Clonal Derivation Research Articles

Relationships Among Pathogenic and Nonpathogenic Isolates ofFusarium oxysporumBased on the Partial Sequence of the Intergenic Spacer Region of the Ribosomal DNA

Using PCR, we amplified and sequenced approximately 1,000 bp of the 5' end of the intergenic spacer (IGS) of the rDNA in 15 isolates of Fusarium oxysporum and one isolate of F. subglutinans. Isolates were selected to represent diversity in our collection based on differences in pathogenic race, vegetative compatibility group (VCG), mitochondrial DNA (mtDNA) haplotype, IGS haplotype, and DNA fingerprint. The objective of this research was to clarify the origin of virulence within F. oxysporum, the relationship between pathogenic and nonpathogenic strains, and the evolution of the different races of F. oxysporum f. sp. melonis. Bootstrapped parsimony analysis of the partial IGS sequence data identified a phylogenetic tree with highly significant branches. The two F. oxysporum f. sp. melonis VCGs, 0131 and 0134, were separated into distinct lineages. Race was not distinguished by significant IGS sequence differences within the pathogen VCGs. One exception was a race 1 isolate which was associated with VCG 0131 but, based on both mtDNA and IGS haplotype, had greater affinity with VCG 0134. Two IGS sequence types were found in this race 1 isolate, one suggesting an affiliation with VCG 0131 and the other similar to isolates in VCG 0134. This may have resulted from past somatic or sexual interactions between F. oxysporum f. sp. melonis, VCGs 0131 and 0134. Nonpathogens that were vegetatively compatible with the pathogen were not closely related to the pathogen based on IGS sequence data. Thus, nonpathogens and pathogens may share common alleles at vegetative compatibility loci by coincidence rather than because of recent clonal derivation from a common ancestor.

Clonality in Myeloproliferative Disorders

The myeloproliferative disorders are a group of hematologic diseases that are believed to arise from somatic mutations in an early hematopoietic stem cell. This statement is based on the demonstration of monoclonal involvement of terminally differentiated myeloid and lymphoid elements. The techniques for establishing clonal derivation of cells are discussed and the application of these techniques to myeloproliferative diseases is reviewed. The evidence for limited myeloid involvement, lineage heterogeneity, in some patients with myeloproliferative disorders is summarized.

Isolation and characterization of an IL-1-dependent IL-4-producing bovine CD4 + T cell clone

An Ag-specific interleukin 1 (IL-1)-dependent bovine CD4 + Th cell clone, termed 300B1, was isolated and found to resemble the previously described IL-1-dependent murine CD4 + Th2 cell clone, D10.G4.1. Both the 300B1 and the D10.G4. 1 T cell clones proliferated to bovine (Bo) IL-1β human (Hu) IL-1α and IL-1β, and murine IL-1α when cells were costimulated with concanavalin A (Con A). Proliferation of the 300B1 clone, when costimulated with Con A, appeared to be IL-1 -specific in that proliferation could not be promoted by BoIL-2, HuIL-3, HuIL-4, HuIL-5, or HuIL-6. The 300B1 clone produced interferon-γ (IFN-γ), but not IL-2 following stimulation with either Con A, Con A plus phorbol 12-myristate 13-acetate or Ag plus antigen-presenting cells. Upon stimulation with Con A, the 300B1 clone expressed IL-4 mRNA and produced an automne growth factor (AGF) that could be inhibited by anti-HuIL-4 but not by anti-HuIL-2 Ab. The clonal derivation of the 300B1 clone was confirmed by isolating five 300B1 subclones, all of which produced IFN-γ and an AGF but not IL-2. Collectively, these results suggest the IL-1-dependent bovine 300B1 Th cell clone produces IL-4, but not IL-2, as an AGF. Furthermore, the bovine Th cell clone appeared to share many characteristics of previously described murine Th2 cell clones except that the bovine clone produced IFN-γ.

The molecular biology of parathyroid disease

Advances in molecular genetics have shed important new light on the understanding of the basis for human tumors. The application of these methods has allowed for characterization of endocrine neoplasms at a level of resolution that was not previously possible. A variety of molecular techniques have been applied to the study of parathyroid tumors at the DNA level. Studies of the clonal derivation of adenomas and hyperplasia suggest that these entities arise through fundamentally different mechanisms. The gene for parathyroid hormone (PTH) has been cloned and mapped within the human genome. In a small subset of parathyroid tumors, a rearrangement of the PTH gene has been described which may have contributed to their pathogenesis. A separate gene has been identified which appears to be responsible for the humoral hypercalcemia of malignancy. Chromosomal deletions which appear to be involved in the pathogenesis of multiple endocrine neoplasia type 1 have also been found in sporadic parathyroid adenomas. Characterization of tumors at the DNA level may make it possible to correlate specific genetic abnormalities with the biologic behavior of different parathyroid neoplasms and may be useful in distinguishing between adenoma, hyperplasia, and carcinoma.

Clonal derivation of a rat muscle cell strain that forms contraction-competent myotubes.

A muscle cell strain capable of forming contracting myotubes was isolated from an established rat embryo cell line. The myogenic cells, termed rat myoblast omega or RMo cells, have a diploid complement of chromosomes (n = 42). In the presence of mitogen-containing growth medium, RMo cells proliferated with a cell generation time of about 12 hours. In mitogen-depleted medium, RMo cells withdrew from the cell cycle and formed myotubes that spontaneously contracted. Differentiated RMo cells produced creatine kinase isozymes in a ratio characteristic of skeletal muscle cells. RMo cells were easy to cultivate. Cells proliferated and differentiated equally well on gelatin-coated or noncoated culture dishes, at clonal or mass culture densities, and in all basal media tested. In most experiments, growth medium consisted of horse serum-containing medium supplemented with either chicken embryo extract or FGF activity; cells proliferated equally well in medium containing unsupplemented calf serum. RMo cells differentiated if growth medium was not replenished regularly. Alternatively, differentiation was induceable by incubation in mitogen-depleted medium consisting of basal medium supplemented either with 10(-6) M insulin, 0.5% serum, or 50% conditioned growth medium. RMo cells were competently transformed with cloned exogenous genes. Because it forms functional myofibrils, the RMo cell line constitutes a useful model system for studying the cell biology and biochemistry of proteins involved in contractile apparatus assembly and muscle disease.

Functional and phenotypic characteristics of bovine natural cytotoxic cells

Bovine natural cytotoxic (NC) cell activity of peripheral blood leukocytes (PBL) was studied in a chromium release assay, using xenogeneic tumor cells (YAC-1, P815) and herpes virus-infected bovine fibroblasts. The activity pattern resembled that of murine natural killer cells, in that YAC-1 cells were readily lysed, whereas P815 cells were resistant. However, 10–16 h of incubation were usually required to give appreciable cell lysis. Low, but consistent NC-activity was expressed against virus-infected fibroblasts. Using various biophysical and biochemical cell-separation methods, it was found that the effector cell active against both the xenogeneic tumor cells and virus-infected fibroblast belonged to the mononuclear phagocyte system. The indications are that at least two subpopulations of the blood monocytes, differing perhaps in maturation or clonal derivation, express NC-activity.

The apparent inducibility of tumour marker glycoproteins in a melanoma cell line selected for growth in low levels of glucose.

It has been reported previously that malignancy in hybrid cells is associated with changes in two glycoproteins of molecular mass 90,000 and 100,000. In the current paper the characteristics of a clonal derivation of a murine melanoma cell line (PG19G-), selected for growth in low levels of glucose, are described. The cells had considerably reduced amounts of the 90 K and 100 K glycoproteins as judged by lectin affinity-labelling. When the level of glucose in the medium is raised the two glycoproteins become detectable after a few hours. This inducibility is specific for D-glucose and D-mannose at concentrations in excess of 300 mg/l and is inhibited by cytochalasin B but not by cytochalasin A. The 90 K glycoprotein differs from the 100 K in that its level varies with the glucose concentration even in the parent cell line PG19 and in other tumour lines. This is not the case for the 100 K glycoprotein, the synthesis and/or glycosylation of these two glycoproteins are, therefore, governed by separate control mechanisms. The relationship of the glucose-sensitive proteins to the glucose transporter is discussed.

Biological characteristics and viral susceptibility of a stable dog kidney cell line.

A cell line derived in 1956 from normal dog kidney is described. The cells are epithelial, contact-inhibited, and can be maintained in the same culture vessels for period of more than 2.5 yr. Karyologically, the cells are hypodiploid with a modal number of 72 as opposed to the diploid number of 78. The karyotype indicates male origin of the cells and clonal derivation of extant cultures due to the presence of two marker chromosomes in all metaphases observed. At the 159th passage the dog kidney (DK) cells did not produce tumors in athymic rats. At least 13 viruses of various types produced transmissible cytopathogenic effects in the DK cells, including all of the human influenza viruses investigated.

T-Cell Growth Factor and the Culture of Cloned Functional T Cells

Publisher Summary Detailed studies have indicated that the production of T-cell growth factor (TCGF) is augmented by macrophage-derived lymphocyte-activating factor (LAF). Since the effects of LAF are concentration dependent, and because T-cell clonal expansion is TCGF-concentration dependent, the magnitude of the resultant T-cell immune response is necessarily dictated by the available quantities of these lymphokines. that Minor alterations in the production or action of LAF and TCGF may result in major changes in T-cell proliferation that may have physiological and pathological relevance. A T-cell proliferative response to antigedlectin is actually mediated by a soluble protein TCGF. The T-cell proliferative response results from the interaction of at least three distinct cell types. As antigen or lectin is introduced to a mixed become “activated.” This results in at least three responses: release of LAF from monocytes and macrophages; under the influence of LAF, the release of TCGF by a specific T-cell subset; and binding of TCGF by separate T-cell subsets resulting in the proliferative response. Three signals are involved in the elicitation of TCGF release from TCGF producer cells: lectin or antigen cell membrane binding, Zr gene products, and LAF. The relative importance of these signals and their sequence of application to the T-cell have yet to be delineated. This chapter discusses clonal derivation and maintenance of functional T-cell lines. The latter include the clonal derivation of T-cell lines, cytolytic T-cell lines, helper T-cell lines, suppressor T-cell lines, and neoplastic T-cells and TCGF. Because LAF and TCGF both appear to interact with their target cells by means of specific membrane-binding sites, alterations of the active sites of these hormone-like factors, or their receptors, may lead to alterations in immunocyte function.

Hodgkin's disease: unfolding concepts concerning its nature, management and prognosis.

The evolution of our concepts concerning the fundamental nature and natural history of Hodgkin's disease is reviewed. Recent evidence establishes that it is indeed a malignant neoplasm, albeit a curious one, the giant cells of which display aneuploid karyotypes, often with marker chromosomes indicating their clonal derivation, and the functional and surface marker properties of cells of the mononuclear phagocyte series. The disease tends initially to spread by contiguity from one lymph node chain to others with which direct lymphatic channel communications exist. Essentially all patients, even those with limited disease, have some degree of impairment of cell-mediated immunity, apparently due to inhibition of T-lymphocyte function. Lymphangiography, computed tomography, and laparotomy with splenectomy have greatly improved the accuracy of clinical staging. Total lymphoid megavoltage radiotherapy and/or multidrug combination chemotherapy now permit the eradiction of disease of all stages, resulting in a dramatic improvement in prognosis and an increasing frequency of permanent cure of this once inevitably fatal condition.

Clonal derivation of viruses.

Clonal derivation of viruses.