Biomedical subjects
Publications and source records attributed to K A Smith.
A male patient with a de novo proximal interstitial deletion of the short arm of chromosome 1 (46XY, del(1)(p13p22.3) is described with multiple anomalies and developmental delay. This patient's clinical manifestations are compared to previously reported patients with deletions of chromosome 1p.
Treatment with recombinant rat stem cell factor (rSCF) protects mice from the lethal effects of irradiation. Mice treated with a single dose of rSCF prior to irradiation of up to 1150 rads [given as a split dose (1 rad = 0.01 Gy)] resulted in > 80% long-term survival, whereas a single injection given after the last dose of irradiation was not radioprotective. The combination of pre- and posttreatment (-20 h, -2 h, and +4 h) with rSCF resulted in 100% survival of otherwise lethally irradiated mice. Using this optimum schedule of rSCF administration, a radioprotective factor of 1.3-1.35 was achieved. The major cause of death in the control animals was massive bacteremia consisting of enteric organisms. The rSCF-treated animals had a much lower frequency of septicemia, due primarily to a rapid hematopoietic recovery of bone marrow function not evident in control animals.
Interleukin-2 (IL-2) and its receptor complex have become one of the most studied members of a growing family of protein hormones characterized by structural similarities in both ligands and their receptors. Structure-function studies of IL-2 have been complicated by the multimeric nature of its receptor. Two receptor subunits (55- and 75-kDa type I cell surface proteins) can participate to form the high affinity binding site. Although the IL-2 is apparently unique in some respects, similar subunit cooperativity has now been shown to be a common feature for other members of this receptor family. The availability of cell lines expressing the individual IL-2 receptor subunits has allowed detailed analysis of subunit binding characteristics. Results regarding the relationship of molecular recognition at each subunit to the mechanism of ligand binding at the high affinity site, however, have led to different interpretations. In this study we have employed previously prepared C-terminal IL-2 mutant proteins to examine receptor binding at all three classes using a variety of equilibrium and kinetic techniques. These results indicate that the high affinity IL-2 receptor complex includes the p55/p75 heterodimer prior to IL-2 binding and that both receptor subunits participate simultaneously in ligand capture.
The maintenance and differentiation of hematopoietic stem cells is influenced by cells making up the hematopoietic microenvironment (HM), including bone marrow-derived stromal cells. We and several other investigators have recently demonstrated the molecular basis of abnormal HM observed in the steel mutant mouse and cloned the normal cDNA products of this gene (termed SCF, KL, or MCF). In this report, we focus on the human counterpart of the mouse Steel (Sl) gene. Alternative splicing of the human SCF pre-mRNA transcript results in secreted and membrane-bound forms of the protein. To investigate the role of these two forms of human SCF, we targeted an immortalized stromal cell line derived from fetal murine homozygous (Sl/Sl) SCF-deficient embryos for gene transfer of various human cDNAs encoding SCF. We report that stable stromal cell transfectants can differentially process the two forms of human SCF protein product. We also demonstrate that both soluble SCF and membrane-bound SCF are active in increasing the number of human progenitor cells in the context of stromal cell cultures, although in a qualitatively different manner. Hence, the membrane-bound form of SCF may play an important role in the cell-cell interactions observed between stromal and hematopoietic cells both in vitro and in vivo.
Previous analyses by fluorescence in situ hybridization of structures present 20-30 cell generations after the primary events of mammalian gene amplification have shown that tens of megabases of DNA separate each copy of the selected gene in chromosomal arrays that contain up to 15 copies. Since these structures are very unstable, it is necessary to study amplified DNA as soon as possible after it has been formed to relate the structures observed to the primary mechanisms that generated them. Previously, new amplifications of the CAD gene were analyzed in colonies of 10(5) N-(phosphonoacetyl)-L-aspartate-resistant Syrian hamster BHK cells. CAD is on the p arm of chromosome B9 and the amplified genes were usually found in large extensions of B9p, with one copy in its normal position. We now report that dividing drug-resistant cells have been physically separated from static drug-sensitive cells, to allow the amplified structures to be observed only a few cell generations after they have been formed. The most informative results are that about one-third of the newly formed chromosomes carrying amplified CAD genes are dicentric and that about half of these carry two B9q arms. These observations reveal that recombination between the p telomeric regions of two B9 sister chromatids is an important primary event of amplification in this system. The resulting dicentric chromosomes can then enter bridge-breakage-fusion cycles that provide the means to increase the number of CAD genes per cell in successive generations by an asymmetric distribution at each cell division.
Stem cell factor (SCF) stimulates the growth of burst-forming unit-erythroid (BFU-E) and colony-forming unit granulocyte-macrophage (CFU-GM) by binding to a specific cell surface receptor. The receptor for SCF is encoded by the protooncogene c-kit. After immunizing mice with the human erythroleukemia cell line OCIM1, we obtained a monoclonal antibody (MoAb) that recognizes the human c-kit receptor. This MoAb, designated SR-1, blocks binding of 125I-human SCF to the c-kit receptor, and neutralizes the biologic effects of SCF in hematopoietic colony assays. With few exceptions, c-kit expression was identified on all hematopoietic and lymphoid cell lines tested by indirect immunofluorescent analysis using SR-1 and by binding studies with 125I-SCF. SR-1 recognizes a small fraction of normal bone marrow mononuclear cells, and these cells have the morphologic appearance of blasts. Colony assays show that BFU-E and CFU-GM display the c-kit receptor. SR-1 does not cross-react with murine c-kit protein, indicating that the binding epitopes of the human and murine c-kit receptors are antigenically distinct. This MoAb may be useful to characterize the spectrum of cells that display the c-kit receptor and to further define the role of SCF in hematopoiesis.
Suction-assisted lipectomy is the most commonly performed surgical aesthetic procedure in North America today. The procedure is not without significant morbidity, as death as well as serious nonfatal complications have been reported. Thromboembolic disease as a complicating factor of various types of surgical procedures and trauma has been well documented in the literature. Stasis, injury, and hypercoagulation--the limbs of Virchow's triad--contribute to predisposition for morbidity. The effects of stasis and injury are experienced with most operative procedures. We questioned whether suction-assisted lipectomy, in the appropriately selected and managed patient, would demonstrate a predisposition toward a hypercoagulable state and subsequent thromboembolic disease. In our group of ten female patients who underwent large-volume liposuction, a carefully selected assay of hematological factors demonstrated alterations consistent with a controlled response to tissue injury, but did not demonstrate a predisposition to a hypercoagulable state or subsequent increased risk of thromboembolic sequelae.
Fluorescent in situ hybridization was used to localize the adenylate deaminase 2 (AMPD2) genes and flanking sequences on the chromosomes of the Chinese hamster line GMA32 and to study the distribution of additional copies of these genetic sequences in amplified mutants selected at several early stages of the amplification process. The synteny of AMPD2 genes and MDR1 genes, located on chromosomes 1, was demonstrated; in GMA32 the existence of a rearrangement positioning the two AMPD2 genes at different distances from the telomeres was disclosed. Using this structural marker, we showed that the amplified copies distribute along only one of the chromosomes 1. Their organization in different cells of clonal mutant populations at a very early stage of amplification was extremely heterogeneous; classes of organization could be recognized however. Their quantitative distribution at this stage and in cells which went through 10 more division cycles suggests an evolution pathway common to the mutant clones under study: as a rule, tandems of few units of identical and very large size (47 Mb) appear to be the first detected product of amplification; this organization is progressively overtaken by structures with more units of reduced and irregular size, while, in a growing number of cells, clusters of much shorter units can be observed. The nature of segregative amplification mechanisms operating in these processes and the possible involvement of replicative ones are discussed.
Flavone acetic acid (FAA) causes significant regression of larger established tumors in murine systems in vivo, but is only slightly toxic in vitro. This in vivo effect is thought to be indirect, or immunological, rather than a direct cytotoxic effect on tumor cells. Using the WHFIB fibrosarcoma, which grows both in vivo and in vitro, and the murine endothelial cell line B10, we have studied the effect of FAA on the survival of tumor and endothelial cells in vitro. The times taken for 1 mg ml-1 FAA to reduce survival to 0.1 surviving fraction were 63 hr for B10 and greater than 85 hr for WHFIB in vitro. WHFIB tumors in vivo were more sensitive than tumor cells in vitro, a single dose of 150 mg kg-1 FAA inducing a tumor growth delay of 10 days at treatment size + 2 mm. As FAA is more toxic to tumor-bearing animals than to those which are non-tumor bearing the effect of tumor conditioned medium on the cytotoxicity of FAA toward B10 cells was studied; no enhanced effect was seen. As FAA is only weakly cytotoxic in vitro to endothelial cells, and even less so to tumor cells, sublethal effects of FAA on endothelial cell function in vitro were studied. The permeability of monolayers of human unbilical vein endothelial cells (HUVEC) in vitro is transiently increased by FAA. Also, procoagulant activity of HUVEC is induced by FAA and this activity is further enhanced in the presence of a factor isolated from Meth-A tumor cells.
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In this study eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) were given in a cumulative manner, every 6 weeks, starting with 10 mg, then 100 mg, 1000 mg and 10,000 mg EPA daily to mild to moderate essential hypertensive black patients. The corresponding DHA doses were 3, 33, 333 and 3333 mg. A control group was given olive oil as placebo for the entire 24 weeks. The placebo group had lower diastolic and systolic blood pressures after 24 weeks than the EPA and DHA group. No effect was seen on plasma triglycerides, cholesterol, HDL-cholesterol and gamma-glutamyltranspeptidase at any stage of the trial. In the EPA group plasma free-EPA increased significantly from 1000 mg onwards and plasma free-arachidonic acid (AA) decreased after 1000 mg EPA. No other plasma free essential fatty acid changed during the trial, although the HDL:cholesterol increased slightly but non-significantly with an increase in EPA and DHA. No significant changes in diet pattern or body mass was observed. It is therefore concluded that EPA and DHA supplementation had no beneficial effects in mild to moderate essential hypertensive black patients except for a lowering of plasma AA.
In the past year there have been significant advances in understanding the role of interleukin-2. Its role in the activation of T cells by antigen-presenting cells, the structure-activity relationships between interleukin-2 and its receptor and the subsequent signaling have all become clearer. The creation of mice with a specific defect in the interleukin-2 gene has given us a clearer idea of its role in vivo. Recent studies also suggest that interleukin-2 may finally find a role in immunotherapy.
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21 patients with advanced malignant melanoma were treated with interferon alpha 2a at 9MU daily with vindesine every 21 days. No patient had received previous chemotherapy. The overall response rate was 24% with a median survival time of 33 months in 18 patients. The four complete remissions were maintained for 20, 18, 15 and 11 months, while the single partial remission continues at 18 months after the start of treatment. Side-effects were generally mild or moderate and did not lead to cessation of therapy. This combination provides an active outpatient regimen for advanced melanoma and produces durable remissions.
Interleukin 2 (IL-2), a T lymphocyte product released upon antigen stimulation, has been used for cancer therapy in high doses for more than five years. More recently, its potential as a stimulant of cell-mediated immunity in infectious diseases, particularly those caused by intracellular microbes, has become appreciated. Drawing on the extensive information available as to the structure, cellular and molecular effects of IL-2, this review focuses on its use in patients with lepromatous leprosy and AIDS in low, physiologic doses. The data indicate that IL-2 is effective in stimulating cell-mediated immunity without systemic toxicity.
These studies demonstrated that the inbred BALB/c mouse strain can be optimized for the assessment of vaccinia virus virulence, growth, and spread from the site of inoculation and immune protection from a lethal vaccinia virus challenge. The studies established that manipulation of the vaccinia virus genome generated mutants exhibiting a wide range of attenuated phenotypes. The nine NYCBH vaccinia virus mutants had intracranial 50% lethal doses that ranged from 2 to greater than 7 log10 units. The decreased neurovirulence was due to decreased replication in brain tissue. Three mutants had a decreased ability to disseminate to the lungs, brains, livers, and spleens of mice after intranasal infection. One mutant had a decreased transmission from mice infected by tail scarification to naive cage mates. Although the mutants, with one exception, grew to wild-type titers in cell culture, they showed a growth potential on the scarified skin of mice that was dramatically different from that of the wild-type virus. Consequently, all of the mutants had significantly compromised immunogenicities at low virus immunization doses compared with that of the wild-type virus. Conversely, at high immunization doses most mutants could induce an immune response similar to that of the wild-type virus. Three Wyeth vaccine strain mutants were also studied. Whereas the thymidine kinase, ribonucleotide reductase, and hemagglutinin mutants had a reduced virulence (50% lethal dose), only the thymidine kinase mutant retained its immunogenicity.
Percutaneous nicotine administration induces predominant sudorific and rubiform responses in the skin which may be accompanied by subtle piloerection, hyperalgesia and pruritus (although these signs are not overtly manifest). These dermal responses are complex and mechanisms have been proposed for the direct nicotine-stimulation of sweat glands, piloerection and vasoconstriction. These reactions are accompanied by secondary activation and release of vasodilator peptides which produce a predominating vasodilator tone following topical administration, this response masking the direct axon reflex-mediated vasoconstriction.
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