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Biomedical subjects

K A Smith

Publications and source records attributed to K A Smith.

At least 163 records · Page 9Linked to original sources
PubMed1989

The effect of chronological and theoretical birth order on sexual attitudes and behaviors.

A survey designed to evaluate the impact that both chronological and theoretical birth order may have on sexual attitudes and behaviors was completed by 441 respondents (221 male, 220 female). The attitudes and behaviors investigated were aggressive sexual behavior, social relationship attitudes, emotional relationship attitudes, total heterosexual behavior, and total orgasmic behavior. The data revealed no difference between male and female in terms of total sexual behavior. However, males had significantly higher sexual aggression, social attitudes, emotional attitudes, and total orgasm scores. These results suggest that while there may be equity between the sexes in terms of heterosexual behavior, there are still significant differences between the sexes in terms of specific behaviors and attitudes. No differences were found on any of the scales based on either chronological or theoretical birth order. These results suggest that sexual behavior and attitudes may be influenced to a greater degree by biological and cohort factors than by sibling position.

Adolescent↗
PubMed1989

Interleukin 2: prototype for a new generation of immunoactive pharmaceuticals.

Molecular biological techniques have revealed the interleukin-2 receptor to be a dimer composed of one alpha-subunit and one beta-subunit which interact noncovalently in a cooperative manner. Site-directed mutagenesis, in conjunction with structural analysis, is beginning to clarify the relationship between structural components of the receptor and their function, and Thomas Ciardelli and Kendall Smith explain why this is bringing drug developers closer to the design of IL-2 agonists and antagonists.

Adjuvants, Immunologic↗
PubMed1989

Vascular markers for murine tumours.

Damage to the vasculature may represent an important component of several forms of cancer therapy. Methods for studying the structure and function of the vasculature of experimental mouse tumours are required. In this paper several relatively simple methods are described for the histological examination of the vascular structure of murine tumours. The methods have been applied to cryostat sections of two sarcomas and two carcinomas. Immunoperoxidase staining with polyclonal antibodies to laminin highlights the vascular basement membrane of sarcomas, but has limited use with carcinomas, while the monoclonal antibody MECA-20 is a good marker for the endothelial cells of all vessels in all four tumours tested. The presence of endothelial cells in normal tissues can also be demonstrated by the use of enzyme-histochemical techniques for alkaline phosphatase, 5'-nucleotidase and nucleotide diphosphatase (ADPase), but only one of these methods (ADPase) works consistently in tumours. The relative merits of these methods are discussed and in all cases related to the staining pattern obtained with normal mouse tissues. The significance of these methods for vascular targeting is also discussed.

Alkaline Phosphatase↗
PubMed1989

The fractionated response of mouse stroma after X-rays and neutrons: influence of early vs late expression of damage.

The sparing effect of dividing a radiation treatment into many small fractions is different for acutely responding tissues and those which have delayed expression of injury. We have used the Tumour Bed Effect assay (TBE) to investigate the influence of the time of damage expression on the response of the normally quiescent subcutaneous stroma. Implanted tumour cells provide an angiogenic stimulus which forces the vasculature to proliferate. The subcutaneous stroma of the mouse dorsum was irradiated with one to 32 equal fractions of X-rays, followed by a top-up dose of neutrons. CaNT cells were implanted into the treated site either within 3 days or not until 6 months after irradiation, to stimulate the expression of latent injury. The dose-response curves obtained for tumour growth in irradiated sites were much steeper at 1 to 3 days than at 6 months, suggesting some sort of repair of damage during this period. There was no suggestion that repair occurred preferentially after low doses per fraction and the alpha/beta ratio remained unchanged when the expression of stromal injury was delayed. The time of damage expression therefore seems unlikely to explain the difference in the alpha/beta ratios measured for early and late responding tissues. Rather, it seems to be determined by the proliferative status of the tissue at the time of irradiation.

Animals↗
PubMed1989

Flavone acetic acid induces a coagulopathy in mice.

The effects of flavone acetic acid (FAA) on the coagulation properties of plasma from tumour-bearing and non-tumour-bearing mice have been investigated. The study was carried out primarily on CBA mice and the CaNT tumour, although substantiating data are included for two other tumours grown in the WH strain. FAA was injected at a range of single doses up to a maximum of 300 mg kg-1, and clotting properties of the plasma were measured in vitro at various times after FAA administration. Platelet numbers and the concentration of fibrin degradation products (FDP) in the plasma were also determined. Following a dose of 300 mg kg-1, the clotting times were significantly reduced at 15-30 min in both tumour-bearing and non-tumour-bearing mice of both strains. Detailed studies on coagulation in the CBA strain (+/- CaNT tumour) indicate that in tumour-bearing animals the initial decrease in clotting time is followed 4-6 h later by an increase in clotting time, thrombin time and FDP levels. Platelet counts of tumour-bearing mice also decreased significantly over this period. Similar experiments in non-tumour-bearing mice did not show these late effects. All the data from the coagulation tests on mice with CaNT tumours are consistent with the hypothesis that intravascular coagulation occurs following treatment with FAA, and that vascular occlusion in tumours, as a results of FAA-induced coagulopathy, may contribute to tumour regression.

Animals↗
PubMed1989

Identification and Partial Characterization of the Denaturation Transition of the Light Harvesting Complex II of Spinach Chloroplast Membranes.

Differential scanning calorimetry was employed to investigate the structure of spinach (Spinacia oleracea) chloroplast membranes. In a low ionic strength Hepes-buffered medium, major calorimetric transitions were resolved at 42.5 degrees C. (A), 60.6 degrees C (B), 64.9 degrees C (C(1)), 69.6 degrees C (C(2)), 75.8 degrees C (D), 84.3 degrees C (E), and 88.9 degrees C (F). A lipid melting transition was also commonly seen at 17 degrees C in scans starting at lower temperatures. The D transition was demonstrated by four independent methods to derive from denaturation of the light harvesting complex associated with photosystem II (LHC-II). Evidence for this conclusion was as follows: (a) the endotherm of the isolated LHC-II (74.0 degrees C) was very similar to that of D (75.8 degrees C); (b) the denaturation temperature of the 27 kilodalton LHC-II polypeptide determined in intact chloroplast membranes by thermal gel analysis was identical to the temperature of the D transition at pH 7.6 and after destabilization by shifting the pH to 6.6 or by addition of Mg(2+); (c) analysis of the stability of the LHC-II complex by electrophoresis in native gels demonstrated that the complex dissociates during the D transition, both at pH 7.6 and 6.6; and (d) the 77 Kelvin fluorescence maximum of LHC-II in chloroplasts was seen to shift to lower wavelengths (indicating gross denaturation of LHC-II), at the temperature of the D transition when examined at either of the above pHs. With this identification, five of the eight major endotherms of the chloroplast membrane have now been assigned.

Journal Article↗
PubMed1989

Identification and Partial Characterization of the Denaturation Transition of the Photosystem II Reaction Center of Spinach Chloroplast Membranes.

Sensitive differential scanning calorimetry was employed to investigate thylakoid membrane structure. Calorimetric scans of chloroplast membranes suspended in a low ionic strength Hepesbuffered medium revealed endothermic transitions centered at the following temperatures ( degrees C): A (42.5), B (60.6), C(1) (64.9), C(2) (69.6), D (75.8), E (84.3), and F (88.9). The B transition was demonstrated by several different methods to originate from denaturation of the photosystem II reaction center complex. Evidence for this conclusion is as follows: (a) the isolated reaction center complex denatures near the temperature of the B transition; (b) inorganic phosphate destablizes the isolated reaction center complex and the B endotherm to a similar extent; (c) heat inactivation of the photosystem II-mediated 1,5-diphenylcarbazide --> dichloroindophenol photoreaction occurs at the temperature of the B transition and is influenced in a manner similar to B by the presence of phosphate; (d) thermal gel analysis indicates that the 43 and 47 kilodalton polypeptides of the photosystem reaction center complex denature at the temperature of the B transition, both in the presence and absence of phosphate; (e) low temperature (77 Kelvin) fluorescence reveals that a change in photosystem II emission at 695 nanometers occurs during the B transition; and (f) ioxynil, a specific inhibitor of photosystem II, selectively stabilizes the B endotherm. With the identification of the B transition established, the origins of six of the eight major transitions of the chloroplast membrane have now been determined.

Journal Article↗
PubMed1989

Cell-associated pullulanase from Bacteroides thetaiotaomicron: cloning, characterization, and insertional mutagenesis to determine role in pullulan utilization.

We have cloned a pullulanase gene from Bacteroides thetaiotaomicron. The pullulanase expressed from this clone in Escherichia coli was cell associated and soluble and had a molecular mass of 72 kilodaltons by gel filtration. Maxicell analysis of proteins coded by the cloned insert showed that a 71.6- to 73.2-kilodalton doublet was associated with pullulanase activity. Thus, the pullulanase is probably a monomer. The cloned pullulanase produced maltotriose as an end product of pullulan digestion. In B. thetaiotaomicron the pullulanase activity was cell associated. Approximately 80% of the activity was soluble, and 16 to 18% was membrane associated. The molecular mass of the soluble pullulanase was 77 kilodaltons by gel filtration. To determine whether the cloned pullulanase gene was essential for pullulan utilization, we used directed insertional mutagenesis to inactivate the B. thetaiotaomicron pullulanase gene. The pullulanase specific activity of the mutant was approximately 45% of that of wild-type B. thetaiotaomicron. However, the pullulanase-negative insertional mutant 95-1 was still able to grow on pullulan at a rate similar to that of wild-type B. thetaiotaomicron. Thus, there must be a second pullulanase in B. thetaiotaomicron.

Bacteroides↗
PubMed1989

Low-density lipoprotein transport in blood vessel walls of squirrel monkeys.

Transmural accumulations of low-density lipoprotein (LDL) were examined in the blood vessel walls of four squirrel monkeys. Vascular wall concentrations of LDL were measured using quantitative autoradiography after 125I-labeled LDL circulation for 30 min. Profiles of relative tissue concentration from different sections in the same region were similar to each other, and there was little animal-to-animal variation. Concentrations were highest near the luminal endothelium, lower near the medial-adventitial border, and lowest within the media. Profiles from different regions fell into three groups: 1) aortic samples had steep intimal concentration gradients and near-zero media concentrations; 2) the iliac, femoral, popliteal, and common carotid arteries had higher intimal concentrations than group 1 but had similar concentrations deep within the media; and 3) the cerebral and coronary arteries, inferior vena cava, and pulmonary artery had intimal concentrations that were similar to group 2, but the concentrations deep within the media were greater than either groups 1 or 2. Arterial bifurcation profiles from the inner wall and the outer walls were similar to each other and to profiles from the upstream and downstream areas. Out of 280 total sites examined, 15 examples of profiles with substantially increased concentrations near the luminal endothelium were found scattered throughout the cardiovascular system, demonstrating that there are focal regions throughout the cardiovascular system which have greatly increased 125I-LDL transendothelial permeability.

Animals↗
PubMed1989

Application of the electrochemistry of cytochrome c to the measurement of superoxide radical production.

An electrochemical sensor for the measurement of the superoxide anion produced as a result of the respiratory burst of neutrophils is described. The reduction of cytochrome c by the superoxide ion was monitored using a surface-modified gold electrode. The current produced was shown to be superoxide-specific and proportional to the number of neutrophils in both purified neutrophil preparations and plasma.

Cytochrome c Group↗
PubMed1988

Induction of antigen-specific parasiticidal cytotoxic T cell splenocytes by a major membrane protein (P30) of Toxoplasma gondii.

Infection with Toxoplasma gondii has become a major cause of morbidity in patients with AIDS. To investigate the mechanisms responsible for immune responses to toxoplasma Ag we used a highly purified membrane protein (P30) of T. gondii to stimulate an in vitro Ag-specific cytotoxic T cell response. P30 immune mouse splenocytes reduced extracellular T. gondii plaque-forming units by more than 50% when incubated at an E/T ratio of 10:1 or greater. By using a [3H]uracil radioisotope release assay, the effect of the immune splenocytes was determined to be a direct parasite lytic mechanism. The immune splenocytes were P30 Ag specific and of the Thy 1.2, Lyt2,3+ (CD4-, CD8+) phenotype, specific for mouse cytotoxic T cells. Opsonization of the parasites with monoclonal P30-reactive mAb did not enhance parasiticidal activity. Culture supernatants obtained during the 2-h cytotoxic assay were not parasiticidal, and anti-asialo-GM1 antibody plus C did not destroy the parasiticidal activity of the P30 responder cells. Accordingly, we have identified an Ag-specific subset of CD4-, CD8+, P30 responder T cells that are directly parasiticidal to extracellular T. gondii, and that exhibit cytotoxicity independent of antibody opsonization, lymphokine secretion, NK cell activity, and, apparently, MHC involvement as well.

Animals↗
PubMed1988

Interleukin-2: inception, impact, and implications.

Interleukin-2 (IL-2), the first of a series of lymphocytotrophic hormones to be recognized and completely characterized, is pivotal for the generation and regulation of the immune response. A T lymphocyte product, IL-2 also stimulates T cells to undergo cell cycle progression via a finite number of interactions with its specific membrane receptors. Because T cell clonal proliferation after antigen challenge is obligatory for immune responsiveness and immune memory, the IL-2-T cell system has opened the way to a molecular understanding of phenomena that are fundamental to biology, immunology, and medicine.

Cell Cycle↗
PubMed1988

A design approach to the structural analysis of interleukin-2.

A semi-synthetic protein design approach has been employed for the structural investigation of a putative helical region at the C-terminus of Interleukin-2. With crystallographic or NMR derived conformational data as yet unavailable, we have relied only on primary sequence information and computer-assisted modelling to direct the analysis. By employing both chemical peptide synthesis and recombinant DNA methods, the C-terminus of IL-2 was modified according to a strategy designed to stabilize helical secondary structure. A semi-synthetic protein incorporating 12 simultaneous amino acid replacements was constructed, which possessed potentiated biological activity and displayed a far UV circular dichroism spectrum comparable to a hybrid protein with the authentic sequence. By comparison, another hybrid protein containing a C-terminal region designed to contain helix breaking residues was totally devoid of bioactivity. These findings provide evidence that the modelling method correctly identified a helix necessary for the formation of a bioactive tertiary fold. Moreover, by employing semi-synthesis it was possible to circumvent the difficulties associated with the preparation, purification and analysis of multiple recombinant proteins, and also to avoid the unreliability of total chemical synthesis for proteins greater than 100 residues.

Amino Acid Sequence↗
PubMed1988

Validation of the fluorescent dye Hoechst 33342 as a vascular space marker in tumours.

The DNA-binding fluorescent dye Hoechst 33342 (H33342) has been used in a series of investigations of the vascular parameters of two murine tumours. This dye has been shown, to have a short half-life in the circulation (T1/2 less than 2 min), but is stably bound for at least 2 h after it enters cells. It can be used in morphometric studies on frozen sections to determine the effective vascular volume, the capillary fraction and the size distribution of blood vessels in each tumour. These latter two parameters cannot be deduced from the less labour intensive techniques using radioactive isotopes. The effective vascular volume perfused in 1 min by H33342 was compared with the volume perfused in 30 min with 51Cr labelled erythrocytes. Similar volumes were estimated with the two techniques in a murine carcinoma and in a sarcoma. Both techniques showed that the vascular volume decreased in larger tumours. The H33342 analysis of vessel size showed the decrease in capillary vessels in the carcinomas was even greater, falling from 70% in small tumours to 20% in larger tumours. The deteriorating vascular network in larger tumours is associated with an increasing fraction of necrotic tissue. Experiments in which the isotopes and dye were co-injected suggest that at 40 mgkg-1 the dye may rapidly lead to a partial shutdown of the tumour vascular bed. This is less marked with 20 mg kg-1. In spite of this effect there is in general a close correlation between the volumes perfused by labelled red blood cells and the fluorescent dye.

Adenocarcinoma↗
PubMed1988

cAMP antagonizes interleukin 2-promoted T-cell cycle progression at a discrete point in early G1.

T lymphocytes are stimulated to proliferate in an autocrine/paracrine manner by the lymphokine interleukin 2 (IL-2). In seeking further insight into the mechanisms by which IL-2 induces progression of T cells through the G1 phase of the cell cycle, studies were performed with agents that increase cellular adenosine 3',5'-cyclic monophosphate (cAMP), a well-known inhibitor of lymphocyte growth. The addition of dibutyryl-cAMP, cholera toxin, forskolin, or 3-isobutyl-1-methylxanthine to an IL-2-dependent murine T-cell line evoked a dose-related suppression of S-phase transition without affecting cellular viability. Moreover, elevation of cAMP levels led to an accumulation of uniformly small cells, suggesting an arrest in early G1. Consistent with these findings, dibutyryl-cAMP inhibited the incorporation of both [3H]-uridine and [3H]thymidine by IL-2-stimulated, synchronized normal human T cells. Furthermore, maximal inhibition occurred during early G1, as indicated by experiments where the addition of dibutyryl-cAMP was delayed with respect to IL-2 stimulation. Quantitative flow cytometric analysis of RNA and DNA content of IL-2-stimulated cells affirmed that increased cAMP inhibits RNA accumulation and S-phase transition. In addition, exposure of IL-2-dependent, asynchronously proliferating normal human T cells to dibutyryl-cAMP resulted in uniform growth arrest in early G1, the point at which cycling T cells accumulate when they are deprived of IL-2. These results indicate that increased cAMP inhibits G1 progression stimulated by IL-2 and provide a rationale for the use of cAMP analogues as pharmacologic probes for the dissection of molecular events occurring during IL-2 signaling and T-cell G1 transit. They also suggest the possibility of therapeutic immunosuppression by a combination of agents that act at different stages of the T-cell cycle.

Bucladesine↗