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Biomedical subjects

J M Robinson

Publications and source records attributed to J M Robinson.

At least 91 records · Page 5Linked to original sources
PubMed1991

A simple method for ultrastructural enzyme cytochemistry on small volumes and numbers of isolated cells.

A simple technique is described for ultrastructural enzyme cytochemistry using small volumes and numbers of single cells. Small aliquots (1-5 microliters) of single cells suspended in a glutaraldehyde fixative are incubated on poly-L-lysine-coated glass coverslips. Cells in contact with the coated coverslip become immobilized to that support. All changes of solutions are accomplished without repeated centrifugation.

5'-Nucleotidase↗
PubMed1991

Use of hearing aids by patients with closed mastoid cavity.

Twenty-five patients who had undergone a closed-cavity tympanomastoidectomy in our Unit and wore a hearing aid in the operated ear were reviewed, and information was recorded on the use of the aid, and the patients' impression about it. The information obtained was analysed and compared with similar data from 39 hearing aid users of similar age with no history of ear surgery. Eighty per cent of the patients with a closed mastoid cavity were satisfied with the aid, and no significant difference was found between the two groups regarding the impression about the aid (chi square 3.06, p = 0.08), or the problems with it, which, in most of the cases, were related to several changes of mould (chi square 2.19, p = 0.13). The various recorded parameters are discussed, and it is concluded that the patients with a closed mastoid cavity can tolerate a hearing aid in the operated ear at least as well as the control subjects with no ear surgery.

Attitude↗
PubMed1991

A novel intracellular compartment with unusual secretory properties in human neutrophils.

Human neutrophils contain a novel intracellular compartment that is distinct from the previously characterized azurophil and specific granules. This compartment is distinguished by the presence of cytochemically detectable alkaline phosphatase activity. The alkaline phosphatase-containing compartments are short rod-shaped organelles that rapidly undergo a dramatic reorganization upon cell stimulation with either a chemoattractant or an active phorbol ester. Biochemical analysis shows that in unstimulated neutrophils the majority of the alkaline phosphatase activity is intracellular, but after stimulation essentially all of this activity becomes associated with the cell surface. The exocytotic pathway is unusual in that these small organelles fuse to form elongated tubular structures before their association with the plasmalemma.

Alkaline Phosphatase↗
PubMed1991

Biochemical and cytochemical studies on enzymes that dephosphorylate inositol (1,4,5)-trisphosphate in neutrophils.

Guinea pig neutrophils contain membrane-bound and soluble phosphatases that catalyze the dephosphorylation of inositol (1,4,5)-trisphosphate [Ins(1,4,5)P3]. The activities were 5.1 +/- 0.2 and 1.3 +/- 0.2 (SD; n = 5) nmoles phosphate (Pi) released/min/10(7) cell equivalents, respectively. The membrane-bound enzyme dephosphorylated many substrates (e.g., beta-glycerophosphate), exhibited alkaline pH optima, and was inhibited by levamisole. In contrast, the soluble phosphatase was specific for Ins(1,4,5)P3, exhibited a neutral pH optimum, and was insensitive to levamisole. A cerium-based ultrastructural cytochemical procedure was employed to identify the subcellular sites of the membrane-bound activity. Staining was observed on the exterior of the plasmalemma and in a population of granules. Staining in the granules was observed only in permeabilized cells. Treatment of neutrophils with p-diazobenzenesulfonate (DBSA) (4.0 mM) for 20 min at 37 degrees C blocked the cytochemical reaction on the cell surface using beta-glycerophosphate as the substrate, but did not affect the staining of the granules on subsequent permeabilization. In biochemical studies, this treatment with DBSA inhibited the membrane-bound activity by c. 50% but did not affect the soluble phosphatase. Therefore, the membrane-bound phosphatase is, in fact, an alkaline phosphatase that resides in locales not accessible to Ins(1,4,5)P3 generated during cell stimulation. Breakdown of Ins(1,4,5)P3 generated during cell stimulation, therefore, would be catalyzed by the soluble enzyme.

Alkaline Phosphatase↗
PubMed1991

Rapid-freezing cytochemistry: preservation of tubular lysosomes and enzyme activity.

We show that tubular structures present in phorbol ester-stimulated macrophages are sensitive to commonly used chemical fixatives (i.e., they usually become fragmented during fixation). These structures are well preserved in macrophages that are physically fixed by rapid-freezing and subsequent freeze-substitution in osmium-acetone. We have developed methods that combine rapid-freezing, freeze-substitution, and enzyme cytochemistry for preservation of these tubular structures and for detection of endocytosed material (i.e., horseradish peroxidase). This method of rapid-freeze cytochemistry may be useful in other situations where chemical fixation does not adequately preserve cell structures, particularly of membrane compartments.

Animals↗
PubMed1990

Utility of staurosporine in uncovering differences in the signal transduction pathways for superoxide production in neutrophils.

Neutrophils exhibit an intense phosphorylation of a 47 kDa protein and release large quantities of superoxide (O2-) upon stimulation with phorbol 12-myristate 13-acetate (PMA) or fMet-Leu-Phe (fMLP). Antagonists of protein kinases (e.g., 200 microM 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7); 15 nM staurosporine) inhibited these phenomena when the stimulus was PMA (Badwey, J.A. et al. (1989) J. Biol. Chem. 264, 14947-14953). In this paper, we now report that while neutrophils treated with 15 nM staurosporine and PMA release little O2-, cells in the presence of these compounds can be stimulated to release near normal quantities of O2- by the subsequent addition of fMLP. Surprisingly, staurosporine (15 nM) reduced the incorporation of 32P into the 47 kDa protein in fMLP stimulated cells at least as effectively as H-7, yet, while the staurosporine treated cells released substantial amounts of O2-, the cells treated with H-7 did not. These data suggest that a stimulatory pathway exists in neutrophils that contains a protein kinase 'distinct' from that which is activated when PMA is the stimulus and that this pathway may enable the O2- producing system to become functional with little or no phosphorylation of the 47 kDa protein. They further suggest that the steps which are sensitive to H-7 in the signal-transduction pathways utilized by PMA and fMLP may be different.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1990

Fluoride-mediated activation of the respiratory burst in electropermeabilized neutrophils.

Electropermeabilization creates small pores in the plasma membrane allowing the introduction of low-molecular-weight modulatory components, such as ions and nucleotides, into the cytosol. The present study investigates fluoride-mediated stimulation of the signal transduction pathway that activates the respiratory burst in electropermeabilized neutrophils. In marked contrast to intact (i.e., non-electropermeabilized) neutrophils, cells permeabilized by this technique demonstrated an immediate and potent stimulation of the superoxide (O2-)-generating NADPH oxidase in response to the addition of fluoride. Furthermore, permeabilization of neutrophils in the presence of exogenously added ATP enhanced the rate of F(-)-mediated O2- production. Fluoride-stimulated O2- production in electropermeabilized neutrophils was antagonized by GDP beta S and dependent upon the presence of Mg2+ in the medium, but was insensitive to pertussis toxin treatment, consistent with the hypothesis that fluoride activates a G protein, probably Gp, by interacting with the nucleotide-binding site on the G alpha subunit. In addition, electropermeabilized neutrophil O2- release triggered by F- was blocked by staurosporine and H-7, indicating that this pathway proceeds largely through protein kinase C activation. However, nucleotide-enhanced O2- production was only partially blocked by these inhibitors, suggesting that under such conditions ATP either competes with the inhibitor-protein kinase interaction or affects the signaling pathway(s) in such a way that protein kinase C may no longer be necessary for the activation of NADPH oxidase.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1990

Mg.ATP primes superoxide-generating responses in electropermeabilized neutrophils.

The present study utilizes an electropermeabilized cell system to determine the effect of Mg.ATP on neutrophil superoxide (O2-)-generating responses stimulated by suboptimal concentrations of fMLP, GTP gamma S and PMA. Permeabilization in the presence of exogenously added Mg.ATP was neither sufficient to initiate O2- release nor necessary for stimulated O2- production. However, the inclusion of Mg.ATP in the permeabilization medium primed the O2(-)-generating responses mediated by suboptimal concentrations of these stimuli. The site of action of Mg.ATP is intracellular. Moreover, the fact that Mg.ATP primes responses stimulated by fMLP, GTP gamma S and PMA suggests that the modulatory effect is at the level of protein kinase C.

Adenosine Triphosphate↗
PubMed1990

Predicting urodynamic dysfunction from clinical features in incontinent elderly women.

In a retrospective survey of 296 elderly women referred with urinary incontinence and in whom urodynamic studies were completed, multiple logistic regression was used to determine the optimal set of clinical features to predict the urodynamic dysfunction. Three formulae were obtained expressing the predicted probabilities (p) of the urodynamic dysfunctions as functions of the clinical variables. By choosing the cut-off values of p which gave equal weighting to sensitivity and specificity, we found that clinical features (symptoms, signs, residual volume) are helpful in diagnosing the presence of unstable detrusor and voiding dysfunction, but not incompetent urethra.

Aged↗
PubMed1990

Nitrite photoreduction in vivo is inhibited by oxygen.

It was hypothesized previously that an O(2) inhibition of NO(2) (-) photoreduction would reflect a competition between O(2) and NO(2) (-) for electrons from ferredoxin at the site of plastid nitrite reductase. In order to test this in vivo, intact spinach (Spinacia oleracea L.) leaf chloroplast and mesophyll cell isolates held in high light were aerated with streams of 20% O(2)/80% N(2) (250 micromolar O(2) in aqueous solution) or, alternatively, streams of 100% N(2). Bicarbonate plus CO(2) and NO(2) (-) were supplied to reaction mixtures at levels just sufficient to promote maximal assimilations of CO(2) and NO(2) (-). In chloroplast isolates, there was a 9 to 30% O(2) inhibition of NO(2) (-) reduction while there were high rates of CO(2) fixation. In spinach and soybean (Glycine max) leaf cell isolates, NO(2) (-) photoreduction rates were 10 to 55% inhibited by O(2) at near ambient levels. It is possible that O(2) may compete, albeit weakly, with NO(2) (-) (nitrite reductase) for equivalents derived from reduced ferredoxin. Also, O(2) may oxidize sulfhydryl groups on nitrite reductase which are involved in substrate binding and/or activation.

Journal Article↗
PubMed1990

Photosynthesis and photosynthate partitioning in n(2)-fixing soybeans.

Leaf area, chlorophyll content, net CO(2) photoassimilation, and the partitioning of fixed carbon between leaf sucrose and starch and soluble protein were examined in Glycine max (L) Merr. cv Williams grown under three different nitrogen regimes. One group (Nod+/+) was inoculated with Bradyrhizobium and watered daily with a nutrient solution containing 6 millimolar NH(4)NO(3). A second set (Nod+/-) was inoculated and had N(2) fixation as its sole source of nitrogen. A third group (Nod(-)) was not inoculated and was watered daily with a nutrient solution containing 6 millimolar NH(4)NO(3). The mean net micromole CO(2) uptake per square decimeter per hour of the most recently matured source leaves was similar among the three groups of plants, being about 310. Mean leaf area of the source leaves, monitored for net photosynthesis was also similar. However, the mean milligram of chlorophyll per square decimeter of Nod+/- test leaves was about 50% lower than the other groups' leaves and indicated nitrogen deficiency. Thus, Nod+/- utilized their chlorophyll more efficiently for photosynthetic CO(2) uptake than the plants of the other treatments. The ratio of foliar carbohydrate:protein content was high in Nod+/- but low in the plants from the other two treatments. This inverse relationship between foliar protein and carbohydrate content suggests that more fixed carbon is diverted to the synthesis of protein when nitrogen availability is high. It was also found that Nod+/- sequestered more storage protein in their paraveinal mesophyll than plants of the other treatments. This study indicates that when inorganic nitrogen regimes are used to control photosynthate partitioning, then both leaf carbohydrate and leaf protein must be considered as end products of carbon assimilate allocation.

Journal Article↗
PubMed1990

Computerized tomography mini-series: an alternative to standard paranasal sinus radiographs.

Computerized tomography (CT) imaging provides detailed information on the paranasal sinuses that is not available from standard radiographs. The planning and safety of endoscopic sinus surgery is greatly improved by CT imaging. Frequently standard radiographs are ordered because of the reduced cost. The four-slice CT Mini-series outlined in this paper provides high resolution bone and soft tissue imaging of the key surgical areas. The Mini-series comprises three precisely placed coronal slices and one axial slice. An average Mini-series takes 20 min to perform at a cost of $NZ66.00. Standard radiographs take up to 30 min to perform at a cost of $NZ38.00.

Endoscopy↗
PubMed1990

Localization of cerium-based reaction products by scanning laser reflectance confocal microscopy.

Scanning laser confocal microscopy was utilized to visualize sites of hydrogen peroxide release from stimulated neutrophils and lysosomal acid phosphatase in these and other cells using cerium in the detection systems. Imaging of the cerium-containing reactions was achieved by employing the reflectance mode of this instrument. Localization of these products at the light microscope level was direct and did not require other reactions to generate a visible product. This new approach to cerium cytochemistry should prove useful for many applications.

Acid Phosphatase↗
PubMed1989

1,2-dioctanoyl-sn-glycerol can stimulate neutrophils by different mechanisms. Evidence for a pathway that does not involve phosphorylation of the 47-kDa protein.

Neutrophils treated with 1,2-dioctanoyl-sn-glycerol (DiC8) are known to release large quantities of superoxide (O2-) and to exhibit an intense phosphorylation of two proteins with molecular masses of approximately 47 and 49 kDa. In this paper, we report that O2- release from guinea pig cells stimulated with a near optimal amount of DiC8 (2.0 microM) is markedly inhibited (greater than or equal to 70%) by antagonists of protein kinase C (i.e. 150 nM staurosporine; 200 microM 1-(5-isoquinolinylsulfonyl)-2-methyl-piperazine (H-7], whereas that from cells stimulated with an optimal amount of DiC8 (7.8 microM) is not (approximately 25% inhibition). However, staurosporine (150 nM) effectively reduced the level of phosphorylation of the 47- and the 49-kDa proteins to that observed in unstimulated cells when either amount of DiC8 (i.e. 2.0 or 7.8 microM) was utilized. Thus, neutrophils stimulated with 7.8 microM DiC8 in the presence of staurosporine release large quantities of O2- without an enhanced phosphorylation of the 47- and the 49-kDa proteins. In contrast, these antagonists of protein kinase C effectively blocked O2- release from neutrophils stimulated with an optimal amount of phorbol 12-myristate 13-acetate (PMA), and the percentage of inhibition was not affected by increasing the concentration of PMA 160-fold. These data show that DiC8 and PMA, both activators of protein kinase C, can have distinct effects on O2- release by neutrophils. Moreover, they suggest that DiC8 (or a metabolite) under certain circumstances may function in a stimulatory pathway for O2- release that is independent of protein kinase C. Differences in the morphology of neutrophils stimulated with PMA and DiC8 are presented. Ancillary data on human neutrophils are also provided.

Alkaloids↗
PubMed1989

Paradoxical effects of retinal in neutrophil stimulation.

Retinal stimulates the activity of phospholipase C and superoxide (O2-) release in neutrophils. The latter response is comparable in magnitude to that observed when phorbol 12-myristate 13-acetate (PMA) is the stimulating agent. Cells treated with retinal, however, do not undergo degranulation, nor do they exhibit the formation of intracellular vesicles, as is commonly observed with other agents (e.g. Lochner, J. E., Badwey, J. A., Horn, W., and Karnovsky, M. L. (1986) Proc. Natl. Acad. Sci. U. S. A. 83, 7673-7677). Retinal promotes redistribution of the activity of protein kinase C from a soluble to a particulate fraction in neutrophils, and this redistribution precedes O2- release. Superoxide release stimulated with retinal, however, is largely insensitive to inhibitors of protein kinase C (1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7); staurosporine). These compounds substantially block both O2- release and the phosphorylation of two proteins with molecular masses of about 47 and 49 kDa when the stimulus is PMA. The data indicate that retinal and PMA elicit the formation of active protein kinase C complexes of different natures, or that the mechanism of stimulation of O2- release by retinal does not involve this kinase. The significance of these observations to the common use of retinoids as inhibitors of protein kinase C is discussed.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1989

Bone pâté repair of the eroded incus.

The authors report on a series of 25 patients who, in the process of undergoing tympanoplasty, were noted to have erosion of the long process of the incus. This was repaired with bone pâté in an attempt to restore ossicular continuity. The methods and results are presented and the prospects for the future are discussed.

Ear Diseases↗