GLP-I-7-37) or -(7-36)-amide peptides, both of which are potent insulin secretagogues. The alpha TC1 cell line produced glucagon-related peptides in a relatively uniform pattern by immunocytochemistry, and electron microscopy revealed typical alpha-type (glucagon) secretory granules. Although the cell line was derived from an islet tumor producing only glucagon, the alpha TC1 cell line also produced insulin in addition to the glucagon peptides.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenoma, Islet Cell↗
PubMed1989

Transgenic mice as probes into complex systems.

The transfer of genetic information into mouse embryos to stably alter the genetic constitution of mice is affording new insights into and opportunities in a wide variety of biological problems. Higher eukaryotes are composed of many interacting cells and organs. The properties of individual cell systems are often discernible only by studying natural or induced disruptions in their functions. Transgenic mice represent a new form of perturbation analysis whereby the selective expression of novel or altered genes can be used to perturb complex systems in ways that are informative about their development, their functions, and their malfunctions. The utility of this strategy is illustrated by recent research into immunological self-tolerance, oncogenes and cancer, and development.

Animals↗
PubMed1989

Directed expression of NGF to pancreatic beta cells in transgenic mice leads to selective hyperinnervation of the islets.

Nerve growth factor (NGF) is implicated in the differentiation of neurons in both the central and peripheral nervous systems. As a new approach to its role in neuronal development, we have used transgenic mice to selectively overexpress NGF in an innervated peripheral tissue, the islets of the pancreas. In two lines of mice, directed expression of NGF in the beta cells elicits a dramatic increase in the innervation of the islets, but not the surrounding exocrine tissue, by one class of sympathetic neurons. In contrast, the innervation by sensory and parasympathetic neurons appears unchanged. The results indicate that expression of NGF by a target tissue during neuronal development selectively influences the characteristics of its innervation.

Animals↗
PubMed1989

Induction of angiogenesis during the transition from hyperplasia to neoplasia.

It is now well established that unrestricted growth of tumours is dependent upon angiogenesis. Previous studies on tumour growth, however, have not revealed when or how the transition to an angiogenic state occurs during early tumour development. The advent of transgenic mice carrying oncogenes that reproducibly elicit tumours of specific cell types is providing a new format for studying multi-step tumorigenesis. In one of these models, transgenic mice expressing an oncogene in the beta-cells of the pancreatic islets heritably recapitulate a progression from normality to hyperplasia to neoplasia. We report here that angiogenic activity first appears in a subset of hyperplastic islets before the onset of tumour formation. A novel in vitro assay confirms that hyperplasia per se does not obligate angiogenesis. Rather, a few hyperplastic islets become angiogenic in vitro at a time when such islets are neovascularized in vivo and at a frequency that correlates closely with subsequent tumour incidence. These findings suggest that induction of angiogenesis is an important step in carcinogenesis.

Adenoma, Islet Cell↗
PubMed1989

Specific chromosomal abnormalities characterize fibrosarcomas of bovine papillomavirus type 1 transgenic mice.

In the BPV1.69 line of transgenic mice, the bovine papillomavirus type 1 genome elicits both benign dermal fibroblastic proliferation (fibromatoses) and malignant fibrosarcomas. Because these lesions arise only with time, nonviral factors appear to be involved. We have karyotyped several primary tumors as well as a series of low-passage cell lines derived both from fibromatoses and from fibrosarcomas. The fibrosarcomas, but not the preneoplastic fibromatoses, show consistent abnormalities of one or both of two chromosomes, chromosome 8 (trisomy or duplication) and chromosome 14 (monosomy or translocation). The chromosomal abnormalities are not a direct consequence of the viral integration, which we have mapped to chromosome 15 by in situ hybridization. These results suggest that transgenic mice can be used to study the role(s) of cytogenetic changes in tumorigenesis and may direct the search for genes involved in tumor progression.

Animals↗
PubMed1989

cDNA sequence of neuroendocrine protein 7B2 expressed in beta cell tumors of transgenic mice.

The cDNA for a widely distributed neuroendocrine protein called 7B2 has been cloned from beta cell tumors of transgenic mice and sequenced. As deduced from the cDNA sequence, 7B2 is a secretory protein of 186 amino acids, nearly identical to its human and porcine homologs. The presence of several pairs of basic residues in the carboxyl terminal portion of the protein suggests that 7B2 can undergo proteolytic maturation in secretory granules and thus generate potential bioactive peptides. 7B2 mRNA is about 1.5 kilobase long and is apparently transcribed from a single gene per haploid genome. The use of tissue-specific promoters to express oncogenes in rare cell types of transgenic mice is a powerful tool for immortalization and expansion of these cells, and it facilitates the isolation and the study of rare proteins such as 7B2.

Adenoma, Islet Cell↗
PubMed1989

Cell-heritable stages of tumor progression in transgenic mice harboring the bovine papillomavirus type 1 genome.

Tumorigenesis of dermal fibroblasts in a line of transgenic mice carrying the BPV-1 genome was found to involve distinct proliferative stages. Cell cultures derived from normal skin, from benign proliferative fibromatoses, and from malignant fibrosarcomas each evidenced distinguishable, cell-heritable characteristics. The latent viral genome was transcriptionally inactive in normal-appearing skin and was activated in the dermal fibromatoses. Fibrosarcoma cells grew continuously in culture, formed domelike foci, and had a more rounded, anaplastic appearance. Independent cultures derived from the fibromatoses varied in their proliferative characteristics, which correlated well with the levels of viral gene expression. In contrast, progression to malignancy was not accompanied by a further increase in transgene activity, which strongly implicated cellular genetic changes in the later stages of tumorigenesis.

Animals↗
PubMed1989

Tumorigenic latency and separable stages during fibrosarcoma development in transgenic mice carrying papillomavirus genomes.

Transgenic mice have been established carrying the genomes of bovine papillomavirus type 1 (BPV-1), and human papillomaviruses types 5 and 18. Transcriptional dormancy is characteristic of all three viral genomes in transgenic mouse lines maintained for 2-4 years. Only BPV-1, which induces both dermal and epidermal pathology in its natural host, has been found to elicit abnormalities when carried in transgenic mice. The BPV-1 genome acts in these mice as a tissue specific oncogene, in that it elaborates the development of skin fibrosarcomas. Three abnormal stages are evident: two distinct and successive stages of a proliferative hyperplasia (a mild and an aggressive fibromatoses), and the solid tumors (fibrosarcomas). Analysis of tissue biopsies and of derivative cell cultures from each pathology confirms that this pathway is composed of separate stages. Notably, progression from hyperplasia to neoplasia is accompanied by specific cytogenetic changes, which appear necessary in addition to the actions of the BPV oncogenes. The reproducibility of the tumorigenic pathway induced by the BPV genome is providing inroads into the molecular genetic and biochemical mechanisms of tumor development.

Animals↗
PubMed1989

Tissue specificity of oncogene action: endothelial cell tumours in polyoma middle T transgenic mice.

Newborn mice inoculated with murine polyoma (Py) virus develop tumours in a wide range of tissues. To investigate viral oncogenesis we have generated transgenic mice carrying either the Py large T antigen (LT) gene or the Py middle T antigen (MT) gene linked to Py early regulatory sequences. Some Py LT mice develop pituitary tumours, while Py MT mice develop multifocal tumours of the vascular endothelium. These haemangiomas are lethal to the animals and can be passaged in vivo. Transgene RNAs and protein are present in both haemangiomas and the testes of these mice, and the Py middle T protein in both tissues is complexed to a cellular tyrosine kinase. The expression of complexed middle T protein in both tumorigenic endothelial cells and in unperturbed testes implies that endothelial cells may be particularly susceptible to the action of the middle T oncogene. The idea that oncogenes may exhibit a tissue specificity in their action is supported by other transgenic mouse models of oncogenesis and by studies of human tumours.

Animals↗
PubMed1988

Hybrid insulin genes reveal a developmental lineage for pancreatic endocrine cells and imply a relationship with neurons.

Insulin appears in the developing mouse pancreas at embryonic day 12 (e12). Transgenic mice harboring three distinct hybrid genes utilizing insulin gene regulatory information first express the transgene product two days earlier, at e10, in a few cells of the pancreatic bud. Throughout development and postnatal life, all of the insulin-producing (beta) cells coexpress the hybrid insulin gene. In addition, islet cells containing glucagon, somatostatin, pancreatic polypeptide, and the neuronal enzyme tyrosine hydroxylase coexpress the transgene when they first arise. Similarly, coexpression of these normally distinct islet cell markers occurs during differentiation of the four endocrine cell types. The transgene product also appears transiently during embryogenesis in cells of the neural tube and in neural crest. The results suggest a common precursor for the endocrine cells of the pancreas. Moreover, they imply a relationship between neural and pancreatic endocrine tissue.

Animals↗
PubMed1988

McrA and McrB restriction phenotypes of some E. coli strains and implications for gene cloning.

The McrA and McrB (modified cytosine restriction) systems of E. coli interfere with incoming DNA containing methylcytosine. DNA from many organisms, including all mammalian and plant DNA, is expected to be sensitive, and this could interfere with cloning experiments. The McrA and B phenotypes of a few strains have been reported previously (1-4). The Mcr phenotypes of 94 strains, primarily derived from E. coli K12, are tabulated here. We briefly review some evidence suggesting that McrB restriction of mouse-modified DNA does occur in vivo and does in fact interfere with cloning of specific mouse sequences.

5-Methylcytosine↗