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Biomedical subjects

A King

Publications and source records attributed to A King.

At least 181 records · Page 10Linked to original sources
PubMed1997

Structure of the complex of an Fab fragment of a neutralizing antibody with foot-and-mouth disease virus: positioning of a highly mobile antigenic loop.

Data from cryo-electron microscopy and X-ray crystallography have been combined to study the interactions of foot-and-mouth disease virus serotype C (FMDV-C) with a strongly neutralizing monoclonal antibody (mAb) SD6. The mAb SD6 binds to the long flexible GH-loop of viral protein 1 (VP1) which also binds to an integrin receptor. The structure of the virus-Fab complex was determined to 30 A resolution using cryo-electron microscopy and image analysis. The known structure of FMDV-C, and of the SD6 Fab co-crystallized with a synthetic peptide corresponding to the GH-loop of VP1, were fitted to the cryo-electron microscope density map. The SD6 Fab is seen to project almost radially from the viral surface in an orientation which is only compatible with monovalent binding of the mAb. Even taking into account the mAb hinge and elbow flexibility, it is not possible to model bivalent binding without severely distorting the Fabs. The bound GH-loop is essentially in what has previously been termed the 'up' position in the best fit Fab orientation. The SD6 Fab interacts almost exclusively with the GH-loop of VP1, making very few other contacts with the viral capsid. The position and orientation of the SD6 Fab bound to FMDV-C is in accord with previous immunogenic data.

Antibodies, Monoclonal↗
PubMed1997

Transgenic mouse model for studying the transcriptional activity of the p53 protein: age- and tissue-dependent changes in radiation-induced activation during embryogenesis.

The p53 tumor suppressor protein is a sequence-specific transcriptional activator of target genes. Exposure of cells to DNA damage results in accumulation of biochemically active p53, with consequent activation of p53-responsive promoters. In order to study how the transcriptional activity of the p53 protein is regulated in vivo, a transgenic mouse strain was generated. These mice harbor the p53-dependent promoter of the mdm2 gene, fused to a lacZ reporter gene. Induction of lacZ activity by DNA damage (ionizing radiation) was monitored in embryos of different p53 genotypes. The transgenic promoter was substantially activated in vivo following irradiation; activation required functional p53. The activation pattern became more restricted with increasing embryo age, as well as with the state of differentiation of a given tissue. Generally, maximal p53 activation occurred in rapidly proliferating, relatively less differentiated cells. A striking extent of haploinsufficiency was revealed-induction of promoter activity was far less efficient in mice carrying only one wild-type p53 allele. This suggests that normal levels of cellular p53 are limiting, and any further reduction already compromises the p53 response significantly. Thus, the activation potential of p53 is tightly controlled in vivo, both spatially and temporally, and an important element in this control is the presence of limiting basal levels of activatable p53.

Alleles↗
PubMed1997

Human histocompatibility leukocyte antigen (HLA)-G molecules inhibit NKAT3 expressing natural killer cells.

The crucial immunological function of the classical human major histocompatibility complex (MHC) class I molecules, human histocompatibility leukocyte antigen (HLA)-A, -B, and -C, is the presentation of peptides to T cells. A secondary function is the inhibition of natural killer (NK) cells, mediated by binding of class I molecules to NK receptors. In contrast, the function of the nonclassical human MHC class I molecules, HLA-E, -F, and -G, is still a mystery. The specific expression of HLA-G in placental trophoblast suggests an important role for this molecule in the immunological interaction between mother and child. The fetus, semiallograft by its genotype, escapes maternal allorecognition by downregulation of HLA-A and HLA-B molecules at this interface. It has been suggested that the maternal NK recognition of this downregulation is balanced by the expression of HLA-G, thus preventing damage to the placenta. Here, we describe the partial inhibition of NK lysis of the MHC class I negative cell line LCL721.221 upon HLA-G transfection. We present three NK lines that are inhibited via the interaction of their NKAT3 receptor with HLA-G and with HLA-Bw4 molecules. Inhibition can be blocked by the anti-NKAT3 antibody 5.133. In conclusion, NK inhibition by HLA-G via NKAT3 may contribute to the survival of the fetal semiallograft in the mother during pregnancy.

Cells, Cultured↗
PubMed1997

Expression of killer cell inhibitory receptors on human uterine natural killer cells.

The establishment of the human placenta in early pregnancy is characterized by the presence of large numbers of natural killer (NK) cells within the maternal decidua in close proximity to the fetally-derived invading extravillous trophoblast which expresses at least two HLA class I molecules, HLA-G and HLA-C. These NK cells have an unusual phenotype, CD56(bright) CD16, distinguishing them from adult peripheral blood NK cells. They may control key events in trophoblast migration and therefore placentation. Human NK cells in peripheral blood express receptors for polymorphic HLA class I molecules. This family of receptors, known as killer cell inhibitory receptors (KIR), are expressed on overlapping subsets of NK cells to give an NK cell repertoire which differs between individuals. Using a panel of monoclonal antibodies to several members of the KIR family and analysis by flow cytometry, we have found that KIR are expressed by decidual NK cells. There is variation in both the percentage of cells expressing a particular receptor and the density of receptor expression between decidual NK cells from different individuals. Comparison of NK cells from decidua and peripheral blood of the same individual showed that NK cells from these two different locations express different repertoires of KIR. Receptors are present in individuals who do not possess the relevant class I ligand, raising the possibility that these NK receptors may be involved in recognition of the allogeneic fetus by the mother at the implantation site.

Adult↗
PubMed1997

Cytokine production and cytokine receptor expression by cells of the human first trimester placental-uterine interface.

There is currently a great deal of interest in the role that cytokines may play in placental development. A variety of cytokines have been demonstrated at the placental-uterine interface, but the exact cellular sources of production have not yet been identified due to the complex tissue topography of the implantation site. In the present study, we have used the enzyme-linked immunosorbent assay (ELISA) to investigate cytokine production in vitro by cells isolated with a high degree of purity from the placenta and decidua. The cytokines assayed were EGF, interleukin 1 beta (IL-1 beta), IL-2, IL-3, interferon alpha (IFN-alpha), IFN-gamma, tumour necrosis factor alpha (TNF-alpha) and transforming growth factor beta 1 (TGF-beta 1). A necessary starting point in the elucidation of cytokine function is to define those cell populations which express relevant cytokine receptors as this would identify the potential cell targets. For this reason, cytokine receptor expression by placental and uterine cells using immunohistology and flow cytometry has been investigated, those being: IGF-1r, PDGF-r alpha/beta, IL-1rII, IL-6r, IL-7r, IFN-gamma r, TNF-rp80 and endoglin. Taken in conjunction with previously published data on other cytokine receptors, a comprehensive picture can begin to be constructed of the putative cytokine-responding cells present in the human placental-uterine interface.

Cells, Cultured↗
PubMed1997

Transfection of the inner cell mass and lack of a unique DNA sequence affecting the uptake of exogenous DNA by sperm as shown by dideoxy sequencing analogues.

PURPOSE: The purpose of this study was to determine whether exogenous DNA internalized into blastocysts after transference from DNA-carrier sperm are localized at the inner cell mass or trophoblast cells and to identify differences in uptake of exogenous DNA fragments by sperm due to unique DNA sequences. METHODS: Mouse blastocysts at the hatching stage were exposed to migrating human sperm cells carrying exogenous DNA fragments synthesized from the E6-E7 conserved gene regions of human papillomavirus (HPV) types 16 and 18. After an interaction period of 2 hr, the transfected blastocysts were washed several times to remove extraneous sperm and the blastocysts were dissected into groups of cells derived from the inner cell mass and trophoblasts. The cells were analyzed by polymerase chain reaction (PCR) for the presence of HPV DNA fragments. In the second part of the experiment, thawed donor (N = 10) sperm cells were pooled, washed, and divided into two fractions. The first (control) fraction was added with formalin and further divided and added with a 35S-radiolabeled G, A, T, or C sequencing mixture. The second fraction was similarly treated but the formalin step was omitted from the treatment. After an hour of incubation at 37 degrees C, the sperm specimens were washed several times by centrifugation and DNA extracted by the GeneReleaser method. The extracted DNA were processed on sequence gels, and the autoradiographs analyzed. RESULTS: Mouse blastocysts transfected by carrier sperm with DNA from HPV types 16 and 18 showed localization of the HPV DNA to both the inner cell mass and trophoblast cells. Negative controls consisting of untreated human sperm and untreated mouse blastocysts did not reveal any evidence of HPV DNA. The positive sperm control generated expected DNA fragments from HPV types 16 and 18. In the second experiment, the intensities of the DNA fragments in the G, A, T, and C columns from low to high molecular weights were not different from the positive control bands. Band intensities of the four sequencing columns were similar. Formalin pretreatment of the sperm inhibited uptake of the DNA fragments from the smallest to the largest DNA molecules. CONCLUSIONS: Exogenous DNA taken into blastocysts are localized to both the inner cell mass and trophoblast cells. Only live sperm exhibited the capacity to carry various sizes of exogenous DNA, suggesting the involvement of active cell membrane mechanism in the transference process. The results showed that DNA fragments terminating in any of the four nucleotides were equally taken up by the sperm cell. Fragments of DNA produced by the sequencing reaction failed to identify a unique DNA sequence that would facilitate or inhibit the sperm from taking up exogenous DNA.

Animals↗
PubMed1997

Silane-coated silica particle colloid processing of human sperm.

PURPOSE: The purpose of this study was to determine differences in the quality of human sperm processed through different lots of silane-coated silica particle colloid solutions. The objectives were to compare (a) sperm kinematic parameters, (b) the sperm acrosome status, (c) the membrane integrity of the head and tail regions, (d) the DNA normality, and (e) the heat-inducible hyperactivation motility after processing sperm through either a Silane-coated silica particle colloid solution, a Percoll solution, or a simple centrifuge sperm wash (control). METHODS: Sperm cells were derived from pooled cryopreserved-thawed specimens of several donors (n = 10). The pooled sperm were divided and processed through either the centrifuge wash, the 90:47% two-layer Percoll, or one of three lots of silane-coated silica particle colloidal solutions from three vendors. Aliquots of sperm cells were analyzed using the Hamilton-Thorn HTM-C motility analyzer for differences in kinematics and hyperactivation. Sperm were also analyzed for membrane integrity at both head and tail regions, normal morphology, acrosome status, and viability. Sperm undergoing apoptosis were determined using the acridine orange stain. Processed sperm were also incubated at 40 degrees C for 4 hr and the quality of the sperm was assessed using the heat-induced hyperactivation and motility parameter. RESULTS: The data showed that after sperm processing, the number of sperm recovered was higher for the three lots of colloids (silane-coated silica particle colloid solutions) compared with Percoll processing. Total sperm motility was higher in the colloidal washes compared with the control. There were no differences in motility between Percoll- and colloid-processed sperm. In contrast, the percentages of sperm exhibiting progressive motility or hyperactivation varied among the different lots of colloid solutions. The Percoll wash solution yielded the highest percentage of sperm with intact tail membranes, whereas some lots of colloid solutions disrupted sperm head membranes. The percentages of sperm undergoing apoptosis varied for the different lots of colloid solutions. There was a marked increase in hyperactivation associated with one colloid solution after heat induction. CONCLUSIONS: The results demonstrated variability in the different lots of silane-coated silica particle colloid solutions for processing sperm. Each lot of colloid solution excelled at improving different sperm parameters. The silane-coated silica particle colloid solutions were shown to be effective in recovering motile sperm compared with Percoll but the types of motility and sperm quality varied for the different lots of colloid solutions. Due to the variability in lots of silane-coated silica colloid solutions, reported studies based on only one lot or one source of colloid solution may be difficult to interpret. Furthermore, it may be advantageous to select the best lot of silane-coated silica particle colloid solution to produce the highest number of sperm exhibiting the ideal parameters for use in assisted reproduction technologies.

Acridine Orange↗
PubMed1997

Perioperative nurses' guide to the library.

Nurses have a professional responsibility, both morally and ethically, to provide the most current and informed patient care possible. Nurses should maintain competence in nursing, exercise informed judgment, participate in activities that contribute to the ongoing development of their profession's body of knowledge, and participate in their profession's efforts to implement and improve standards of nursing care. Nurses should stay current with new information and share this information with coworkers to help disseminate new knowledge. Such practices help advance both the body of knowledge for perioperative nursing and the profession of nursing. Nurses are encouraged to make at least two visits each year to the library. During this time, they can take the opportunity to review current periodicals, recent publications, do a CINAHL search, and browse the Internet. Nurses need to be familiar with library resources, so go to the library. Share your findings with colleagues and students. Acquire the information-seeking skills that will best serve you and your patients in the twenty-first century.

Catalogs, Library↗
PubMed1997

Antibiotics: effect on cryopreserved-thawed human sperm motility in vitro.

OBJECTIVE: To analyze the motility and fertilizing capacity of sperm treated with different antibiotics. DESIGN: Prospective comparative study. SETTING: Clinical and academic research environment. PATIENT(S): Pooled cryopreserved donor sperm (n = 14). INTERVENTION(S): Sperm were washed with Percoll and resuspended in HEPES-buffered human tubal fluid medium containing either amoxicillin, ofloxacin, ciprofloxacin hydrochloride, nitrofurantoin monohydrate, doxycycline hyclate, cefuroxime axetil, or control medium. MAIN OUTCOME MEASURE(S): Sperm kinematic and fertilizing parameters. RESULT(S): Sperm hyperactivation was decreased in physiologic concentrations of ciprofloxacin hydrochloride and doxycycline hyclate over the course of 48 hours. At pharmacologic concentrations, ciprofloxacin hydrochloride, cefuroxime axetil, and nitrofurantoin monohydrate adversely affected motility with decreased rapid progression. Cessation of motility occurred in cefuroxime axetil and nitrofurantoin monohydrate. Sperm hyperactivation was also absent. Cefuroxime axetil decreased the percentage of intact acrosomes. In contrast, physiologic doses of ciprofloxacin hydrochloride or ofloxacin enhanced sperm fertilizing capacity. CONCLUSION(S): Ciprofloxacin affected hyperactivation by altering membrane properties, whereas doxycycline inhibited the capacitation process. Cessation of motility in cefuroxime axetil was linked to disrupted sperm head membranes. Sperm motility and fertilizing capacity were decreased in nitrofurantoin because of decreased metabolism. The positive effect of ofloxacin on fertilizing capacity did not involve changes in acrosome.

Analysis of Variance↗
PubMed1997

Human uterine NK cells have a similar repertoire of killer inhibitory and activatory receptors to those found in blood, as demonstrated by RT-PCR and sequencing.

The expression of natural killer (NK) cell receptors specific for HLA class I molecules has been studied in CD56bright, CD3- NK cells isolated from the pregnant uterine mucosa, the decidua. RT-PCR was performed on cDNA from uterine NK cells with primers designed to amplify members of the killer inhibitory receptor (KIR)/killer activatory receptor (KAR) gene family. Sequencing of the PCR products revealed that uterine NK cells express KIR/KAR which have two or three extracellular immunoglobulin superfamily (Ig-SF) domains. NK receptors for both groups of HLA-C alleles were found. KIR, characterised by a long cytoplasmic tail containing the immune receptor tyrosine-based inhibitory motif (ITIM), and KAR, characterised by a short cytoplasmic domain with a transmembrane region containing a charged lysine, were both identified. Different individuals appear to have a distinct but overlapping repertoire of KIR/KAR. No new members of this NK receptor gene family were identified in the uterine CD56bright NK cells. Similar findings were obtained from non-pregnant endometrial tissues representative of different stages of the menstrual cycle. Immunohistology confirmed that the KIR protein products were expressed by decidual NK cells. These results reveal that NK receptors for trophoblast HLA class I molecules are present in maternal uterine NK cells. Fetal trophoblast cells infiltrating the decidua express HLA-G and HLA-C gene products. This suggests that maternal recognition of the fetus may be mediated by an NK allorecognition system.

CD56 Antigen↗
PubMed1997

Immunology of human placental implantation: clinical implications of our current understanding.

In the early 1950s, Medawar proposed the concept of "the fetus as an allograft". Since then, the immunological relationship between the mammalian fetus and its mother during pregnancy has been considered to be similar to that between a transplanted allograft and its recipient. Because of this analogy, it has been assumed that implantation of the fetal placenta in the uterus would similarly be controlled by a maternal immune response mediated by T cells recognizing paternally derived alloantigens expressed by the placenta. Surprisingly, recent evidence suggests that implantation might predominantly involve a novel allogeneic recognition system based on natural killer cells rather than T cells. The cellular and molecular basis of this local immune interaction between the fetal placenta and maternal uterus is now the focus of intense research interest. Because aberrant implantation can cause a variety of clinical problems including miscarriage, intrauterine growth retardation and pre-eclampsia, an understanding of the immunological mechanism by which this process is controlled could lead to the development of regimes to treat these important obstetric conditions.

Female↗
PubMed1997

The incorporation of mannoproteins in the cell wall of S. cerevisiae and filamentous Ascomycetes.

In yeast, glucanase extractable cell wall proteins are anchored to the plasma membrane at an intermediate stage in their biogenesis via a glycosylphosphatidylinositol (GPI) moiety before they become anchored to the wall glucan via a beta 1,6-glucan linkage. The mechanism of the membrane processing step of cell wall proteins is not known. Here, we report that Ascomycete filamentous fungi involved in food spoilage such as Aspergillus, Paecilomyces and Penicillium, also contain GPI membrane-anchored proteins some of which are processed by an endogenous phospholipase C activity. Furthermore, similar to the situation in yeast, their cell walls contain mannoproteins which are linked to the glucan backbone through a beta 1,6-glucan linkage. Interestingly, one mould which contains a significant amount of non covalently linked beta 1,6-glucosylated cell wall proteins, is much more sensitive towards beta 1,3-glucanases and membrane perturbing peptides than the others.

Ascomycota↗
PubMed1997

The effect of formal feedback sessions on test security for a clinical practice examination using standardized patients.

OBJECTIVE: To determine the effects of formal feedback on test security for a clinical practice examination administered to successive rotations over a year's time. METHOD: A seven case clinical performance examination was administered to ten rotations of students throughout an academic year in a required Ambulatory Care Clerkship. Three of the cases were developed at Saint Louis University School of Medicine (SLU); four of the cases were developed by the National Board of Medical Examiners (NBME). Immediately following each examination, the examinees participated in a feedback session for just the three SLU cases: checklists were displayed, and details of the cases were discussed. No feedback on the four NBME cases was provided. Student performance in successive rotations on the three cases in which students were given detailed feedback by the faculty was compared with performance on the four cases in which no feedback was given. SAMPLE: Data were collected from 119 students. RESULTS: The interaction of case and group was not significant, indicating no difference in the trend over the ten groups between the three cases for which feedback was provided and the four cases for which it was not. The results showed no significant increasing trend over the ten rotation groups for either cases where feedback was given or withheld. CONCLUSION: The results suggest that the formal feedback sessions did not pose a threat to test security.

Journal Article↗
PubMed1997

The relationship between chromosome structure and function at a human telomeric region.

We have sequenced a contiguous 284,495-bp segment of DNA extending from the terminal (TTAGGG)n repeats of the short arm of chromosome 16, providing a full description of the transition from telomeric through subtelomeric DNA to sequences that are unique to the chromosome. To complement and extend analysis of the primary sequence, we have characterized mRNA transcripts, patterns of DNA methylation and DNase I sensitivity. Together with previous data these studies describe in detail the structural and functional organization of a human telomeric region.

Base Sequence↗
PubMed1997

Lymphocytes of human term decidua decrease cell adhesion to a plastic substrate.

Although human decidual lymphocytes have been widely studied, their function and possible interaction with trophoblast are still unclear. Here we show that whereas human early (EDL) and term (TDL) decidual lymphocytes were unable to kill human trophoblast by necrosis (assessed by the 51Cr-release assay) or apoptosis (DNA fragment assay), TDL but not EDL decreased trophoblast adhesion to a plastic substrate as determined by a [3H]thymidine assay. This effect, however, was not selective for trophoblast, as TDL also decreased the adhesion to plastic of human decidual stromal cells and HeLa cells. Our results suggest that TDL may play a role in placental detachment during parturition by decreasing trophoblast or decidual stromal cell adhesion.

Apoptosis↗
PubMed1997

Fluconazole disc diffusion testing for the routine laboratory.

The increasing incidence of resistance to the antifungal agent, fluconazole, prompted the need for a rapid, reliable and easy-to-use susceptibility test. We have developed a disc diffusion test for fluconazole against Candida spp. suitable for a clinical laboratory. Disc diffusion tests on six different media were compared with MIC values. On the basis of correlation coefficient with MICs (r = -0.95), quality of growth and zone edge definition, Yeast Nitrogen Base agar with glucose (YNBG) produced the best results. Further studies on YNBG showed that the method is reliable for Candida albicans and for resistant isolates with no zone of inhibition, but results for the slower growing and uncommon species must be interpreted with some caution. Implementation of this test in the clinical laboratory has provided a much needed therapeutic service for clinicians within the hospital. It has also reduced the reliance on the reference laboratory for susceptibility results and the consequent costs involved.

Candida↗