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Biomedical subjects

A K Wilson

Publications and source records attributed to A K Wilson.

34 records · Page 2Linked to original sources
PubMed1991

An increase or a decrease in myosin II phosphorylation inhibits macrophage motility.

Myosin II purified from mammalian non-muscle cells is phosphorylated on the 20-kD light chain subunit (MLC20) by the Ca2+/calmodulin-dependent enzyme myosin light chain kinase (MLCK). The importance of MLC20 phosphorylation in regulating cell motility was investigated by introducing either antibodies to MLCK (MK-Ab) or a Ca2+/calmodulin-independent, constitutively active form of MLCK (MK-) into macrophages. The effects of these proteins on cell motility were then determined using a quantitative chemotaxis assay. Chemotaxis is significantly diminished in macrophages containing MK-Ab compared to macrophages containing control antibodies. Moreover, there is an inverse relationship between the number of cells that migrate and the amount of MK-Ab introduced into cells. Interestingly, there is also an inverse relationship between the number of cells that migrate and the amount of MK- introduced into cells. Other experiments demonstrated that MK-Ab decreased intracellular MLC20 phosphorylation while MK- increased MLC20 phosphorylation. MK- also increased the amount of myosin associated with the cytoskeleton. These data demonstrate that the regulation of MLCK is an important aspect of cell motility and suggest that MLC20 phosphorylation must be maintained within narrow limits during translational motility by mammalian cells.

Animals↗
PubMed1991

Evaluation of the electroinjection method for introducing proteins into living cells.

The introduction of impermeant probes such as antibodies and other proteins into living cells without compromising physiological function is an important approach for studying cellular regulatory mechanisms. Many techniques including direct microinjection, liposome-mediated delivery, fusion of red cell ghosts, and osmotic lysis of pinocytic vesicles have been used to introduce proteins into intact cells. We have used a modification of the voltage-discharge technique to introduce antibodies and other proteins into living physiologically responsive pheochromocytoma and other cultured cells. In this technique, called electroinjection, a single discharge of relatively low field strength is used to transiently permeabilize the plasma membrane. Our experiments demonstrate that electroinjection permits the introduction of large amounts (microM) of probe into 2-5 x 10(6) cells simultaneously without compromising cell viability or physiological responsiveness when performed under carefully defined conditions. They also demonstrate that electroinjection results in a single population of loaded cells and that protein incorporation is a function of field strength, capacitance, molecular weight of the protein, and the concentration of the protein in the electroinjection buffer. Interestingly, a significant fraction of the protein electroinjected into cells is trapped in the plasma membrane when cells are shocked at high capacitance. These results demonstrate that electroinjection appears to be an efficient method for loading exogenous proteins into cells while maintaining the integrity of the physiological properties of the cell.

Adrenal Gland Neoplasms↗
PubMed1991

Okadaic acid, a phosphatase inhibitor, decreases macrophage motility.

Cellular locomotion results from a series of spatially and temporally integrated reactions. The coordinated regulation of these reactions requires sensitive intracellular signaling mechanisms. Because protein phosphorylation reactions represent important signaling mechanisms in mammalian cells, we investigated the effect of okadaic acid, a phosphoprotein phosphatase inhibitor, on protein phosphorylation and macrophage motility. Okadaic acid was applied to rat alveolar macrophages, and motility was quantitated by a directed chemotaxis assay. Okadaic acid inhibits macrophage motility in a dose-dependent fashion; the concentrations for 50 and 100% inhibition were 3 and 25 microM, respectively. Protein phosphorylation studies demonstrated a 2.5-fold increase in total protein phosphorylation in macrophages treated with 25 microM okadaic acid. These experiments also demonstrated a dose-dependent increase in the phosphorylation of the 20-kDa light chain of myosin. Moreover, 25 microM okadaic acid 1) maximally increased myosin light chain phosphorylation by 6.6-fold, 2) raised the level of myosin associated with the cytoskeleton from a basal level of 47.0 to 96.7% of the total myosin, and 3) induced profound morphological changes as visualized by scanning electron microscopy. These data correlate an increase in protein phosphorylation with a decrease in macrophage motility. Furthermore, they suggest that phosphoprotein phosphatase inhibition may prevent motility by uncoupling coordinated processes, such as cytoskeletal reorganization, that are essential for macrophage motility.

Animals↗
PubMed1990

A dominant mutation in Arabidopsis confers resistance to auxin, ethylene and abscisic acid.

We have screened a large population of M2 seeds of Arabidopsis thaliana for plants which are resistant to exogenously applied indole-acetic acid (IAA). One of the resistant lines identified in this screen carries a dominant mutation which we have named axr2. Linkage analysis indicates that the axr2 gene lies on chromosome 3. Plants carrying the axr2 mutation are severe dwarfs and display defects in growth orientation of both the shoot and root suggesting that the mutation affects some aspect of gravitropic growth. In addition, the roots of axr2 plants lack root hairs. Growth inhibition experiments indicate that the roots of axr2 plants are resistant to ethylene and abscisic acid as well as auxin.

Abscisic Acid↗
PubMed1990

The aux1 Mutation of Arabidopsis Confers Both Auxin and Ethylene Resistance.

Mutagenized populations of Arabidopsis thaliana seedlings were screened for plants capable of root growth on inhibitory concentrations of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid. Four of the mutant lines recovered from this screen display a defect in root gravitropism as well as hormone resistance. The aerial portions of these plants are similar to wild-type in appearance. Genetic analysis of these four mutants demonstrated that hormone resistance segregated as a recessive trait and that all four mutations were alleles of the auxin-resistant mutation aux1 [Maher HP, Martindale SJB (1980) Biochem Genet 18: 1041-1053]. These new mutants have been designated aux1-7, 1-12, 1-15, and 1-19. The sensitivity of wild-type and aux1-7 roots to indole-3-acetic acid, 2,4-dichlorophenoxyacetic acid, and ethylene was determined. The results of these assays show that aux1-7 plants require a 12-fold (indole-3-acetic acid) or 18-fold (2,4-dichlorophenoxyacetic acid) higher concentration of auxin than wild-type for a 50% inhibition of root growth. In addition, ethylene inhibition of root growth in aux1-7 plants is approximately 30% that of wild-type at saturating ethylene concentrations. These results indicate that aux1 plants are resistant to both auxin and ethylene. We have also determined the effect of ethylene treatment on chlorophyll loss and peroxidase activity in the leaves of aux1 and wild-type plants. No difference between mutant and wild-type plants was observed in these experiments, indicating that hormone resistance in aux1 plants may be limited to root growth. Our studies suggest that the AUX1 gene may have a specific function in the hormonal regulation of gravitropism.

Journal Article↗
PubMed1989

Clearance of a persistent human enterovirus infection of the mouse central nervous system by the antiviral agent disoxaril.

Enteroviruses can cause persistent central nervous system (CNS) infections in agammaglobulinemic individuals. Because these infections are rarely cured by passive administration of antibody, a chemotherapeutic approach would be advantageous. In this study, the efficacy of the antienterovirus (and antipicornavirus) drug disoxaril was demonstrated in a murine model of persistent enterovirus infection. Disoxaril is a hydrophobic antiviral compound that blocks picornavirus uncoating. The W-2 strain of human poliovirus type 2 (PV2) persists in the CNS of immunosuppressed mice and causes late paralysis. Mice were inoculated intracerebrally with PV2, immunosuppressed with cyclophosphamide, and treated intragastrically with disoxaril at 50, 100, or 200 mg/kg per day in two divided doses beginning on postinfection day 20. At 200 mg/kg per day, disoxaril significantly decreased the incidence of clinical disease, i.e., paralysis and death. Assays for virus revealed more rapid clearance of virus from the CNS in the drug-treated group. No drug-associated toxicity was observed. Residual isolates of virus were not drug-resistant, suggesting that the appearance of drug resistance during prolonged treatment may not be a clinical problem.

Animals↗
PubMed1989

Factors affecting the placement of preschool-aged deaf children.

The placement in and movement among preschool programs of 131 children with severe and profound hearing losses was monitored over four years. Longitudinal data were obtained on the children's linguistic and social development. In the first year of the study, most of the children were enrolled in auditory/oral (A/O) programs. By the final year, most were enrolled instead in total communication (TC) programs; similar movement occurred from individual to classroom programs. Children in A/O individual programs had the most hearing, highest intelligence and most favorable family profile, followed by children in A/O classes, TC individual programs, and TC classes. Scores on measures of spoken language followed the same pattern. On measures of receptive language and mother-child communication, children in A/O individual programs and those in TC classes had similar scores. Children in TC individual programs came next, and last were children in A/O classes. These findings have implications for the choice of programs for preschool deaf children.

Child, Preschool↗
PubMed1988

An evaluation of recent trends in preschool programming for hearing-impaired children.

A longitudinal study was conducted of 139 children with severe and profound hearing losses. The children were between 3 and 5 years of age at the commencement of the 4-year study. The relationship of several background and educational variables with the linguistic, academic, and social development of the children was investigated. Age, hearing loss, and intelligence were related to most of the dependent measures. Relationships involving communication mode (auditory/oral vs. total communication) varied with the measure being considered. The variable of program type (individual vs. group) interacted with the degree of hearing loss and with communication mode. Although early intervention was related to certain dependent measures, no relationships were found that involved direct instruction by parents or integration.

Achievement↗
PubMed1984

Characterization of genetic changes occurring in attenuated poliovirus 2 during persistent infection in mouse central nervous systems.

Genomic changes occurring in the attenuated W-2 strain of poliovirus 2 during persistent infection of the central nervous system of immunosuppressed mice were analyzed. The RNase T1 oligonucleotide fingerprints of 34 different viruses, isolated from the brains and spinal cords of paralyzed and nonparalyzed mice during a 105-day period, were used to quantitate and compare the mutations occurring in each isolate. Although mice were inoculated with plaque-purified virus, genetically distinct viruses were recovered from the central nervous system. The number of oligonucleotide changes occurring in isolates from paralyzed mice generally was greater than that observed in isolates from nonparalyzed mice. However, differences in the extent of mutation in isolates from the two groups of mice did not appear to be related to the level of virus replication. In paralyzed mice, the number of oligonucleotide changes on average was greater in viruses isolated during the first 60 days of the infection than in the last 45 days. The number of oligonucleotide changes was essentially constant throughout the infection, however, in viruses isolated from the brains of nonparalyzed mice. In addition, several specific oligonucleotide changes were found only in viruses isolated from paralyzed animals.

Animals↗